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Veterinary Laboratory Manual

9th edition
Veterinary Laboratory Service
Edition Year Under the Title

1st edition 1945 Instructions for the Collection and Despatch of Material for Pathological and
Bacteriological Examination.
2nd edition 1947 Instructions for the Collection and Despatch of Material for Pathological and
Bacteriological Examination.
3rd edition 1950 Instructions for the Collection and Despatch of Material for Laboratory
Examination
4th edition 1961 Collection and Despatch of Material for Laboratory Examination

5th edition 1974 Collection and Despatch of Material for Laboratory Examination

6th edition 1981 Collection of Material for Laboratory Examination

7th edition 1985 Collection of Material for Laboratory Examination

8th edition 1995 Laboratory Specimen Submission Manual

9th edition 2006 Veterinary Laboratory Manual

Prepared by staff of Elizabeth Macarthur Agricultural Institute, Menangle


and Regional Veterinary Laboratories, Orange and Wollongbar

ISBN 0 7305 6656 0

VETERINARY LABORATORY MANUAL II


INTRODUCTION
This 9th edition of the Manual is published for the first time on the Web. Its aim is to provide
accessible, up-to-date information to laboratory clients.

The general format of the previous edition, edited by Graeme Eamens, has been retained.

Many laboratory staff have contributed to this revised Manual. Also, field veterinarians have provided
suggestions, which have been incorporated.

The Manual includes information about specimen collection and submission in general, as well as for
specific diseases.

For further information about tests for diseases of commercial livestock, please contact your nearest
Regional Veterinary Laboratory or Customer Service at 1800 675 623.

VETERINARY LABORATORY MANUAL III


TABLE OF CONTENTS
INTRODUCTION .....................................................................................................................................iii
VETERINARY LABORATORY DIRECTORY ....................................................................................... ix
SPECIMENS (GENERAL) ...................................................................................................................... 1
CONDITIONS FOR ACCEPTANCE ................................................................................................................ 1
LABORATORY CHARGES.............................................................................................................................. 1
SPECIMEN SUBMISSION FORM ................................................................................................................... 1
COLLECTION AND LABELLING OF SPECIMENS ......................................................................................... 3
PACKING OF SPECIMENS............................................................................................................................. 4
TRANSPORT OF SPECIMENS....................................................................................................................... 5
DISEASE SURVEILLANCE ............................................................................................................................. 6
SPECIAL REQUIREMENTS FOR TESTING OF STOCK FOR OVERSEAS EXPORT................................... 7
SPECIAL REQUIREMENTS FOR TESTING OF STOCK FOR INTERSTATE MOVEMENT .......................... 8
CHECKLIST OF EQUIPMENT FOR CLINICAL AND NECROPSY EXAMINATIONS ..................................... 9
SPECIMENS (BY DISCIPLINE)............................................................................................................ 11
BACTERIOLOGY .............................................................................................................................................. 11
STANDARD CONTAINERS AND EQUIPMENT ............................................................................................ 11
COLLECTION OF SPECIMENS .................................................................................................................... 11
STORAGE OF BACTERIOLOGICAL SPECIMENS PRIOR TO DESPATCH ................................................ 13
DESPATCH OF SPECIMENS ....................................................................................................................... 14
BIOCHEMISTRY ............................................................................................................................................... 14
STANDARD CONTAINERS AND EQUIPMENT ............................................................................................ 14
COLLECTION OF SPECIMENS .................................................................................................................... 14
STORAGE OF SPECIMENS PRIOR TO DESPATCH................................................................................... 15
DESPATCH OF SPECIMENS ....................................................................................................................... 15
GENETICS .................................................................................................................................................... 15
STANDARD CONTAINERS AND EQUIPMENT ............................................................................................ 15
COLLECTION OF SPECIMENS .................................................................................................................... 15
STORAGE OF SPECIMENS AND DESPATCH OF SPECIMENS ................................................................ 16
GROSS PATHOLOGY ...................................................................................................................................... 16
STANDARD CONTAINERS AND EQUIPMENT ............................................................................................ 16
COLLECTION OF SPECIMENS FOR GROSS PATHOLOGY ...................................................................... 16
STORAGE OF SPECIMENS PRIOR TO DESPATCH................................................................................... 16
HISTOPATHOLOGY ......................................................................................................................................... 16
STANDARD CONTAINERS AND EQUIPMENT ............................................................................................ 16
COLLECTION OF SPECIMENS .................................................................................................................... 17
HAEMATOLOGY............................................................................................................................................... 18
BLOOD .......................................................................................................................................................... 18
BLOOD FILMS............................................................................................................................................... 18
PREPARATION OF BLOOD FILMS .............................................................................................................. 18
STORAGE AND DESPATCH OF SPECIMENS ............................................................................................ 18
PARASITOLOGY .............................................................................................................................................. 18
STANDARD CONTAINERS AND EQUIPMENT ............................................................................................ 18
COLLECTION OF SPECIMENS .................................................................................................................... 19
STORAGE AND DESPATCH OF SPECIMENS ............................................................................................ 19
SEROLOGY....................................................................................................................................................... 19
STANDARD CONTAINERS AND EQUIPMENT ............................................................................................ 20
COLLECTION OF SPECIMENS .................................................................................................................... 20
STORAGE AND DESPATCH OF SPECIMENS ............................................................................................ 21
TYPES OF SEROLOGICAL TESTS .............................................................................................................. 21
TOXICOLOGY ................................................................................................................................................... 24
STANDARD CONTAINERS AND EQUIPMENT ............................................................................................ 24
COLLECTION OF SPECIMENS .................................................................................................................... 24
STORAGE OF SPECIMENS PRIOR TO DESPATCH................................................................................... 24
VIROLOGY ........................................................................................................................................................ 25
DIAGNOSIS OF VIRAL DISEASE ................................................................................................................. 25
STANDARD CONTAINERS AND EQUIPMENT ............................................................................................ 27
COLLECTION OF SPECIMENS .................................................................................................................... 27
STORAGE AND DESPATCH OF SPECIMENS ............................................................................................ 28
SPECIMENS (BY DISEASE OR SYNDROME).................................................................................... 31
DISEASES OF LIVESTOCK ............................................................................................................................. 31
ABORTION (GENERAL)................................................................................................................................ 31
ABORTION IN CATTLE................................................................................................................................. 31
ABORTION IN SHEEP AND GOATS ............................................................................................................ 31
ABORTION IN HORSES ............................................................................................................................... 32

VETERINARY LABORATORY MANUAL IV


ABORTION IN PIGS ...................................................................................................................................... 32
ACETONAEMIA............................................................................................................................................. 32
ACTINOBACILLOSIS AND ACTINOMYCOSIS ............................................................................................. 32
ACTINOBACILLUS SEMINIS INFECTION IN RAMS .................................................................................... 32
AFLATOXICOSIS .......................................................................................................................................... 33
AKABANE DISEASE IN SHEEP AND CATTLE............................................................................................. 33
ALGAL POISONING ...................................................................................................................................... 33
ALPHA MANNOSIDOSIS OF CATTLE.......................................................................................................... 33
ANAPLASMOSIS........................................................................................................................................... 33
ANAEMIA....................................................................................................................................................... 33
ANNUAL RYEGRASS TOXICOSIS (ART) .................................................................................................... 33
ANTHELMINTIC RESISTANCE..................................................................................................................... 34
ANTHRAX...................................................................................................................................................... 36
ARSENIC POISONING.................................................................................................................................. 36
ARTHRITIS AND POLYARTHRITIS .............................................................................................................. 37
ARTHROGRYPOSIS AND HYDRANENCEPHALY....................................................................................... 37
ASPERGILLOSIS .......................................................................................................................................... 37
ATAXIA .......................................................................................................................................................... 37
ATROPHIC RHINITIS OF SWINE ................................................................................................................. 38
BABESIOSIS ................................................................................................................................................. 38
BALANO POSTHITIS IN RAMS..................................................................................................................... 38
BENIGN FOOTROT....................................................................................................................................... 38
BETA MANNOSIDOSIS OF GOATS ............................................................................................................. 38
'BIG KNEE' IN GOATS .................................................................................................................................. 38
BLACK DISEASE........................................................................................................................................... 38
BLACKLEG .................................................................................................................................................... 38
BLOAT ........................................................................................................................................................... 38
BLUE-GREEN ALGAL POISONING.............................................................................................................. 39
BORDER DISEASE IN SHEEP ..................................................................................................................... 39
BOTULISM .................................................................................................................................................... 39
BOVINE LEUKOSIS ...................................................................................................................................... 39
BOVINE LEUCOCYTE ADHESION DEFICIENCY (BLAD) ........................................................................... 39
BOVINE MALIGNANT CATARRH ................................................................................................................. 39
BOVINE VIRUS DIARRHOEA ....................................................................................................................... 40
BRACKEN FERN POISONING...................................................................................................................... 40
BRUCELLOSIS BOVINE ............................................................................................................................... 40
BRUCELLOSIS OVINE.................................................................................................................................. 40
BRUCELLOSIS PORCINE ............................................................................................................................ 40
CALCULI........................................................................................................................................................ 41
CAMPYLOBACTER ABORTION OF SHEEP ................................................................................................ 41
CAMPYLOBACTER ENTERITIS ................................................................................................................... 41
CAMPYLOBACTERIOSIS OF CATTLE......................................................................................................... 41
CAMPYLOBACTERIOSIS OF PIGS.............................................................................................................. 42
CAPRINE ARTHRITIS ENCEPHALITIS (CAE).............................................................................................. 42
CARDIOMYOPATHY AND WOOLLY HAIRCOAT (CWH) SYNDROME ....................................................... 42
CASEOUS LYMPHADENITIS (CLA) ............................................................................................................. 42
CHLAMYDIAL INFECTIONS ......................................................................................................................... 43
CITRULLINAEMIA ......................................................................................................................................... 43
CLOVER DISEASE........................................................................................................................................ 43
COBALT DEFICIENCY .................................................................................................................................. 43
COCCIDIOSIS ............................................................................................................................................... 44
COCCIDIOSIS – HEPATIC IN RABBITS....................................................................................................... 44
COLIBACILLOSIS.......................................................................................................................................... 44
COMPLEX VETERBRAL MALFORMATION (CVM) ...................................................................................... 44
CONGENITAL ABNORMALITIES.................................................................................................................. 45
CONTAGIOUS EQUINE METRITIS (CEM) ................................................................................................... 45
CONTAGIOUS OPHTHALMIA....................................................................................................................... 45
CONTAGIOUS PUSTULAR DERMATITIS .................................................................................................... 45
COPPER DEFICIENCY ................................................................................................................................. 45
COPPER POISONING OF SHEEP AND CATTLE ........................................................................................ 46
CORYNEBACTERIUM EQUI IN HORSES OR PIGS .................................................................................... 46
CRYPTOSPORIDIOSIS................................................................................................................................. 46
CYANIDE POISONING.................................................................................................................................. 47
CYSTICERCOSIS.......................................................................................................................................... 47
DEFICIENCY OF URIDINE MONOPHOSPHATE SYNTHETASE (DUMPS) ................................................ 47
DERMATOPHILOSIS .................................................................................................................................... 47
DIARRHOEA.................................................................................................................................................. 47
DRENCHING MORTALITIES ........................................................................................................................ 48
DWARFISM IN DEXTER CATTLE................................................................................................................. 48
ENCEPHALOMYELITIS AND ENCEPHALITIS ............................................................................................. 48

VETERINARY LABORATORY MANUAL V


ENCEPHALOMYOCARDITIS (EMC) OF PIGS ............................................................................................. 48
ENTEROTOXAEMIA ..................................................................................................................................... 48
ENZOOTIC ATAXIA....................................................................................................................................... 49
ENZOOTIC BOVINE LEUKOSIS (EBL)......................................................................................................... 49
ENZOOTIC HAEMATURIA OF CATTLE ....................................................................................................... 49
ENZOOTIC PNEUMONIA OF PIGS .............................................................................................................. 49
EPERYTHROZOONOSIS OF SHEEP........................................................................................................... 50
EPHEMERAL FEVER.................................................................................................................................... 50
EPIDIDYMITIS ............................................................................................................................................... 50
EQUINE BABESIOSIS................................................................................................................................... 50
EQUINE INFECTIOUS ANAEMIA (EIA) ........................................................................................................ 50
EQUINE VIRAL ARTERITIS (EVA)................................................................................................................ 50
ERYSIPELAS................................................................................................................................................. 51
EXOTIC DISEASES....................................................................................................................................... 51
EXUDATIVE EPIDERMITIS IN PIGS............................................................................................................. 51
FACIAL ECZEMA .......................................................................................................................................... 52
FACTOR VIII DEFICIENCY ........................................................................................................................... 52
FACTOR XI DEFICIENCY ............................................................................................................................. 52
FASCIOLIASIS IN SHEEP AND CATTLE ..................................................................................................... 52
FISTULOUS WITHERS ................................................................................................................................. 52
FLEECE ROT ................................................................................................................................................ 52
FOCAL SYMMETRICAL ENCEPHALOMALACIA (FSE) ............................................................................... 53
FOOT ABSCESS IN SHEEP ......................................................................................................................... 53
FOOTROT IN SHEEP.................................................................................................................................... 53
FOOTROT IN GOATS AND CATTLE ............................................................................................................ 56
FUNGAL INFECTIONS (OTHER THAN MYCOTOXICOSES)....................................................................... 56
GENERALIZED GLYCOGENOSIS................................................................................................................ 57
GENETIC DISEASES .................................................................................................................................... 57
GOITRE ......................................................................................................................................................... 58
GRAIN POISONING ...................................................................................................................................... 58
GRASS TETANY ........................................................................................................................................... 58
HAEMAGGLUTINATING ENCEPHALOMYELITIS VIRUS (HEV) INFECTION OF PIGS.............................. 59
HAEMATURIA ............................................................................................................................................... 59
HAEMOGLOBINURIA.................................................................................................................................... 59
HAEMOPHILIA .............................................................................................................................................. 59
HEEL ABSCESS - OVINE ............................................................................................................................. 59
HELIOTROPE POISONING .......................................................................................................................... 59
HENDRAVIRUS INFECTION......................................................................................................................... 59
HEPATOSIS DIETETICA............................................................................................................................... 60
HERPESVIRUS VULVOVAGINITIS/BALAN0POSTHITIS IN GOATS........................................................... 60
HISTOPHILUS OVIS/ HAEMOPHILUS SOMNUS INFECTIONS .................................................................. 60
HYPOCALCAEMIA ........................................................................................................................................ 60
HYPOGLYCAEMIA........................................................................................................................................ 61
HYPOMAGNESAEMIA .................................................................................................................................. 61
HYPOPHOSPHATAEMIA.............................................................................................................................. 61
ILL THRIFT .................................................................................................................................................... 61
INFECTIOUS BOVINE RHINOTRACHEITIS (IBR) AND INFECTIOUS PUSTULAR VULVO
VAGINITIS (IPV) ............................................................................................................................................ 62
INFERTILITY IN CATTLE, SHEEP, PIGS ..................................................................................................... 62
INHERITED DEFECTS.................................................................................................................................. 62
INHERITED CONGENITAL MYOCLONUS ................................................................................................... 62
IRON TOXICITY SYNDROME IN PIGLETS .................................................................................................. 63
ITCHMITE IN SHEEP .................................................................................................................................... 63
JAUNDICE ..................................................................................................................................................... 63
JOHNE'S DISEASE ....................................................................................................................................... 63
KIKUYU POISONING .................................................................................................................................... 66
LEAD POISONING ........................................................................................................................................ 66
LEPTOSPIROSIS .......................................................................................................................................... 66
LICE RESISTANCE TO INSECTICIDES IN SHEEP ..................................................................................... 68
LISTERIOSIS................................................................................................................................................. 68
LIVER FLUKE INFECTION............................................................................................................................ 68
LUPINOSIS.................................................................................................................................................... 69
LYME DISEASE............................................................................................................................................. 69
LYSSAVIRUS ................................................................................................................................................ 70
MALIGNANT OEDEMA ................................................................................................................................. 70
MALNUTRITION ............................................................................................................................................ 70
MANGE.......................................................................................................................................................... 70
MANNOSIDOSIS ........................................................................................................................................... 71
ALPHA-MANNOSIDOSIS .............................................................................................................................. 71
BETA-MANNOSIDOSIS ................................................................................................................................ 71

VETERINARY LABORATORY MANUAL VI


MAPLE SYRUP URINE DISEASE (MSUD) ................................................................................................... 71
MASTITIS (BOVINE) ..................................................................................................................................... 72
MASTITIS (CAPRINE) ................................................................................................................................... 73
MASTITIS (OVINE)........................................................................................................................................ 73
MASTITIS METRITIS AGALACTIA SYNDROME IN SOWS.......................................................................... 73
MENANGLE VIRUS INFECTION................................................................................................................... 73
METABOLIC DISEASES ............................................................................................................................... 74
MUCOSAL DISEASE..................................................................................................................................... 74
MULBERRY HEART DISEASE ..................................................................................................................... 74
MUSCULAR DEGENERATION, NUTRITIONAL ........................................................................................... 74
MYCOPLASMOSIS ....................................................................................................................................... 75
MYCOTIC DERMATITIS................................................................................................................................ 76
MYCOTOXICOSIS......................................................................................................................................... 76
NASAL GRANULOMA OF CATTLE .............................................................................................................. 76
NECROBACILLOSIS ..................................................................................................................................... 76
NEOPLASMS................................................................................................................................................. 77
NEOSPOROSIS ............................................................................................................................................ 77
NERVOUS DISORDERS ............................................................................................................................... 77
NITRATE-NITRITE POISONING ................................................................................................................... 78
OEDEMA DISEASE OF PIGS ....................................................................................................................... 78
OPHTHALMIA................................................................................................................................................ 78
ORGANOCHLORINE AND ORGANOPHOSPHATE POISONING................................................................ 78
OSTEOCHONDROSIS IN PIGS .................................................................................................................... 79
OSTEOMALACIA, OSTEOPOROSIS............................................................................................................ 79
OSTERTAGIOSIS.......................................................................................................................................... 79
OVINE BRUCELLOSIS.................................................................................................................................. 79
OXALATE POISONING ................................................................................................................................. 79
PAPULAR STOMATITIS OF CALVES........................................................................................................... 79
PARAKERATOSIS OF SWINE ...................................................................................................................... 79
PARAMPHISTOMIASIS................................................................................................................................. 80
PARASITES (EXTERNAL) ............................................................................................................................ 80
PARASITES (INTERNAL).............................................................................................................................. 80
PARVOVIRUS INFECTION IN PIGS ............................................................................................................. 84
PASTEURELLOSIS ....................................................................................................................................... 85
PEPSINOGEN ESTIMATIONS FROM SERUM OR PLASMA....................................................................... 85
PERINATAL MORTALITIES IN CATTLE....................................................................................................... 85
PERINATAL MORTALITIES IN SHEEP AND GOATS .................................................................................. 85
PESTIVIRUS INFECTION ............................................................................................................................. 86
PHALARIS POISONING................................................................................................................................ 87
PHOSPHORUS DEFICIENCY....................................................................................................................... 87
PHOTOSENSITIZATION ............................................................................................................................... 87
PIGLET ANAEMIA ......................................................................................................................................... 87
PINKEYE ....................................................................................................................................................... 87
PLANT POISONING ...................................................................................................................................... 87
PNEUMONIA ................................................................................................................................................. 87
POISONING (CHEMICAL)............................................................................................................................. 88
POISONING (PLANT).................................................................................................................................... 88
IDENTIFICATION OF SUSPECT POISONOUS PLANTS ............................................................................. 88
POLIOENCEPHALOMALACIA (PEM) ........................................................................................................... 89
POLYARTHRITIS .......................................................................................................................................... 89
POMPE'S DISEASE ...................................................................................................................................... 89
PORCINE COLITIS........................................................................................................................................ 89
PORCINE ENTEROVIRUS ENCEPHALOMYELITIS .................................................................................... 91
PORCINE ENZOOTIC PNEUMONIA ............................................................................................................ 91
PORCINE PLEUROPNEUMONIA ................................................................................................................. 91
PORCINE INTESTINAL HAEMORRHAGE SYNDROME.............................................................................. 91
PORCINE MYOCARDITIS (PMC) ................................................................................................................. 92
PORCINE PROLIFERATIVE ENTEROPATHY.............................................................................................. 92
PORCINE STRESS SYNDROME (PSS) ....................................................................................................... 92
PREGNANCY TOXAEMIA............................................................................................................................. 93
PSEUDOCOWPOX ....................................................................................................................................... 93
PROTOPORPHYRIA ..................................................................................................................................... 93
PYELONEPHRITIS (BOVINE) ....................................................................................................................... 93
PYOGENIC INFECTIONS ............................................................................................................................. 93
PYRROLIZIDINE ALKALOIDOSIS ................................................................................................................ 93
Q FEVER ....................................................................................................................................................... 94
RED GUT IN SHEEP ..................................................................................................................................... 94
RHODOCOCCUS EQUI PNEUMONIA IN HORSES ..................................................................................... 94
RHODOCOCCUS EQUI LYMPHADENITIS IN PIGS .................................................................................... 94
RINGWORM (DERMATOMYCOSIS) ............................................................................................................ 94

VETERINARY LABORATORY MANUAL VII


ROCK FERN POISONING ............................................................................................................................ 94
ROTAVIRUS INFECTION.............................................................................................................................. 95
RYEGRASS STAGGERS .............................................................................................................................. 95
SALMONELLOSIS......................................................................................................................................... 95
SALT POISONING IN PIGS .......................................................................................................................... 95
SARCOSPORIDIOSIS................................................................................................................................... 95
SCABBY MOUTH .......................................................................................................................................... 95
SCOURING.................................................................................................................................................... 96
SELENIUM DEFICIENCY .............................................................................................................................. 96
SEMEN EXAMINATION ................................................................................................................................ 96
STRYCHNINE POISONING .......................................................................................................................... 98
SWAINSONA POISONING............................................................................................................................ 98
SWINE DYSENTERY .................................................................................................................................... 98
TAPEWORM INFESTATION IN SHEEP ....................................................................................................... 99
TEAT LESIONS ............................................................................................................................................. 99
TETANUS ...................................................................................................................................................... 99
TICK FEVER OF CATTLE ............................................................................................................................. 99
TOXAEMIC JAUNDICE ............................................................................................................................... 100
TOXOPLASMOSIS IN CATS ....................................................................................................................... 100
TOXOPLASMOSIS IN SHEEP AND GOATS .............................................................................................. 100
TOXOPLASMOSIS IN OTHER ANIMALS ................................................................................................... 100
TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHY (TSE).................................................................. 101
TRICHOMONIASIS OF CATTLE ................................................................................................................. 101
TUBERCULOSIS ......................................................................................................................................... 101
ULCERATIVE SPIROCHAETOSIS IN PIGS................................................................................................ 102
ULCERS, GASTRIC IN PIGS ...................................................................................................................... 102
UREA POISONING...................................................................................................................................... 102
URINE EXAMINATION ................................................................................................................................ 102
DISEASES OF POULTRY............................................................................................................................... 105
STANDARD CONTAINERS AND EQUIPMENT .......................................................................................... 105
COLLECTION OF SPECIMENS .................................................................................................................. 105
STORAGE AND DESPATCH OF SPECIMENS .......................................................................................... 105
SEROLOGICAL TESTS AVAILABLE FOR POULTRY ................................................................................ 105
AVIAN ENCEPHALOMYELITIS (AE)........................................................................................................... 106
BIG LIVER AND SPLEEN DISEASE ........................................................................................................... 106
LEUKOSIS ................................................................................................................................................... 107
MAREK'S DISEASE..................................................................................................................................... 107
MYCOPLASMOSIS (AVIAN) ....................................................................................................................... 107
PASTEURELLOSIS (AVIAN) ....................................................................................................................... 108
PULLORUM DISEASE ................................................................................................................................ 108
RETICULOENDOTHELIOSIS (RE) ............................................................................................................. 108
TUBERCULOSIS (AVIAN) ........................................................................................................................... 108
DISEASES OF CAGE BIRDS, AVIARY BIRDS AND RACING PIGEONS..................................................... 109
STANDARD CONTAINERS AND EQUIPMENT .......................................................................................... 109
COLLECTION OF SPECIMENS .................................................................................................................. 109
STORAGE AND DESPATCH OF SPECIMENS .......................................................................................... 109
DISEASES OF BEES ...................................................................................................................................... 110
DISEASES OF FISH........................................................................................................................................ 111
STANDARD CONTAINERS AND EQUIPMENT .......................................................................................... 111
HISTORY AND DIAGNOSIS........................................................................................................................ 111
COLLECTION, STORAGE AND DESPATCH OF SPECIMENS.................................................................. 112
SOME DISEASES OF FISH, CRUSTACEANS AND SHELLFISH ALREADY ENCOUNTERED
IN NSW........................................................................................................................................................ 113
FEEDS AND PASTURE ANALYSIS .................................................................................................. 114
WATER TESTING............................................................................................................................... 116

VETERINARY LABORATORY MANUAL VIII


VETERINARY LABORATORY DIRECTORY
Phone ‘Customer Service’ at 1800 675 623 to contact any of the following veterinary laboratories

Location Address Contacts


Menangle Regional Veterinary Laboratory Ph: 02 4640 6327
Elizabeth Macarthur Agricultural Institute Fax: 02 4640 6400
‘Camden Park’
Woodbridge Road
Menangle NSW 2568

Postal address: (PMB 8, Camden 2570)

Officer in Charge: Keith Walker


Pathologists: Steven Hum
Rod Reece
Richard Luong

Elizabeth Macarthur Agricultural Institute Ph: 02 4640 6333


(Main Reception) Fax: 02 4640 6300

Orange Regional Veterinary Laboratory Ph: 02 6391 3858


Agricultural Research and Veterinary Centre Fax: 02 6391 3899
Forest Road
Orange NSW 2800

Officer in Charge: Graham Bailey


Pathologists: Patrick Staples
Erica Bunker
Wollongbar Regional Veterinary Laboratory Ph: 02 6626 1103
Wollongbar Agricultural Institute Fax: 02 6626 1276
1243 Bruxner Highway
Wollongbar NSW 2477

Officer in Charge: Graeme Fraser


Pathologists: John Boulton
Paul Gill

VETERINARY LABORATORY MANUAL IX


SPECIMENS (GENERAL)
All NSW Department of Primary Industries specimens are unlikely to be admissible as
Veterinary Laboratories operate according to evidence in criminal prosecutions and civil
ISO 17025, and are accredited by the National actions.
Association of Testing Authorities (NATA).
Details on the scope of this accreditation can INFORMATION ANALYSIS
be found on the NATA website at Test results and findings may be provided
http://www.nata.asn.au. to other authorised staff and used for
statistical, surveillance, extension,
CONDITIONS FOR ACCEPTANCE certification and regulatory purposes in
accordance with Department policies. The
The Veterinary Laboratories examine information assists disease and residue
specimens from all livestock, poultry, native control programs and underpins marked
and feral animals and birds, fish and bees. access for agricultural products. The source
In relation to pets, cage birds, racing dogs of the information will remain confidential
and horses, it is policy to examine only unless otherwise required by law or
specimens where the health of the flock, regulatory policies.
herd, aviary etc., is involved or where public
health is likely to be involved or for import
LABORATORY CHARGES
and export purposes.
NSW Department of Primary Industries
Specimens must be accompanied by a operates the Veterinary Laboratories on a
Specimen Submission Form signed by the cost recovery basis. Submitters will
submitter. The submitter thereby accepts normally be invoiced soon after the final
responsibility for payment of laboratory report is issued. The laboratory fees for
charges for the tests requested. Where the most common tests can be downloaded
submitter expects the work to be funded from
(entirely or partially) by NSW Department of http://www.dpi.nsw.gov.au/agriculture/vetm
Primary Industries, the reason for this anual/submission/lab-charges#fees.
request and the source of funds (where Laboratory fees for tests not listed can be
known) should be clearly and obtained by calling Customer Service at
conspicuously stated. 1800 675 623.

Specimens are generally only accepted on Laboratory charges are subsidised for
referral from a veterinarian. Exceptions testing in the following circumstances*:
include WormTest kits, which can be • Notifiable disease
submitted by primary producers. The • Emergency (exotic) disease
Genetics section, at EMAI also accepts • National TSE Surveillance Program
bovine hair samples collected by cattle • Mortality investigations
owners for genetic disease testing where
certification is not required. *See ‘Disease Surveillance’ below for more
details.
CASES LIKELY TO INVOLVE CRIMINAL
INVESTIGATIONS Because it is often difficult for submitters to
Examination is not undertaken on specify the precise tests that will be
specimens likely to involve criminal required, many find it useful to indicate a
prosecutions (e.g. in cases of suspected maximum laboratory fee on the Specimen
malicious poisoning). If a case is likely to Advice Form. The Pathologist will consult
involve police matters or matters controlled with the submitter before exceeding this
by other regulatory authorities, those bodies amount.
should be contacted by phone and
consulted before any action is taken. Often SPECIMEN SUBMISSION FORM
they have special procedures to be followed
with respect to sampling/examination, and Field veterinarians should supply all
with respect to sealing, custody, care and pertinent details of the history, clinical signs
transport of exhibits. They may also have and necropsy findings. This information will
analyses performed by the Government allow the laboratory veterinarian interpret
Analytical Laboratories. the laboratory results and suggest
additional testing as appropriate. All
Laboratory Reports on samples submitted information supplied will be included in the
by field veterinarians as routine diagnostic Laboratory Report.

VETERINARY LABORATORY MANUAL 1


contact between the laboratory and the
NSW Department of Primary Industries' veterinarian. Provide your email address if
Specimen Submission Form facilitates the you would prefer to receive your results by
orderly collection of this information. email.

Specimen Submission Forms are supplied DISEASE SUSPECTED


in pads of 100 pages. Alternatively, the For Diagnostic cases, give the first two
Form can be downloaded for printing from disease possibilities you suspect.
the NSW Department of Primary Industries
web site at SPECIMENS SUBMITTED
http://www.agric.nsw.gov.au/reader/das- Give a full list of the specimens submitted
vettesting. together with the examinations requested
on each. The number of containers, swabs
An official Specimen Submission Form and blood samples, should be clearly listed
should be used with all specimens so that the laboratory staff can check to be
submitted. sure that all the specimens submitted have
been received. Use the Specimen
Special forms may also be available for Submission Key List in addition to the
special investigations (eg footrot, ovine or Sample Submission Form if there are
bovine Johnes disease,). These forms insufficient lines on the latter.
provide a check list for information required
by the laboratory in these investigations. TESTS REQUESTED
Be as specific as possible, e.g. Brucella
Specimen Submission forms should be CFT and ELISA. This is especially
completed using black or blue ballpoint important for regulatory/ movement/ export
pens, as this ensures good photocopies. testing. It should be clear which test should
Be as concise as possible. Long narratives be applied to each sample. Your request is
waste your time and ours. The space equivalent to an order for services and you
available on the form indicates the level of will be charged for all tests you have
detail required. For more complex requested. If you wish the laboratory to hold
investigations, a second sheet may be a specimen for possible testing later, write
attached. Such detailed advice is best 'HOLD' beside this specimen.
emailed to the laboratory for inclusion in the
laboratory report. REASON FOR TEST
Tick the most appropriate checkbox.
The following provides guidance on
completing each section of the form: SPECIES
Include the species, breed, age and sex of
OWNER affected animals.
Provide the normal trading name and full
address, including the property name of the For species, select from the following:
owners, e.g. A. Brown, A. Brown & Sons, cattle, sheep, pigs, poultry, goats, horses,
Mia Mia Pastoral Co. fish, bees, deer. If others, state clearly.

Please print clearly. The same name must For breed, give breed or predominant breed
be used in all specimen submissions from a in crosses.
property to allow previous submissions from
the property to be traced. For sex, select from the following: male,
female, castrate, spayed, mixed group,
RURAL LANDS PROTECTION BOARD unknown.
and RLPB Property ID
The affected property's RLPB (Rural Lands NUMBER OF ANIMALS
Protection Board) district and its RLPB We use the numbers provided to calculate
property ID is required for statistical and the epidemiological statistics, 'period
reporting purposes. The Property ID is the prevalence' and 'cumulative mortality'. For
Property Identification Code (PIC), tail tag the statistics to have meaning, the following
number, or Assessment Number. conventions should be followed:

SUBMITTER At risk: The total number of animals or


The full name and address (including units (eg litters, pens) at the start of the
postcode) should be printed. The nominated period. Please specify the unit.
submitter's telephone and facsimile
numbers should also be given to allow

VETERINARY LABORATORY MANUAL 2


Affected: The total number of animals or
units that have been affected (sick or dead) Histopathology
during the nominated period eg year, Representative samples of affected tissue
month, lifetime. Please specify the period. with adjacent normal tissue. Tissues should
be in 1cm thick slices in 10x their volume of
Dead: The total number of animals or units buffered formalin solution.
dead during the same nominated period.
Parasitology
Further information eg daily mortality or Approximately 30g of faeces for faecal egg
prevalence can be provided in the 'History' count.
section.
Serology
HISTORY AND CLINICAL FINDINGS Full 10ml plain blood tube.
Include details of husbandry, nutrition,
clinical signs, treatments, and necropsy Toxicology
findings. Approximately 50ml of ingesta, faeces or
fresh tissue.
PREVIOUS REFERENCES
The laboratory reference number (eg. Virology
W04/01762), or date of any previous Full 10ml plain and EDTA blood tubes.
submissions relating to the condition from 30ml of fresh chilled tissue eg heart, spleen
this owner should b given, This allows the or swabs of lesions or tissues in PBGS.
duty pathologist to check previous reports,
offer more informed comment, and maintain NB All specimens should be clearly
continuity between individual submissions labelled with the animal identification.
from the same owner.
NSW Department of Primary Industries
POSTMORTEM FINDINGS does not supply specimen containers or
An accurate summary of significant consumables.
postmortem findings will help us provide
interpretation of laboratory test results. SPECIMENS (BY DISEASE OR
SYNDROME)
COLLECTION AND LABELLING OF For details of the most appropriate
SPECIMENS specimens and tests for a wide variety of
animal diseases, see Specimens (by
SPECIMENS (BY DISCIPLINE) disease or syndrome) in this Manual, with
For detailed information on collection and subsections:
handling of specimens for each laboratory • Diseases of livestock
discipline,see Specimens (by discipline) • Diseases of poultry
in this Manual. • Diseases of caged birds
• Diseases of bees
The following is a brief guide: • Diseases of fish
Bacteriology In general, it is better to send too many
Swabs of tissues eg heart blood, intestinal rather than too few samples. Specimens
content, in transport medium. will be held in the laboratory under
30 ml of chilled lesion, fluid or tissue eg appropriate conditions pending the results
liver, lung, intestine in a screw-capped of initial tests. The pathologist should be
container. given sufficient history and specimens to
allow the application of other examinations
Biochemistry which may be suggested from the history or
Full 10ml plain and LiHep blood tube. lesions. Such additional testing will be
undertaken only after consultation with the
Genetics submitter.
50 tail hairs with roots.
When investigating difficult or unusual
Gross Pathology disease problems, we encourage field
Representative samples of affected tissue veterinarians to consult with NSW
with any adjacent normal tissue. Department of Primary Industries laboratory
veterinarians to decide on the most
Haematology appropriate sampling and testing strategies.
Full 10ml EDTA blood tube and blood
smear.

VETERINARY LABORATORY MANUAL 3


Care must be taken in the collection and NB Virtually all specimens sent by
despatch of samples to the laboratory to veterinarians to laboratories are transported
ensure that you, your staff, personnel as ‘Diagnostic specimens’ and must be
handling the specimens during transit and packed to comply with IATA Packing
the staff at the Laboratory who have to Instruction 650.
unpack and handle the specimens are not
subjected to any risk of infection. For postage, IATA Packing Instruction
650 applies, since carriage by air may be
PACKING OF SPECIMENS involved. For carriage by road, other
regulations apply but, in general, are
Submitters are responsible for correctly satisfied by packing for carriage by air.
packing and consigning specimens in
compliance with current International Air IATA PACKING INSTRUCTION 650
Transport Association (IATA) regulations, (UN 3373 DIAGNOSTIC SPECIMENS)
which are updated annually in the IATA
publication ‘Dangerous Goods This instruction covers the labelling and
Regulations’. Incorrect packing may result packing of Category B infectious
in refusal by a courier to carry the substance as defined by IATA.
consignment and/or in legal action. ‘Diagnostic specimens’ is the Proper
Shipping Name.
NSW Department of Primary Industries
does not supply specimen containers or NB Virtually all specimens sent by
consumables. On request, recyclable veterinarians to laboratories are transported
specimen containers or eskies will be as ‘Diagnostic specimens’ and must be
returned to submitters. There is a small packed to comply with IATA Packing
charge for this service. Instruction 650.
IATA divides infectious substances into See Guidelines for Packing Diagnostic
Category A (IATA Packing Instruction Specimens below for practical advice on
602) and Category B (IATA Packing packing to comply with IATA Packing
Instruction 650). Packing Instruction 602 is Instruction 650.
more stringent than Packing Instruction
650. A number of companies sell suitable
• Within each Category, infectious packing, but by far the most common outer
substances are given a UN Number container is the foam esky enclosed within
and Proper Shipping Name a rigid cardboard box.
appropriate for their hazard
classification. The Consignment Note* must include the
following details:
Category A (IATA Packing Instruction • Proper Shipping Name ie ‘Diagnostic
602): An infectious substance which is specimens’
transported in a form that, when exposure • Net quantity* and
occurs, is capable of causing permanent • UN number ie ‘UN 3373’
disability, life-threatening or fatal disease to *Maximum 4 kg of solid or 4 L of liquid
humans or animals. material (excluding coolant). A
• UN 2814 (Infectious substance, maximum of 1 L within each primary
affecting humans*). container.
• UN 2900 (Infectious substance,
affecting animals only). A UN 3373 DIAGNOSTIC SPECIMENS
diamond-shaped label must be applied
*Proper Shipping Name. next to the Consignment Note. These labels
can be obtained from RVLs or from
Examples include Hendravirus, Bacillus couriers.
anthracis (cultures only), and exotic
pathogens such as FMD virus and Nipah *Pre-addressed Consignment Notes
virus. See IATA indicative list of Category A carrying the above endorsements, and also
substances. the account number of NSW Department of
Primary Industries (to charge the cost of
Category B (IATA Packing Instruction freight to the laboratory).are available from
650): An infectious substance which does your Regional Veterinary Laboratory.
not meet the criteria for Category A.
• UN 3373 (Diagnostic specimens) IATA PACKING INSTRUCTION 602
(UN2814 INFECTIOUS SUBSTANCE

VETERINARY LABORATORY MANUAL 4


AFFECTING HUMANS or UN2900 watertight secondary container with
INFECTIOUS SUBSTANCE AFFECTING sufficient absorbent material to
ANIMALS ONLY) absorb all fluid from the primary
container(s).
This instruction covers the labelling and A sturdy plastic bag will suffice as a
packing of Category A Infectious secondary container - it must be
Substance as defined by IATA (see sealed watertight.
above). It includes those agents that cause ii. Indelibly label the secondary
serious communicable disease. container with the owner's name.
iii. Put Specimen Submission Form(s)
Examples include Hendravirus, Bacillus into a separate plastic bag.
anthracis (cultures only), and exotic
pathogens such as FMD virus and Nipah Outer container (usually an esky)
virus. See IATA indicative list of Category A i. Put specimen secondary
substances. container(s), Specimen Submission
Form(s), cooler bricks and tight
IATA Packing Instruction 602 is more packing into the outer container
stringent than 650, and requires: (usually an esky).
• Person packing to have been trained Crumpled newspaper will suffice for
by an IATA-approved trainer within the tight packing.
past two years. ii. If the esky is foam, seal it into a
• Completed Shipper’s Declaration cardboard box.
for Dangerous Goods form with the iii. Complete the Consignment Note
package. and stick it onto the outer container.
iv. Stick a UN 3373 DIAGNOSTIC
GUIDELINES FOR PACKING SPECIMENS diamond-shaped
DIAGNOSTIC SPECIMENS label adjacent to the consignment
(IATA PACKING INSTRUCTION 650) note. These labels are available
from RVLs or from some couriers.
‘Dangerous Goods Regulations’, published
annually by the International Air Transport Call Customer Service (1800 675 623 if you
Association (IATA), should be consulted for need the Account Number of NSW
advice on specimen packing requirements. Department of Primary Industries for a
particular courier (to charge the cost of
NB Virtually all specimens sent by freight to the laboratory). Pre-addressed
veterinarians to laboratories are transported Consignment Notes, available from your
as ‘Diagnostic specimens’ and must be Regional Veterinary Laboratory, include this
packed to comply with IATA Packing Account Number
Instruction 650.
NB Suspect ANTHRAX submissions
The following notes are provided to assist must have a clear warning under the lid
submitters. of the outer packing. This warning must
be on top of samples and not packed in
Primary containers with the samples.
i. Tighten the lids of all primary
specimen containers. For TRANSPORT OF SPECIMENS
histopathology containers, tape the
lid to prevent loosening. For glass Specimens (other than those for export
slides, wrap together between stiff testing), should be submitted to the
cardboard. Regional Veterinary Laboratory which
ii. Clearly label each primary specimen services your region. Depending on the
container or slide with the animal tests specified, the samples may be sent by
identification. that laboratory to another appropriate
iii. Check that the specimens tally with Departmental testing centre. This may be
those identified on the Specimen another RVL or one of the Central
Submission Form. Laboratories at EMAI, Menangle.
iv. Estimate the total net weight of
specimens and record this on the A map showing the location of laboratories
Consignment Note. is available at
http://www.dpi.nsw.gov.au/agriculture/vetm
Secondary container(s) anual/contact .
i. Seal all primary containers from
each property/problem into a Generally specimens should not be sent by
overnight courier on Friday or at the

VETERINARY LABORATORY MANUAL 5


weekend. Weekend delivery attracts a left confined for long periods
heavy surcharge from all courier awaiting or during transport.
companies. Before calling your courier to
pick up specimens on a Friday, call the DISEASE SURVEILLANCE
laboratory to discuss options.
NSW Department of Primary Industries
The cost of transport of submissions to the collects epidemiological data on the
laboratory is included in the test charges. prevalence of animal disease by monitoring
Submitters can therefore charge the cost of the presenting problem and result of testing
freight to the NSW Department of Primary of material submitted to its laboratories.
Industries’ Account Number on the This data is used to satisfy the
Consignment Note. requirements of our trading partners and to
direct research and extension efforts of our
COURIERS staff. This passively-collected data is
Most submissions are sent to our supplemented by data collected while
laboratories by overnight courier. testing healthy animals for export or
Commercial couriers may use either road or interstate movement and by active
air transport and specimens should surveillance programs funded by industry or
therefore be packed in accordance with by the Department.
the more stringent IATA requirements
for air transport. For notifiable diseases (including
emergency diseases) and for mortality
NSW Department of Primary Industries has investigations, NSW Department of Primary
contracts or agreements with several large Industries encourages veterinarians to
courier companies as well as with local submit samples to their laboratories by
companies. Some companies offer same offering some 'free' testing.
day delivery. Call the laboratory to discuss
transport options before contacting a NOTIFIABLE DISEASES
courier company. A full list of the currently notifiable diseases
is available at
http://www.dpi.nsw.gov.au/agriculture/vetm
FORWARDING LIVE OR DEAD ANIMALS anual/submission/disease-surveillance.
The laboratory should be consulted before
live or whole dead animals are forwarded. Veterinarians and livestock owners have an
We will determine the feasibility of the obligation under various Acts of Parliament,
operation and make arrangements for to report any suspected occurrence of
receiving the animals. these diseases to NSW Department of
Primary Industries staff.
All costs associated with freight and
carcase disposal will be borne by the Generally, no fee will be charged for tests
submitter unless prior arrangements have conducted at a NSW Department of
been made. Primary Industries laboratory to exclude an
endemic notifiable disease. For emergency
It is the responsibility of the sender to meet (exotic) disease, all testing to exclude that
with the requirements not only of the disease and to establish an alternative
transport authority, but also with the diagnosis will be free of charge to the
requirements of welfare of the animal submitter. Full details of the eligibility
during transport. The following should be criteria have been circulated to
used as guidelines: veterinarians in NSW.
i. Containers must meet the
requirements of the transport For endemic diseases, submission of
authority. specimens to a NSW Department of
ii. The containers should be large Primary Industries' laboratory accompanied
enough for the animal to stand, to lie by a fully completed Specimen Submission
down fully extended and to turn Form nominating the suspected disease is
round. a suitable form of notification.
iii. Suitable bedding should be
provided. For emergency (exotic) disease, contact
iv. Animals should be placed in the the Animal Disease Watch Hotline 1800
container and delivered to the 675 888 or the local Senior Field Veterinary
terminal or railway station shortly Officer.
before the estimated time of
departure. Animals should not be

VETERINARY LABORATORY MANUAL 6


NATIONAL TSE SURVEILLANCE Submission Form. Specific
PROGRAM (NTSESP) requirements for testing of stock for
Full details of this program are available at each overseas export destination
http://www.animalhealthaustralia.com.au/pr can be obtained from the Animal
ograms/adsp/adsp_home.cfm. See also Quarantine Inspection Service
Transmissible spongiform encephalopathy (AQIS) [phone (02) 8334 7434; fax
in this Manual.. Veterinarians are (02) 8334 7430).
encouraged to submit specimens from adult
sheep and cattle with progressive iii. The name of the official veterinarian
neurological disease. The laboratory supplying the information on export
charges for this testing are charged to the testing requirements should be
NTSESP. Veterinarians and owners receive advised and stated on the Specimen
incentive payments. Submission Form. This will allow the
laboratory to check on any changes
MORTALITY INVESTIGATIONS in requirements, where necessary,
and to prepare quotations for the
NSW Department of Primary Industries
cost of laboratory services.
subsidizes laboratory fees for the
investigation of mortalities in livestock. To
iv. The name of the agent or shipping
qualify for this subsidy:
company involved should be
• The animals involved must be from a supplied on the Specimen
commercial livestock herd or flock. Submission Form with
Horse mortalities are eligible. For telephone/facsimile numbers.
poultry more than 100 birds must have
died. v. The Officer in Charge of the
• All owner and property details must be Regional Veterinary Laboratory,
provided on a fully completed Menangle or his/her appointed
Specimen Submission Form. nominee should be advised of the
expected dates of sampling and
NB Foetuses and neonates up to one day shipment of the animals prior to the
of age are not eligible for this subsidy. submission of the specimens.
When using this subsidy, submitters pay vi. The vendor of the stock should be
the first $80 of the laboratory fee and NSW listed as the owner of the stock
Department of Primary Industries up to rather than the agent or shipping
$140 of the residual. (These values are company. This allows different lots
GST inclusive) to be traced, when animals are
being purchased from several
properties.
SPECIAL REQUIREMENTS FOR
TESTING OF STOCK FOR vii. Full legal animal identification details
OVERSEAS EXPORT should be cross-matched to sample
identification and documented.
The following requirements should be
fulfilled: viii. It is particularly important for export
i. Intending exporters (owners and/or testing that samples of good quality
agents) must advise the Animal and adequate quantity be submitted
Quarantine Inspection Service and that they be correctly labelled.
(AQIS) [phone (02) 8334 7434; fax
(02) 8334 7430] of their intent to ix. For all overseas export testing the
export, at least four (4) weeks prior samples should be forwarded
to the commencement of testing, directly to:
otherwise there can be no
guarantee that testing will be Officer in Charge
completed and certification provided Regional Veterinary Laboratory
to allow export to proceed on Elizabeth Macarthur Agricultural Institute
schedule. Field veterinarians should Woodbridge Road
check to ensure this requirement MENANGLE NSW 2568
has been met.
The Regional Veterinary Laboratory and the
ii. The veterinarian submitting material Central Laboratories at EMAI, Menangle
is totally responsible for determining offer almost all required export tests for
the tests required. This information livestock. The RVL Menangle co-ordinates
must be included on the Specimen

VETERINARY LABORATORY MANUAL 7


most export testing in NSW and can be transport of the animals, prior to the
contacted by all field veterinarians directly submission of the specimens.
for enquiries and assistance.
iii. The vendor of the stock should be
Alternatively, certain tests can be listed as the owner of the stock,
performed within Regional Veterinary rather than the agent or transport
Laboratories at Orange, and Wollongbar for company. This allows different lots
export certification. to be traced, when animals are
being purchased from several
SPECIAL REQUIREMENTS FOR properties.
TESTING OF STOCK FOR
iv. Full legal animal identification must
INTERSTATE MOVEMENT be documented.
i. Advice on requirements for
interstate movement should be v. It is particularly important that
obtained from the District samples of good quality and
Veterinarian of the local Rural Lands adequate quantity be submitted, and
Protection Board. Nomination of that they be correctly labelled. This
tests to be performed is the will expedite testing.
responsibility of the authorised
official submitting veterinarian. vi. For interstate movement, send
specimens to the relevant Regional
ii. In cases where large numbers of Veterinary Laboratory for the district
animals are to be tested for of origin of the stock.
interstate movement (i.e. greater
than 200), the Officer-in-Charge of Those tests unavailable at Regional
the local Regional Veterinary Veterinary Laboratories will be undertaken
Laboratory or his/her appointed at EMAI following referral of specimens
nominee should be advised of the from the Regional Veterinary Laboratories.
expected dates of sampling and

VETERINARY LABORATORY MANUAL 8


CHECKLIST OF EQUIPMENT FOR CLINICAL AND NECROPSY EXAMINATIONS
Suggest Checklist for Clinical and Necropsy Examinations
1 Personal • Rubber boots
• Overalls
• Gloves
i. disposable
ii. post mortem gloves or gauntlets
• Post mortem apron (optional)
• Towel and soap
• Wet weather gear
2 Animal Husbandry • Halter
• Nose grips
• Bleeding choke rope
• Twitch
Pig handler
• Drugs
i. Rompun 2%
ii. Xylocaine 2%, etc
3 Clinical • Stethoscope
• Thermometers
• Obstetrical gloves
4 Clinico-pathological • Nitrate and cyanide test

5 Clinical Specimens • Vacutainers - plain, heparin and EDTA


• Vacutainer needle holders and needles (18G)
• Sterile containers (100 ml)
• Sterile bottles
• 1 oz Macartney - 25 ml
• Bijou Macartney - 5 ml
• Swabs
• Transport media
• Amies charcoal transport medium in bottles of commercial swab
packs
• Phosphate buffered glycerol saline (PBGS) in bottles
• Microscope slides (and spreaders for blood films)
• Syringes (1-20 ml) and needles (14-26 G)
• Scalpel handle and disposable blades
• Scissors
• Non-sterile faecal containers (50-100 ml)
• Vaginal mucus pipettes
• Ethanol/iodine for skin asepsis
• Biopsy instruments and small bottles of fixative
6 Euthanasia • Rifle and ammunition
• Euthanasia solution and syringe/needle
• Killing knife
7 Necropsy • Knives
• 20 cm skinning 2.5 cm wide straight blade
• 20 cm skinning 2.5 cm wide curved blade
• 15 cm boning knife pointed straight
• Steel - 30 cm Butcher's
• Footrot secateurs
• 7.5 cm blades
• Handsaw 30 cm blade and replacement blade
• Scissors
• Mayo straight 16 cm
• Mayo straight 14 cm
• Double sharp 11 cm fine
• Pointed/1 rounded end and knob (for gut running)

VETERINARY LABORATORY MANUAL 9


Suggest Checklist for Clinical and Necropsy Examinations
• Bone forceps
• Toothed forceps
• 20 cm
• 18 cm
• 13 cm
• Scalpel handles
• No. 4
• Scalpel blades
• No. 24 and/or No. 20
• Rib cutters
• Meatsaw/hacksaw
• Cleaner/hatchet
• Buckets - plastic (2)
• Trays
• plastic (large)
• plastic (small)
• Sterile scissors and forceps for virological tissue samples
8 Pathological • Sterile containers
Specimens • 100 ml
• 25 ml
• 5 ml
• Plastic bags 0.1 mm thick
• large
• medium
• small
• Rubber bands
• Microscope slides and diamond pencils, or markable slides and
pencil
• Histopath jars with 10% neutral buffered formalin
• Large 500 ml
• Medium 250 ml
• Small 125 ml
• Special fixatives (e.g. Bouins) if required
• Ball of string
9 Decontamination • Container and 20 litres water
/Disinfection • Disinfectant – one litre Lysol or similar
• Nail scrubbing brush (for cleaning instruments)
• Long handled scrubbing brush (for cleaning boots)
• Roll paper towel
• Plastic garbage bags - suitable container for contaminated 'sharps'
10 Clerical • Clip board
• Postmortem and Specimen Submission Forms
• Specimen collection handbook
• Pencil
• Marking pens
• Black biro
• Adhesive labels
• String-tie labels
11 Storage • Large metal esky with a blood vacutainer box and small foam esky
inside
• Frozen cold bricks, dry ice or portable car fridge
• Serum storage plastic disposable tubes - 5 ml

VETERINARY LABORATORY MANUAL 10


SPECIMENS (BY DISCIPLINE)

BACTERIOLOGY Microscope slides


Use slides of standard size and ensure they
are clean. Some commercially available
A bacterial disease can only be diagnosed
slides are not washed and are not suitable
when a pathogenic organism can be
for making satisfactory smears. Slides with
demonstrated, either on smear, culture, using frosted ends are preferred for labelling
molecular techniques or in tissue sections, in
purposes. Ensure slides are packed such
association with pathological changes. Many
that they don’t break in transit eg slide
pathogenic organisms are present in normal holder, wrapped in tissue/paper with
animals, e.g. Clostridium perfringens in
sufficient padding.
intestinal contents, so that recovery of the
organism alone may not necessarily be
Milk sample bottles
significant. In other cases e.g. E coli, there are
Sterile 30 ml Macartney bottles or any
many serotypes, but only a few are commonly
equivalent sterile, wide-mouthed, screw-
pathogenic. topped, plastic container. e.g. sterile 30 ml
universal containers.
STANDARD CONTAINERS AND
EQUIPMENT Pipettes for collecting vaginal mucus
Polystyrene artificial insemination pipettes,
Sterile plastic bottles and jars
approximately 50. cm long x 2mm internal
These are available commercially in a
diameter and 2 ml capacity. These are
range of sizes from about 10 to 100 ml.
available commercially.
They are used for fluids, discharges, small
lesions, etc.
Pipettes for collecting preputial
scrapings from bulls
Plastic bags
See
For larger specimens of organs or foetuses.
http://www.dpi.nsw.gov.au/agriculture/vetm
The primary receptacle must be leakproof
anual/specimens-by-
and not contain more than 500g. Plastic
discipline/bacteriology/02-bull-vd-coll.pdf
bags may be suitable. A minimum bag
thickness of 0.1 mm should be used. All
Plastic pipettes made from polystyrene or
bags should be securely sealed by
polypropylene with suggested dimensions
ziplocking or double folding and application
of external diameter 9.5 mm, internal
of stout rubber bands or equivalent, the bag
diameter 6.4 mm, 61.0 cm in length,
should be packed inside a second bag
straight except for a bend at the end that is
which also contains sufficient absorbent
held by the operator during collection; the
material to absorb fluid should the inner bag
other end, which contacts the preputial
leak and securely sealed as above. Plastic
surfaces, is bevelled. They should be fitted
bags must not be used if bone, horns or
to 90 ml firm rubber bulbs for suction while
other sharp objects may puncture the bag.
scraping.
Swabs
NB Do not use non sterile containers,
Swab transport systems for the recovery of
gloves or fragile containers when submitting
organisms are available commercially. They
material for bacteriological examination.
consist of plain cotton wool sterile swabs,
Aspirates submitted in syringes with
which are inserted into a semisolid
needles will not be examined.
transport medium (such as Stuart Transport
Medium or Amies Charcoal Transport
Medium) particularly for fastidious bacteria, COLLECTION OF SPECIMENS
before submission to the laboratory. There Specimens should be collected aseptically
are different transport mediums available and submitted promptly in individual sterile
commercially, such as swab/transport non-leaking, screw-topped, or plastic jars.
medium packs for general purposes,
swab/charcoal transport medium for more If small, the entire lesion or organ should be
sensitive organisms, swab/transport submitted. If the lesions are large or
systems modified for anaerobe recovery. widespread, submit a portion of the affected
Some testing involving special stains or tissue containing the lesion and
antigen testing may require dry swabs or surrounding area. Alternatively an aspirate
specific transport conditions of the lesion can be taken and transferred
to a small sterile container and submitted

VETERINARY LABORATORY MANUAL 11


Smears of intestinal mucosa and
If a septicaemia/bacteraemia is suspected, pathological lesions
submit portions of liver, spleen, heart blood Firmly smear suspect area with slide. A
and lung, in separate containers. number of smears can often be made on
each slide. AIR DRY and wrap separately
Swabs or place in slide protective holders for
Are generally inferior to the above. In the transport (commercially available).
case of exudates, a smear(s) should be
made and submitted with the swab. for Always leave one (frosted) end of the slide
culture should be submitted as soon as clean for handling and labelling. Smears
possible in transport medium (such as should be clearly labelled,
Stuart Transport Medium or Amies
Charcoal Transport Medium) is preferred., Intestinal contents
Stuart Transport Medium is unsuitable for Should be expressed into sterile, wide-
contagious equine metritis (Taylorella mouthed, non-leaking containers, avoiding
equigenitalis) transport if the swabs are to gross contamination of the sample or the
be processed more than 24 hours after container (preferred collection method).
collection. Swabs of intestinal contents should be
placed in a transport medium (such as
Aborted foetuses and foetal membranes Stuart Transport Medium or Amies
For foetuses from smaller animals (e.g. Charcoal Transport Medium).
sheep, goats, pigs) and smaller foetuses
from the larger animals, the entire foetus Milk samples
together with the foetal membranes should Wash the udder and the teats thoroughly
be enclosed in separate plastic bags and and dry with a paper towel. Swab the teats
submitted chilled. . are generally inferior to with 70 per cent alcohol. The first couple of
the above. In the case of exudates, a squirts should be discarded unless used for
smear(s) should be made and submitted a field test and, about 10 ml milk squirted
with the swab. into a sterile 30 ml Macartney bottle or
sterile 30 ml plastic universal container held
In the case of the large foetuses which nearly horizontally.
cannot be delivered to a laboratory, a post-
mortem examination should be conducted Avoid touching the mouth of the bottle with
and the appropriate specimens should be the teat. Code the samples numerically.
collected aseptically into sterile jars (see
‘Abortion’). The samples must be submitted chilled
using either crushed ice or cooling bricks in
Impression smears of foetal membranes an insulated container. Milks must be
should be taken prior to despatch of refrigerated if transport is slightly delayed,
specimens. or frozen if transport is delayed more than
2-3 days.
Portions of the placenta or cotyledons
should also be taken and fixed in buffered Preputial material from bulls
formalin prior to despatch. Preputial scrapings submitted unchilled in
selective transport medium for T foetus
Faecal samples and rectal swabs (InPouch TF) See
Collect approximately 30 g faeces direct http://www.dpi.nsw.gov.au/agriculture/vetm
from the rectum into a sterile, wide- anual/specimens-by-
mouthed non-leaking container. Do not discipline/bacteriology/02-in-pouch-
overfill the container. (preferred collection guide.pdf
method) Rectal swabs, heavily impregnated
with faeces, should be submitted in Selective transport medium for T foetus
transport medium such as Stuart Transport (InPouch TF), together with instructions for
Medium or Amies Charcoal Transport the collections and despatch of specimens
Medium. will be forwarded from the laboratory on
request.
For Johne's faecal culture, see the section
‘Johne's disease’. Preputial samples should not be frozen or
refrigerated. Keep at temperatures between
NB. If a parasitological examination is also 18oC and 30oC.
required, a duplicate sample should be
collected. Semen samples

VETERINARY LABORATORY MANUAL 12


Must be collected aseptically avoiding If specimens for Campylobacter culture
contamination by preputial material. will not reach the laboratory within 6-8
Samples collected using an artificial vagina hours, then sealed pipettes from which
are often grossly contaminated unless they air bubbles are excluded should be
are cultured shortly after collection. used (Atmospheric oxygen is lethal to
Campylobacter fetus). Plastic pipettes
From 0.5 to 1.0 ml is sufficient for can be sealed adjacent to the mucus
bacteriological examination. Attempts to using a pair of pliers previously heated
collect large volumes by repeated in a flame. on dry ice is recommended.
ejaculation increase the risk of Consult with your courier if you intend
contamination. to use dry ice for submission of
samples.
Urine samples
For routine cultural examinations to be Bacteria with particular requirements
meaningful, urine samples should reach the Mycoplasma, Spirochaetes (including
laboratory within a few hours of collection. If Leptospira, Brachyspira), Chlamydia,
there are likely delays before or during obligate anaerobic bacteria (including
transport, specimens must be chilled, but Dichelobacter nodosus) have special
should be processed by the laboratory requirements. If culture is required, consult
within 72 hours of collection. with your Regional Veterinary Laboratory.

When direct examination for leptospires is STORAGE OF BACTERIOLOGICAL


required, submit a fresh urine specimen SPECIMENS PRIOR TO DESPATCH
specifically for leptospiras motility
examination at the laboratory within 20 In general, all samples for bacteriological
minutes of collection. Otherwise, for examination, except smears, should be
leptospire morphology submit a separate kept chilled (2-8oC) but not frozen, from the
urine sample of at least 20 ml, preserved time of collection until they are received in
with 0.25 ml of undiluted formalin or 1.5 ml the laboratory. This will ensure the
of 10% formalin. minimum growth of contaminants. Insulated
containers with frozen icebricks or plastic
Vaginal mucus samples bags of crushed ice should be taken in the
Mucus can be collected from the ventral car to all investigations and specimens
fornix of the vagina and the external os of placed therein as soon as possible.
the cervix by guiding a plastic artificial
insemination pipette by hand (per rectum) Delays in transport over weekends and
and applying GENTLE suction to the public holidays must be considered.
external end. Specimens are better held refrigerated until
the sender is sure the transport to the
To diagnose bovine venereal laboratory will not be delayed.
campylobacteriosis (BVC) in abortion or
infertility cases: Samples of vaginal mucus for
i. (Preferred method): Vaginal mucus bacteriological examination for
samples are collected for Campylobacter fetus should be submitted
demonstration of C fetus subsp on dry ice in an insulated container. Consult
venerealis antibodies by BVC ELISA with your courier.
test. PBST diluent and instructions
for collection of mucus samples are Specimens best stored at room
available from the laboratory. See temperature (not refrigerated)
http://www.dpi.nsw.gov.au/agricultur The following samples should be stored
o o
e/vetmanual/specimens-by- and transported between 18 C and 37 C (ie
discipline/bacteriology/01-bvc-elisa- not refrigerated or frozen):
guide.pdf. Vaginal mucus is • Samples submitted in Campylobacter
collected using a plain, sterile cotton Transport Medium (CETM)
swab and placing the cotton swab • Samples submitted in Trichomonas
end into phosphate buffered saline Transport Medium (In Pouch)
plus Tween (PBST) for transport. • Swabs submitted in Amies Charcoal
The swabs, PBST and instructions medium for Contagious Equine Metritis
are provided by the laboratory. (CEM)
ii. For isolation of Campylobacter, • Swabs submitted in Amies Charcoal
direct inoculation to Campylobacter medium for or Atrophic rhinitis.
transport media (CTM) is ideal.

VETERINARY LABORATORY MANUAL 13


• Tissues for fungal culture (temperatures and disposable plastic containers have
of less than 15oC can be detrimental to eliminated this problem.
fungal survival).
Standard containers for Biochemistry are:
DESPATCH OF SPECIMENS • Blood vacuum tubes (10 ml),
siliconized, without anticoagulant: For
See section ‘Packaging and sending blood and serum samples.
specimens to the laboratory’. In addition, • Blood vacuum tubes (10 ml) with EDTA
when forwarding specimens for or heparin: For plasma samples.
bacteriological examination the following • Serum vials: 5 ml vials or bottles with
procedures should be adopted: leak-proof closures.
• All specimens for transport should be • Bottles, jars, plastic bags - for tissues,
packed in accordance with IATA organs, etc., as listed for Bacteriology.
requirements for diagnostic specimens
(Packing Instructions 650) so as to
prevent leakage and risk of accidental COLLECTION OF SPECIMENS
exposure of personnel handling the Blood and serum samples
container. Blood samples should be collected
• Specimens for transport from cases of aseptically into the appropriate 10 ml
suspected tuberculosis or anthrax vacuum tube.
should always be submitted individually
in containers separate from all other Contamination of the sample by soil,
specimens and in accordance with faeces, hair, etc., must be avoided.
IATA requirements for infectious
substances (Packing Instructions 602). Where whole blood will not arrive at the
Pack such that on opening the outer laboratory within 24 hours of collection,
packaging, a note advising ‘Suspect sera should be separated from the clot as
TB’ or ‘Suspect Anthrax’. soon as possible and frozen or chilled in 5
• The specimen advice form is best ml serum vials. Chilling should also
sealed in a plastic bag to avoid continue in transit.
contamination or damage from water.
• Check to ensure that specimens are Care should be taken to avoid haemolysis
submitted at the temperature (see "Serology - Avoiding Haemolysis of
appropriate for the disease condition, Samples"). Haemolysis will seriously
i.e. chilled, or at room temperature. interfere with the level of serum
Check with your courier. magnesium, phosphate, total protein,
albumin and enzyme levels.

BIOCHEMISTRY Sera for Vitamin A or E analysis must be


protected from heat and light. Separated
sera or plasma must be placed into 5 ml
NSW Department of Primary Industries’ containers wrapped in foil, then forwarded
laboratories do not carry out any biochemical chilled to the laboratory.
analysis; all such testing is outsourced to
laboratories that are accredited by NATA to Plasma samples
conduct biochemical tests. At least 2 ml of plasma should be submitted
frozen in 5 ml screw capped containers.
In some cases it is more appropriate to apply a
test on fresh samples in the field rather than The blood sample should be collected in
submit samples to a laboratory, e.g. urine the appropriate blood vacuum tube. Where
analysis, where commercially available kits will whole blood will not arrive at the laboratory
provide a more reliable answer on the spot, within 24 hours of collection, plasma should
rather than the same test applied in the be separated by centrifugation as soon as
laboratory 24 to 48 hours later. possible and preferably within 4 hours of
collection. The plasma should be removed,
free of all cells and then kept chilled or
STANDARD CONTAINERS AND frozen. Chilling should also continue in
EQUIPMENT transit.
Contamination of glassware and other
Plasma for Vitamin A or E analysis must be
collection equipment by traces of the
protected from heat and light. Separated
element of interest has been an issue in the
sera or plasma must be placed into 5 ml
past. Vacuum blood tubes and needles,

VETERINARY LABORATORY MANUAL 14


containers wrapped in foil, then forwarded Muscular degeneration, nutritional
chilled to the laboratory. Selenium deficiency
Serum enzymology
Tissue samples Vitamin A and E
At least 50 g of tissue should be submitted Vitamin B12
frozen in a screw capped container. Care
must be taken to avoid contamination by GENETICS
soil, faeces, ruminal or intestinal contents
when collecting tissue for biochemical The field veterinarian should check the
analysis. Specimens (By Disease or Syndrome)
section of this manual for details on the
Different organs must be submitted in specimens required for diagnosis of the
separate containers. If other examinations specific disease suspected.
are required, e.g. bacteriology, pathology,
then duplicate samples as required for the Where DNA based tests have been
other examinations should be submitted in developed for genotyping at a defined locus
separate containers. the preferred sample is hair roots. DNA can
also be isolated from anticoagulant-treated
Tissues for vitamin A or E analysis blood and from semen; however the tests
These should be protected from heat and will be significantly more expensive than
light; wrap in foil or brown paper and with those that exploit hair roots as a
submit chilled to the laboratory. source of DNA. Only with prior arrangement
with the laboratory can other tissue
samples be the utilised as the source of
STORAGE OF SPECIMENS PRIOR
DNA for routine tests.
TO DESPATCH
Samples of tissues must be chilled or Where DNA tests are not available,
frozen as appropriate until despatch. diagnosis may be possible by histological
examination of suitable tissues and findings
DESPATCH OF SPECIMENS used to complement historical observations,
breeding data and clinical findings.
Specimens should be forwarded in an
insulated container with an icebrick. To STANDARD CONTAINERS AND
prevent frozen samples from becoming
EQUIPMENT
heated during transit, always ensure there
are several ice bricks to ensure samples Vacuum blood tubes (5mL or 10mL, with
arrive at least chilled: anticoagulant EDTA or heparin)
• Always seal the specimen advice form For blood samples.
in a separate plastic bag.
Paper envelope or pre-labelled ‘ziplock’
GUIDE TO INTERPRETATION OF plastic bags
BIOCHEMICAL PARAMETERS IN SHEEP For tail hairs.
AND CATTLE
Normal values are supplied by the laboratories COLLECTION OF SPECIMENS
to which we outsource and are included in our Hair
reports to submitters. Suggested normal Tease out 20 to 25 hairs growing at the
values for some analytes are included under distal end of the tail. Select hairs that are
specific diseases in the section ‘Specimens (by free of visible faecal contamination. Tie a
disease or syndrome)’ of this manual. Where knot in the hair shafts approximately one
values supplied by laboratories differ from quarter of the distance from their proximal
these suggested values, the former should be end. Firmly grasp the knotted hairs and pull
used. with a quick action. Ensure roots are
present at the proximal end of the hair
The ‘Serum enzymology’ section includes shafts. If desired, cut off and discard the
information on enzyme changes see in various distal half of the shafts. Place the shafts
conditions. (with roots) in an envelope labelled with the
animals identity, and post to EMAI. See
Refer to relevant section of the manual. See: http://www.dpi.nsw.gov.au/agriculture/vetm
Acetonaemia anual/specimens-by-disease-
Copper deficiency syndrome/diseases_of_livestock/dpi-samp-
Hypomagnesaemia coll-prot.pdf
Hypocalcaemia
Hypophosphataemia

VETERINARY LABORATORY MANUAL 15


Use of forceps or artery clamps may delivered. Any dead animal should arrive in
facilitate collection of suitable samples from time to allow a post mortem examination on
newly-born calves. the same day.

Clearly state the breed of the subject and STANDARD CONTAINERS AND
DNA test required. EQUIPMENT
Blood Specimens for gross pathology should be
Fill evacuated blood tubes, to end of draw, submitted in glass or plastic screw-top
from the coccygeal vessels or the jugular containers or strong plastic bags. When live
vein. Immediately after drawing the sample or dead animals are submitted, the
ensure mixing of anticoagulant and blood containers used should prevent
by repeated but gentle inversion of the contamination of the environment with
tubes. Label each tube with the identity of possible pathogens.
the subject.
COLLECTION OF SPECIMENS FOR
Semen GROSS PATHOLOGY
A semen straw or 0.5mL of raw semen in a
serum vial. When submitting whole animals, ensure
that they are typical of the syndrome being
STORAGE OF SPECIMENS AND investigated.
DESPATCH OF SPECIMENS When the necropsy is done in the field and
Samples of tissues other than hairs must be a laboratory opinion is sought on gross
chilled until despatch. changes detected, then a large portion of
the tissue or organ should be submitted
Do not freeze blood samples. chilled but not frozen, containing the lesion
and adjoining normal tissue for comparison.
Clean, dry hairs are stable at ambient
temperature for an extended period of time. If material is submitted for macroscopic
examination, small portions in buffered
Specimens other than hairs should be formalin should also be taken and
forwarded in an insulated container with an submitted for histopathological examination.
icebrick. Use sufficient icebricks to ensure
samples arrive at least chilled. Refer to STORAGE OF SPECIMENS PRIOR
Guidelines for Packaging Specimens for TO DESPATCH
more detail.
Tissue samples should be kept chilled from
Always seal the specimen advice form in a the time of collecting until they are received
separate plastic bag. at the laboratory.

NB Do not freeze samples being


submitted for either gross or
GROSS PATHOLOGY histopathological examination
Provided that the field veterinarian can be sure
that the specimens will be delivered to the
Regional Veterinary Laboratory promptly, there HISTOPATHOLOGY
are advantages in submitting the whole animal
or whole affected organs. In general, the histopathologist will only be
able to offer comment on the significance of
Submission of the animal, live or dead, should lesions when an appropriate range of
be considered when there is a: specimens has been submitted, together with a
• High mortality or morbidity from an good history and a description of the clinical
unknown cause. and necropsy findings.
• Continuing problem and previous
examinations have not established a STANDARD CONTAINERS AND
diagnosis. EQUIPMENT
All animals either live or dead, submitted to the (Neutral) Buffered Formalin
laboratory must be accompanied by the
appropriate specimen advice form with all Buffered formalin solution is the
relevant details. The laboratory should be recommended fixative because of its
advised by phone that the animal is being stability, longer life of preserved tissues and

VETERINARY LABORATORY MANUAL 16


a negligible fixation deposit. It is prepared Ensure that the mucosal surface of tubular
as follows: organs (eg gut, uterus, bladder) is exposed
by excision of the wall before immersion in
Commercial Formalin * 100 ml fixative.
NaH2PO4.2H2O 4.0 g Handle only the edges of delicate tissues
Na2HPO4 6.5 g so that histological detail is not lost.

Water to 1 litre Spinal cords


* Commercial formalin is Formalin B.P. 34- After exposure, the spinal cord should be
38% formaldehyde. It should be used neat lifted by grasping the dura with tissue
in this formulation. forceps. Each spinal nerve root is then
severed as it is exposed.
Buffer salts are available commercially in
prepared packs. The dura is then opened longitudinally in
the dorsal midline and the cord and dura
submitted whole in a large container of
Other fixatives buffered formalin.
Other fixatives for special purposes are
available from the laboratory. To prevent fixation of the cord in a curled
position, Transect the cord at 10 to 20 cm
Containers intervals, leaving a part of the dura intact at
These must have a tightly fitting lid to each transection site, to keep the cord
prevent leakage. segments connected. The cord will then be
Containers must have sufficiently wide fixed in short, straight segments. Open the
mouths to allow tissue to be easily dural to allow better penetration of the
withdrawn after fixation. fixative.

Don’t squeeze large tissue samples into Brain


containers. There are several methods of exposing the
brain for removal. Convenient methods for
COLLECTION OF SPECIMENS large animals (eg longitudinal and
transverse craniotomy) are used in the
Tissues collected at necropsy should be National TSE Surveillance Program
preserved immediately. (http://www.animalhealthaustralia.com.au/pr
ograms/adsp/adsp_home.cfm)
If fresh organs are being submitted for
bacteriological or gross pathological It is preferable to submit the brain whole in
examination, then a small block of the a wide mouth container with 10 times its
organ (sliced 0.5 cm and no more than 1 volume of buffered formalin solution. After
cm thick) should be fixed in buffered 24 hours fixation, the formalin solution can
formalin at the time of collection, for be changed to hasten fixation.
histopathology.
Ideally, bovine and equine brains should be
Always submit an adequate range of tissue immersed in at least 6 litres of buffered
specimens. formalin solution (e.g. in plastic buckets) for
at least 48 hours, then submitted in a large
Preserved tissue should be submitted in at container of fresh buffered formalin
least 10 times its volume of fixative. solution.
It is essential to avoid distortion of the brain
Decomposing or frozen tissues are not during fixation. Compression of brain
suitable for histopathological examination surface resting on the base of the container
can be avoided by adding neat formalin
Organs until the brain floats.
Organs smaller than 1 cm in thickness can
be fixed whole. If the brain cannot be submitted whole, it
may be cut transversely into two or three
Organs greater than 1 cm thick must be pieces. Always cut transversely so that all
sectioned. Tissue for fixation should be brain sections can be examined (grossly
sliced 0.5 cm (and no more than 1 cm) and histologically) for bilateral lesions. Do
thick. They should include part of the lesion not cut the brain longitudinally.
and adjacent apparently healthy tissue.

Gastrointestinal tract and other tubular


organs

VETERINARY LABORATORY MANUAL 17


HAEMATOLOGY NB Do not chill blood films or expose
them to formalin vapour.
BLOOD Blood films can be submitted in the same
For red and white cell counts, haemoglobin container as chilled specimens, provided
estimation and derived parameters, collect the blood films are securely packed and
blood in EDTA (purple top) tubes. well wrapped for insulation against chilling.

EDTA blood may not be suitable for


differential white cell count and morphology PARASITOLOGY
if blood films are not made within 1 hour of
blood collection. Faecal egg counts and egg type (by larval
differentiation) are a guide to the size and type
BLOOD FILMS of worm burden. Faecal egg counts are
influenced by faecal consistency and bulk, host
Fresh blood films should be made from
resistance, stage of pregnancy and effects of
EDTA blood immediately after collection.
lactation.
They should then be air dried.
Starvation will increase egg counts and
Leucocytes degenerate when blood stands
inappetence may cause the count to increase
for a few hours.
by up to 30 to 40 times. Diarrhoea will reduce
egg counts.
Use dry slides, free of dirt and grease.
Avoid contamination when preparing blood
Faecal egg counts will vary according to the
films.
parasite species involved and whether the
worm burden consists of sexually mature
PREPARATION OF BLOOD FILMS parasites.
i. Use a glass slide as the spreader
The spreading edge must be The history should provide details of recent
smooth (i.e. no chips out of it) and anthelmintic treatments including drench
clean. Spreaders can be re-used if resistance status, stocking rate, pasture
cleaned and dried after each use. availability, swampy areas, grazing rotation.
ii. Place a small drop of blood
(thoroughly but gently mixed) in the STANDARD CONTAINERS AND
centre of the slide about 1 cm from EQUIPMENT
the end. Grasp the spreader with
thumb and middle finger using the Jars
index finger to put light pressure on Plastic (5 ml, 20 ml, 70 ml) screw top
the spreader. containers for insects, parasites, aliquots of
iii. Place the spreader beyond the drop gut washings, skin scrapings.
and at 45o to the slide. Draw it back
until it touches the drop and the Parasite Faecal Collection Kits
blood spreads evenly across the Collection kits are available from your
slide almost to the edges. Regional Veterinary Laboratory:
iv. Quickly and smoothly push the • WormTest kits contain 10 containers for
spreader right to the end of the slide monitoring a single mob or grazing
at the 45o angle. group in a herd.
v. Wave the slide quickly in the air to • DrenchTest kits include 50 containers
dry it. Label at the thick end with the for one control and four test drench
owner's name and animal ID (if groups. Use additional WormTest kits
necessary) using a lead pencil. for extra drench groups.
• Horse WormTest kits are suitable for 5
STORAGE AND DESPATCH OF horses.
SPECIMENS • DrenchRite® kits for bulk faecal
collection allow farmers to test for
Blood should be kept chilled, not frozen, resistance with a single muster.
whilst being held prior to despatch and
during transport to the laboratory. Blood Vacuum blood tubes (with
should be submitted to the laboratory anticoagulant)
chilled in insulated containers. Avoid For heartworm antigen and microfilaria
contact between the blood sample and tests in dogs (generally for export
icebricks. purposes).

VETERINARY LABORATORY MANUAL 18


Vacuum blood tubes (plain) When identification of parasitic cysts is
For serology of liver fluke infection in cattle. required, tissues should be submitted
chilled in a jar.
COLLECTION OF SPECIMENS
Blood
Faecal samples Collect a full tube of blood. For tubes with
For individual animals at least 30 g of fresh anticoagulant ensure blood is gently mixed
faeces should be collected (preferably from with anticoagulant. Plain blood tubes
the rectum) and placed directly into a jar. should be allowed to clot at room
temperature.
The jar should be filled and the lid closed
tightly in order to minimise exposure of
STORAGE AND DESPATCH OF
worm eggs to air which would allow egg
development. SPECIMENS
Faecal samples should be kept cool, but
Wormtest Kit samples should be collected not frozen. Even prolonged chilling at 5oC
from 10 animals, including apparently kills the eggs of most species and makes
healthy and affected animals in each mob. samples unsuitable for larval culture for
Complete instructions for collection of strongyle egg identification.
samples are provided with each WormTest
kit. Faecal samples should be submitted as
soon after collection as possible, chilled in
Gastrointestinal tract an insulated container, with icebricks
The total gastrointestinal tract should be wrapped in newspaper to prevent freezing.
submitted direct to the laboratory when it
can be delivered by the submitter, double Gastrointestinal tract washings should be
bagged in strong clear plastic bags. Only preserved in 5% formalin.
submit unpreserved chilled
gastrointestinal tracts if delivery to the Also see specific parasitic diseases:
laboratory is assured within 36- 48 • Anthelmintic resistance in sheep
hours. The tract should not be opened and • Babesiosis
each part should be tied off at the • Coccidiosis
appropriate junction, i.e. between the • Cryptosporidiosis
stomach (abomasum) and small intestines
• Cysticercosis
and between the small and large intestines.
• Drenching mortalities
Alternatively the organs can be washed by • Equine babesiosis
the field veterinarian and aliquots (from • Fasciolosis in sheep and cattle
washings of known volume) submitted to • Ostertagiosis
the laboratory. Contact the laboratory for • Paramphistomiasis
instructions on the procedure. • Parasites internal/external/resistance
• Sarcosporidiosis
Insects and snails for identification • Tapeworms
These should be submitted in 70% alcohol • Tick fever
in a small leak proof container. • Toxoplasmosis
• Trichomoniasis of cattle
Mites • Total worm counts (interpretation)
Acute skin lesions should be scraped with a • Worm egg counts (interpretation)
scalpel until blood is produced. Moisten the
scalpel with liquid paraffin. Scrape
particularly at the edge of any visible lesion.
Before scraping, hair or wool must be SEROLOGY
clipped as close to the skin as possible.
Scrapings should be sealed in a small, wide Serology is available for a range of bacterial
mouthed bottle and submitted unpreserved. and viral diseases, as well as for some
chlamydial, mycoplasmal, rickettsial, protozoan
Parasites for identification and metazoan diseases. Such tests may be
Flukes, tapeworms and roundworms should available in Regional Veterinary Laboratories,
be washed in water and preserved in 5 per or only in Central Veterinary Laboratories at
cent formalin. Always include the head of EMAI, Menangle.
the tapeworm.
A serological test is used to show the presence
or absence of antibody to a specific
aetiological agent or group of agents. The

VETERINARY LABORATORY MANUAL 19


presence of antibody indicates exposure to the Use a separate sterile needle to avoid
organism, which may be due to a current mechanically transmitting infectious agents
clinical condition or to an earlier unrelated from one animal to another.
infection.
Avoiding haemolysis of samples
In tests where results are expressed in titres, Haemolysis occurs as a result of poor
the best evidence of infection is the collection techniques, contaminated
demonstration of a four-fold titre rise between equipment or poor handling of the sample
samples collected early in the clinical episode once it is collected.
and those collected 2-3 weeks later. A titre
variation of less than four-fold (i.e. one dilution) Common causes of haemolysis include:
is within the normal variation of a serological • Use of non sterile containers for
test and is not significant. collection or storage.
• Contamination by faecal and other
Since many animals in endemic areas may material due to faulty aseptic
have antibody to a given organism, a single techniques.
sample from an affected animal may not allow • Contamination of the sample by water.
the serological test to be interpreted. • A slow flow from the needle, due to
obstruction of the needle, or failure to
A single serum sample is particularly useful in insert into mid-vein.
eliminating a diagnostic possibility. Frequently, • Forcibly expelling blood through a
presence of antibody in the acute phase or needle.
absence in the convalescent phase will • Heating of samples, usually in car boots
eliminate a diagnostic possibility. or through back windows of car, or after
prolonged exposure to direct sunlight
Haemolysed or contaminated samples often during collection.
give unreliable results in a complement fixation • Freezing.
test (CFT): the serum may be anti
complementary or give non-specific low titre NB. Pig blood haemolyses quickly. Serum
positives, particularly in sheep and goat CFT's. should always be separated from the clot
within 4 hours of collection.
Poor quality samples will give poor quality
results, necessitating retesting of the animals Labelling of samples
involved (see Samples must be labelled serially (e.g. from
Avoiding haemolysis of blood samples). 1 to 30) with a water proof pen, preferably
on an adhesive label. Keep a key list which
STANDARD CONTAINERS AND correlates sample numbers with animal
EQUIPMENT identification. Do not label the stopper,
which is removed during testing.
Blood vacuum tubes, 10 ml
These should be silicone coated, without
The specimen advice form submitted with
anticoagulant.
the samples should list clinical details
beside each sample number. This will allow
Sterile screw capped containers, 5 ml
the laboratory to offer an informed comment
For serum samples.
on the results and perhaps apply other
relevant tests.
COLLECTION OF SPECIMENS
Blood samples should be collected DO NOT label samples with tag numbers,
aseptically, using a 10 ml blood vacuum names, etc., as this leads to confusion and
tube. At least 5 ml of blood should be errors in reading numbers in the laboratory.
collected. When a range of serological tests It also makes it very difficult for the
is required (particularly when viral serology laboratory to ensure all sample are present
and non-viral serology is to be undertaken), or to check on missing or broken samples.
duplicate samples should be collected.
DO NOT label containers with water soluble
Contamination of the container and stopper ink. It smudges when wet and may rub off
should be avoided. Blood and faecal if samples are chilled or frozen.
material should be removed prior to
despatch, to reduce the risk of Serum can be frozen, provided there are no
contamination of laboratory staff handling blood cells present in it.
the specimens.

VETERINARY LABORATORY MANUAL 20


STORAGE AND DESPATCH OF The level of ELISA-reactive antibodies can
SPECIMENS be measured in a qualitative and
quantitative way. The quantitative measure
Samples should be allowed to clot before is relative to control sera. This may be in
transporting them over any distance. Clots terms of:
may not retract readily in cold weather or if i. Absorbance (Optical Density, OD) of
they are chilled too soon after collection. the test serum, or,
Samples should be held in a warm room ii. ELISA ratio - compares the OD of
until the clot retracts. the test serum to that of a negative
control, but taking into account the
Once the clot has retracted, blood samples performance of one or more positive
must be held chilled to reduce control sera on the plate to give a
contamination, haemolysis and autolysis. valid test. Depending on the test,
ELISA ratios of 2 or 3 are common
If the laboratory will not get the samples cut-off points, or,
within 48 hrs of collection, decant the serum iii. ELISA value or ELISA units -
into a 5 ml sterile, screw capped plastic depending on the test, this can be
container and submit the serum sample used to reflect either:
only. If virological examination of the clot is • the OD of the test sample relevant
also needed (e.g Pestivirus antigen to a standard curve, originating
detection), submit clot separately. from a range of positive and
negative controls, or,
TYPES OF SEROLOGICAL TESTS • a mathematical formula which
expresses the OD of the test
Complement Fixation Test (CFT)
sample as a percentage of the
The test quantifies complement-fixing
positive control, with subtraction of
antibodies to a range of antigens. Test
the negative control OD from each,
serum, complement and antigen are
ie: (test OD - neg OD) / (pos OD -
incubated then sensitised erythrocytes are
neg OD) x 100
added. If there is specific antibody present,
an antigen-antibody complex is formed
(Agar Gel Immunodiffusion (AGID)
which binds complement and sensitised
Syn: Gel Diffusion Precipitin Test (GDPT)
erythrocytes are not lysed. Results are
given in titre form, reflecting a doubling
Specific antibody is detected by placing test
serial dilution of serum from 1:4 upwards.
serums and control serum in wells in a gel
The titre indicates the dilution at which 50%
around a central antigen well. Migration of
or more of the erythrocytes are not lysed.
antigen and antibody towards each other
results in a visible line of precipitation
Enzyme-Linked Immunosorbent Assay
where an antigen/antibody complex is
(ELISA)
formed. A GDPT reaction reflects the
The ELISA detects specific antibodies in
position of a specific line of reaction
serum which are allowed to bind to antigen
relevant to the serum and antigen wells, on
on a solid phase. To detect the bound
a limited scale of trace, 1, 2 or 3. A reading
serum antibodies, the test uses a second
of 2 indicates a line midway between the
antibody system (directed against
serum and antibody wells. A trace and 1
antibodies of the animal species under test)
reaction are located closer to the serum
to which is linked an enzyme. This enzyme
well, whereas a 3 reaction is located closer
catalyses a colour reaction, and the amount
to the antigen well. Trace is the weakest
of colour which develops reflects the level
reading, and 3 reflects the highest level of
of original serum antibody. This colour is
antibody detected.
measured quantitatively and is termed the
serum absorbance or optical density.
Serum Agglutination Test (SAT)
Tests for agglutinating antibody in a tube-
The second antibody is usually species-
based test, where clearing of the tube due
specific, but may react with closely related
to agglutination is used to determine an
animal species at a different concentration.
endpoint. Results are expressed as a titre,
Thus each ELISA is geared towards one
and the dilution series used (and therefore
test animal species.
the titre) reflects the standard protocol for
An absorbed ELISA (e.g. Johne's ELISA)
the test required. In some tests, the
involves the above steps, but also includes
endpoint readings are converted to
treatment of serum to absorb non-specific
international units rather than titres.
antibodies before testing.

VETERINARY LABORATORY MANUAL 21


Microscopic Agglutination Test (MAT) Rapid Plate Test (RPT)
A test used for Leptospiral antibody which An agglutination assay used for some
measures the ability of sera at varying poultry pathogens, and performed with
dilutions to give an endpoint of 50% diluted serum on a solid base.
agglutination of one of a range of live
leptospiral serovars. Latex Agglutination Test (LAT)
Antigen coated-latex beads are reacted
Indirect Fluorescent Antibody Test with diluted sera, with a positive test giving
(IFAT) a lattice of latex beads due to agglutination.
A slide-based assay using serum at varying
dilutions to show specific binding to the test Rose Bengal Test (RBT)
antigen on the slide. The fluorescent- A spot agglutination test performed on a
labelled second antibody provides a solid base and used as a screening test for
measurable signal of bound antibody from Br. abortus. Gives a qualitative (+/-) result
the test serum. only.

VETERINARY LABORATORY MANUAL 22


SUMMARY OF AVAILABLE SEROLOGICAL TESTS FOR VARIOUS NON-VIRAL DISEASES
Host Disease Agent Available Diagnostic Test
Cattle Leptospirosis L pomona MAT (serovar specific)
L hardjo
MAT* (serovar specific)
Other Lepto serovars include:
copenhageni, tarassovi,
grippotyphosa, canicola,
australis, zanoni
Campylobacteriosis Campylobacter fetus venerealis ELISA (vaginal mucus)
Johne's disease M paratuberculosis ELISA, CFT*
Salmonellosis S Typhimurium SAT (serogroup specific)
S Dublin
Chlamydiosis/SBE C psittaci CFT
Brucellosis B abortus ELISA, RBT*, SAT*
Liver fluke F hepatica ELISA
Q fever C burnetti CFT*
Neosporosis Neospora caninum ELISA
Sheep Brucellosis B ovis CFT, ELISA
Salmonellosis S Typhimurium SAT (serogroup specific)
S Dublin
A. seminis A seminis CFT (at ARI Yeerongpilly)
epididymitis
Johne's disease M paratuberculosis AGID, CFT*
Chlamydiosis C psittaci CFT
Liver fluke F hepatica ELISA
Toxoplasmosis T gondii LAT
Goats Johne's disease M paratuberculosis ELISA, AGID, CFT*
Toxoplasmosis T gondii LAT
Brucellosis B abortus SAT*
Q fever C burnetii CFT*
Horses Babesiosis B equi IFAT*
Leptospirosis L pomona MAT*(serovar specific)
L tarassovi
Brucellosis/ B abortus SAT*
Fistulous withers
Pigs Mycoplasmosis M hyopneumoniae ELISA
M hyorhinis
Leptospirosis L pomona, MAT (serovar specific)
L tarassovi,
L bratislava
Brucellosis B suis RBT, SAT
Poultry Pullorum S pullorum RPT
Mycoplasmosis M gallinarum RPT
M synoviae
Dogs Leptospirosis L canicola MAT (serovar specific)
L copenhageni

Key
AGID Agar gel immunodiffusion test;
CFT Complement fixation test
ELISA Enzyme linked immunosorbent assay
IFAT Indirect fluorescent antibody test;
LAT Latex agglutination test
MAT Microscopic agglutination test
RPT Rapid plate test
RBT Rose Bengal test
SAT Serum agglutination test;
* Commercial test only

VETERINARY LABORATORY MANUAL 23


TOXICOLOGY Separate organs should be placed in
separate containers.
It is not possible to comprehensively screen Body fat is the preferred tissue for
samples for 'poisons' or 'toxins'. It is up to the insecticide residue testing. For biopsy
submitter to consider the history, clinical signs material, a minimum of 2g is required.
and lesions (if any) and identify specific toxins
for analysis. Contact your Regional Veterinary
Blood samples
Laboratory if you are unsure whether a test is At least 8 ml of blood in a blood vacuum
available for a particular toxin. tube, free from contamination with faeces
etc.
In cases of suspected poisoning, it is important
that an effect be demonstrated in the animal. Serum samples
For example, nitrate poisoning is confirmed by At least 2 ml of serum should be submitted.
demonstrating the presence of nitrate in the
serum or blood of the animal, not by Blood smears
demonstrating the presence of nitrate in Thick air dried smears should be prepared,
pasture plants in the paddock. taking care to leave one end of the slide
clean. They should be dry before being
It is important that the history provided includes wrapped in paper.
details of treatment with any suspected toxic
compound, particularly in relation to the Suspected toxic material
strength of the preparation and the time since Suspected material, feedstuffs or plants
treatment or access to the material. should NOT be sent unless appropriate
specimens from affected animals have also
STANDARD CONTAINERS AND been sent.
EQUIPMENT
At least 50 g of material should be
Blood vacuum tubes, 10 ml
forwarded.
For blood samples
Plant material
Leakproof 5 ml tubes or bottles
Examination for nitrate nitrite and cyanide
For serum samples.
are best performed in the field.
Bottles and jars
Plants for identification should be pressed
For tissue samples
and dried. Refer section on "Poisoning -
plant" for submission of plants for
Slides
identification.
These must be clean. Some brands
available have not been washed and are
Ingesta
not suitable unless they are washed.
Nitrate and nitrite disappears rapidly from
ingesta, and thus ingesta is of no value in
COLLECTION OF SPECIMENS diagnosing nitrate/nitrite poisoning.
Particular care should be taken in collecting
and packaging specimens for toxicology, For other chemical poisons, eg. arsenic,
because there is often a possibility that lead, at least 250 g of ingesta should be
infectious agents are involved and these submitted in an air tight, leakproof
can create hazards to staff handling the container.
material in a laboratory. Therefore avoid
contaminating the outside of any submitted STORAGE OF SPECIMENS PRIOR
containers with tissues and ensure they are TO DESPATCH
leakproof.
Tissues should be frozen. Herbage and
Tissue samples ingesta samples for toxin examination
At least 100 g of tissue should be collected, should also be frozen. Other animal
taking care to avoid contamination with soil, specimens should be chilled.
faeces or intestinal contents.

VETERINARY LABORATORY MANUAL 24


INTERPRETATION OF ARSENIC, LEAD AND COPPER CONCENTRATIONS IN CATTLE AND
SHEEP
Analyte Sample Units Deficient Normal Toxic
Arsenic Liver mg/kg (wet wt) < 0.5 *>8
Lead EDTA blood µmol/L < 1.2 > 1.2
Kidney mg/kg (wet wt) <4 > 25
Faeces mg/kg (wet wt) < 10 > 25
Copper Liver mg/kg (wet wt) <4 ** 20-70 † > 100
Kidney mg/kg(wet wt) 4-6 >8

* Liver arsenic may be in range 2 to 8 mg/kg if several days elapsed since toxic exposure.
** Typical liver copper for cattle is > 20 and sheep > 40.
† In sheep, liver copper may increase up to 200 mg/kg (wet wt) before poisoning occurs

NB
i. Interpret concentrations between normal and toxic according to clinical and pathological
findings.
ii. Concentrations based on dry wt are approximately 5 times the above (wet wt) values.
iii. Conversion from mg/kg to SI units is as follows:
• As mg/kg x13.3 = As μmol/kg
• Pb mg/kg x 4.8 = Pb μmol/kg
• Cu mg/kg x 0.0157 = Cu mmol/kg

VIROLOGY
Presence of antibody can be a result of
clinical disease, unapparent infection,
Virological examinations involve demonstration passive immunity or vaccination.
of a pathogenic virus or detection of antibody
to virus. Findings must be interpreted in the Tests variously demonstrate group specific
light of history, clinical findings, lesions, etc. antibody, type specific antibody or a cross
reaction.
It is not possible to screen for a wide range of
viruses. Submitters should forward specimens Viral serology can be applied with the
to be tested for specific viruses. If there is any following limitations:
doubt about the availability of a test, the
laboratory should be contacted for advice.
Single Serum
• Useful only to eliminate a diagnostic
DIAGNOSIS OF VIRAL DISEASE possibility. Presence of antibody in the
The diagnosis of a viral disease can be acute phase may eliminate the
based on Histopathology, Virus isolation, diagnostic possibility; absence of
Virus Serology, or Virus or Virus Antigen antibody in the convalescent phase
Detection. eliminates the diagnostic possibility.
• Accurate interpretation often difficult
HISTOPATHOLOGY because time of collection may be
Cytological changes can indicate a viral critical. A negative test on serum < 3
aetiology. weeks after a suspected viral condition
leaves the possibility that the animal
VIRUS ISOLATION was infected but had not yet produced
Cultivation and identification of virus grown detectable antibody.
in tissue culture or eggs inoculated with
specimen. Paired Sera
• A change from negative to positive
NB Failure to cultivate the virus does not antibody status is known as
rule out a viral aetiology. seroconversion and indicates infection.
• A four-fold rise in antibody titre between
Cultivation of a virus does not necessarily acute and convalescent samples can
mean it caused the disease process. also indicate infection by the specific
pathogen.
VIRAL SEROLOGY
Detection and quantitation of virus specific However both the above may also be due
antibody in the serum of an infected animal. to cross reaction or booster immunisation.
Plasma is also an acceptable sample for They may also indicate stress induced
most serological tests. reactivation of latent infection.

VETERINARY LABORATORY MANUAL 25


Serum is usually titrated in two fold serial
The relationship between virus and dilutions beginning at 1/4 or 1/10. The virus
antibody varies between diseases. In some, neutralising antibody titre is the reciprocal
virus does not occur in animals with of the serum dilution that will neutralise a
detectable antibody, e.g., Akabane, standard amount of virus (usually
Ephemeral Fever whereas in others, virus 100TCID50).
and antibody can be present in the one
animal, e.g., EIA, IBR, CAE, EBL and Two serial dilutions rise (i.e. 4-fold) in titre
occasionally Pestivirus. in the convalescent serum sample
compared to the acute serum sample is
NSW Department of Primary Industries considered significant. This occasionally
offers the following types of viral serology occurs as a result of technical or statistical
tests: variation, so greater differences can be
• Agar Gel Immunodiffusion Test (AGID) accepted with greater confidence.
• Virus Neutralisation Test (VNT)
• Haemagglutination Inhibition (HI) Test Haemagglutination Inhibition (HI) Test
• Enzyme Linked Immunosorbent Assay HI tests are used to detect antibodies to
(ELISA) viruses which agglutinate erythrocytes. HI
tests depend on antibody binding a fixed
Agar Gel Immunodiffusion (AGID) Test amount of antigen, preventing it from
AGID involves diffusion of viral antigen and causing haemagglutination. Sera is tested
antibody towards each other through a gel. in two-fold dilutions, and results expressed
When they combine, they precipitate in the as a titre (negative, 2, 4, 8, .......) etc. More
gel. This produces a visible line where the accurate measurement of flock status can
concentrations of antigens and antibody are be gained by repeat sampling of the flock to
balanced. An excess of antigen or antibody determine if a rise in titre (e.g. a rise in titre
can alter the location and appearance of of two dilutions or more is arbitrarily
the precipitin line. considered significant) has occurred.

Each test sample for viral antibody is tested Enzyme linked immunosorbent assay
against a known viral antigen to examine (ELISA)
the relationship of any precipitin line formed ELISA results can express the detection of
with an adjacent standard reference line antibody in a qualitative (+ or -) or a
formed between known positive serum and quantitative way. The three common means
virus antigen. This reference line is of expressing a quantitative result are
optimised to give a strong visible precipitin absorbance (optical density), ELISA ratio
line centrally located between the antigen and ELISA value (unit).
and positive serum wells (termed a 3
reaction - see below). Absorbance (Optical Density)
The amount of antibody in a sample is
For viral serology, a specific precipitin line proportional to the absorbance (optical
formed by the tested serum sample is density, OD) given by it. The OD may
recorded as 1, 2, 3, or >3 to describe its be standardised against one or more
relative position to the serum well and the serum controls on the ELISA plate.
antigen well: ELISA ratio
The ELISA ratio indicates the strength
Description / Position of Precipitin Line of the sample OD compared to that of a
Result
Turn on end of reference line negative 1 serum control on the same
Line, closer to serum well ELISA plate.
2 (E/R = OD test/OD neg
Line, midway between serum well and antigen well control) 3
Line, closer to antigen well ELISA value
>3 or ELISA units
The sample OD is fitted to a curve
A 1, 2, 3 or >3 antibody reaction is a determined by the performance of
positive test for antibody, and indicates that control positive and negative samples,
an animal has been infected with the and the result given as an ELISA value
specific virus (or a related virus). (eg 0 100). This is often a
measurement of how the sample OD
Strength of viral antibody levels in the test compares with that of a high positive
generally does not reflect the severity and control (taken as a value of 100).
stage of infection with any certainty. Percent inhibition
The results of blocking or competitive
Virus Neutralisation Test (VNT) ELISAs are expressed as the reduction
in OD (as a %) relative to the OD given

VETERINARY LABORATORY MANUAL 26


by a reference positive (often Can be used to detect viral antigen or virus
monoclonal) antibody. particles. Results are only qualitative (+ or
), eg:
ELISA ODs, ratios or values may be placed • Rotavirus detection
into categories
(negative/inconclusive/positive) determined Haemagglutination (HA) Test
by previous experience of the performance Can be used to screen for
of the test. haemagglutinating viruses, eg:
• Newcastle Disease
VIRUS OR VIRUS ANTIGEN DETECTION • Avian Influenza
• Egg Drop Syndrome
The following types of test are available for
the direct detection of virus or viral antigen: On poultry tissue or allantoic fluid from
• Electron microscopy (EM) eggs. Results can be given as a titre
• Agar Gel Immunodiffusion (AGID) Test derived from a doubling dilution of the
• Enzyme linked Immunosorbent Assay sample (e.g. 2, 4, 8, ...etc).
(ELISA)
• Latex Agglutination Test To determine which virus is responsible, HA
• Haemagglutination (HA) Test positive samples can be tested in the
presence of specific antibody in a direct (or
Electron microscopy (EM) reverse) HI test. The results are qualitative
Allows detection and identification of virus (+ or - ).
particles on a morphological basis e.g.
Rotavirus, Poxvirus. STANDARD CONTAINERS AND
EQUIPMENT
Agar Gel Immunodiffusion (AGID) Test
Antigen detection is useful where virus Blood vacuum tubes, silicone coated
infected tissues contain soluble virus For serum samples and blood clots (for
antigens that visibly precipitate with specific isolation of, or detection of antigen to, some
antibody, eg: viruses). Please do not use serum
• Parvovirus in mummified pig foetuses separator tubes.
• Pestivirus in gut scrapings of cases of
mucosal disease Blood vacuum tubes with heparin or
• Rotavirus in faeces. EDTA
For blood for virus isolation (of mainly the
In such cases, the sample is tested against arboviruses) or preparation of buffy coat
a known positive serum and the relationship samples for antigen detection.
of any precipitin line formed with a standard
reference line, originating from an adjacent Sterile bottles for samples of tissues,
positive antigen, is examined. organs
Sterile 5 ml vials: For serum and fluid
A test sample giving a line of similar samples.
position and intensity, which is continuous Swabs and PBGS bottles: For discharges,
through a turn of identity with the reference etc. Plain sterile swabs are used and
line is classed as +++ for viral antigen. immediately placed in small bottles of
phosphate buffered gelatin saline
The results are recorded in terms of relative (PBGS). These are available from the
strength (+, ++, +++, or XS) against the Regional Veterinary Laboratories.
reference line.
Sterile instruments
Enzyme linked Immunosorbent Assay An adequate range of sterile scissors and
(ELISA) tissue forceps should be available.
Can be used to detect either virus antigen
or intact virus particles. Results can be The containers and equipment listed are
expressed qualitatively (+ or ) or suitable for the routine collection of tissues
quantitatively (OD's or ELISA ratios or for virus examination. Containers and
ELISA values). eg: equipment for special requirements will
• Pestivirus detection in the Pestivirus normally be supplied by the laboratory, after
antigen capture ELISA (PACE). prior arrangements have been made.

Latex Agglutination Test COLLECTION OF SPECIMENS


Specimens for virus isolation must be
collected by aseptic techniques, using

VETERINARY LABORATORY MANUAL 27


sterile instruments and sterile containers. carefully, avoiding contamination from other
Avoid contamination from other tissues as sites.
well as that from extraneous sources.
Submissions for virus isolation should be Each swab should then be transferred to
accompanied by serum from the affected PBGS at room temperature, then chilled
animal(s) whenever possible. (but never frozen) for transport to the
laboratory.
Blood
This should be collected using the Faeces
appropriate blood vacuum tube, avoiding At least 10 g of fresh faeces should be
contamination of the sample. A separate collected into a clean jar.
needle should be used for each animal.
STORAGE AND DESPATCH OF
Avoid contamination of the outside of the SPECIMENS
tube by blood, soil and faeces, as this
creates difficulties in handling the All virological specimens should be chilled
specimens within the laboratory. prior to and during transport. If more than
48 hours is to elapse between collection
Serum and receipt at the laboratory, specimens
If blood samples cannot arrive at the except blood samples should generally be
laboratory within 2 days, the serum should frozen.
be poured off aseptically into a sterile 5 ml
vial. The clot should also be submitted as it For isolation of Arboviruses (especially
may be required for virus isolation or Bluetongue) and Herpesviruses (e.g. IBR),
antigen detection. specimens should never be frozen. These
viruses have extremely poor survival at
o
Tissues and organs 20 C with just a single freeze, and tissues
These must be collected aseptically, using for their isolation must be kept chilled.
separate sterile instruments and containers
for each tissue or organ. Contamination All specimens should be clearly labelled
between tissues must also be avoided. and sent in a leakproof container. Check
that screw caps are tight, especially after
Portions of tissue (no greater than 2 cm x 2 freezing.
cm x 2 cm) should be taken.
All samples should be packed in insulated
Scabs containers with sufficient icebricks to
Scabs and underlying tissue should be ensure that they are still cold when received
submitted in a sterile bottle. at the laboratory. However, care should be
taken to prevent direct contact between
Swabs coolant bricks and specimens, which may
Swabs from excretions, exudates, mucosal otherwise become frozen.
surfaces and orifices should be taken
SUMMARY OF AVAILABLE VIROLOGICAL TESTS FOR VARIOUS DISEASES OR DISEASE
SYNDROMES

Section A: Recognised Viral Diseases


Host Disease Virus Available Diagnostic
Test
Cattle Ephemeral fever (EF) Rhabodovirus VNT, (ELISA), (VI)
Bovine pestivirus Pestivirus Antigen ELISA, AGID,
Mucosal disease (MD) VI
Infertility Antibody ELISA, VNT
Abortion
Congenital abnormalities
Bovine malignant catarrh Bovine herpesvirus Nil; use histopath
Ovine herpesvirus 2 PCR at AgWest
(OHV2)
Infectious bovine Bovine herpesvirus – Type VNT, VI
rhinotracheitis (IBR) 1
Infectious pustular vulvovaginitis Bovine herpesvirus – Type VNT, VI
(IPV) 1
Encephalitis Bovine herpesvirus – Type VNT, VI
1

VETERINARY LABORATORY MANUAL 28


Host Disease Virus Available Diagnostic
Test
Mammilitis Bovine herpesvirus – Type EM
2
Bovine papular stomatitis Parapox virus EM
(pseudocowpox)
Congenital Akabane virus AGID, VNT, (VI)
arthrogryposis/hydranencephaly Aino virus
(AG/HE) Palyam virus
Pestivirus also causes AG
Calf scours Rotavirus LA, EM
Coronavirus EM
Enzootic bovine leucosis (EBL) EBL virus (Retrovirus) ELISA, AGID
Bluetongue Bluetongue virus (BTV) AGID, ELISA, VNT, VI
(various types) NB. AGID is group
specific; VNT is type
specific
Abortion Palyam virus AGID, (VNT, VI)
congenital abnormalities (various types)
(see also Pestivirus)
Enzootic haemorrhagic AGID, (VNT, VI)
disease of deer (EHD)
virus (EHD1 EHD5)
Ibaraki disease EHD2 AGID, (VNT, VI)
Sheep Contagious pustular dermatitis Parapox virus EM, Histopath
(Orf, scabby mouth)
Arthrogryposis/hydranencephaly Akabane virus AGID, VNT, ELISA
(AG/HE)
Ovine pestivirus (Border Pestivirus Antigen ELISA, AGID,
disease) VI,
Antibody ELISA, VNT
Bluetongue Bluetongue virus (BTV) AGID, ELISA, VNT, VI
(various types) NB. AGID is group
specific; VNT is type
specific
Pigs Encephalomyocarditis virus Cardiovirus VI, (VNT)
(EMC)
Swine Pox Poxvirus EM, Histopath
Haemagglutinating encephalitis Coronavirus (HI)
virus infection (HEV)
(Vomiting and wasting disease)
Piglet scours Rotavirus LA, EM
Parvovirus disease Parvovirus AGID
(for antigen and
antibody)
Goats Caprine arthritis encephalitis Retrovirus ELISA, (VI, AGID)
(CAE, big knees)
Vaginitis Caprine herpesvirus VI, (VNT)
Horses Equine infectious anaemia (EIA) Retrovirus AGID
Equine rhinopneumonitis Equine herpesvirus –Type VI, (VNT)
Viral abortion 1
Poultry Avian encephalomyelitis (AE) Picornavirus ELISA, Histopath
Avian influenza Orthomyxovirus AGID, ELISA, HA, VI,
Fowl plague/ Histopath
Duck influenza
Avian leukosis Retrovirus Histopath
Egg drop syndrome (EDS 76) Haemagglutinating HI, (VNT, HA, VI)
adenovirus 127 (group II)
Fowl adenovirus (FAV) Adenovirus (group I) AGID, VNT, Histopath
acute inclusion body hepatitis (1 serotype of 12) AGID is group specific;
(IBH) of chickens < 14 d VNT is serotype
specific

VETERINARY LABORATORY MANUAL 29


Host Disease Virus Available Diagnostic
Test
Fowl Pox (FP) Avipoxvirus EM, (VI), Histopath
Turkey haemorrhagic enteritis Adenovirus (group III) AGID, Histopath
(THE)
Haemorrhagic enteritis
(HE) of turkeys
Herpesvirus of turkeys Herpesvirus (VI), (HVT)
Inclusion body hepatitis (IBH) Adenovirus (group I) VI, VNT, Histopath
(Many serotypes)
Infectious bronchitis (IB) Coronavirus (VI), Histopath
Infectious bursal disease (IBD) Birnavirus ELISA, (AGID),
Histopath
Infectious laryngotracheitis (ILT) Herpesvirus Antigen ELISA,
Histopath, (VI)
Marek's disease Herpesvirus (AGID, VI), Histopath
Newcastle disease (ND) Paramyxovirus HI, ELISA, HA, VI,
Histopath
Reticuloendotheliosis (RE) Retrovirus VNT, (VI), Histopath
Fish Epizootic haematopoietic Iridovirus VI
necrosis (EHN)

Crustacea White spot syndrome (WSS) White spot syndrome virus PCR

Section B: Syndromes with Possible Viral Involvement

Host Disease Possible Virus Available Diagnostic Test


Cattle Respiratory disease Parainfluenza 3 (PI3) ELISA, (VNT)
Respiratory syncytial (RSV) ELISA, (VNT)
Pestivirus AGID, (ELISA, VI, VNT)
Rhinovirus *
Adenovirus AGID
Sheep Respiratory disease PI3 ELISA, (VNT)
Poultry Big liver spleen syndrome ? AGID, Histopath
(BLS)

Key
AGID Agar gel immunodiffusion test
ELISA Enzyme linked immunosorbent assay
EM Electron microscopy
HA Haemagglutination test
HI Haemagglutination inhibition test
LA Latex agglutination test
VI Virus isolation
VNT Virus neutralisation test
* available by arrangement

VETERINARY LABORATORY MANUAL 30


SPECIMENS (BY DISEASE OR SYNDROME)

DISEASES OF LIVESTOCK If a full term calf is examined in the field,


and dystocia is indicated, there is little value
in laboratory investigation.
ABORTION (GENERAL)
Notes on the individual species should be Specimens required
consulted. Whenever possible, the aborted
foetus together with the foetal membranes Aborted foetuses
should be submitted to the laboratory. i. The whole foetus and foetal
Every opportunity should be taken to membranes submitted chilled, or:
emphasise to owners the importance of ii. Portions of foetal stomach contents,
collecting the foetus and foetal membranes. placenta, liver, lung, spleen;
iii. Clear serous body fluid (pericardial,
Where it is not feasible to submit the foetus, thoracic or peritoneal) or heart blood
the autopsy must be complete, and a range submitted chilled in separate sterile
of tissues and body fluids submitted to containers, and,
eliminate cause of perinatal death other iv. Sections of cotyledons, brain,
than infectious agents e.g. dystocia. myocardium, lung, liver, and
kidney in buffered formalin for
Diagnosis histopathology.
Based on bacteriology, virology, serology Cows
and histopathology findings together with i. Sera from the aborting animals and
the previous clinical history of the dam. 10-15 animals from the same group.
Indicate on the Specimen
Infectious abortions can be distinguished Submission Form which animals
from non-infectious abortion by measuring have aborted.
elevated levels of species-specific IgG in
foetal fluid (blood, peritoneal, pleural or ABORTION IN SHEEP AND GOATS
pericardial fluid). This test is available for Common causes in New South Wales
cattle and pigs. include toxoplasmosis, campylobacteriosis,
listeriosis, brucellosis, Border disease,
ABORTION IN CATTLE Akabane virus and pasteurellosis. Other
Common infectious causes of abortion in causes include chlamydiosis and conditions
New South Wales are leptospirosis, and causing clinical illness in the ewe or doe,
campylobacteriosis. Other infectious e.g. salmonellosis, hypocalcaemia,
causes include listeriosis, fungal and viral pregnancy toxaemia, septicaemia. Habitual
infections of the dam and foetus. Pestivirus, abortion can occur in Angora does.
Akabane and Palyam virus possibly cause
abortion. Pestivirus may be associated with Specimens required
abortion at any stage of gestation; akabane
and palyam are predominantly associated Aborted foetuses
with abortions of the last trimester of i. Foetus and foetal membranes
pregnancy. These three viral infections may submitted whole in an insulated
cause congenital abnormalities. container. They should be chilled,
not frozen. or
In some regions (e.g. North Coast, Illawarra ii. Portions of foetal stomach contents,
and South Coast), the most commonly placenta, liver, lung, spleen
diagnosed infectious cause of abortion is iii. Clear serous body fluid (pericardial,
neosporosis. Characteristic lesions are thoracic or peritoneal) or heart blood
seen in sections of fixed brain and submitted chilled in separate sterile
myocardium. Neospora caninum, a containers, and,
sporozoan with a canid-bovine transmission iv. Sections of cotyledons, brain,
cycle, has recovered from tissues of myocardium, lung, liver, and
affected foetuses. kidney in buffered formalin for
histopathology.
Non-infectious causes include plant Ewes and does
poisonings and congenital abnormalities. i. Sera from the aborting animals and
up to 10 flock mates submitted
chilled. Indicate on the Specimen

VETERINARY LABORATORY MANUAL 31


Submission Form which animals ii. Serum sample from the aborting
have aborted. sow and up to 10 herd mates,
See Abortion in cattle re specimens for submitted chilled.
virology
ACETONAEMIA
ABORTION IN HORSES
Syn: Ketosis
In the past, it has frequently been See also: Pregnancy toxaemia in sheep
impossible to reach a diagnosis because of
an inadequate history and unsuitable Occurs in both beef and dairy cattle
specimens submitted.
Diagnosis
Common infectious agents responsible for History, particularly starvation and/or stress
equine abortion include Streptococcus spp., in late pregnancy and early lactation;
Staphylococcus spp., Escherichia coli, clinical signs, serum biochemistry (beta
Salmonella spp., Klebsiella spp., hydroxy butyrate), demonstration of
leptospirosis, equine herpesvirus and fungi. ketones in urine and milk.

Non-infectious causes may include Specimens required


maternal pyrexia and malnutrition, cord i. Serum sample submitted chilled (for
compression abnormalities, endocrine beta hydroxybutyrate analysis).
dysfunction, endometrial incompetence and
immunological factors. NB Examinations of urine and milk for
ketones and glucose should be carried out
Specimens required in the field.

Aborted foetuses ACTINOBACILLOSIS AND


i. Foetus and foetal membranes ACTINOMYCOSIS
submitted whole and chilled in an
insulated container, or, Diagnosis
ii. Portions of placenta, liver, lung, Demonstration of Actinomyces or
kidney and spleen together with Actinobacillus organisms in pus or tissue
foetal stomach contents submitted sections from early lesions; typical
chilled. histological changes. Granulomatous
iii. Portions of placenta, liver, adrenal lesions containing granules in the pus may
gland, lung, kidney, brain and also be caused by staphylococci and
spleen submitted in buffered Arcanobacter (syn Corynebacterium)
formalin for histopathology. pyogenes.

ABORTION IN PIGS Specimens required


i. At least four pus smears from deep
Common causes of abortion in swine in parts of the lesion. Smears from the
New South Wales include leptospirosis, surface are not satisfactory.
streptococcal infection and toxaemic ii. Affected tissue and specimens of
conditions in the sow. Other possible pus submitted chilled.
causes include genetic factors, oestrogenic iii. Sections of recent lesions in
feeds, erysipelas, encephalomyocarditis buffered formalin for histopathology.
(EMC) virus and vitamin A deficiency.
ACTINOBACILLUS SEMINIS
Abortion should be differentiated from
parturient and immediate post parturient INFECTION IN RAMS
deaths due to hypoxia and managerial Diagnosis
factors such as injury or stress, smothering History, clinical examination (particularly for
and chilling. orchitis in young rams). Demonstration of A
seminis on culture of semen.
NB Parvovirus infection causes foetal
mummification and parturient deaths; it Specimens required
does not normally cause abortion. i. Semen sample collected aseptically,
submitted chilled.
Specimens required ii. Serum sample, submitted chilled (to
i. Aborted foetuses and membranes eliminate Brucella ovis as a cause).
submitted chilled. iii. Testes, epididymes and accessory
sex glands including ampullae and
seminal vesicles submitted chilled.

VETERINARY LABORATORY MANUAL 32


ALGAL POISONING
NB The CF test on serum is not a reliable
test for A. seminis. See: Blue-green algal poisoning'

AFLATOXICOSIS ALPHA MANNOSIDOSIS OF


CATTLE
Aflatoxins can cause a range of clinical
signs including nervous signs, diarrhoea, See: Mannosidosis
hepatic disorders and abortions.
ANAPLASMOSIS
Diagnosis
Based on clinical findings, histopathology, See: Tick fever
serum enzymology, and aflatoxin analysis
where appropriate. ANAEMIA
This can arise from acute or chronic blood
Specimens required loss, by increased erythrocyte destruction
i. Serum for enzymology. or by impaired erythrocyte production.
ii. Suspected foodstuff for aflatoxin
analysis and mycology. Diagnosis
iii. Specimens appropriate to the The diagnosis of clinical anaemia should be
particular clinical syndrome should made in the field. The diagnosis of the
be submitted. Irrespective of the cause requires the submission of the
syndrome, sections of liver and appropriate range of specimens.
kidney in buffered formalin should
be submitted to check for toxic Many conditions, including haemonchosis,
changes. fascioliasis and enzootic haematuria are
best diagnosed in the field. The following
AKABANE DISEASE IN SHEEP AND disease conditions should be considered.
CATTLE • Parasitism, including fascioliasis and
other helminthiases.
See also: Congenital abnormalities,
• Enzootic haematura in cattle.
abortion, arthrogryposis and
hydranencephaly' • Babesiosis, anaplasmosis and
eperythrozoonosis.
Diagnosis • Plant poisonings, including bracken
The diagnosis of Akabane disease relies on fern, pyrrolizidine alkaloids, rape.
history, pathological findings and the • Copper and cobalt deficiency in
demonstration of antibodies to Akabane ruminants.
virus in foetal fluids or serum samples taken • Iron deficiency anaemia in piglets.
from calves or lambs which have not fed • Coccidiosis, sarcosporidiosis.
from the mother.
Specimens required
Specimens required Depending on clinical and post mortem
i. At least 2 ml of foetal fluid findings, appropriate tissues should be
(preferably pericardial or pleural sent. These should include:
rather than heart blood or peritoneal i. Liver, kidney, spleen, skeletal and
fluid which are more contaminated) cardiac muscle and red bone
for virus serology, marrow from the femur in buffered
formalin.
or, from a neonate, serum if the animal ii. Blood films for haematology.
has not sucked. iii. At least 5 ml EDTA blood for
haematology.
ii. Serum sample from the dam, iv. Faeces for parasitology
submitted chilled. (This is only useful v. Serum chilled for copper etc.
in eliminating a diagnosis of
Akabane or Aino infection) ANNUAL RYEGRASS TOXICOSIS
iii. Brain, spinal cord and muscle in (ART)
buffered formalin for histopathology.
Toxicity of Wimmera ryegrass to cattle and
NB Specimens should also be submitted sheep in WA and SA is associated with a
to exclude pestivirus infection, which can plant toxin of the tunicaminyluracil antibiotic
cause arthrogryposis (AG) and CNS complex produced in galls in the rye grass
lesions. seed head by parasitic bacteria (Clavibacter
formerly Corynebacterium rathayi)

VETERINARY LABORATORY MANUAL 33


transported by invading plant nematodes IN VITRO LARVAL DEVELOPMENT
(Anguina agrostis). This disease has not yet ASSAY (LDA)
been recorded in NSW, but a similar
disease involving the same toxin and In vitro larval development assay (LDA,
Clavibacter bacterium has been seen in DrenchRite™) is especially useful for initial
cattle and some sheep in late 1990 along drench resistance testing on a property as it
the Darling flood plains, originating from requires little on-farm effort. However, it is
infected "blowaway grass" (Agrostis only able to quantify resistance to
avenacea). Clinical signs of ART include benzimidazole (BZ), levamisole (LEV),
neurological disturbances characterised by combination (BZ/LEV) and indications for
ataxia and collapse with tetanic and clonic macrocyclic lactones (ML, ie
convulsions, ending in death, and abortions avermectin/milbemycins). It can not be
in ewes. used for increased dose levels or
combinations.
Diagnosis
History, clinical findings, gross pathology, FAECAL EGG COUNT REDUCTION TEST
histopathology. Evidence of yellow bacterial (FECRT)
galls on annual ryegrass plant
heads/seeds; identification of Clavibacter Faecal egg count reduction test (FECRT)
sp in feed. Elimination of other diseases requires considerable on-farm effort.
associated with neurological signs and CNS FECRT can be performed using either:
oedema (e.g. enterotoxaemia). i. A DrenchTest kit, or,
ii. Several WormTest kits (one for each
Specimens required anthelmintic and one for a control
i. Fixed liver and brain in buffered group).
formalin
ii. Suspect grass or grass seed (e.g. FECRT anthelmintics
showing bacterial galls). Where nematode resistance is suspected,
the trial should contain groups treated with
ANTHELMINTIC RESISTANCE a range of anthelmintics. Traditionally 5
groups (BZ, LEV, BZ/LEVcombination,
Repeated treatment of populations of ivermectin (IVM) and an untreated control
parasites with anthelmintics selects group) were used. However, with recent
individuals that have innate or acquired extensive spread of resistance in these
resistance to the drugs. Ultimately chemical groups, other drenches including
treatment becomes ineffective because a naphthalophos (NAP, Rametin ™), and its
large proportion of the parasites in the combinations with BZ and LEV, closantel
population are resistant. (CLS), and other MLs such as abamectin,
and moxidectin should be considered.
It is now common for sheep to harbour at
least one parasite species that is resistant Since 2002 half-dose ivermectin and 1/3-
to one of the major drench groups. It is dose CLS have been recommended for the
becoming disturbingly common for worms early detection of emerging resistance. Use
to be resistant to anthelmintics in all of the of special formulations of triple or quadruple
major drench groups. combinations (Triton™ and Q-drench™,
respectively) should be included if possible.
Drench resistance is common and often
highly developed in goats. Only a limited Groups may be included where treatments
range of anthelmintics are registered for are given at greater than the recommended
use in goats. Due to their altered drug dose rates to fully evaluate the available
metabolism goats may require higher dose anthelmintics. In practice, only double dose
rates than recommended for sheep. LEV or double dose LEV plus single BZ
have been shown to be effective where the
For more information see: single dose is not effective. However, it is
http://www.agric.nsw.gov.au/reader/das- emphasized that the use of double dose
vettesting/2566 rate or combinations of anthelmintics are
not general recommendations; they are
Diagnosis used to obtain a prompt answer to possibly
History, failure to respond to treatment, in a serious and immediate problem on
vitro larval development assay (LDA, specific farms.
DrenchRite™), or on-farm trial using a
faecal egg count reduction test (FECRT). FECRT pre-trial check sampling (Day -
10)

VETERINARY LABORATORY MANUAL 34


To avoid wasted effort, it is of value to All trial sheep should be sampled between
arrange for a pre-trial collection of faeces to 10 and 14 days after treatment. Faecal
obtain an estimate of the egg count of the samples should be collected from the
flock. If this is not possible, faecal samples rectum of sheep within the first hour after
may be collected from the control group at yarding and before much handling has
day 0 to assess if egg content in faeces is occurred. A minimum of 5 g (10 to 15
sufficient to continue the trial. Objective pellets) should be collected from each
assessment of the efficacy of an sheep.
anthelmintic is not reliable where the mean
faecal egg count of the control group is less Transportation of faeces
than 200 epg. Some authorities even Samples in individually sealed containers
suggest 300 epg. Subjective evaluation of filled as close to the top as possible to
the data may however be possible below exclude air, should be transported to the
this figure. laboratory as soon as possible. The
samples should be kept cool in an esky with
FECRT treatment (Day 0) freezer bricks wrapped in newspaper to
Selection of trial animals avoid freezing. Storage or transport of
Trial sheep should be preferably be young faeces at or below 4oC may affect the
sheep at weaning, 3-6 months of age, of hatching of H. contortus eggs in a faecal
even body weight and bred on the property. culture.
Undrenched weaners are preferred, but if
not possible they should NOT have FECRT laboratory testing
received a drench during the previous 6 Larval differentiation
weeks or 8-12 weeks (for persistent ML Pooled faecal cultures and larval
anthelmintics) to avoid testing an already differentiations should always be carried
‘selected’ worm population. out on the control group and preferably
each of the test groups. This will allow
Number of trial animals determination of the species composition
Each group should contain 15 sheep and species resistance levels. This is
allowing that faeces may not be collected important as some species have different
from all animals at the subsequent visit. (A susceptibility to different drenches (eg
minimum of 10 sheep per group from which Levamisole may still be effective as a
collections are made is necessary). narrow spectrum against Haemonchus, but
not as a broad spectrum against other
Randomisation of trial animals species).
Sheep should be allocated to treatment and
control groups randomly. The best method Calculation of FECR results
of randomisation is based on pretreatment The arithmetic mean (‘average’) on epg of
egg counts; however, as these are usually samples collected 10-14 days after
not available, the sheep should be drafted treatment, together with a 95% confidence
into as many groups as there are interval, most accurately describes the
anthelmintics to be tested plus one range of in efficacy of an anthelmintic. The
untreated control group. For example, if 4 arithmetic mean is the most stringent test of
drenches are to be tested, the first 5 sheep a percentage reduction in faecal egg output
in the race are allocated at random to 5 and therefore is a more conservative
groups; the next 5 are similarly allocated measure of an anthelmintic's performance.
until each group contains 15 sheep. Each Geometric means are sometimes requested
group should be identified with a different by trial sponsors, but arithmetic calculations
colour marker (coloured ear tag is often the should always be conducted as well.
most convenient).
Each set of results will include the following
Anthelmintic treatment of trial animals information per group for each genus
Anthelmintics should be administered present (provided a larval differentiation has
accurately by either a calibrated syringe or been conducted):
a drench gun previously calibrated. In most • Arithmetic mean (‘Average’)
situations, sheep should be orally dosed • Range (Highest and lowest counts)
according to the weight of the heaviest • Standard deviation
animal in the group. If the group is an even • Mean percent reduction in epg (i.e. =
line of sheep the dose received on a mg/kg FECR)
basis will not vary greatly. • 95% confidence interval of FECR
(Upper and lower limit of percent
FECRT sampling (Day 10-14) reduction for 95% confidence interval)
Collection of faeces

VETERINARY LABORATORY MANUAL 35


Interpretation of FECRT results ii. the lower 95% confidence limit of the
Resistance should be regarded as being % reduction in egg count is less
present if: than or equal to 90%.
i. the FECR is < 95%, and,

An example of simple test groups without larval differentiation:


Control BZ Levam BZ+Levam
Arithmetic mean 155 76 190 23
Highest 520 160 800 80
Lowest 40 0 0 0
Std Dev 168 72 287 31
% Reduction * 51 *0 * 85
Upper 95% conf. 82 69 96
Lower 95% conf. *0 *0 * 43

Comment: Interpret these results with caution due to low egg counts in the controls. However it
appears that:
i. BZ resistance(*) is present.
ii. LEV resistance (*) is present.
iii. BZ + LEV resistance (*) is present.

ANTHRAX ii. From animals other than sheep and


cattle, smears from any affected
Anthrax is a notifiable disease. Fees for organs, e.g. swellings in throat and
tests undertaken to confirm or exclude a lymph nodes of pigs.
diagnosis of anthrax are paid by NSW
Department of Primary Industries. NB Always clearly label specimens as
‘Suspect Anthrax’
Caution Anthrax is a zoonotic
disease, causing serious illness and Pack securely and forward separately from
sometimes death in humans. other specimens
Where anthrax is suspected, a post mortem Advise the laboratory that you are
should not be undertaken as it will cause submitting the samples and place a warning
the vegetative forms of Bacillus anthracis to under the lid of the outer packaging. This
sporulate. These spores will remain a will ensure special biosecurity precautions
source of infection for long periods. are undertaken at the laboratory
A diagnosis of anthrax can be made from
ARSENIC POISONING
smears of peripheral blood obtained by
cutting the ear of the unopened carcase. Diagnosis
Where anthrax is suspected as a result of History; clinical and post mortem findings,
changes seen at necropsy then impression with gastritis or enteritis; detection of a
smears of tissue should be submitted in significant concentration of arsenic in the
addition to a range of fresh and fixed tissue ingesta or tissues of the dead animal. In
to establish an alternative diagnosis if chronically exposed animals, detection of
necessary. arsenic in milk or urine.

Diagnosis Specimens required


Demonstration of Bacillus anthracis in blood Each specimen should be packed in a
and tissues. separate container, clearly labelled and
submitted chilled.
Specimens required i. 250 g ruminal or stomach contents
i. Blood smears from peripheral blood submitted chilled for toxicology
vessels. Smears should be sent (Reinsch test).
from more than one animal. ii. 50 g of liver from the dead animal,
• Make thick, air-dried blood smears, submitted chilled for toxicology.
leaving one end of the slide clean. NB Arsenic analysis of tissue is
• Fix blood smears with methanol not routinely undertaken, due to the
prior to submission. costs. In most cases, a positive
• Do NOT fix smears by heat or Reinsch test (indicates >1mg
other agents. arsenic/kg) on rumen contents from
animals with a suggestive history and

VETERINARY LABORATORY MANUAL 36


lesions is adequate qualitative arsenic poisoning has been
confirmation. confirmed in the animal.
iii. 50 ml of milk or urine for arsenic v. Specimens required to allow for a
analysis (only where chronic differential diagnosis if arsenic is not
exposure is suspected). the cause.
iv. 50 g of suspect source material for
toxicology, to be examined only if

Interpretation of tissue and secreta arsenic concentrations


Analyte Sample Units Normal Possibly toxic Toxic
Arsenic Liver, kidney mg/kg (wet wt) < 0.5 0.5-8 > 8*
Arsenic Milk mg/kg (wet wt) < 0.25 0.25-1.5 > 1.5
Arsenic Urine mg/kg (wet wt) < 0.25 2-100

Interpretation depends on the interval between exposure and sampling. Maximum concentrations
of arsenic in tissues occur about eight hours after ingestion, and animals that survive for 2 to 3
days may have liver levels as low as 2 mg/kg. Conversely, clinically normal stock, chronically
exposed to arsenic (eg via arsenic contaminated feed over a long period or, in the past stock
regularly treated in arsenical dips) may have liver levels up to 8mg/kg and milk levels to 1.5 mg/kg.

ARTHRITIS AND POLYARTHRITIS ii. Serum sample from the dam,


submitted chilled for serology.
Diagnosis iii. Brain and spinal cord in buffered
Based on clinical signs, bacteriological, formalin for histopathology.
pathological and serological examination. It NB. Arthrogrypotic animals may
is often difficult to demonstrate the have CNS lesions only in the spinal
presence of organisms in chronic lesions. cord.
iv. Sections of extensor muscles in
Specimens required buffered formalin for histopathology.
i. Aseptically collected joint fluid,
chilled.
ASPERGILLOSIS
ii. Affected, unopened joints, with the
joint capsule intact, submitted See: Fungal infections'
chilled.
iii. Acute and convalescent serum ATAXIA
samples from affected sheep or
cattle, submitted chilled for Diagnosis of causes of 'ataxia' requires a
chlamydial serology. careful clinical examination to determine
whether the condition is due to a nervous,
NB There is no routine serological test muscular, skeletal, circulatory or respiratory
available for Erysipelas. defect.

Diagnosis
ARTHROGRYPOSIS AND
Based on clinical examination supported by
HYDRANENCEPHALY history and laboratory examination to define
Causes include infections of the dam with the specific cause.
Akabane virus, Aino virus, Palyam virus
and Pestivirus (Mucosal disease and Specimens required
Border disease), as well as inherited i. Brain, spinal cord and peripheral
defects. nerves in buffered formalin for
histopathology.
Diagnosis ii. Sections of skeletal and heart
Based on clinical signs and gross muscle in buffered formalin for
pathology. Diagnosis of the cause depends histopathology.
on history and samples from the dam and iii. Sections of lung and other organs
affected animals. with lesions, in buffered formalin for
histopathology.
Specimens required iv. At least 3 ml of serum, free of cells
i. Sample of foetal pericardial or and haemolysis, submitted chilled
pleural fluid submitted chilled for for serum enzymology and
virus isolation and serology, or, a biochemistry.
blood sample from the neonatal
animal if it has not sucked.

VETERINARY LABORATORY MANUAL 37


v. 30 g of liver and kidney, submitted
chilled in separate copper free 'BIG KNEE' IN GOATS
containers for copper analysis.
See: Caprine arthritis-encephalitis (CAE)'
ATROPHIC RHINITIS OF SWINE
BLACK DISEASE
Syn: Enzootic/progressive atrophic rhinitis
Clostridium novyi is present soon after
Diagnosis death in a large proportion of livers of
Clinical signs, gross pathology, normal sheep and cattle, so that
bacteriology. Can be diagnosed clinically demonstration of the organism is not
when large numbers of pigs with snout always diagnostic.
deformities are present. If bacteriological
confirmation of toxigenic P multocida is Diagnosis
needed, collect nasal swabs from up to 10 Demonstration and recovery of Cl novyi
clinically affected pigs (preferably weaners from necrotic foci; histopathology. Often
and growers). Despatch swabs in transport associated with the presence of immature
media to reach the laboratory within 24 liver fluke, Fasciola hepatica.
hours of collection.
Specimens required
If possible: i. At least three (3) impression smears
• Swab nares with alcohol to reduce from cut surface of lesion for
contamination. staining with specific fluorescent
• Use mini tipped, aluminium-shafted antisera.
swabs, especially when sampling ii. Portion of liver, submitted chilled for
weaner and grower pigs. bacteriology, taken from a freshly
dead animal.
• Advise RVL of intended sampling, so
iii. Portion of liver containing lesions, in
fresh selective media can be available
buffered formalin for histopathology.
in the laboratory.

Specimens required BLACKLEG


i. Snout swabs transported at ambient Diagnosis
temperature in Amies Charcoal Must be based on a careful post mortem
transport medium. examination with demonstration of either
Clostridium chauvoei or Clostridium
BABESIOSIS septicum in the lesions of haemorrhagic
myositis.
See: Tick fever
Specimens required
BALANO POSTHITIS IN RAMS These must be taken from a freshly dead
Occurs more commonly in the Border animals:
Leicester breed, but also reported in Dorset i. At least three (3) smears of
Horn and other breeds. Usually associated haemorrhagic muscle exudate for
with vulvitis in ewes. bacteriology.
ii. Portion of affected muscle, collected
Diagnosis aseptically, and submitted chilled for
Clinical findings, bacteriology in early cases bacteriology.
to assist in attempting to determine iii. Portion of affected muscle in
aetiology. buffered formalin for histopathology.

Specimens required BLOAT


i. Swabs from the lesion submitted in
Diagnosis
Amies transport medium for
Clinical examination, post mortem findings,
bacteriology.
including injection of eyes, protrusion of
ii. Scabs from the lesion submitted
tongue and numerous large haematomata
chilled for bacteriology.
in the frontal sinuses.
BENIGN FOOTROT Specimens required
See: Footrot Bloat must be diagnosed on history and
gross pathology.
BETA MANNOSIDOSIS OF GOATS
See: Mannosidosis

VETERINARY LABORATORY MANUAL 38


Specimens can be taken to eliminate other demonstration of the toxin in serum, liver
causes of sudden death, e.g. and gut contents.
enterotoxaemia, hypomagnesaemia.
Specimens required
BLUE-GREEN ALGAL POISONING i. Frozen serum, liver, stomach
contents, small intestinal contents
The toxic blue green algae, Microcystis and large intestinal contents for
cyanea and Anabaena circinalis are mainly toxin ELISA.
a problem in late summer and autumn. ii. Suspect food material (approx 30g)
for toxin ELISA.
Diagnosis
Presence of algae as a green scum on the
BOVINE LEUKOSIS
water. The clinical syndrome includes
sudden death or jaundice and See Enzootic bovine leukosis'
photosensitisation. Evidence of a
hepatotoxin and gastrointestinal irritation. BOVINE LEUCOCYTE ADHESION
DEFICIENCY (BLAD)
Specimens required
i. 500 ml of scum and water for (Holstein/Friesian calves)
examination for toxic algae.
ii. Serum from affected animals for This defect was disseminated throughout
serum chemistry/enzymology the international Holstein herd from a
iii. Sections of liver and kidney in founder bull in the USA whose descendants
buffered formalin. have been used extensively in the
international Holstein herd. The disease is
BORDER DISEASE IN SHEEP characterised by progressive leucocytosis
emanating from a deficiency of an adhesion
Diagnosis factor on the surface of leucocytes that is
Clinical signs including abortion and the essential for leucocytes to migrate from
birth of lambs with abnormally hairy birth blood vessels to sites of infection.
coats, which may or may not show nervous Consequently, affected calves suffer with
signs. Demonstration of Pestivirus in repeated bacterial infections.
affected lambs and antibody in the ewe.
AI centres select against BLAD
Specimens required heterozygotes when screening bulls for
i. Affected lambs submitted whole entry to their centres.
foetal membranes are also required
if it is an anteparturient or parturient Specimens required
death. See
ii. Blood sample, submitted chilled on http://www.dpi.nsw.gov.au/agriculture/vetm
the clot, from the affected lamb and anual/specimens-by-disease-
its mother for serology. syndrome/diseases_of_livestock/dpi-samp-
iii. Heparinised blood sample for coll-prot.pdf
antigen ELISA.
iv. Portions of fresh lung, spleen and For diagnosis of the disease and for
mesenteric lymph node collected heterozygote detection:
aseptically and kept chilled for
antigen ELISA or virus isolation. i. 20 to 25 hairs, with roots attached,
v. Brain and spinal cord, fixed in from the distal end of the tail.
buffered formalin for histopathology.
BOVINE MALIGNANT CATARRH
BOTULISM
Syn: Malignant catarrhal fever
There are five (5) major types of botulism
(A to E). Types C and D are the usual Diagnosis
causes of botulism in cattle; type C in Sporadic incidence, clinical signs,
horses and wild birds. histopathology, PCR for ovine herpesvirus-
2 (OHV-2).
Diagnosis
Usually based on clinical signs, which Specimens required
include flaccid paralysis, and the absence i. Brain and sections of sections of
of gross and histopathological lesions. liver, kidney, adrenal gland, bladder
Important cases may be confirmed by the and nasal mucosa in buffered
formalin for histopathology.

VETERINARY LABORATORY MANUAL 39


ii. 5 ml of EDTA blood for PCR testing Specimens required
for ovine herpesvirus-2 (OHV-2) Cows
iii. 5 ml of blood on clot submitted i. Uterine discharges or vaginal
chilled for examination for pestivirus mucus, submitted chilled for
(mucosal disease). In dead animals, bacteriology.
alternatively submit spleen and ii. Milk samples from the lactating cow,
kidney chilled for pestivirus. submitted chilled for bacteriology
iv. Ocular and nasal swabs submitted and serology.
in phosphate buffered gelatin saline iii. 2 ml of serum for serology. Samples
(PBGS) for examination for can be submitted on the clot if they
infectious bovine rhinotracheitis will be tested within 48 hours of
(IBR). collection.

BOVINE VIRUS DIARRHOEA Bulls


i. 0.5 to 1 ml of semen collected
See: Pestivirus aseptically, and submitted chilled for
bacteriology.
BRACKEN FERN POISONING ii. 2 ml of serum, submitted chilled for
serology.
See also: Enzootic haematuria of cattle

In ruminants, principally a disease of cattle, Aborted foetuses


iii. Specimens as required for diagnosis
affecting the haematopoietic system and
of abortion in cattle (see Abortion in
bladder wall. In adult sheep, long term
cattle).
intake of bracken fern by adult sheep can
induce "bright blindness", a progressive
retinal degeneration. BRUCELLOSIS OVINE
Diagnosis
In horses, pigs and other monogastrics Characterised by infertility in rams due to
rarely encountered, but associated with epididymitis. Abortion in ewes is not a
nervous signs from thiamine deficiency common finding.
attributed to a thiaminase in the plant.
The diagnosis in rams must be based on
Diagnosis the result of a clinical examination and
Clinical signs, haematology, histopathology. serology, supported in some cases by
There is no specific diagnostic test in the bacteriological examination of semen.
live animal for bracken fern poisoning.
Specimens should be submitted to Specimens required
eliminate other diseases. i. Serum samples submitted chilled for
serological testing. Samples can be
Specimens required (for cattle) submitted on the clot if they will be
Live animal tested within 48 hours of collection.
i. 5 ml of blood in EDTA for ii. Semen samples collected
haematology. aseptically and submitted chilled for
ii. Blood films for haematology. bacteriology.
iii. Serum sample, chilled for iii. Testes, epididymes and accessory
phosphorus estimation (see sex glands, including ampullae and
Hyophosphataemia). seminal vesicles, submitted chilled
for bacteriology and histopathology.
Dead animal
i. Sections of liver, kidney, red femoral
BRUCELLOSIS PORCINE
shaft bone marrow and affected
tissues in buffered formalin for Brucella suis infection was last diagnosed
histopathology. in NSW in 1968.
ii. From cases of haematuria, sections
of bladder wall with lesions, Diagnosis
submitted in buffered formalin, for History and serology. The agglutination test
histopathology. is of use mainly as a herd test, as there is a
tendency for infected pigs to cease to react
BRUCELLOSIS BOVINE although they still remain infected and false
positives can also occur.
NSW reached Bovine Brucellosis free
status in 1992. Specimens required

VETERINARY LABORATORY MANUAL 40


i. Serum samples (not blood) i. Fresh, chilled small intestine for
submitted chilled for serology (SAT bacteriology.
and RBT). ii. Formalin-fixed small intestine for
ii. Lesions, submitted chilled for histopathology.
bacteriology.
iii. Aborted foetuses submitted whole CAMPYLOBACTERIOSIS OF
for bacteriology CATTLE
CALCULI Syn: Bovine venereal campylobacteriosis
(BVC); formerly bovine vibriosis
Diagnosis
Evidence of uraemia/urinary retention, Bovine venereal campylobacteriosis (BVC)
cystitis and/or swelling and cavitation of is caused by Campylobacter fetus subsp
kidney pelvis with pressure effects on venerealis and is characterised by infertility
cortex and medulla. Demonstration of and early embryonic death. Abortion occurs
calculus at critical location. Chemical in a small percentage of infected cows.
analysis of the calculus may indicate the
cause of the calculus formation. NSW Campylobacter fetus subsp fetus (formerly
Department of Primary Industries subsp intestinalis) also causes abortion in
outsources this testing. cattle.

Specimens required Diagnosis


i. Diagnosis should be made on clinical History of reduced breeding efficiency in
and post mortem findings. herds where natural breeding is practised.
ii. Calculi for chemical analysis. Clinical signs commonly observed are
repeated returns to service, with prolonged
CAMPYLOBACTER ABORTION OF interservice intervals and sporadic
SHEEP occurrence of mid to late term abortions.
Usually attributed to Campylobacter fetus Abortion and infertility due to C fetus subsp
subsp fetus (formerly subsp intestinalis); venerealis can be confirmed by
occasionally Campylobacter jejuni demonstration of specific antibodies in the
implicated. vaginal mucus of affected cows by ELISA.
C fetus subsp venerealis can also be
Diagnosis isolated from the prepuce of infected bulls.
Late term abortions or birth of weak or dead In a herd situation, demonstration of
full term lambs. Typical liver lesions in antibody in vaginal mucus is the preferred
some full term lambs. Bacteriology. method of diagnosis as it can be difficult to
recover the organism from the prepuce.
Specimens required
Specimens as required for the diagnosis of C fetus subsp venerealis and C fetus subsp
abortion in sheep. fetus can be isolated from aborted foetuses.

CAMPYLOBACTER ENTERITIS Specimens required


Campylobacter jejuni and occasionally Cows
Campylobacter coli have been associated i. Vaginal mucus samples collected for
with enteritis and diarrhoea in calves, demonstration of C fetus subsp
lambs, pigs, dogs, poultry and man. venerealis IgA antibodies by ELISA
However, both may be found in otherwise test. PBST diluent and instructions
normal intestinal tracts of sheep and cattle for collection of vaginal mucus
(C jejuni) or pigs, poultry and man (C coli). samples for BVC ELISA test are
available from the laboratory. See
The role of other Campylobacters such as http://www.dpi.nsw.gov.au/agricultur
e/vetmanual/specimens-by-
C sputorum and C hyointestinalis in the
discipline/bacteriology/01-bvc-elisa-
aetiology of diarrhoea and intestinal
guide.pdf
pathology in pigs is uncertain (see Porcine
proliferative enteropathy).
ii. Abortion investigation: animals
which have aborted can be sampled
Diagnosis
from 1 week to 3 months after the
Bacteriology, histopathology.
abortion.
Specimens required

VETERINARY LABORATORY MANUAL 41


iii. Infertility investigation:
representative sampling of herd; at Affected calves can be identified at birth by
least 10 vaginal mucus samples their distinctive woolly haircoat, (also
collected from infertile heifers or described as ‘fuzzy’, ‘wirey’ or ‘curly’).
cows. This can be done when Some CWH calves have protruding eyes
pregnancy testing reveals infertility. and keratoconjunctivitis (‘pinkeye’). Cardiac
arrhythmias are usually detectible.
Bulls
i. Preputial scrapings submitted in Death from heart failure occurs between
Campylobacter transport enrichment birth and 12 weeks of age. Most calves with
medium (TEM) for bacteriology. CWH die suddenly. Owners occasionally
TEM is supplied from the laboratory see an affected calf collapse and die,
together with instructions for usually following exertion Some CWH
collection of samples. See calves show clinical signs of progressive
http://www.dpi.nsw.gov.au/agricultur heart failure.
e/vetmanual/specimens-by-
discipline/bacteriology/02-bull-vd- Diagnosis
coll.pdf Clinical signs, history. and gross cardiac
ventricular fibrosis, with or without
Aborted foetuses mineralisation. Microscopic examination is
i. Specimens as required for diagnosis necessary to accurately demonstrate the
of abortion in cattle (see Abortion in extent of the heart muscle changes.
cattle).
CWH cases can be misdiagnosed as
CAMPYLOBACTERIOSIS OF PIGS selenium deficiency (cardiac white muscle
disease, nutritional myopathy), particularly if
See: Porcine proliferative enteropathy; the woolly haircoat is overlooked. Calves
Campylobacter enteritis' with CWH are not selenium-deficient.

CAPRINE ARTHRITIS Any woolly-coated Poll Hereford calf that


ENCEPHALITIS (CAE) dies during the first few months of life
should have a postmortem examination to
Syn: ‘Big knees’, Caprine retrovirus check for evidence of cardiomyopathy. It is
infection not sufficient to diagnose CWH in a woolly-
coated calf without postmortem
Diagnosis confirmation of the cardiomyopathy.
Viral serology; clinical history of nervous
signs in young kids, big knees in adults Specimens required
associated with lameness, ill thrift, weight i. Whole heart, eyes, liver, kidney and
loss, chronic progressive pneumonia, hard skeletal muscle (including
udder and mastitis, histopathology of diaphragm, tongue, psoas muscle)
affected tissues. in buffered formalin.
ii. Fresh liver and kidney for selenium
Specimens required estimation.
i. Serum samples chilled for viral iii. 2 x 10 ml of EDTA blood (1.5 mg
serology. EDTA/ml of blood) for DNA isolation
ii. Portions of affected tissues in (from affected calf, dam and sire).
buffered formalin for histopathology These DNA samples will be held
(carpal joint, brain, spinal cord, pending development of a
lung). diagnostic DNA test (not yet
available).
CARDIOMYOPATHY AND WOOLLY iv. 1 x 10 ml lithium heparin blood for
HAIRCOAT (CWH) SYNDROME glutathione peroxidase (GSHPx).
(Poll Hereford calves)
CASEOUS LYMPHADENITIS (CLA)
Dd: Cardiac white muscle disease Diagnosis
(nutritional myopathy, selenium deficiency). Recovery of Corynebacterium
pseudotuberculosis from lesions.
CWH is a congenital genetic disease, with
an autosomal recessive mode of Specimens required
inheritance presumed to involve defective i. Smears from early lesions or from
structural protein(s) within desmosomal the periphery of active lesions for
complexes. bacteriology.

VETERINARY LABORATORY MANUAL 42


ii. Portions of tissues containing early
lesions or from the periphery of Diagnosis
active lesions, submitted chilled for Clinical signs, histopathology, confirmed by
bacteriology. DNA analysis,
iii. Portions of tissue containing early
lesions or from the periphery of Specimens required
active lesions, in buffered formalin See
for histopathology. http://www.dpi.nsw.gov.au/agriculture/vetm
anual/specimens-by-disease-
CHLAMYDIAL INFECTIONS syndrome/diseases_of_livestock/dpi-samp-
coll-prot.pdf
Syn: Chlamydiosis, sporadic bovine
encephalomyelitis (SBE), chlamydial (In order of preference)
arthritis, chlamydial abortion of sheep,
avian chlamydiosis Diagnosis of the disease
i. 20 to 25 hairs, with roots attached,
Caution Chlamydiosis is a zoonotic from the distal end of the tail.
disease, causing serious illness and ii. Whole brain in buffered formalin for
sometimes death in humans. histopathology.
Diagnosis Heterozygote detection
Based on clinical findings with i. 20 to 25 hairs, with roots attached,
demonstration of group specific antibody from the distal end of the tail.
rise between 'acute' and 'convalescent'
serum samples in association with disease.
CLOVER DISEASE
Demonstration of Chlamydia by FAT on
smears of fresh tissues or fluids. Diagnosis
Based on flock history, reproductive
Specimens required performance of older ewes in the flock and
i. Serum samples taken in the acute demonstration of cystic endometritis.
and convalescent stages of disease
(3 to 4 weeks apart) from individually Specimens required
identified and observed animals, i. Reproductive tracts including
submitted chilled for chlamydial ovaries from 15 to 20 ewes which
serology. did not lamb at the previous
ii. Depending on the clinical condition, lambing. Submit chilled, as it is often
specimens as required for the difficult to determine endometrial
investigation of nervous disorders, cysts grossly in fixed tissue.
arthritis or abortion.
COBALT DEFICIENCY
Interpretation of Titres
The demonstration of a four fold rise in CFT In NSW, cobalt deficiency is confined to
antibody titres, within 3 to 4 weeks of the restricted geographical areas. Signs have
clinical episode, is considered significant. been seen in sheep and cattle.

Diagnosis
CITRULLINAEMIA
History of ill thrift, in young animals
(Holstein/Friesian calves) particularly after elimination of other likely
causes, e.g. parasitism, deficiency of other
Affected calves are clinically normal at birth, minerals, malnutrition. Response to cobalt
become depressed by 24 hours, develop supplements or injections of vitamin B12,
signs of neurological dysfunction by 48 preferably in young, weaned animals.
hours. Typical clinical signs are seemingly Evidence of a mild to moderate macrocytic
aimless wandering, tongue protrusion, anaemia associated with weepy eyes.
chewing on fixed objects, head pressing
and ultimately collapse and death between Specimens required
the third and fifth day of life. ii. 10 ml of blood, clotted in vacuum
The mutation responsible for this defect tubes, from each of 3 to 5 animals in
was imported into Australia by a US-born the affected group, submitted chilled
bull whose descendants were selected for for biochemistry.
high EBVs for butter fat content of milk. The iii. Specimens should be submitted to
mutation is now relatively rare in the eliminate other causes of ill thrift.
Australian Holstein herd due to selection
against heterozygotes by AI centres.

VETERINARY LABORATORY MANUAL 43


COCCIDIOSIS liver with multiple yellow or white nodular
lesions.
Occurs mainly in young sheep, cattle, goats
and pigs. Lesions occur in the mucosa of Diagnosis
the intestinal tract. Characterised in Gross pathology, histopathology.
ruminants by scouring, anaemia and ill Demonstration of Eimeria stiedae.
thrift. In pigs, it has been shown to be an
important cause of scouring in 10 21 day Specimens required
old suckers in some herds. i. Portion of affected liver in buffered
formalin for histopathology.
Diagnosis ii. Oocysts of E stiedae may be
Clinical signs, impression smears of demonstrable in the exudate of
intestine (acute), demonstration of large dilated bile ducts and gall bladder
numbers of coccidia in faeces (chronic),
and histopathology. In acute coccidiosis,
COLIBACILLOSIS
oocysts are often not yet present in faeces,
and impression smears of affected intestine See also: Oedema disease, scouring
and histopathology are recommended.
Diagnosis
In pigs, oocysts may be difficult to detect in Bacterial evidence of Escherichia coli
faeces during the diarrhoeal phase, and a enteritis, enterotoxaemia or septicaemia.
definitive diagnosis can be made by Demonstration of pathogenic strains of E
examination of small intestinal smears of a coli.
typically affected pig.
Specimens required
Specimens required i. Preferably the whole animal
i. Faecal sample for oocyst count. submitted chilled.
ii. In piglets, and other animals with ii. Sections of liver, kidney, jejunum,
acute coccidiosis, obtain 5-10 colon and mesenteric lymph node,
impression smears of ileum and submitted chilled, for bacteriology.
jejunum and air dry. iii. Sections of lung, liver, kidney, small
iii. Portions of intestines (e.g. ileum, intestine and mesenteric lymph node
jejunum) showing lesions in buffered submitted in buffered formalin for
formalin for histopathology. histopathology.
iv. In animals showing nervous signs,
COCCIDIOSIS – HEPATIC IN brain submitted in buffered formalin
RABBITS for histopathology.

Characterised by mortalities with gross


pathology typically revealing an enlarged

E coli serotypes commonly associated with colibacillosis in pigs in NSW


0149:K91,K88ac: 0141:K85ac: 0141:K85ab: 08:K87,K88ac:
H10 H4 or HNM H4 or HNM H19 or HNM
Neonatal mortality + +
Neonatal diarrhoea + +
Weaner mortality + + + +
Weaner diarrhoea + + + +
Weaner nervous signs + +
Weaner oedema disease + +

POULTRY disease is characterised by shortening of


There is no real information on the the cervical and thoracic vertebra and
prevalence of pathogenic serogroups in symmetrical arthrogryposis in the fore and
poultry in Australia. Typing of poultry clinical sometimes rear legs.
isolates is not being undertaken at this time.
The mutation responsible for this defect has
COMPLEX VETERBRAL been traced to a founder US Holstein sire
MALFORMATION (CVM) whose descendants have been used
extensively worldwide. AI centres select
Affected calves are generally aborted, but against CVM heterozygotes when
in rare instances may be born alive. The screening bulls for entry to their centres.

VETERINARY LABORATORY MANUAL 44


refrigeration in Amies charcoal transport
Specimens required medium for bacteriology:
See
http://www.dpi.nsw.gov.au/agriculture/vetm i. Stallions 3 swabs per animal, from
anual/specimens-by-disease- the following:
syndrome/diseases_of_livestock/dpi-samp- • prepuce and base of penis
coll-prot.pdf (smegma)
• urethral fossa and urethral sinus
Diagnosis of the disease and • terminal urethra
heterozygote detection
i. 20 to 25 hairs with roots attached. ii.Non-pregnant mares - 3 swabs per
From the distal end of the tail animal, from the following sites:
• cervix
CONGENITAL ABNORMALITIES • clitoral sinus
See also Akabane, abortion, arthrogryposis • clitoral fossa
and hydranencephaly, pestivirus, genetic
diseases. iii. Pregnant mares - 2 swabs per
animal from the following sites:
In general, the occasional abnormality or • clitoral sinus
freak is of little interest and there is no • clitoral fossa
benefit in submitting such cases.
However, increased incidence of congenital CONTAGIOUS OPHTHALMIA
abnormalities or repeated occurrence over
See: Ophthalmia
a number of years warrant further
investigation. Advice should be sought from
the laboratory in these cases. CONTAGIOUS PUSTULAR
DERMATITIS
Diagnosis Syn: Scabby mouth, contagious ecthyma,
Clinical signs. It is important to eliminate orf
infectious causes before incriminating
environmental, chemical or other Diagnosis
teratogenic causes or inherited defects. A Clinical signs, demonstration of virus
detailed history, particularly in relation to particles on electron microscopy.
events during the first half of pregnancy Differentiate from Staphylococcal infections
may be helpful. The breeding history of the and Dermatophilosis.
animals may be important in some cases.
Specimens required
Specimens required i. Scabs with underlying tissue
i. Affected animals submitted whole, scrapings, submitted in a sealed
together with foetal membranes if container, e.g. a Macartney bottle,
available. for electron microscopy.
ii. Serum sample from mother of
affected animal, submitted chilled for
serology.
COPPER DEFICIENCY
iii. Body fluid (preferably pericardial or Diagnosis
pleural) from the foetus or serum History, clinical signs, serum and liver
sample from the neonate prior to copper levels, response to therapy.
sucking, submitted chilled for
serology and examination for Specimens required
immunoglobulins. Sample collection equipment must be free
of copper. Vacuum blood collection tubes
CONTAGIOUS EQUINE METRITIS are satisfactory for blood testing.
(CEM)
NB In all cases of suspected copper
Diagnosis deficiency, clotted blood is preferred to liver
Recovery of Taylorella (formerly or kidney.
Haemophilus) equigenitalis from the genital i. 10 ml of blood, clotted in vacuum
tract. tubes, from each of 3 to 5 animals in
the affected group, submitted chilled
Specimens required for biochemistry.
Swabs should be taken from each of the ii. Brain and spinal cord, in buffered
following sites and submitted without formalin, from cases of suspected
enzootic ataxia.

VETERINARY LABORATORY MANUAL 45


Interpretation of copper concentrations (cattle and sheep)
Analyte Sample Minimum Special Units Species Deficient Normal
amount requirement
Copper Serum or 1 ml OK on clot µmol/L Cattle < 7.5 7,5-16
hep plasma Sheep < 7.5 7.5-20

COPPER POISONING OF SHEEP Diagnosis


AND CATTLE Clinical signs, typical post mortem findings,
histopathology, analysis of tissues.
See also: Pyrrolizidine alkaloidosis'
NB. The diagnosis can be confirmed
Chronic copper poisoning in ruminants of histologically. Histochemical methods for
the hepatogenous and/or phytogenous form staining for copper in liver sections although
is quite common. The terminal acute phase not quantitative, are quicker, easier and
is characterised by liver necrosis, cheaper to perform than tissue analysis.
haemolytic anaemia, haemoglobinuria and
jaundice. Specimens required
Sheep of all ages and preruminant calves i. Sections of liver & kidney, in
accumulate copper in the liver when dietary buffered formalin for histopathology.
copper is in excess of 30 mg copper /kg dry ii. 30 g kidney and liver, in separate
weight of feed consumed (adult cattle copper free jars, submitted chilled
require and therefore tolerate 100 mg and unpreserved for copper
copper /kg of feed). Calf milk replacers estimation.
should contain <20 mg copper / kg.

Interpretation of copper concentrations (cattle and sheep)


Analyte Sample Minimum Special Units Species Deficient Normal Elevated
amount requirement
Copper Liver 30g Fresh mg/kg Cattle <4 20-70 > 100
(wet wt) Sheep <4 40-70 > 200
Copper Kidney 30g Fresh mg/kg Cattle, 4-6 >8
(wet wt) sheep

Toxicological results must be interpreted in associated with gastrointestinal or


relation to history, clinical findings and respiratory disease, affecting particularly
histopathology. See Toxicology for young animals. Ruminants, pigs, horses,
suggested normal and toxic values. Kidney mice, birds, and man are common hosts.
is considered superior to liver for Occasionally detected in young cats, dogs,
biochemistry in acute and chronic copper rabbits, guinea pigs and other mammals, as
poisoning: Kidney copper concentrations well as in reptiles and fish.
rise faster in acute toxicosis. Liver
concentrations of 200 mg/kg (wet wt) can Signs most commonly seen are anorexia
be found in sheep dying from other causes, and diarrhoea in calves, lambs, kids,
especially sheep affected with pyrrolizidine piglets, foals and deer calves. Disease is
alkaloidosis, which promotes liver copper spread by infectious oocysts, which are
accumulation. shed in faeces or respiratory secretions,
and have high environmental resistance.
CORYNEBACTERIUM EQUI IN Host debilitation and/or concurrent viral or
HORSES OR PIGS bacterial infection may predispose to
infection. Immuno-compromised or
See: Rhodococcus equi pneumonia in immuno-incompetent animals are at high
horses risk of infection.

See: Rhodococcus equi lymphadenitis in Diagnosis


pigs' Identification of oocysts in faeces, by
examination of faecal smears with acid-fast
CRYPTOSPORIDIOSIS stains or phase contrast microscopy of
faecal flotations. Identification of organisms
Cryptosporidium spp. are coccidial in the brush border of mucosal surfaces on
parasites that are not host specific, are not histopathology. Antigen capture ELISA on
susceptible to conventional anti-coccidial faeces of calves (where available), although
treatment and to date have no available
efficacious specific chemotherapy. They are

VETERINARY LABORATORY MANUAL 46


this technique may be less sensitive than Gross pathology and dissection of a viable
microscopic examination. scolex from the fluid filled cyst, or
histopathology of degenerate cysts.
Specimens required
i. Faecal sample for preparation of Specimens required
smears and examination by MZN or i. Fresh tissue with lesions, submitted
similar stain, or by floatation and chilled for parasitology.
examination by phase contrast ii. Lesions, in buffered formalin, for
microscopy. histopathology
ii. Formalin-fixed abomasum/stomach
(peptic gland epithelium) and small DEFICIENCY OF URIDINE
intestine from freshly-killed MONOPHOSPHATE SYNTHETASE
mammals, reptiles or fish.
iii. For birds, formalin-fixed small (DUMPS)
intestine, caecum, colon and bursa, (Holstein/Friesian cattle)
or epithelium of the upper
respiratory tract and lung from Deficiency of uridine monophosphate
freshly-killed cases. synthetase results in foetal death during the
first trimester of pregnancy. The mutation
CYANIDE POISONING responsible for the defect has been
recorded in North American Holsteins, in a
Syn: Prussic acid poisoning. sire line that, fortuitously, was not used
widely in the international Holstein herd. It
See Poisoning (plants) for information on is not known if exists in the Australian
collection of specimens for plant Holstein/Friesian herd.
identification.
Specimens required
Diagnosis See
Clinical signs, post mortem appearance of http://www.dpi.nsw.gov.au/agriculture/vetm
blood and mucosae, odour of bitter anual/specimens-by-disease-
almonds, access to suspected materials or syndrome/diseases_of_livestock/dpi-samp-
fodders, chemical demonstration of cyanide coll-prot.pdf
in plant material or rumen ingesta.
Heterozygote detection
Specimens required i. 20 to 25 hairs, with roots attached,
Plant specimens can often be identified by from the distal end of the tail.
the local District Agronomist.
DERMATOPHILOSIS
Cyanogenetic glycosides in plants
hydrolyse during transit so that laboratory Syn: Mycotic dermatitis, ‘lumpy wool’
analysis is often misleadingly negative.
Herbage and ingesta should be kept Diagnosis
frozen from the time of collection to the Gross appearance of the lesions and
time of analysis; chilling is not sufficient. demonstration of Dermatophilus
Immersion in 1.3% mercuric chloride also congolensis microscopically. The causal
stops hydrolysis. organism may be isolated from scrapings or
a biopsy specimen.
i. Sample of rumen ingesta in an
airtight container, submitted frozen Specimens required
for toxicology. i. Sample of wool or hair showing
ii. Blood smears to check for nitrate lesions or scabs for gross
poisoning. examination and demonstration of D
iii. Samples of suspected plant material congolensis in impression smears.
for identification and check for ii. Skin biopsy from the area with
cyanide and nitrate. Submit frozen lesions, submitted chilled for
for cyanide toxicology. bacteriology.
iv. Fresh and formalin-fixed specimens iii. Skin biopsy from the area with
to establish an alternative diagnosis lesions, in buffered formalin for
when necessary histopathology.

CYSTICERCOSIS DIARRHOEA
Diagnosis See Scouring

VETERINARY LABORATORY MANUAL 47


DRENCHING MORTALITIES Clinical History and Post-mortem Report
form.
Diagnosis
Based on history, clinical signs and in some Diagnosis
cases, histopathology. Clinical signs, histopathology. Virology and
bacteriology to establish an aetiological
Specimens required diagnosis.
Product information sheets from the
manufacturer should be checked to ensure Specimens required
the appropriate specimens are submitted. ii. Brain and portion of spinal cord, in
i. Sections of liver, kidney and lung, in buffered formalin for histopathology.
buffered formalin for histopathology. iii. Sample of cerebrospinal fluid
Brain and/or spinal cord if any collected aseptically, submitted
neurological signs evident. chilled for bacteriology.
ii. Other samples as required from the iv. Section of brain and spinal cord
product information sheet of the collected aseptically, submitted
manufacturer chilled for virology.
v. Serum samples for virus serology.
DWARFISM IN DEXTER CATTLE vi. Specimens as required for the
Syn: Dexter chondrodysplasia, congenital differential diagnosis of nervous
lethal chondrodysplasia (Dexter bulldog). disorders.

Congenital lethal chondrodysplasia (Dexter ENCEPHALOMYOCARDITIS (EMC)


bulldog) is the homozygous form of Dexter OF PIGS
chondrodysplasia. Affected calves are
Encephalomyocarditis is caused by a
usually aborted before the seventh month
picornavirus virus. The disease is a
of gestation, with extreme shortening of
zoonosis affecting a wide range of
limbs and vertebral column, gross
mammals including mice, wombats and
craniofacial defects (relatively large head
elephants.
with retruded muzzle, cleft palate and
protruding tongue), and a large abdominal
Diagnosis
hernia.
History of sudden death, typical white areas
on heart and throughout myocardium, virus
Specimens required
isolation. Associated with abortion in some
See
instances.
http://www.dpi.nsw.gov.au/agriculture/vetm
anual/specimens-by-disease-
Specimens required
syndrome/diseases_of_livestock/dpi-samp-
i. Fresh heart and spleen collected
coll-prot.pdf
separately and aseptically submitted
chilled for virology.
Heterozygote detection
ii. Sections of heart, liver, kidney, lung,
i. 20 to 25 hairs, with roots attached,
spleen and whole brain in buffered
from the distal end of the tail.
formalin for histopathology.
iii. Section of liver, kidney, spleen, lung
Please provide pedigree of the subject with
and heart blood submitted chilled for
the hair sample.
bacteriology.
ENCEPHALOMYELITIS AND
ENTEROTOXAEMIA
ENCEPHALITIS
Difficulty is often encountered in
See also Haemagglutinating encephalitis establishing a diagnosis of enterotoxaemia
virus (HEV) of pigs, porcine enterovirus in sheep, cattle, goats and deer. The
encephalomyelitis, transmissible following points must be considered:
spongiform encephalopathy (TSE)
• Clostridium perfringens is a normal
For adult sheep and cattle with bowel inhabitant and toxin may be
progressive neurological disease, present in the gut content of healthy
please submit specimens as required by animals.
the National TSE Surveillance Program • Cl perfringens is a common
(http://www.animalhealthaustralia.com.au/pr postmortem invader of parenchymal
ograms/adsp/adsp_home.cfm), tissues and will be present by 8 hours
accompanied by a completed NTSESP after death.

VETERINARY LABORATORY MANUAL 48


• Toxin may diffuse from the gut if the for glycosuria should be carried out
autopsy is delayed. In chronic cases, in the field.
e.g. in focal symmetrical
encephalomalacia, toxin is usually no NB Indicate vaccination history with all
longer present. submissions of suspected enterotoxaemia..
• Typical kidney lesions develop in sheep
within a few hours of death, they are ENZOOTIC ATAXIA
not present at the time of death.
Syn Copper deficiency'
• Type D toxin is stable in gut contents in
vitro without preservative, but other
toxins are very unstable and have ENZOOTIC BOVINE LEUKOSIS
disappeared within 12 hours regardless (EBL)
of preservation, cooling, etc.
A retrovirus infection of cattle which can be
spread by blood transmission from rectal
Diagnosis
palpation, vaccination, tattooing or gouge
A positive diagnosis rests on several of the
dehorning where repeatedly-used
following being present:
equipment is employed. Congenital
• History of sudden death transmission also occurs but is less
• Necropsy findings: important. The virus has been eliminated
• glycosuria from dairy herds in NSW but beef cattle
• pulmonary oedema and congestion remain infected at a low prevalence.
• fibrin clot in pericardial sac
• epicardial haemorrhages Like other retrovirus infections (AIDS, CAE,
• yellow creamy intestinal contents EIA) antibody indicates current (not just
NB: some animals may show minimal previous) infection. Affected animals usually
gross changes e.g. sudden death with develop antibody within 2-3 months of
some small intestinal inflammation. spread.
• histopathological lesions in brain
and kidney (e.g. demonstration of Diagnosis
protein leakage from blood vessels Clinical signs, confirmed by virus serology,
into perivascular spaces of histopathology and haematology.
cerebrum).
• abnormal numbers of Cl Specimens required
perfringens like organisms in i. Serum or preserved milk samples
smears of intestinal mucosa taken (bromopol preservation as used for
from multiple sites from abomasum milk cell counts) for virus serology
to colon, with particular attention to (ELISA or GDPT)
the ileum. ii. Formalin fixed sections of affected
• demonstration of epsilon toxin in lymph nodes, spleen, thymus,
gut contents by counter- intestine or other organs showing
immunoelectrophoresis (CIEP). gross lesions for histopathology.

Specimens required ENZOOTIC HAEMATURIA OF


i. At least 20 smears from the CATTLE
intestinal mucosa at sites from the
abomasum to the colon with See also Bracken fern poisoning
particular attention to the ileum.
Sites selected should be from areas Diagnosis
showing haemorrhages and also Chronic intermittent haematuria and
from nearby non inflamed areas anaemia. Gross pathology and
which may show a heavier flora. 4-5 histopathology of the urinary bladder.
sites can be smeared on one slide.
ii. Samples of small intestinal content Specimens required
at least 10 ml from sheep and 40 ml i. Sections of urinary bladder with
from cattle should be collected suspect lesions, submitted in
from several sites in the small buffered formalin for histopathology.
intestine, particularly from areas
where the contents are a yellow ENZOOTIC PNEUMONIA OF PIGS
colour and creamy consistency.
See Porcine enzootic pneumonia'
Submit chilled for toxicology.
iii. Sections of liver and kidney, and
whole brain in buffered formalin for
histopathology. Urine examination

VETERINARY LABORATORY MANUAL 49


EPERYTHROZOONOSIS OF SHEEP iii. Genital organs, including the
accessory sex glands, submitted
The taxonomy of the causative organism chilled for bacteriology and
has been revised and it is now known as histopathology.
Mycoplasma ovis.
EQUINE BABESIOSIS
Diagnosis
Demonstration of Mycoplasma ovis and This disease caused by the protozoon Babesia
anaemia in sheep. equi was introduced to Australia in infected
imported horses and transmitted locally by
The presence of the organism is insufficient poor injection technique. The disease probably
to establish the diagnosis as it can be found no longer exists in Australia.
in normal sheep. Other causes of anaemia
must be excluded. Diagnosis
Clinical signs of depression and anorexia
Specimens required with intermittent fever. Anaemia,
i. Blood films from 10 affected and at haemoglobinuria and subcutaneous
least 20 apparently normal animals oedema may be present. In chronic cases
in the flock for examination for M there may be loss of condition and mild
ovis (See Specimens for anaemia. Demonstration of the organism in
Haematology - preparation of blood thick and thin blood smears, serology.
films').
ii. Blood samples from affected Specimens required
animals in EDTA vacuum tubes for i. Thick blood smears dried under
haematology. cover (not by heat or exposure to
sunlight) for parasite examination.
EPHEMERAL FEVER ii. Blood films for haematology.
iii. 5 ml of blood in EDTA vacuum tube
Diagnosis for haematology.
General symptomatology and occurrence in iv. At least 2 ml of serum for serology
the herd and district. Virus antibody rise (B equi IFAT).
between acute and convalescent serum
samples.
EQUINE INFECTIOUS ANAEMIA
Specimens required (EIA)
i. Paired serum samples (with a 2 to 3 Diagnosis
week interval) from individually Clinical signs of recurrent febrile attacks
identified animals collected in the associated with anaemia over a period.
very early acute and convalescent Haematology, serology (AGID - Coggins
phases submitted chilled or frozen test) and histopathology. A positive AGID is
for serology. conclusive evidence of EIA; however, the
AGID may occasionally be negative early in
NB. Generally virus serology is not carried the disease course.
out unless paired serum samples are
submitted. Specimens required
i. At least 2 ml of serum for viral
EPIDIDYMITIS serology (AGID - Coggins test).
ii. 5 ml of blood in EDTA vacuum tube
See also Brucellosis-ovine, Actinobacillus
and blood smears for haematology.
seminis, Histophilus ovis
iii. Sections of liver, kidney, spleen and
lymph nodes in buffered formalin for
Diagnosis
histopathology.
Clinical signs and presence of lesions.
Recovery of causal organism from semen
or reproductive organs. Evidence of EQUINE VIRAL ARTERITIS (EVA)
infection with specific pathogens, e.g. See also Exotic diseases'
Brucella ovis, Actinobacillus seminis and
Histophilus ovis. Recent identification of seropositive horses
and a semen-culture positive stallion in
Specimens required Australia suggest this organism has been
i. Serum sample, submitted chilled for established in the horse population at a low
serology. level. Clinical disease has not yet been
ii. Semen sample collected aseptically, recorded.
submitted chilled for bacteriology.

VETERINARY LABORATORY MANUAL 50


EVA is usually subclinical, but clinical i. Portion of lesion(s) collected
disease overseas is characterised by aseptically and forwarded in sterile
pyrexia lasting 1-5 d, subcutaneous containers for bacteriology.
oedema of limbs and ventral abdomen, ii. Portion of lesion(s) in buffered
rhinitis and conjunctivitis with nasal and formalin for histopathology.
ocular discharge and photophobia, a skin
rash, anorexia, depression, dyspnoea and Septicaemia
diarrhoea. Up to 70% of pregnant mares i. Sections of liver, kidney, lung,
abort during the febrile stage. Affected spleen, collected aseptically and
animals usually recover. forwarded in separate sterile
containers for bacteriology.
Virus is spread by respiratory and venereal ii. Portions of above tissues in buffered
transmission. Virus is present in semen, formalin for histopathology.
respiratory secretions and also in urine, iii. From acute clinical cases,
faeces and aborted foetuses. sample of citrated blood collected
aseptically, submitted chilled for
Diagnosis bacteriology.
Serology, virus isolation, histopathology.
Abortion
Specimens required As this is associated with maternal pyrexia
i. Serum rather than foetal or placental infection,
ii. Blood containing anticoagulant, bacteriological examination of aborted
nasal swabs or washings in PBGS, foetuses is usually of little assistance in the
tissues of aborted foetuses (lung, diagnosis of Erysipelas abortion.
liver, spleen, thymus, lymph node),
semen for virus isolation. From adult EXOTIC DISEASES
horses at necropsy, fresh lung,
spleen and lymph nodes draining Guidelines on the collection and submission
oedematous areas. of specimens for exotic diseases can be
found in 'Exotic Diseases of Animals: A field
Send chilled unless delays of more than Guide for Australian Veterinarians'. Geering
24 h are anticipated in transit; if so, WA, Forman AJ and Nunn MJ (1999). This
forward frozen on dry ice. comprehensive book was published by
CSIRO Publishing and distributed to all
iii. Tissues from foetuses or adult Australian veterinarians.
horses as above, fixed in neutral-
buffered formalin for histopathology. On suspicion of an exotic disease,
veterinarians must immediately notify
NSW Department of Primary Industries;
ERYSIPELAS
the exotic disease hotline phone number
Erysipelothrix rhusiopathiae causes arthritis is 1800 675 888. The Officer manning this
(often post-marking) and post-dipping phone will advise on what actions need to
lameness (due to laminitis) in sheep and be taken. In some circumstances,
arthritis, urticaria, valvular endocarditis and veterinarians on a suspect property will be
fatal septicaemia in pigs. The resultant asked to remain until appropriate
pyrexia in sows can result in abortion. arrangements can be made to collect
samples and disinfect equipment and
Diagnosis samples off the property.
Clinical signs and history. Gross pathology.
Isolation of E rhusiopathiae from affected Ausvetplan is a comprehensive set of
tissues. There is no routine serological test manuals dealing with all aspects of exotic
at present available for Erysipelas. disease management in Australia. The
manuals are revised regularly and the
Specimens required current versions can be downloaded from
Arthritis the Animal Health Australia website at
i. Whole joint unopened and chilled. If http://www.animalhealthaustralia.com.au/.
this is not possible, forward
aseptically-collected synovial EXUDATIVE EPIDERMITIS IN PIGS
membrane (most desirable) and
synovial fluid (of lesser value) in Syn: Greasy pig disease
separate sterile containers.
Diagnosis
Urticaria and endocarditis Typical skin lesions on face and body.
Isolation of Staphylococcus hyicus.

VETERINARY LABORATORY MANUAL 51


detected in Holstein bulls in Australian
Specimens required artificial breeding centres.
i. Live or fresh chilled dead piglet for
bacteriology and histopathology. Diagnosis
ii. Smears and swabs from skin
lesions, submitted in Amies charcoal History of extended bleeding following
transport medium. dehorning, at parturition, or into milk at the
onset of lactation.
FACIAL ECZEMA
Specimens required
Diagnosis See
History of moist, humid weather conditions; http://www.dpi.nsw.gov.au/agriculture/vetm
abundant, high humus containing pasture; anual/specimens-by-disease-
clinical signs, gross pathology and syndrome/diseases_of_livestock/dpi-samp-
histopathology. Demonstration of numbers coll-prot.pdf
of spores of Pithomyces chartarum on
pasture. i. 20 to 25 hairs, with roots attached,
from the distal end of the tail.
Specimens required
i. Sections of liver and kidney in
FASCIOLIASIS IN SHEEP AND
buffered formalin for histopathology.
ii. At least 1 kg of pasture taken from CATTLE
the lower 5 to 7.5 cm of pasture See Liver fluke infection
growth and including the dead
leaves and leaf residue, submitted FISTULOUS WITHERS
chilled for fungal examination.
The aetiology is not fully known, but it has
FACTOR VIII DEFICIENCY often been associated with Brucella abortus
infection. Staphylococci and Streptococci
(Hereford cattle) are often present in the lesion.
Syn: Haemophilia A. Diagnosis
Clinical signs, serology.
A sex-linked inherited defect that usually
presents as a fatal hemorrhage event Specimens required
following castration. In one herd when a i. Serum sample from affected animal
“bleeder” bull was used over heterozygous for Brucella abortus serology (RBT
cows, some daughters were observed to and CFT).
bleed excessively when ear marked. It ii. Material from the depths of the
would be reasonable to expect dehorning lesion submitted chilled for
and parturition may also present life- bacteriology.
threatening challenges to homozygous
heifers. Dehorning would also present a
life-threatening challenge to bulls that FLEECE ROT
inherited the mutation from their clinically A dermatitis of the skin, caused either by
normal heterozygous dam. prolonged wetting of the skin or other skin
damage. The serous exudate results in
The condition has been recorded in matting of the wool. The wool may be
Hereford cattle in Australia, but may arise in discoloured due to the growth of
any breed due to new mutations. chromogenic bacteria.

Diagnosis Diagnosis
A history of severe haemorrhages in Clinical signs, bacteriological and
grandsons of a sire should raise a suspicion macroscopic examination of affected wool.
of haemophilia A.
Specimens required
FACTOR XI DEFICIENCY i. Sample of affected wool, submitted
for gross examination
(Holstein cattle) ii. Skin biopsy from affected area
submitted in buffered formalin for
An autosomal recessive defect reported in histopathology.
Holsteins in North America and Europe.
The mutation responsible for the defect has
been defined. The mutation has been

VETERINARY LABORATORY MANUAL 52


FOCAL SYMMETRICAL FOOTROT IN SHEEP
ENCEPHALOMALACIA (FSE) Includes virulent and benign footrot
A chronic form of enterotoxaemia.
For the purposes of the Stock Diseases
Diagnosis Act and the NSW Footrot Strategic Plan,
Clinical signs, histopathology of the brain. all forms of non-benign footrot are
treated as virulent footrot.
Specimens required
i. Brain in buffered formalin for For full discussion of footrot classification
histopathology. (including colour photographs) refer to
ii. Specimens as required to AgFact - Footrot in sheep and goats.
investigate nervous disorders in (http://www.agric.nsw.gov.au/reader/sheep-
sheep. footrot/a0956.pdf?). This may take a little
while to download.
FOOT ABSCESS IN SHEEP
Further information (Policy, AgFacts):
Including: Toe abscess, heel abscess http://www.agric.nsw.gov.au/reader/sheep-
(infectious bulbar necrosis) footrot

TOE ABSCESS A. VIRULENT FOOTROT IN SHEEP


Caused by a range of pyogenic bacteria, Caused by strains of Dichelobacter
usually involving the front feet and often nodosus (formerly Bacteroides nodosus)
associated with shelly toe. More common in with a range of virulence. Outbreaks of
sheep that have been run in wet conditions severe lameness can occur under wet
for some time. pasture conditions when the mean daily
temperature exceeds 9-10oC. Early lesions
The lesion involves soft tissues of the toe resemble benign footrot, but progress to
region, often tracking up to break out at the severe interdigital dermatitis and
coronet in front of the foot. Toe abscess underrunning of the sole and hard horn of
can also cause underrunning and break out axial and abaxial wall within 3-4 weeks.
near the heel, but the granulation Little or no pus present even in severe
associated with the initial infection will be lesions. The lesions will persist under dry
found near the toe. environmental conditions and can affect a
high percentage of the mob. Usually more
HEEL ABSCESS than one foot affected. Not all chronically-
Caused by infection with Fusobacterium infected sheep show lameness.
necrophorum in association with other
bacteria. Occurs in fat, heavy sheep Diagnosis
especially ewes in late pregnancy and History, clinical examination of a sufficient
rams. Outbreaks often seen 7-10 days after number of sheep in the flock (to detect
susceptible sheep have been run in wet, underrunning of the hard horn of the hoof
muddy conditions (wet muddy yards, and to be confident of the range of lesions
grazing cereal crops sown in ploughed present). Bacteriology (clinical diagnosis
ground, wet pastures). Can occur in may be reinforced by demonstration of D
younger sheep if they are put in wet, muddy nodosus-like organisms in smears).
areas.
Specimens required
Usually affects a hind foot, but if severely
challenged can affect 2 or 3 feet. Lesions i. Smears of affected feet lesions for D
seen as swelling and tenderness in the nodosus detection
affected claw with severe lameness, and
commonly affect the interphalangeal joint. Smears of affected feet can be used to
Suppurative lesions break out in the heel detect the presence of D nodosus, but are
area with discharging sinuses and unable to distinguish between virulent and
granulation tissue. Affected foot may be benign strains. In addition, false negative
deformed. smears can occur.

Diagnosis Smears are taken from an active site in the


History and clinical findings. lesion this is at the junction of healthy and
diseased tissue. Where underrunning is
Specimens required evident, the horn of the hoof may have to
None; best diagnosed in the field. be cut away to reveal the active sites.
When the diseased tissue is pared away, a

VETERINARY LABORATORY MANUAL 53


distinct band of active necrosis can be Usually affects a number of sheep in the
observed along the healthy/ infected flock, of all age groups.
border. This area is usually a couple of
millimetres wide and is characterised by a Can cause marked lameness, particularly in
greyish to yellow grey exudate. This rams and heavy sheep. Affected sheep
exudate should abound in D. nodosus and show necrosis and inflammation of the
is the material of choice for diagnostic interdigital skin, involving part or all of the
purposes. soft horn of the axial wall. A small
percentage may have some underrunning
Smears should be prepared by spreading a of the soft horn of the heel. The lesion
small amount of exudate thinly on a resolves quickly if sheep are moved to dry
microscope slide (using a swab stick, match pasture or if footbathed in formalin or
or point of a knife). ZnSO4. Cannot be differentiated from early
virulent footrot, but does not progress to
Smears may be prepared from the moist underrunning of the hard horn of the hoof.
necrotic interdigital skin if there is no
underrunning of the heel or sole. When Diagnosis
preparing the smear, avoid drawing blood, History, clinical examination of a sufficient
as this renders examination of the smear number of sheep in the flock. In some
more difficult. flocks, a second examination 10 to 14 days
later will be necessary to check for
NB progression of the disease, and differentiate
• Smears should be made from 5-10 from virulent footrot. Bacteriology.
affected animals.
• All slides should be numbered clearly The disease is best diagnosed in the field,
and serially, with a corresponding but in some cases, laboratory culture for
description of the animal/lesion on the virulence testing may be necessary.
specimen advice sheet. Use of a
diamond pencil for numbering is Specimens required
recommended, since legal action may i. See Virulent Footrot.
result from the examination findings. If ii. Smears of affected feet lesions for D
slides with frosted glass ends are nodosus detection
available, a dark lead pencil to identify iii. Hoof samples for culture (using
smears is also satisfactory. special footrot kit in field)
• The differential diagnosis of footrot
and foot abscess in sheep is best CLASSIFICATION OF DISEASE BASED
made on clinical examination, with ON CLINICAL EXAMINATION
laboratory confirmation where For full discussion of footrot classification
necessary. (including colour photographs) refer to
AgFact - Footrot in sheep and goats.
ii. Hoof samples for culture (using (http://www.agric.nsw.gov.au/reader/sheep-
special footrot kit in field) footrot/a0956.pdf?). This may take a little
while to download.
Culture of lesions and subsequent testing of
D nodosus isolates may assist the field In ovine footrot, a distinction between
veterinarian to differentiate between benign virulent and benign can be made provided
and early virulent footrot. Under certain there are no recent treatments or
conditions isolation and demonstration of environmental conditions which would mask
virulent strains of D nodosus in the the full expression of disease due to the
laboratory can be performed using a special infecting strain of D nodosus.
footrot kit for sample collection. In such
cases, the Regional Veterinary Laboratory Examination of sheep after a period of
should be contacted. spread (wet pasture conditions/mild
temperature) is more reliable to assess
B. BENIGN FOOTROT IN SHEEP flock status. However, additional factors will
Caused by infection with benign strains of D also influence disease spread and
nodosus. Seen in sheep grazing wet expression, including host resistance and
pastures under mild weather conditions. stocking rate.

VETERINARY LABORATORY MANUAL 54


FOOTROT SCORING SYSTEM
Score Description
0 normal
1 non-specific inflammation and/or necrosis of the interdigital skin (IDS)
2 inflammation of the IDS which is due to infection with footrot
3 any lesion in any claw which results in underrunning of the soft horn of the heels or
sole
4 underrunning of any hard horn of the claw

In all circumstances where lameness is sufficient number of affected sheep (score 2


occurring, it is recommended that sufficient or greater) to assist in the diagnosis.
sheep be examined to ensure that virulent Based on the examination of at least 100
footrot is not present. In most mobs, this will sheep, the prevalence of score 4 lesions
involve the examination of at least 100 relative to the number of footrot-affected
sheep. In some mobs, it may require sheep can then be used to make the
examination of more sheep to detect a following classifications:

% of affected sheep with score 4 lesions Disease Classification


1% or more Virulent footrot
< 1% May be virulent or benign; refer (i) to (iv) below

i. Disease is also considered • In these circumstances a


virulent if: negative gelatin gel test
• Chronic cases of severe may be used to support a
underrunning (score 4) are present field diagnosis of benign
irrespective of prevalence, footrot
particularly in areas of the State
where the microclimate is iii. Disease is probably benign but
unfavourable for full disease reinspection is advisable if:
expression, in individual mobs • After eradication programs on
subsequent to eradication or in properties a very low prevalence of
recently purchased mobs of sheep, underrunning is seen in affected
or, sheep the following spring, where
• Score 4 lesions are present in conditions have been conducive for
obviously recent cases, or, footrot and no treatments
• The veterinarian is confident of a undertaken since the eradication
diagnosis of virulent footrot and can program.
justify this (with adequate records).
A negative gelatin gel test may be used
ii.Caution in footrot diagnosis is to support a field diagnosis of benign
indicated and reinspection footrot.
necessary if a proportion have
score 3 lesions, irrespective of iv. Virulent footrot can be excluded
the flock prevalence. This may be: if:
• Partial expression due to recent • Other identifiable causes of
virulent infection lameness are demonstrated and
• Inadequate environment there is no evidence of D. nodosus
• Over expression of benign footrot in in any sheep examined (minimum
a very favourable environment of 100)
• On reinspection 2-3 weeks • After the mob has been through
later, recent virulent conditions favourable for the
infections will have expression of footrot and
progressed under - no recent treatments have
favourable conditions been undertaken and
• If the environment is - at least 100 adult sheep are
inadequate, virulent examined and
infections will persist at or - 25% or more sheep have D.
about the same level but nodosus infections and
benign cases will have - none have foot scores greater
regressed without than score 2.
treatment

VETERINARY LABORATORY MANUAL 55


A negative gelatin gel test may be used • ‘Footrot - technical information manual’,
to support a field diagnosis of benign published by Coopers Animal Health
footrot. Australia (for use with the NSW Footrot
Strategic Plan) 2nd edn, 1990, edited
ROLE OF THE LABORATORY IN by R.Walker, SFVO Wagga.
FOOTROT DIAGNOSIS • ‘Footrot Strategic Plan 1991’, published
A laboratory test must not be used on its by NSW Farmers, NSW Department of
own to establish a diagnosis of virulent Primary Industries, Rural Lands
footrot. Protection Boards, Stock and Station
Agents Association and University of
Where concern exists on the status of the Sydney.
disease on the property, the producer may • ‘Proceedings of Dept of Animal Health
elect, at the producer's cost, to utilise Footrot Workshop - Gundagai March
laboratory testing as an aid to the flock 1992’, by J.R. Egerton, University of
diagnosis. Sydney.

Culture and virulence testing of D. nodosus FOOTROT IN GOATS AND CATTLE


is expensive and labour intensive, with
cultures requiring 8-18 days to isolate the In goats, the clinical signs are not
organism in sufficient purity for necessarily indicative of the virulence of the
characterisation. organism. Benign sheep strains of D
nodosus can cause severe underrunning
The gelatin gel test is currently used as and lameness in goats; conversely virulent
the standard laboratory test to evaluate sheep strains may only cause interdigital
isolates as being benign or virulent (non- dermatitis in some goats.
benign).
Where sheep and goats are run together
The elastase test is available but is not and spread periods have occurred, then
routinely used for estimating isolate presence of footrot in the goats only is
virulence. suggestive of a benign strain. Virulence of
D nodosus isolates in goats will otherwise
As a minimum, it is anticipated that at least require laboratory testing of those isolates.
100 sheep would have been examined in
the mob and the lesions in individual sheep With rare exceptions, cattle isolates of D
recorded, before specimens are collected nodosus to date have proven to be benign
for laboratory testing. Suitable samples for sheep. However it has been shown that
from a minimum of 5 representative a virulent strain for sheep can be harboured
affected animals should be forwarded by cattle with lesions of foot underrunning
together with an adequate description of and/or discharge.
lesions in the sampled group. This will allow
a minimum of 5 (up to a maximum of 20) Diagnosis
isolates to be examined in the laboratory. See’Virulent footrot in sheep.

The following information must be included Specimens required


in the specimen advice form: See’Virulent footrot in sheep.
• number of sheep examined
• foot scores of affected sheep FUNGAL INFECTIONS (OTHER
• pasture conditions and rainfall over the THAN MYCOTOXICOSES)
past 3-4 weeks
• previous footrot history of the flock Care should be taken in handling
specimens from suspect cases as a
• history of recent introductions
number of fungal infections are
• history of recent treatments
communicable to man.
• age, breed and sex of the sheep
Diagnosis
Laboratory-based tests should be Clinical findings, mycology and
interpreted in relation to clinical findings. In histopathology.
general, a positive test result is meaningful,
but a negative test result is not a guarantee Specimens required
of footrot freedom. A negative gelatin gel Skin conditions
test may be used only to support a field i. Skin biopsy taken from the margin of
diagnosis of benign footrot. an active lesion submitted for
mycology* and histopathology is the
Refer to the booklets: best specimen.

VETERINARY LABORATORY MANUAL 56


ii. Hair and deep skin scrapings from syndrome/diseases_of_livestock/dpi-samp-
the peripheral areas of active coll-prot.pdf
lesions, submitted dry, for
mycology*. (in order of preference)

Systemic conditions Diagnosis of the disease


i. Sections of any organs showing i. 20 to 25 hairs, with roots attached,
lesions submitted for mycology*. from the distal end of the tail.
ii. Sections of liver, spleen and lung ii. Lymphoid tissue and/or whole brain
submitted for mycology*. in buffered formalin for
iii. Sections of affected organs and histopathology.
liver, kidney and spleen in buffered
formalin for histopathology. Heterozygote detection
i. 20 to 25 hairs, with roots attached,
*Samples for fungal culture should not be from the distal end of the tail.
chilled or frozen, as temperatures of less
o
than 15 C can be detrimental to fungal GENETIC DISEASES
survival.
The majority of inherited diseases
Culture of fungi may take up to 3 weeks. recognized in cattle are autosomal
recessive. Due to the practice of “line-
breeding”, some recessive defects may
GENERALIZED GLYCOGENOSIS
become relatively common. In addition,
Syn: Pompe’s disease undue emphasis upon estimated breeding
values (EBVs) to maximize production may
(Beef Shorthorn and Brahman cattle) lead to high levels of inbreeding and an
inevitable increase in prevalence of
The most common clinical presentation is of recessive defects. In contrast, inherited
progressive muscular weakness leading to defects that are autosomal dominant are
collapse and inability to rise, usually, but most likely limited to a single herd, or rarely,
not exclusively, after weaning. There are no to a few associated herds.
pathognomonic gross necropsy findings. Haemophilia A in Herefords is an example
PAS positive cytoplasmic vacuolation of of an X-linked defect, where the clinical
peripheral lymphocytes is indicative of the disease is most common in males.
disease. Similarly, light microscopic findings Potentially there will also be Y-linked
of PAS positive cytoplasmic vacuolation of defects and also those resulting from
lymphoid tissue, liver, kidney or neurons mutations in mitochondrial DNA.
are indicative of generalised glycogenosis.
Autosomal recessive defects are expressed
There are three known “lethal” mutations in in male and female progeny of clinically-
the acidic α-glucosidase gene in cattle, two normal, heterozygous parents. One quarter
in Brahmans (commonly known as E7 and of the progeny of heterozygotes will be
E13) and a third in Shorthorns (commonly affected, half will be heterozygotes like the
known as E18). The E7 mutation is the parents, and the remaining quarter will be
most common, with the E13 mutation homozygous wild-type (normal) - that is,
restricted to descendants of one American they did not inherit the disease-causing
import. mutation.

Submitters are requested to specify breed Registered animals of most cattle breeds
of the subject. Submitters are advised to have been derived from a limited gene pool.
determine (via the Australian Brahman Within modern breeds, herd books are
Breeders’ Association) if a Brahman subject generally closed. Consequently, most
is a descendant of the founder for the rare recessive defects are usually “breed-
E13 mutation. specific” and are caused by founder effects
relating to a single mutation. There are
Diagnosis exceptions where multiple mutations cause
Clinical signs and history, histopathology, the same clinical disease within and across
confirm by DNA analysis. breeds. An affected individual may be
heterozygous for two distinct mutations.
Specimens required The term ‘compound heterozygote’ is used
See to describe these animals. An example
http://www.dpi.nsw.gov.au/agriculture/vetm would be a Brahman calf affected with
anual/specimens-by-disease- generalised glycogenosis if it inherited the
exon7 mutation from one parent and the

VETERINARY LABORATORY MANUAL 57


exon13 mutation from the other parent. and Dwarfism in Dexters, samples should
Similarly, different mutations in the α- be submitted direct to an external reference
mannosidase gene cause α-mannosidosis laboratory – addresses provided. With
in Angus and Galloways. Consequently, an DNA-based assays, hair root samples are
affected Angus/Galloway calf could be preferred as they avoid potential problems
heterozygous for both of the mutations. In created by haemopoietic chimerism, which
like manner, a Poll Hereford cross Poll can occur when using tests based on
Shorthorn calf may be affected with maple analysis of DNA from cellular elements of
syrup urine disease if it inherits the “breed- blood. Bovine co-twins commonly share
specific” mutations from the parents. haemopoietic stem cells, resulting in DNA
in cellular elements of blood containing
DNA tests are available for diagnosis of the nucleotide variants from both twins that
following genetic diseases, or for detection may be different to the DNA variants found
of heterozygotes. With CVM in Holsteins in gametes from an individual co-twin.

Breed of Cattle Disease


Angus α-Mannosidosis
Murray Grey α-Mannosidosis
Galloway α-Mannosidosis
Salers ß-Mannosidosis
Holstein/Friesian Citrullinaemia
Factor XI deficiency
DUMPS
Bovine Leucocyte Adhesion Deficiency (BLAD)
Complex Vertebral Malformation (CVM)*
Poll Hereford Maple Syrup Urine Disease
Inherited Congenital Myoclonus
Limousin Protoporphyria
Beef Shorthorn Generalised glycogenosis (Pompes Disease)
Brahman Generalised glycogenosis (Pompe's Disease)
Dexter Chondroplasia (dwarfism) / Congenital lethal chondrodysplasia (Dexter
bulldog syndrome)**

* Consign samples direct to Dr. Van Haeringen, Laboratorium B.V. PO Box 408, 6700 AK,
Wageningen, The Netherlands.
** Consign samples direct to Reprogen, Faculty of Veterinary Science, University of Sydney,
PMB 3, Camden, New South Wales 2570

See Specimens (by disease) for details of specimen collection for diagnosis of a specific disease.

GOITRE History of access to grain and


demonstration of excess grain in ingesta
Diagnosis with a pH of the ruminal content of less than
Clinical findings, histopathology. 5. The rumen pH must be taken in the field
and is only reliable
Specimens required if taken shortly after death. Elimination of
Thyroid glands submitted in buffered bloat, enterotoxaemia as possible causes.
formalin for histopathology.
Specimens required
GRAIN POISONING i. Serum or eye fluid for D-lactate.
ii. Sections of wall of forestomachs in
Diagnosis
buffered formalin for histopathology.

Interpretation of D-lactate concentration


Analyte Sample Units Normal Acidosis
D-lactate Serum, aqueous humour mM/L < 0.4 >1.0

There is no specific diagnostic test GRASS TETANY


available for grain poisoning. Specimens
should be taken to eliminate other possible See Hypomagnesaemia
causes, e.g. enterotoxaemia.

VETERINARY LABORATORY MANUAL 58


HAEMAGGLUTINATING
ENCEPHALOMYELITIS VIRUS (HEV) HELIOTROPE POISONING
INFECTION OF PIGS See: Pyrrolizidine alkaloidosis
Syn: Vomiting and wasting disease
HENDRAVIRUS INFECTION
HEV is responsible for two overlapping Syn: Equine morbillivirus infection
diseases of young pigs:
• Encephalomyelitis (with nervous signs) Hendravirus infection was first recognised
of pigs < 10 days old in 1994 in Australia, when it caused an
• Vomiting and wasting disease (with outbreak of acute, fatal respiratory disease
wasting and failure to thrive) of pigs that killed 14 horses. During this outbreak
from 7-21 days old. the horse-trainer also died and there was a
non-fatal infection of another person closely
Diagnosis involved with the sick horses. To December
Histopathology; paired virus serology; virus 2004, four more sporadic outbreaks
isolation. Differentiate from enterovirus reported have involved the death of six
encephalomyelitis, other neonatal diseases horses and one human, and one non-fatal
e.g. hypoglycaemia, bacterial human infection. One human death
meningoencephalitis; differentiate from occurred 14 months after exposure at post
other causes of runting (bacterial, parasitic, mortem examination of an affected horse.
nutritional, toxic). Consider exotic diseases
in differential diagnosis of nervous disease: Flying foxes (fruit bats) are the subclinical
Aujeszky's disease, Teschen disease, reservoir of Hendravirus. All known cases
Japanese encephalitis. have occurred in Queensland and
coincided with the flying fox breeding
Specimens required season in the later months of the year.
i. Fixed brain, cervical spinal cord,
gasserian ganglion (trigeminal nerve Hendravirus is a member of the
ganglion), samples of other visceral paramyxovirus family of viruses, and is
organs for histopathology. closely related to Nipahvirus, which caused
ii. Fresh, chilled samples of brain, a major outbreak of acute fatal respiratory
cervical spinal cord, SI content and disease in pigs and humans (and dogs) in
a composite of visceral organs (liver, Malaysia in 1999. Like Hendravirus, Nipah
kidney, lung, spleen) collected virus has flying foxes as its natural
aseptically into sterile Macartney reservoir.
bottles for virus isolation.
iii. Serum samples from acute and Hendravirus infection is notifiable. Fees
convalescent phases (14 days for tests undertaken to confirm or exclude a
apart), where possible. diagnosis of Hendravirus infection and to
iv. Range of samples for bacteriology, establish an alternative diagnosis are paid
for differential diagnosis e.g. brain, by NSW Department of Primary Industries.
liver, kidney, small intestine.
v. Blood in sodium fluoride for Caution Hendravirus is a zoonotic
elimination of hypoglycaemia. agent, causing serious illness and
sometimes death in humans.
HAEMATURIA
Veterinarians should take biosecurity
See also: Enzootic haematuria of cattle,
precautions when performing post mortem
bracken fern poisoning, poisoning-plant'
examinations of horses. Suitable personal
protective equipment includes gloves,
HAEMOGLOBINURIA overalls, and mask to prevent inhalation of
See also: Hypophosphataemia, aerosols. Call customer service (1800 675
leptospirosis, tick fever, copper poisoning' 623) for advice.

HAEMOPHILIA Clinical signs include pyrexia, respiratory


distress (dyspnoea, blood-tinged foamy
See: Factor VIII deficiency (Hereford cattle) discharge from mouth and nares),
and Factor XI deficiency (Friesian cattle)' sometimes neurological changes (head-
pressing, muscle fasciculations) and death.
HEEL ABSCESS - OVINE Colic was the initial sign in two recent
cases.
See: Foot abscess in sheep'

VETERINARY LABORATORY MANUAL 59


Post mortem findings include massive HISTOPHILUS OVIS/ HAEMOPHILUS
pulmonary congestion and oedema with SOMNUS INFECTIONS
airways filled with foamy, sometimes blood-
stained fluid. Histophilus ovis and Haemophilus somnus
are closely related organisms sometimes
Further information at the Queensland associated with the following conditions:
Department of Primary Industries' website
at Sheep
http://www2.dpi.qld.gov.au/health/3892.html Epididymitis; gangrenous mastitis; abortion;
. septicaemia, polyarthritis and abscessation
in lambs; pneumonia and nephritis.
Diagnosis
Clinical signs, post mortem findings, Cattle (in Australia)
histopathology, virology. Purulent vaginitis and endometritis;
pneumonia, fibrinopurulent leptomeningitis
Specimens required and ependymitis.
i. Chilled lung for virology.
ii. Fixed lung for histopathology. (Overseas septicaemia, thromboembolic
iii. 10 ml vacuum tubes of clotted and meningoencephalitis, polyarthritis,
LiHep blood for virology. tendonitis, and less commonly, mastitis).

HEPATOSIS DIETETICA Diagnosis


Clinical findings and pathology, including
A disease of pigs, responsive to vitamin histopathology. Isolation of H. ovis/ H.
E/selenium therapy. somnus from affected tissues

Diagnosis NB The organism may be a commensal,


Clinical signs of dullness, anorexia, carried in the urogenital or upper respiratory
staggering and possibly mild ascites before tract of normal animals.
sudden death.
Specimens required
Specimens required i. Samples of affected tissues and
i. Liver, skeletal muscle, portion of swabs in Amies charcoal transport
small intestine with mesentery medium for bacteriology.
attached, and myocardium in
buffered formalin for histopathology. e.g. milk and affected gland (mastitis),
liver, lung, kidney, spleen
HERPESVIRUS (septicaemia), whole affected joint
VULVOVAGINITIS/BALAN0POSTHITI unopened (arthritis) or synovial fluid
S IN GOATS and synovial membrane (arthritis),
vaginal/uterine swabs and exudate
See also: Infectious bovine rhinotracheitis (reproductive), affected epididymis
(IBR), infectious pustular vulvo-vaginitis (epididymitis), whole aborted foetus and
(IPV) membranes (abortion), brain (CNS),
abscess swab and/or abscess content
Diagnosis (abscessation), lung (pneumonia).
Clinical signs and history of mating season,
straining, dysuria, preputial/ vaginal ii. Portions of affected tissues fixed in
swelling with papules, vesicles and ulcers buffered formalin for histopathology.
which heal in 7-10 days (similar to cows
with IPV); Virus isolation and virus serology. HYPOCALCAEMIA
Specimens required Diagnosis
i. Swabs of vaginal/preputial lesion Clinical signs, response to therapy, serum
secretion in PBGS for virus isolation. or eye fluid calcium level - complication or
Best at papule/vesicle stage, as differential diagnosis of hypomagnesaemia,
virus disappears with ulcer ketosis, hypophosphataemia.
formation.
ii. Serum sample submitted chilled for Specimens required
virus serology. i. At least 2 ml of serum, free of cells
and haemolysis, submitted chilled
for calcium and magnesium
estimation.

VETERINARY LABORATORY MANUAL 60


ii. From cadavers, 0.5-1 ml of aqueous aqueous humour (0.45μ membrane
humor, free of blood and cells for filter) to minimise cellular
calcium estimation. Ideally, filter contamination.

Interpretation in cattle and sheep History, clinical signs, response to therapy,


Calcium serum, eye fluid or urinary magnesium
Normal Serum > 2.25 mmol/L concentration; complications of
Aqueous > 1.4 mmol/L hypocalcaemia, differentiation from ketosis.
Deficient Serum < 2.1 mmol/L
Aqueous < 1.1 mmol/L Specimens required
i. At least 2 ml of serum, free of cells
HYPOGLYCAEMIA and haemolysis, submitted chilled
for calcium and magnesium
Diagnosis estimation.
History, clinical signs. ii. From cadavers, 0.5 1 ml of aqueous
humor, free of blood and cells, for
Specimens required magnesium estimation. Ideally, filter
There is no routine laboratory test for blood aqueous humour (0.45μ membrane
glucose concentrations. filter) to minimise cellular
contamination.
HYPOMAGNESAEMIA iii. Alternatively, 1 ml of urine for
magnesium estimation.
Syn: Grass tetany, lactation tetany, milk
tetany of calves

Diagnosis

Interpretation in cattle and sheep


Magnesium
Normal Serum > 0.7 mmol/L
Aqueous > 0.6 mmol/L
Urine > 2 μmol/mosmol

HYPOPHOSPHATAEMIA i. At least 2 ml of serum free from


cells, submitted chilled or frozen for
Diagnosis phosphorus estimation.
History, area information, clinical signs,
blood or serum inorganic phosphate NB Serum phosphorus levels are of little
concentration, response to therapy. value if the sample is haemolysed or
contaminated. Do not send serum on the
Specimens required clot.

Interpretation in cattle and sheep


Phosphorus
Normal Serum > 1.3 mmol/L
Deficient Serum < 1.0 mmol/L

ILL THRIFT • Parasitism, including helminthiasis,


fascioliasis and coccidiosis.
See also: Anaemia', scouring' • Eperythrozoonosis.
• After effects of systemic or chronic
Ill thrift is a vaguely defined condition with a
conditions including pyrrolizidine
variety of causes. Cases of ill thrift
alkaloidosis, sarcosporidiosis and
associated with anaemia or scouring should
chronic infectious diseases
be investigated under the syndromes of
• Viral: EIA in horses, pestivirus
'anaemia' or 'scouring'.
infections in ruminants, EBL in cattle,
CAE in goats.
Other causes of ill thrift not necessarily
associated with anaemia and/or scouring • Specific bacterial diseases e.g.
include: Johne's disease, caseous
lymphadenitis.
• Malnutrition, the commonest cause.
• Nutritional deficiencies of copper
Diagnosis
cobalt, phosphorus or selenium.

VETERINARY LABORATORY MANUAL 61


Based on clinical findings, laboratory ii. Specimens as required for
examination to eliminate specific causes encephalitis, including brain for virus
and response to treatment. isolation and histopathology.

Specimens required INFERTILITY IN CATTLE, SHEEP,


Specimens should be submitted to PIGS
eliminate some of the possible causes only
after malnutrition has been eliminated as a See also: Abortion (general), congenital
contributing factor. The conditions listed abnormalities, copper deficiency,
should operate on a flock basis so that epididymitis, semen examination,
specimens should be submitted from a trichomoniasis of cattle
number of animals in the flock. The range
of specimens to be submitted could include: Diagnosis
i. Faecal samples from 10 to 20 Must be based on field investigations
animals for parasitology. supported by the laboratory where
ii. Blood samples as required to necessary. The first step is to establish the
eliminate specific deficiencies of stage in the reproductive cycle where the
selenium, phosphorus and copper. problem occurs, i.e. at joining due to
iii. Sections fixed in buffered formalin, fertilisation failure (male or female
of liver, kidney, cardiac and skeletal infertility), embryonic loss, abortion or
muscle, small and large intestine or perinatal loss.
other organs depending on gross
pathological indication of aetiology Specimens required
at field necropsy. Those appropriate to the condition or
iv. If indicated, sera and/or portions of conditions suspected. A detailed history
body organs including the above, should be provided to allow consideration of
submitted fresh for bacteriological, other causes. Detailed investigations
virological and/or parasitological should involve prior consultation with the
examination. laboratory.
v. Live unthrifty animal submitted for
post-mortem examination. INHERITED DEFECTS
See congenital abnormalities, genetic
INFECTIOUS BOVINE diseases'
RHINOTRACHEITIS (IBR) AND
INFECTIOUS PUSTULAR VULVO INHERITED CONGENITAL
VAGINITIS (IPV) MYOCLONUS
Diagnosis (Poll Herefords)
Virus isolation and antibody rise between
acute and convalescent serum samples. Stimulus-responsive myoclonic spasms in
bright and alert calves is the striking clinical
Specimens required feature of this disease. The majority of
Respiratory and ocular disease affected calves have hip lesions, most
i. Swabs from nasal cavities and frequent being fractures of the acetabulum,
conjunctivae in phosphate buffered and less commonly, fractures of the head of
gelatin saline (PBGS). the femur. No consistent histological
findings.
Vulvo-vaginitis:
i. Acute and convalescent serum Diagnosis
samples (collected 3 6 weeks apart), History, clinical signs, confirmed by DNA
submitted chilled for virus serology. analysis.
ii. Swabs of vaginal mucus in PBGS.
Specimens required
Abortion See
i. Specimens as required for bovine http://www.dpi.nsw.gov.au/agriculture/vetm
abortion. anual/specimens-by-disease-
syndrome/diseases_of_livestock/dpi-samp-
Encephalitic form coll-prot.pdf
i. Acute and convalescent serum
samples (collected 3 6 weeks apart), Diagnosis of the disease and
submitted chilled for virus serology. heterozygote detection
i. 20 to 25 hairs, with roots attached,
from the distal end of the tail.

VETERINARY LABORATORY MANUAL 62


Eperythrozoonosis, Leptospirosis,
IRON TOXICITY SYNDROME IN Plant poisoning, Pyrrolizidine
PIGLETS alkaloidosis.
iii. In the live animal, serum chilled for
Diagnosis enzymology.
Clinical history of sudden death of 0-7 day
piglets within 1 12 hours of giving iron JOHNE'S DISEASE
injection, or vomiting/ diarrhoea/ dyspnoea/
coma and death up to 3 days; selenium Syn: Paratuberculosis
deficiency in sow.
Diagnosis
Specimens required History, clinical signs, gross pathology and
i. 10 ml of heparinized blood from histopathology. Demonstration of typical
sow(s), submitted chilled for organisms in faecal smears and tissues.
selenium analysis. Isolation of Mycobacterium
ii. Piglet for post mortem examination paratuberculosis from faeces or tissues.
and differential diagnosis. Serology.

ITCHMITE IN SHEEP Absorbed ELISA


Absorbed ELISA are routinely available for
Diagnosis Johne's Disease testing of cattle, goat,
Demonstration of Psorergates ovis mites in alpaca and deer sera. Absorbed ELISA
deep skin scrapings from suspected cases, testing is the most sensitive serological
examined in the field. Differentiation from method for diagnosis and certification for M.
other causes of fleece derangement e.g. paratuberculosis infection and disease. It
lice, tender fleeces. also has a very high specificity.

Specimens required AGID


In cases of dermatitis or skin irritation An AGID is available for goats and sheep -
where mites cannot be seen at field this has a low sensitivity in identifying
examination, the following samples may be infected animals, but its specificity is
sent to the laboratory: extremely high. It is of value only in those
i. Skin scrapings collected using a animals with moderate histological lesions
scalpel blade dipped in paraffin oil, (including all clinical cases); a positive test
submitted with the blade enclosed in is indicative of current histological lesions of
a secure sample container. the disease. ELISA positive goats can be
ii. A skin biopsy from the affected area, further tested by AGID to identify those
submitted in buffered formalin (may infected animals with most advanced
assist in a differential diagnosis). lesions.

NB Wool must be clipped as close to the Smears of faeces and tissues


skin as possible before taking a scraping Smears of faeces and tissues are of limited
(see Parasites - External). sensitivity in subclinical animals, and should
be restricted to clinical cases.
JAUNDICE
Faecal and tissue culture
Jaundice is a clinical sign which may arise Faecal and tissue culture procedures
as a result of a haemolytic crisis or hepatic depend on whether the cattle (C) or sheep
failure. Common causes include plant (S) strain is likely to be involved. The
poisonings, chronic copper poisoning, majority of cattle, goat, deer and alpaca
leptospirosis and eperythrozoonosis. infections are attributed to C strains, while
sheep infections are predominantly due to
Diagnosis S strains. Cattle or goats reared with
Clinical signs, history. Specific causes infected sheep may be infected with S
diagnosed on the basis of laboratory strain; C strain infection in sheep is
examination. relatively uncommon. The C strain can be
cultured by either conventional (solid
Specimens required media) culture or by radiometric broth
i. Sections of liver, kidney and spleen, (Bactec) culture. For the C strain,
submitted in buffered formalin for conventional culture is usually slower than
histopathology. Bactec culture, but is less expensive. The S
ii. Other specimens appropriate to the strain can only be reliably cultured by
disease condition suspected see radiometric (Bactec) culture, and a longer
Copper poisoning,

VETERINARY LABORATORY MANUAL 63


incubation period is used compared to the be clear on the specimen advice form so
C strain. that procedures specific to the S strain (i.e.
longer incubation; different subculture
The current default procedures (based on medium) can be undertaken.
consumer demand) apply to culture
submissions from cattle, goats, alpaca and The culture process is undertaken in
deer: batches, and requires an initial
decontamination phase over 5 working
Species Method days, followed by an incubation phase, and,
Cattle, deer: Bactec if growth occurs, a confirmatory phase. The
Goats, alpacas: Conventional amount of M. paratuberculosis in the
sample correlates with the rate of growth
If the default procedure is not suitable (i.e. (higher numbers reduce the time delay until
the alternate procedure is required), the growth is detected). This in turn modifies
required procedure must be advised on the the time for culture to be completed. The
Specimen Submission Form. In addition, following table outlines expected time
where OJD is suspected and a non-ovine differences between the two strains in
species is being tested by culture, this must current cultural procedures:

C strain S strain
Method Conventional Bactec Bactec
Decontamination 5-7 d 5-7 d 5-7 d
Incubation 20 wks 8 wks* 12 wks
Confirmation by PCR/REA 1-4 wks (if reqd) 1-4 wks 1-4 wks
Confirmation by subculture
Up to 10 wks Up to 10 wks Up to 10 wks
(mycobactin dependency test)
Total time if no growth detected 21 wks 9 wks 13 wks
Total time if growth detected and M.
12-26 wks 10-21 wks 10-23 wks
paratuberculosis confirmed
Total time if growth detected and not
12-24 wks 12-21 wks 12-23 wks
M. paratuberculosis

* If growth is detected late (e.g. at week 8), incubation to week 10 may be required to reach a
satisfactory level for confirmation.

Culture confirmation of M. Examination for the second characteristic of


paratuberculosis M. paratuberculosis as part of the
Confirmation of M. paratuberculosis from confirmatory process from Bactec media is
Bactec growth is undertaken when growth also commenced when growth reaches a
reaches a high level, as this optimizes the high level. This normally requires a sample
confirmation rate. Two separate of growth to be subcultured to solid medium
characteristics of M. paratuberculosis are for evidence of typical colonies with
targeted in the confirmation procedure for mycobactin dependency, and takes up to
maximum specificity, to reach a conclusion 10 weeks. Different solid media are used
of culture positive for M. paratuberculosis. for subculture of the C and S strain. While
When multiple tissues are cultured from an solid media that supports the S strain can
individual, both characteristics need be met be used to cultivate the C strain, it does so
on only one tissue for a conclusion of less efficiently than the preferred C strain
culture positive status for that animal. medium. In contrast, S strain does not grow
on the solid C strain medium. As an
The first characteristic is measured in a alternative to subculture to demonstrate
molecular assay involving a polymerase mycobactin dependency, testing for a
chain reaction (PCR) and restriction second specific molecular target (e.g.
endonuclease analysis (REA) that target IS1311 PCR and REA) can reduce the time
the IS900 gene sequence. Together, these to complete confirmatory testing
can identify the presence of M. procedures, and yet retain maximal test
paratuberculosis DNA with very high specificity.
specificity. Samples of Bactec growth are
collected and then processed by PCR and Confirmation of M. paratuberculosis from
REA in batches, for greatest efficiency. conventional culture for C strains is
equivalent to the second part of the Bactec
confirmatory procedure i.e. subculture for

VETERINARY LABORATORY MANUAL 64


mycobactin dependency. However, ii. Serum sample for ELISA test (cattle,
additional IS900 PCR and REA testing will goats) and/or AGID (goats, sheep).
be undertaken on growth on solid media
typical of M. paratuberculosis in herds *Note: Pooled faecal culture procedures for
where Johne's disease has not previously cattle and goats are currently under
been confirmed. This should be borne in development.
mind when completing the Specimen
Submission Form for samples from herds In slaughtered animals:
without prior confirmation of Johne's i. Fresh, chilled portions of intestine
disease and where conventional culture is and intestinal lymph nodes in
requested. separate sterile containers for
bacteriology. The best samples are
Direct PCR usually those with obvious lesions
Direct PCR on pooled or individual faeces (gut thickening, lymph node
is available for sheep in limited enlargement). Samples from the
circumstances. It is restricted to use for risk following sites per animal are
assessment purposes to determine if high considered ideal, and are typical of
shedder animals are present. those required for MAP certification.
In addition, faeces at slaughter can
Strain typing be useful to assess the risk of
Strain typing can be undertaken on Bactec shedding.
growth if requested. This relies on PCR and
REA targeting the IS1311 gene sequence. Cattle: Intestinal portions of 5 cm x 10
cm
NB: Allergic tests and serological tests such • proximal ileum (30-45 cm anterior
as the CFT are unreliable in the diagnosis to ICV)
of the disease, with false positives and false • terminal ileum (11-20 cm anterior to
negatives occurring frequently. Individual ICV)
culling on the grounds of such tests is often • ileocaecal valve (ICV) and attached
impractical. The Johne's CFT is not segment of ileum
routinely available. It is restricted for • proximal colon
export/certification testing only, and not for • ileal LN
routine diagnostic purposes. • ileocaecal LN
Specimens required Goats and alpaca: Intestinal portions
In the live animal: of 5 cm in length
i. Faeces, submitted chilled for • jejunum
bacteriology
• proximal and terminal ileum (1 m
• Individual faecal culture - volume of apart)
> 5 g per animal
• ileocaecal valve and attached
• Pooled faecal culture - in sheep*, segment of ileum
one pellet from each of (up to) 50
• proximal colon
sheep per pool. In flock sampling,
• ileal LN
choose additional sheep to allow for
animals from which no sample can • ileocaecal LN
be collected (allow 55 sheep for
each pool of 50 pellets). Sheep:
Notes in regard to PFC submissions: • terminal ileum
• Do not take more than one pellet • ileal LN (caudal jejunal LN)
per animal
• Samples must not overfill a 60 mL ii. Sections of the above, in buffered
sterile plastic universal container. If formalin for histopathology.
pellets are l large, submit only
sufficient per pool to fill the 60 mL In cattle and goats, include fixed caecum. In
container. goats and sheep, can include a section of
fixed liver (microgranulomata).
• Serial faecal culture - in sheep -
one pellet from each of (up to) 10
NB It is preferable that the above samples
sheep and submit from these same
be taken in the field. Where this is not
animals on each of three occasions
possible, do not forward entire intestinal
10-14 days apart. This will create a
tract for laboratory examination without
single pool of up to 30 samples.
prior consultation. Unless such samples are
received soon after collection, autolytic
changes can prevent useful examination,

VETERINARY LABORATORY MANUAL 65


particularly in serological reactor animals There is no value at present in examining
with limited lesions. samples of kikuyu for toxins, fungi, etc.

KIKUYU POISONING LEAD POISONING


A disease of cattle causing salivation, Diagnosis
dehydration, depression, abdominal pain History of illness, clinical signs of central
and death associated with grazing on lush nervous system disturbance, biochemical
kikuyu pastures. The cause is unknown, analysis of tissues. In calves, thymic
although mycotoxins have been suggested haemorrhages are a frequent finding at
in some cases. necropsy.

Diagnosis Specimens required


History, clinical signs and histopathology Live animal
(particularly of omasum and reticulum). i. 100 g of faeces.
ii. 10 mL EDTA blood tube.
Specimens required
i. Sections of omasum, reticulum, Dead animal
rumen abomasum and kidney, in i. 50 g of kidney, submitted chilled for
buffered formalin for histopathology. toxicology.
ii. Sections of liver, kidney and small ii. Sections of liver and kidney, and
intestine, submitted in buffered whole brain in buffered formalin for
formalin for histopathology (for histopathology.
differential diagnosis).

Interpretation of lead concentrations (cattle and sheep)


Analyte Sample Minimum Units Normal Possibly Toxic
amount toxic
Lead EDTA blood 5ml µmol/L < 1.2 > 1.2
Lead Kidney (Liver) 50g mg/kg (wet wt) <4 4-25 > 25
Lead Faeces 100g mg/kg (wet wt) < 10 10-25 > 25

Kidney lead concentrations below 4 mg/kg are considered non-significant. Higher concentrations
should be interpreted on the basis of clinical findings and histopathology.

Faecal lead concentrations below 10 mg/kg are considered non-significant. Higher levels could be
significant, depending on the source of lead.

LEPTOSPIROSIS
Serovar Maintenance Host
Leptospirosis is transmissible to man and L. hardjo Cattle
represents an occupational hazard to L. pomona, L. Pigs
veterinarians, abattoir workers, dairy and tarassovi
pig farmers. L. copenhageni Rats
The most commonly encountered serovars
associated with clinical disease are: In the maintenance host, infections by
• Cattle L hardjo, L pomona these specific serovars are likely to be
• Pigs L pomona, L tarassovi endemic, whereas serovar infections in non
• Horses L pomona maintenance hosts are usually associated
• Dogs L copenhageni with sporadic outbreaks of disease. The
most common manifestations of
Other serovars do affect domestic animals leptospirosis in animals are:
in Australia, particularly Northern Australia, • Abortion (all species)
but such infections are rare in NSW. • Milk drop syndrome/ mastitis
(cattle)
Zoonotic infections are usually acquired • Pyrexia/Septicaemia (all species)
from the maintenance host for each • Icterus and/or haemoglobinuria
serovar: (mainly dogs and calves)
• Kidney lesions found at routine
slaughter (pigs, cattle)

VETERINARY LABORATORY MANUAL 66


• Mortalities following pyrexia neutral buffered formalin to 99 mL
and/or icterus/haemoglobinuria urine) added immediately after
collection, from both affected and
Diagnosis recovered cases.
Serology (Microscopic Agglutination Test).
Confirmed by demonstration of leptospires Icterus/haemoglobinuria
via direct microscopic examination of urine Samples as for Pyrexia, but do not collect
or foetal fluids (thoracic, peritoneal, urine for microscopic examination from
pericardial); and/or by demonstration of animals affected with haemoglobinuria.
organisms in kidney by histopathology. Urine shedding of leptospires occurs after
the haemoglobinuric phase.
In general culture of tissues, foetal fluids,
urine or milk is not undertaken, except Kidney lesions in slaughter animals
under specific arrangements with the i. Portions of affected kidneys in
receiving laboratory. buffered formalin for histopathology.

Specimens required Mortalities following pyrexia and/or


Abortion icterus/haemoglobinuria
Samples as outlined in 'Abortion in Cattle' i. Sections of liver and kidney, in
or 'Abortion in Swine'. Whenever possible, buffered formalin for histopathology.
submit serum from dam, including 10 15
sera from cohorts, and fresh, chilled entire Interpretation of Leptospiral titres
foetus with membranes (or chilled and fixed For the MAT, titres are given which reflect
foetal tissues, including serous fluids) for the standard Leptospiral dilution series for
bacteriology and histopathology. sera. For leptospiral MATs, this is a
doubling dilution series starting at 1:25 (L
Milk Drop Syndrome hardjo) or 1:50 (other Leptospires).
i. Serum samples from 10 15 affected
and recovered cases and cohorts, In cattle, an MAT titre to L hardjo of 25 or
submitted chilled for serology. greater may be significant. Indication of
• A convalescent serum sample current infection can only be gained by
collected 10 days or more after demonstrating at least a four-fold (i.e. 2
agalactia often shows a rise in titre dilution) increase.
to L. hardjo.
ii. Sample of urine preserved with Titres following vaccination are generally
formalin (approx. 1 ml of 10% low (less than 400) and do not persist. In
neutral buffered formalin to 99 mls cases of abortions in cattle due to L hardjo,
urine) added immediately after titres at the time of abortion may be
collection, from both affected and equivocal and falling.
recovered cases, for dark ground
microscopy. It is worthwhile using a In L pomona infections and L hardjo milk
diuretic to obtain a sample with large drop syndrome, titres generally rise within 1
numbers of organisms and minimal 3 weeks of the appearance of clinical signs.
contaminants. L pomona is more immunogenic than L
iii. Samples of milk for differential hardjo, so titres to L pomona are often 800
diagnosis of mastitis, submitted as or more, whereas in L. hardjo milk drop
outlined in 'Mastitis'. syndrome, they may be less than 800. L
iv. In certain cases, following hardjo titres of 200 or more are generally
consultation with the Regional significant.
Veterinary Laboratory,
arrangements may be made to Submission of 10 15 sera from animals in
sample cows for culture. This the same group as the affected individuals
involves field-inoculation into will give a more accurate assessment of the
selective liquid media of mid-stream leptospirosis status of the herd. To
urine collected after diuretic determine recent infection, 10 15 paired
treatment. sera (taken 2 4 weeks apart) should be
submitted to detect rising titres (along with
Pyrexia the clinical history of the tested animals). In
i. Serum samples from 10 15 affected an endemically infected herd at any given
and recovered cases and cohorts, time, some titres will be rising, some will be
submitted chilled for serology. static, and others will be falling.
ii. Sample of urine preserved with
formalin (approx. 1 ml of 10%

VETERINARY LABORATORY MANUAL 67


LICE RESISTANCE TO Meningoencephalitis (circling disease)
INSECTICIDES IN SHEEP Clinical signs and brain pathology.

Resistance of sheep lice to pyrethroid Abortion


insecticides is widespread in NSW. No Post mortem findings and bacteriology.
functional organophosphate resistance has
been detected. Septicaemia
Bacteriology and histopathology.
Resistance may be suspected on a history
of treatment failure after correct application Specimens required
of the correct amount of lousicide at the Meningoencephalitis
correct time, either: i. Cerebrospinal fluid and sections of
• As breakdowns within 3-4 months of brain, liver, spleen and kidney,
application of backline treatment in submitted chilled for bacteriology.
sheep treated off-shears or in short ii. Brain in buffered formalin for
wool, or, histopathology.
• Where sheep in long wool have been
thoroughly treated with a pyrethroid Abortion
and failure to control infestation is i. Specimens as required for abortion
obvious within 6 weeks. in the particular species (see
Abortion).
NB Long wool application of pyrethroids
may leave undesirable residues at Septicaemia
shearing. i. Heart blood and sections of liver,
kidney and spleen, submitted chilled
Faulty application of treatment should be for bacteriology.
considered where infestations are localised ii. Sections of liver, kidney, spleen and
to one side or part of the sheep, or where lung, in buffered formalin for
one mob or line is infested and other mobs histopathology.
are free of lice.
LIVER FLUKE INFECTION
Diagnosis
Examination of sites of, and degree of, See also Parasites Internal
infestation in 4-5 affected sheep by
counting lice in 10 partings on each side of Diagnosis
the infested sheep (each side examined in Grazing history, clinical examination,
2 lines of 5 partings). Demonstration of serology, faecal egg count, response to
resistance using an in-vitro laboratory test. treatment. Necropsy findings.

NB. It is more productive to inspect a larger ELISA serology is available at EMAI for the
number of sheep rather than search a detection of infection with both immature
smaller number of sheep more thoroughly. and adult fluke. Using the Institut Pourquier
(France) antibody ELISA recently adopted
A thorough examination of the dip operation (2004) by NSW DPI laboratories, titres
and thoroughness of wetting is appear 2-4 weeks (cattle) after infection
recommended, particularly in the case of and remain high for at least 20 weeks after
shower dipping failures with insecticides. tretment. The false negative rate is low in
cattle (<2%). The test has not been
Specimens required validated for sheep under Australian
conditions.
Testing for OP resistance is not routinely
available; it requires controlled temperature Faecal egg counts are only of value in
and humidity and is a laboratory based test chronic fascioliasis where eggs are being
at the Elizabeth Macarthur Agricultural excreted (patent infection); they are of no
Institute (contact Dr Garry Levot). value in acute and subacute infections.
Submission of live affected sheep would be ELISA serology is more sensitive than
required. faecal egg counts for the detection of
chronic fascioliasis in both cattle (30% more
infected animals detected) and sheep (15-
LISTERIOSIS 20% more infected animals detected).
Listeriosis can be transmitted to man. ELISA serology is the only antemortem
diagnostic method for the detection of acute
Diagnosis and subacute fascioliasis (before fluke eggs
are excreted), which typically occurs in

VETERINARY LABORATORY MANUAL 68


NSW from autumn to early winter and in LUPINOSIS
late spring.
A mycotoxicosis mainly associated with
The ELISA test is reported by the grazing on lupin stubble infected with
manufacturer to be suitable for individual Phomopsis leptostromiformis.
and bulk vat milk samples. The test is
currently (2004) undergoing validation by Diagnosis
NSW DPI. The milk ELISA is a herd test History, clinical signs and histopathology.
and for statistical significance a minimum of
12-15 samples should be submitted. Specimens required
i. Sections of liver and kidney in
Specimens required buffered formalin for histopathology.
At necropsy, fascioliasis should be ii. If nervous symptoms are seen,
diagnosed in the field and laboratory specimens as required for nervous
specimens will not be required. disorders.

i. Serum samples from affected cattle LYME DISEASE


and their cohorts for ELISA serology
Lyme Disease is a multisystem disease
ii. Faeces for fluke egg counts in the
caused by the spirochaetal bacterium
following situations:
Borrelia burgdorferi. It can affect the joints,
• Where serum collection in cattle is heart and nervous system of man, domestic
not feasible, or, animals and wildlife and is known to occur
• In animals where liver fluke has in Europe, Asia and North America, where it
been detected in the herd or flock is primarily transmitted by Ixodid ticks.
and treatment has been given in There is clinical, serological and
the previous 5 months, or pathological evidence of a Lyme Disease-
• Where paired faeces and serology like in cattle and man in eastern coastal
is specifically sought for Australia but the organism has not yet been
comparative study. isolated. Likely vectors here are Ixodes
holocyclus or other Ixodes spp.
Submit separate faecal samples (at
least 30 g of fresh faeces each) from at In animals, Lyme disease usually presents
least 10 animals in the flock or herd, in as lameness associated with polyarthritis;
a screw capped jar. joints are painful and swollen and there
may be a low grade pyrexia. There may
Faecal samples may be tested also be lethargy, stiffness and myalgia, with
individually, or pooled in groups of two inability to rise in severe cases. Chronic
(designed for sheep) or five samples weight loss, abortion and laminitis like signs
(cattle). Pooling reduces costs but can be shown by affected cattle and
sensitivity is compromised. Submitters horses; some horses may have skin
should nominate how they wish the hypersensitivity, uveitis and CNS signs
samples to be tested. resembling encephalitis.
NB For interstate testing for import Diagnosis
of ruminants to Western Australia, Clinical signs, histopathology (typically
which is fluke-free, individual samples reveals a fibrinoid polyarthritis).
from up to 30 animals in each
consignment are required (further Borrelia burgdorferi is usually present in
details are available at Department of only low numbers in affected joints, and
Agriculture, Western Australia web site requires special media for isolation. The
under Livestock movement at organism may be more readily isolated from
http://www.agric.wa.gov.au). blood during the early stages of disease.
For more information, refer to the Agfact Serology can be undertaken for some
A0.9.57 (second edition 1999 revised 2003) animal species outside of the Department's
‘Liver fluke disease in sheep and cattle’: laboratories, by arrangement.
http://www.agric.nsw.gov.au/reader/an-
diseases/a0-9-57web.pdf Differential diagnosis:
• Other causes of arthritis:
Chlamydiosis, mycoplasmal arthritis
(See Arthritis, Chlamydial Infections,
Mycoplasmosis).
• Ephemeral Fever.

VETERINARY LABORATORY MANUAL 69


• Septicaemic and toxaemic conditions Further advice can be obtained from the
caused by other bacteria. Australian Government’s Department of
Health and Aging website at
Specimens required http://www.health.gov.au/internet/wcms/Publi
i. Chilled acute and convalescent shing.nsf/Content/cda-pubs-other-
serum, especially for alternative bat_lyssa.htm.
diagnosis (Chlamydia)
ii. Fixed tissues for histopathology: Further information on this disease can be
joint capsule, brain, heart, peripheral obtained from the CSIRO website at
nerve, kidney, liver, skin from tick http://www.csiro.au/.
bite site (in neutral buffered
formalin). Diagnosis
iii. Culture may be arranged by Clinical signs, histopathology, virology.
consulting RVL staff at EMAI. In
cases with polyarthritis, chilled joint Specimens required
fluid and (in PM material) joint i. Whole bat (chilled)
capsule is sought. In acute cases,
heparinised blood is suggested. MALIGNANT OEDEMA
Diagnosis
LYSSAVIRUS History of mortality, post mortem findings
Syn: Australian bat lyssavirus, pteropid and recovery of pathogenic clostridia from
lyssavirus lesions. Best diagnosed in the field because
of rapid autolysis of samples.
Australian bat lyssavirus is closely related to
classical rabies virus and causes similar Specimens required
progressive neurological disease in naturally- i. Affected tissues from affected
infected bats and humans. animal or freshly dead animal,
submitted chilled for bacteriology.
Flying foxes (fruit bats) and insectivorous ii. At least three impression smears
bats are affected. Subclinical infection from lesions for bacteriology.
apparently occurs in bats, which are the iii. Sections of affected tissues, in
reservoir of the virus. buffered formalin for histopathology.

Australian bat lyssavirus infection is Material must be taken from affected or


notifiable. Fees for tests undertaken to freshly dead animals, otherwise
confirm or exclude a diagnosis of lyssavirus examination is worthless.
infection are paid by NSW Department of
Primary Industries. MALNUTRITION
Caution Lyssavirus is a zoonotic See also: Feeds and pasture analyses
agent, causing fatal infections in humans.
Malnutrition is the most common cause of ill
Clinical signs in bats include behavioural thrift. The differential diagnosis should
changes (aggression/docility, shivering, include parasitism, infectious diseases,
salivation) and paralysis. Where disease is chronic conditions causing organ
suspected, bats should be euthanased and dysfunction (e.g. pyrrolizidine alkaloidosis)
sent to the laboratory for necropsy. All bats and deficiencies of particular nutrients (e.g.
should be handled with care to avoid bites selenium, copper, phosphorus).
and scratches.
Malnutrition should also be considered as a
The general public should be predisposing cause of many conditions, e.g.
discouraged from rescuing or handling pregnancy toxaemia, plant poisoning in
bats (paralysed, aggressive or unusually- drought situations.
docile bats are at high risk of being
clinically-affected). People bitten or MANGE
scratched by bats should consult a General Diagnosis
Practitioner immediately and seek Clinical signs, demonstration of parasites in
prophylactic vaccination. Individuals regularly deep skin scrapings or skin biopsy,
handling potentially-affected bats (wildlife histopathology.
carers, veterinary pathologists) should be
vaccinated against rabies.

VETERINARY LABORATORY MANUAL 70


Cases are best diagnosed in the field History, clinical signs, confirmed by DNA
from direct examination of multiple deep analysis or histopathology .
skin scrapings.
Specimens required
Specimens required See
i. Skin scrapings. http://www.dpi.nsw.gov.au/agriculture/vetm
ii. Skin biopsy from a recently affected anual/specimens-by-disease-
area, in buffered formalin for syndrome/diseases_of_livestock/dpi-samp-
histopathology (in cases where the coll-prot.pdf
disease is suspected but cannot be
confirmed from skin scrapings). (in order of preference)

MANNOSIDOSIS Diagnosis of the disease


i. 20 to 25 hairs, with roots attached,
from the distal end of the tail.
ALPHA-MANNOSIDOSIS ii. Lymphoid tissue and/or whole brain
in buffered formalin for
(Angus, Murray Grey and Galloway cattle)
histopathology.
Affected Angus and Murray Grey calves
either fail to survive the immediate Heterozygote detection
i. 20 to 25 hairs, with roots attached,
postnatal period, or if they do, they show
from the distal end of the tail.
severe, progressive neurological disease
characterised by tremors of the head,
ataxia, and aggression. The phenotype in MAPLE SYRUP URINE DISEASE
Galloways is more severe, with most (MSUD)
affected cases aborted or stillborn.
(Poll Hereford, Hereford and Poll Shorthorn
calves)
Diagnosis
History, clinical signs, confirmed by DNA
An inborn error in branched chain keto acid
analysis or histopathology.
metabolism that results in a progressive
neurological disease that culminates in
Specimens required
death before 5 days of age. At birth,
See
affected calves are clinically normal, but
http://www.dpi.nsw.gov.au/agriculture/vetm
within 24 hours they develop progressive
anual/specimens-by-disease-
neurological dysfunction characterised by
syndrome/diseases_of_livestock/dpi-samp-
uncoordinated limb movements that prevent
coll-prot.pdf
the calves from rising. Between 48 and 72
hours after birth the calves collapse and
(in order of preference, breed of subject
typically are often found in lateral
must be recorded)
recumbency with opisthotonos. Death
intervenes within 5 days of birth.
Diagnosis of the disease
i. 20 to 25 hairs, with roots attached,
There are no pathognomonic gross findings
from the distal end of the tail.
at necropsy. Widespread vacuolation of
ii. Lymphoid tissue and/or whole brain
white matter tracts in the brain and spinal
in buffered formalin for
cord is a common histological manifestation
histopathology.
of MSUD in Hereford and Shorthorn calves.
Diagnosis
Heterozygote detection
History, clinical signs, histopathology,
i. 20 to 25 hairs, with roots attached,
confirm by DNA analysis.
from the distal end of the tail.
Specimens required
BETA-MANNOSIDOSIS See
(Salers cattle) http://www.dpi.nsw.gov.au/agriculture/vetm
anual/specimens-by-disease-
Affected calves are usually born alive, but syndrome/diseases_of_livestock/dpi-samp-
exhibit severe neurological abnormalities. coll-prot.pdf
These include weakness, incoordination, (in order of preference, breed of the subject
head-swaying, splaying of the front legs, must be recorded)
and a poor sucking reflex.
Diagnosis of the disease
Diagnosis

VETERINARY LABORATORY MANUAL 71


i. 20 to 25 hairs, with roots attached, Mixed quarter samples are not
from the distal end of the tail. recommended as excessive dilution
ii. Whole brain in buffered formalin for of the pathogen can occur.
histopathology. • All samples should be taken prior to
antibiotic treatment. If antibiotic
Heterozygote detection treatment has been given,
i. 20 to 25 hairs, with roots attached, resample case at next occurrence
from the distal end of the tail. of clinical signs or at least three
weeks post treatment.
MASTITIS (BOVINE) • Before sample is taken, teat must
be washed and dried thoroughly
Diagnosis and teat end sterilised with 70%
Clinical mastitis alcohol or undiluted teat dip
History, clinical signs and bacteriology solution.
• Discard the first few squirts before
Subclinical mastitis
collecting sample.
Bacteriology, Rapid Mastitis Test, Mixed
Samples should be kept on ice until
quarter milk cell counts (individual cow cell
arrival at the laboratory.
counts) in excess of 250,000 averaged over
a lactation, elevated milk NAGase activity,
Interpretation of results
increased milk electrical conductivity.
Staphylococcus aureus, Streptococcus
uberis, Streptococcus dysgalactiae and
Herd mastitis problem
Streptococcus spp (environmental) and are
Evidence of subclinical mastitis (see
the most common pathogens found in both
above), problems in milking parlour
clinical and subclinical mastitis. In mastitis
management or milking machine operation,
cases occurring at or near calving,
environmental factors such as
Escherichia coli, Enterobacter spp and
contaminated water supply or muddy
other coliforms may be present. Serratia
conditions.
spp and Klebsiella spp can cause persistent
mastitis symptoms.
Bacteriology and antibiotic sensitivity
testing is performed by the Regional
Recovery of milk quality in quarters affected
Veterinary Laboratory. Individual cow cell
by these pathogens may not occur.
counts are available through Herd
Incurable mastitis can be caused by
Recording. Cost of cell counts depends
Pseudomonas aeruginosa. Nocardia spp
upon whether dairy farmer is a herd
and Arcanobacterium (Actinomyces)
recording member or not.
pyogenes. Recovery of the former two
pathogens from mastitis cases should
History required
indicate cull for slaughter of the affected
All mastitis submissions should be
cow especially if Nocardia sp are involved
accompanied by full history of the mastitis
as these pathogens pose a potential human
case. For clinical cases of mastitis this
health risk. Mastitis due to Arcanobacterium
should include breed, age, previous
pyogenes usually necessitates drying off
antibiotic treatment (both for previous
affected quarter after treatment and
clinicals and dry cow treatment), stage of
reassessing damage to the quarter after
lactation, previous mastitis history.
subsequent calving.
If a herd problem of clinical mastitis is
Recurrent cases of mastitis are usually due
occurring, additional information required is
to chronic staphylococcal infection. All
number of cows involved, description of
recurrent cases should be cultured to check
clinical signs and any recent changes in
cause. Antibiotic sensitivity necessary for
farm management or milking parlour
recurrent cases. Of the current S4
management.
antibiotics registered for clinical and
subclinical mastitis, cloxacillin products are
Subclinical mastitis problems require an
effective for all staphylococcal and
intensive investigation of milking parlour
streptococcal infections and neomycin-
operation. Regional veterinary or dairy
based products effective for staphylococcal
livestock officers may provide assistance in
and enviromental mastitis pathogens such
these investigations.
as coliforms. If other antibiotic products are
used, antibiotic sensitivity will be necessary.
Specimens required
i. Approximately 20 ml of foremilk
collected aseptically from affected
quarter into a sterile container.

VETERINARY LABORATORY MANUAL 72


MASTITIS (CAPRINE) MASTITIS METRITIS AGALACTIA
Diagnosis SYNDROME IN SOWS
Bacteriology is the most effective diagnostic Occurs from 12 to 48 hours after parturition
method. Rapid Mastitis Test can be used. with a rapid fall off in milk production,
Cell counts and NAGase are unreliable in toxaemia, constipation, fever and vaginal
herds with caprine arthritis encephalitis discharges in the sow.
(CAE)virus infection.
Diagnosis
History and specimens required The cause or causes of the syndrome are
As for bovine samples. not clearly understood. Management
factors are implicated.
Interpretation of results
Coagulase-negative staphylococci Specimens required
(Staphylococcus epidermidis) are the most Samples of vaginal discharges are of
common cause of subclinical mastitis in limited value because of the wide range of
dairy goats. organisms normally present in the vagina of
the sow.
Clinical mastitis can be caused by
coagulase negative staphylococci, At present, there is little that can be offered
Staphylococcus aureus, Streptococcus spp. in the laboratory in relation to this condition.
and enterobacteria such as Escherichia
coli, Serratia spp and Klebsiella spp. MENANGLE VIRUS INFECTION
Care must be taken with treatment. Menangle virus infection was first recognised
Intramammary antibiotics will have when it caused a single outbreak of
extended witholding periods in goats reproductive disease in pigs in a large
compared to cows. The blue dye in these piggery near Sydney in 1997. A control
antibiotics can be excreted in the milk of program, successfully managed by NSW
treated goats for extended periods of time. Agriculture, subsequently eradicated the
Parenteral antibiotics may be used. virus from the piggery. A large serological
survey failed to find any evidence of infection
MASTITIS (OVINE) in other Australian pigs. No further outbreaks
of the disease have occurred to date.
Ovine mastitis may be acute/severe,
gangrenous, chronic or subclinical. The Flying foxes (fruit bats) near the piggery had
most frequently isolated bacteria are antibodies to the Menangle virus. It is
Staphylococcus aureus, Mannheimia believed that flying foxes are the natural host
(Pasteurella) haemolytica, Arcanobacterium of the virus, and that it was a relatively rare
pyogenes, and Streptococcus spp. event that led to the virus 'jumping' into pigs.
Coagulase-negative Staphylococci, Bacillus
spp, Micrococcus spp and E coli may also Menangle virus is a member of the
be incriminated in subclinical disease. Less paramyxovirus family of viruses, and is
commonly Corynebacterium closely related to Tioman virus, isolated in
pseudotuberculosis, Histophilus ovis, 1999 from fruit bats in Malaysia, but which
Actinobacillus seminis, Pseudomonas spp, has not yet been associated with disease in
Proteus spp and Klebsiella spp are any animal species.
involved. Clostridium perfringens is
associated with gangrenous mastitis (also S Menangle virus infection is notifiable.
aureus and E coli). Mycoplasma spp can Fees for tests undertaken to confirm or
also cause ovine mastitis. exclude a diagnosis of Menangle virus
infection are paid by NSW Department of
Diagnosis Primary Industries.
Bacteriology. Other tests including Somatic
cell count (high variability makes Clinical signs include a severe decline in
interpretation difficult; also epithelial cell farrowing rate, a high incidence of delayed
shedding as a normal phenomenon returns to service and a marked increase in
increases SCC thresholds for identifying the number of mummified and stillborn
mastitic milk), Rapid mastitis test, and piglets, some with severe deformities.
NAGase can be applied on a herd basis. Stillborn piglets had a severe non-
suppurative encephalomyelitis. Although
History and specimens required many pigs were infected, the virus was not
As for bovine samples. highly contagious and thus probably spread

VETERINARY LABORATORY MANUAL 73


by excretion in faeces and urine, rather than
by respiratory aerosols. MUSCULAR DEGENERATION,
NUTRITIONAL
There was also a severe influenza-like
illness in two of the workers at the piggery. Syn: Selenium/vitamin E deficiency, white
The two workers recovered from the illness. muscle disease, nutritional myopathy,
‘muscular dystrophy’’

Diagnosis This disease complex usually occurs as


History, clinical signs, histopathology, defined clinical and pathological syndromes
virology. which bear a several names. They are
thought to have a nutritional pathogenesis
Specimens required involving a dietary deficiency of selenium
i. Chilled stillborn piglets for necropsy, and/or vitamin E.
virology and histopathology.
ii. Fresh heart, lung, spleen, brain and Diagnosis
body fluids collected separately and History, clinical signs, post mortem
aseptically, submitted chilled for examination, histopathology, biochemistry,
virology. response to therapy.
iii. Sections of heart, liver, kidney, lung,
spleen and whole brain in buffered Specimens required
formalin for histopathology. i. 10 ml of heparinised blood
submitted chilled (not frozen) for
METABOLIC DISEASES glutathione peroxidase (GSHPx)
concentrations, from at least 5
See 'hypocalcaemia, hypomagnesaemia, animals in the group.
hyophosphataemia, biochemistry: guide to
biochemical parameters in sheep and cattle Selenium status can be measured directly
by analysis of blood or tissue selenium
MUCOSAL DISEASE concentrations, or indirectly by analysis of
blood glutathione peroxidase concentration.
See Pestivirus, abortion, arthrogryposis and
The latter, as in most enzyme tests,
hydranencephaly'
requires heparinized blood (EDTA is
unsuitable). The correlation between
MULBERRY HEART DISEASE glutathione peroxidase (GSHPx) and
A cause of sporadic mortalities in rapidly selenium concentrations is very high in non-
growing pigs, characterised by pronounced deficient animals. In deficient animals, this
myocardial haemorrhage. Many cases are high mathematical correlation is not
associated with vitamin E and/or selenium present, but of little diagnostic
deficiency, but these are not the sole consequence, i.e. in deficient animals, both
cause. More cases tend to occur under GSHPx and Se concentrations are low but
conditions of heat stress. there is not necessarily a close
mathematical correlation between values.
Diagnosis
History and characteristic necropsy findings ii. Sections of affected and unaffected
of petechial and ecchymotic haemorrhages skeletal muscle and cardiac muscle,
in the heart, and distension of the in buffered formalin for
pericardium by a serofibrinous transudate. histopathology.
Liver necrosis/ rupture/ haemorrhage is a iii. At least 2 ml of serum, free of cells
feature in many cases. and haemolysis, submitted frozen
for serum enzymology.
Specimens required iv. 20 g liver, submitted chilled and
i. Heart, liver and brain, in buffered protected from light immediately
formalin for histopathology. following collection. (See Collection
ii. Kidney and liver submitted chilled or of specimens for biochemistry and
frozen for selenium estimation. Vitamin deficiency).
iii. 10 ml of heparinised blood,
submitted chilled (not frozen) for NB. Whenever possible, submit heparinised
biochemistry from 5 to 10 animals in blood from 3 5 peer animals rather than
the same pen. tissues of dead cases

VETERINARY LABORATORY MANUAL 74


Interpretation of blood glutathione peroxidase (GSHPx) concentrations
The following values should be taken only as a guide:

GSHPx U/g Hb
Interpretation Cattle Sheep
Disease < 10 <10
Production response < 25 <25
Normal > 100 >50

Note that current blood concentration of be toxic to mycoplasmas if left in contact for
GSHPx reflect the selenium intake of the extended periods.
animal at the time the present circulating
erythrocytes were formed. An ‘average’ Diagnosis
time would be the half life of the erythrocyte
for the species and age of animals to be Pigs
examined (e.g. adult cattle approx. 80 days, Culture of some pathogenic mycoplasma
adult sheep approx. 60 days, calves and species can be undertaken using media
lambs approx. 50 days). specific for pig mycoplasmas.
Identification of M. hyopneumoniae in lung
The interpretation of selenium status and tissues or nasal swabs by PCR.
prognosis is further complicated by factors Identification of M. hyopneumoniae herd
which include: infection by ELISA serology.
• Season (winter and summer peaks,
autumn and spring troughs of Se NB: M. hyorhinis is a common inhabitant of
availability). the URT and ears (Eustachian tube), and is
• Soil type (acid soils < pH 5.5 have a common co-inhabitant of pneumonic
50% reduction in Se availability to lungs. It has no pneumonic potential and
plants). can rapidly overgrow M. hyopneumoniae
• Pasture type (legumes and other leafy cultures. It can cause a serofibrinous
plants lower in Se than grasses). polyserositis, or fibrinous arthritis in suckers
• Fertilizer history (competitive or weaners less than 10 weeks old. It
interaction from sulphur results in produces a milder, more sporadic disease
reduced selenium in pastures on than Glasser's disease, and can
superphosphate fertilized soils). occasionally affect young adults. (DD:
• Age of animals preruminant animals Glasser's disease, Streptococcus suis,
more prone to deficiency than Erysipelas).
ruminants.
M. hyosynoviae causes synovitis and
For diagnosis of selenium deficiency in arthritis
sheep, it is recommended that weaners be
sampled after access to good feed (e.g. (DD: Erysipelas; M. hyorhinis polyserositis
clover). Older sheep normally have lower of young pigs; Glasser's disease
levels. Diagnosis should be considered on polyserositis or respiratory disease;
a property basis. Pasteurella spp synovitis; Streptococcus
suis purulent arthritis or polyserositis; other
Streptococci causing purulent arthritis with
MYCOPLASMOSIS
percutaneous infections/abrasions; leg
See also Porcine enzootic pneumonia weakness/osteochondrosis)

Mycoplasma infections cause a wide range Cattle, sheep and goats


of clinical conditions, particularly among Culture of affected tissue/s.
pigs, cattle, sheep, goats, and poultry.
Certain mycoplasmas are part of normal Mycoplasma spp bovine Group 7 is a
mucosal flora, and can outgrow pathogenic frequent cause of polyarthritis in calves. It
mycoplasmas in broth culture procedures. can also cause outbreaks of bovine mastitis
and abortion. This organism is readily
In general, special media is required for cultured, and is occasionally cultivable on
growth of mycoplasmas. Some blood agar cultures without special
mycoplasmas are slow growing, taking up mycoplasmal media. Other Mycoplasma
to 3 weeks to appear in primary culture or in spp are associated with bovine mastitis,
subcultures. eye lesions, reproductive tract infections,
Avoid submission of swabs for and caprine and ovine arthritis and mastitis.
mycoplasmal culture. Swab materials can

VETERINARY LABORATORY MANUAL 75


Specimens required concentrations of toxins in the feed the
animals ingested. Often, because of the
Pigs expense of mycotoxin analysis, the
i. Serum (or heparin plasma) samples diagnosis is made by elimination of other
for M. hyopneumoniae ELISA.. causes.
ii. Fresh chilled or frozen lung for M.
hyopneumoniae PCR (1 cm³). Mycotoxin analysis in feed is a specialised
iii. Nasal swab material collected into area of chemistry not offered by NSW
sterile PBS for M. hyopneumoniae Department of Primary Industries. Our
PCR. Laboratories will outsource this testing on
iv. Fresh chilled joint fluid or membrane request.
for M. hyosynoviae culture.
v. Fresh chilled tissues or joints Specimens required
affected with polyserositis/arthritis i. Sections of liver and kidney, in
for M. hyorhinis culture (suckers, buffered formalin for histopathology
weaners). to demonstrate toxic effects.
vi. Fixed lung for histopathology. ii. 500g of feed for mycotoxin analysis.
iii. Specimens for differential diagnosis,
particularly where nervous system
Cattle, sheep, goats disease is seen clinically, e.g.
i. Depending on the pathogen, fresh Fusarium spp. (see Nervous
tissues, joint fluid, joint membrane, disorders).
ocular fluid, milk for mycoplasmal
culture. NASAL GRANULOMA OF CATTLE
ii. Sections of affected organs in
buffered formalin for histopathology. Diagnosis
Clinical signs, histopathology.

Poultry Specimens required


i. Serum for Mycoplasma i. Section of affected nasal mucosa, in
gallisepticum (MG) and Mycoplasma buffered formalin for histopathology.
synoviae (MS) antibody detection by ii. Section of affected nasal mucosa,
rapid plate test. submitted fresh for bacteriology and
mycology*.
NB Culture for avian mycoplasmas is not
routinely undertaken. Specialised services *Tissues for fungal culture should not be
are available at some other laboratories. refrigerated or frozen (temperatures of less
o
than 15 C can be detrimental to fungal
MYCOTIC DERMATITIS survival); such specimens should be stored
and transported between 18oC and 37oC.
See Dermatophilosis
NECROBACILLOSIS
MYCOTOXICOSIS
Syn: Calf diphtheria, necrotic stomatitis of
The effects of mycotoxins vary according to calves, 'necrobacillosis of lambs'
species and age of the animal, the quantity
of mycotoxin ingested and the period of Diagnosis
exposure. Clinical signs, necropsy findings and
demonstration of Fusobacterium
Mycotoxicoses include aflatoxicosis, necrophorum in lesions. In lambs, multiple
ergotism, facial eczema, Fusarium spp large creamy/caseous necrotic liver and
toxicosis, paspalum staggers, tremorgen lung lesions (and occasionally
intoxication, lupinosis and mouldy corn diaphragmatic lesions) are associated with
poisoning. entry of Fusobacterium necrophorum via
primary lesions in the rumen and/or
Diagnosis abomasum.
History, clinical signs, histopathology and
demonstration of toxic concentrations of Specimens required
mycotoxin in feed. Demonstration of a i. Smears taken from the depth of the
toxigenic species of fungus in the feed is lesions.
not enough, the fungi are ubiquitous and ii. Portion of lesion submitted chilled
only some strains are toxigenic. for bacteriology.
Mycotoxicosis can only be confirmed by
demonstrating biologically effective

VETERINARY LABORATORY MANUAL 76


iii. Sections of affected organs,
submitted in buffered formalin for Dog
histopathology. Clinical signs (neurological disease),
histopathology, serology.
NEOPLASMS
Specimens required
In cases of biopsies from the live animal, a Cattle
detailed description is necessary, especially Aborted foetuses
in those cases where the field veterinarian i. Specimens as required for diagnosis
is seeking a prognosis. of abortion in cattle (see Abortion in
cattle), particularly:
Diagnosis • serous foetal body fluid
History, clinical signs, gross pathology and (pericardial, thoracic or peritoneal)
histopathology. or heart blood for serology
• brain and myocardium (including
The history should include species, age, from autolyzed foetuses) formalin-
sex, breed and full description of the tumour fixed for histopathology
giving its location and the rate of Cows
development of the tumour. i. 10 ml of clotted blood in vacuum
tube for serology. A group of
Specimens required aborted and non-aborted animal
i. Sections of affected organs, in should be tested. In view of the
buffered formalin for histopathology. widespread prevalence of subclinical
neosporosis in cattle in coastal NSW
The sections should be taken both from the areas, seropositivity in an aborted
centre of the lesion and also from around cow does not confirm Neospora
the margin, at the junction of normal and abortion.
abnormal tissue. Dogs
i. Spinal cord and brain and other
NEOSPOROSIS tissue formalin-fixed for
Neospora caninum is a coccidian parasite histopathology.
related to Toxoplasma gondii. The dog is ii. Clotted blood in vacuum tube for
the definitive host, with sexual stages in the serology.
gut resulting in oocysts shed in faeces.
NERVOUS DISORDERS
Cattle (an intermediate host species) are See also Ataxia', encephalomyelitis, and
infected by ingesting feed contaminated by specific disease syndromes, including
sporulated N caninum oocysts shed by transmissible spongiform encephalopathy
dogs or by infected cattle tissues (eg (TSE).
infected foetal membranes), or are infected
transplacentally. Nervous signs can arise from a wide range
of causes, including bacterial, viral,
The asexual proliferative stages of parasitic and protozoon infections, chemical
Neospora caninum (including tissue cysts), and plant poisonings and metabolic and
cause: genetic conditions.
• Abortion in cattle (world-wide,
including coastal NSW) and Laboratory diagnosis of the cause or
congenital neonatal neurological causes will require a range of specimens to
disease in cattle (rarely). Neospora be submitted, together with a detailed
abortion is reported in deer history and clinical examination.
(rarely).
• Neurological disease in the dog For adult sheep and cattle with
(rarely) progressive neurological disease,
please submit specimens as required by
Diagnosis the National TSE Surveillance Program
Cattle (http://www.animalhealthaustralia.com.au/a
History, clinical signs (abortion), ahc/programs/adsp/tsefap/tsefap_home.cf
histopathology, serology. m), accompanied by a completed
NTSESP Clinical History and Post-
Distinctive histological changes are seen in mortem Report form.
brain (necrogranulomatous encephalitis)
and myocardium (non-suppurative Diagnosis
myocarditis) of aborted foetuses.

VETERINARY LABORATORY MANUAL 77


History, clinical signs, histopathology and OPHTHALMIA
biochemistry.
Syn: Pink-Eye
Specimens required
i. Portion of brain, spinal cord and Diagnosis
cerebrospinal fluid, collected Clinical signs, identification of causal
aseptically and submitted chilled for organisms (Moraxella bovis or IBR in cattle,
bacteriology and virology. Branhamella spp or Chlamydia in sheep).
ii. Brain and spinal cord, in buffered
formalin for histopathology. Specimens required
iii. Serum sample from the affected i. Smears and swabs of conjunctival
animal, submitted chilled for surface, submitted chilled for
serology. bacteriology.
iv. At least 3 ml of serum, free of cells ii. Swabs of conjunctival sac in
and haemolysis, submitted chilled or phosphate buffered gelatin saline
frozen for enzymology and (PBGS), together with paired sera,
biochemistry. submitted chilled for virology.
v. Other specimens as required for
specific diseases. Moraxella bovis can be recovered
consistently from the conjunctival sac only
when blood agar plates are inoculated
NITRATE-NITRITE POISONING
immediately swabs are taken.
Diagnosis Arrangements can be made with the
History, clinical signs, particularly laboratory for the supply of blood agar
methaemoglobinaemia, chemical plates in special circumstances where the
confirmation of nitrate or nitrite in the isolation of Moraxella bovis is desirable.
animal.
ORGANOCHLORINE AND
Specimens required ORGANOPHOSPHATE POISONING
Either,
i. 1 ml of serum submitted chilled for There may be some delay (approximately 2
toxicology, or, weeks) in testing samples for
ii. Thick, air dried blood smears for organochlorines and organophosphates.
toxicology. Testing is undertaken at NSW Department
of Primary Industries' laboratory at
NB. Testing of plant material, serum or Wollongbar for a range of OC and OP
blood is best performed in the field. compounds. Currently analytical methods
for pesticides in ingesta or tissues are
Appropriate specimens should also be available for the following chemicals:
taken to eliminate other causes of sudden
death. Organochlorines
Aldrin, Dieldrin, Endrin, Lindane, DDE,
OEDEMA DISEASE OF PIGS DDD, DDT, HCB, Heptachlor, Heptachlor
Epoxide, Alpha Chlordane, Gamma
Diagnosis Chlordane, Alpha BHC and Beta BHC.
Clinical signs, necropsy findings and the
recovery of enteropathogenic Escherichia Organophosphates
coli. Bromophos Ethyl, Chlorfenvinphos,
Chlorpyrifos, Chlorpyrifos Methyl, Diazinon,
Specimens required Dichlorvos, Ethion, Fenitrothion, Fenthion,
Malathion, Methacrifos, Methyl Parathion,
i. Segment of ileum or swab in Parathion, Pirimiphos Methyl, Profenophos,
transport medium for bacteriology. Sulprofos, and others.
ii. Brain and sections of jejunum and
ileum in buffered formalin for Diagnosis
histopathology. Based on clinical findings, response to
iii. Range of fresh and fixed tissues for treatment and toxicology. There are no
differential diagnosis. pathological changes observed at
iv. Moribund or freshly dead animal for necropsy. The history should include details
necropsy and bacteriological of the specific insecticides to which the
examination. animals may have had access (see also
'Specimens for Toxicology').

Specimens required

VETERINARY LABORATORY MANUAL 78


Acute exposures serum or plasma pepsinogen levels (see
i. At least 20 g of stomach contents, Pepsinogen estimations from serum or
liver and brain submitted frozen for plasma').
toxicology.
ii. Sections of liver, kidney, lung, brain Specimens required
and any other organs with gross Live animal
lesions, should be submitted in i. At least 30 g of fresh faeces in a
buffered formalin for differential screwtop jar filled to capacity,
diagnosis on histopathology. submitted chilled for egg count and
larval culture.
Chronic exposures ii. In cattle, serum or plasma
i. At least 20 g of fat submitted frozen pepsinogen may be useful,
for toxicology particularly in growing animals.
ii. Sections of liver, kidney, lung, brain
and any other organs with gross
lesions, should be submitted in Dead animal
buffered formalin for differential i. At least 30 g of fresh faeces as
diagnosis on histopathology. above.
ii. The abomasum and abomasal
OSTEOCHONDROSIS IN PIGS contents whole, with both ends tied
off (also tied at the abomaso-
A condition characterised by joint duodenal and ileocaecal junctions),
abnormalities in rapidly growing pigs, submitted chilled or frozen for total
usually seen in baconers. worm count and abomasal digest for
recovery of histotrophic larvae.
Diagnosis
History, clinical signs and gross pathology.
OVINE BRUCELLOSIS
Specimens required See brucellosis ovine'
Affected limbs, with unopened joints,
submitted chilled for bacteriology, gross OXALATE POISONING
pathology and histopathology.
Diagnosis
History of access to plants containing
OSTEOMALACIA, OSTEOPOROSIS
oxalate, clinical findings, hypocalcaemia,
Disease associated with transient and histopathology.
shifting lameness, bone fractures, perverted
appetite and tendency to bone chewing. Specimens required
Affected bones are light, porous and soft, i. Sections of liver and kidney,
and often the seat of healing fractures. submitted in buffered formalin for
histopathology.
Diagnosis ii. At least 2 ml of serum, free of
Clinical signs, gross pathology, haemolysis and cells, submitted
histopathology. chilled for calcium estimation. (see
also Hypocalcaemia).
Most cases are best diagnosed on clinical
signs and gross pathology. Histological PAPULAR STOMATITIS OF CALVES
examination is performed only in certain
cases, following laboratory consultation Diagnosis
Clinical signs, electron microscopy.
Specimens required
Specimens required
i. Sections of affected bone, submitted i. Digestive tract lesions fixed in
in buffered formalin for buffered formalin for histopathology,
histopathology. if required for differential diagnosis.
ii. Scabs or necrotic debris in a sealed
container, submitted chilled for
OSTERTAGIOSIS
electron microscopy.
See also Parasites internal
PARAKERATOSIS OF SWINE
Diagnosis
Grazing history, age of affected animals, Diagnosis
clinical examination, faecal egg count and Clinical signs differentiation sarcoptic
larval culture, total worm counts. In cattle, mange and exudative epidermitis;
histopathology.

VETERINARY LABORATORY MANUAL 79


can be collected from "struck" wool
Specimens required by placing the wool on a sheet of
i. Portions of oesophagus, tongue and paper in sunlight.
affected skin, submitted in buffered iii. Mange mites - scrape recent
formalin for histopathology. lesions, using a scalpel moistened
ii. 10 mL clotted blood tube or 2 mL with liquid paraffin. Transfer
serum for zinc analysis. scrapings to a small wide mouthed
bottle and submit unpreserved.
PARAMPHISTOMIASIS
PARASITES (INTERNAL)
See also Parasites internal'
See also Ostertagiosis, liver fluke infection,
Diagnosis paramphistomiasis'
Grazing history and associated seasonal
conditions. Clinical findings, demonstration Faecal egg counts and differential larval
of immature paramphistomes in the counts are a guide to the size and type of
duodenum or adult flukes in the rumen and worm burden. The faecal egg count
reticulum. depends on a number of factors including
faecal consistency and bulk, host
Specimens required resistance, stage of pregnancy and effects
Dead animal of lactation, as well as the parasite involved
i. The abomasum and small intestine, and whether the parasites are sexually
unopened and tied off at each end, mature.
submitted chilled or frozen for
parasitology. The history should provide details of all
recent anthelmintic treatments and flock
NB Faecal egg counts are of no value in management, e.g. stocking rates, pasture
diagnosing clinical paramphistomiasis. The availability, swampy areas, paddock
presence of paramphistome eggs indicates movement, etc.
non-pathogenic adult stomach flukes are
present. Adult paramphistomes may confer Further information on internal parasites of
some immunity. sheep or cattle is available on:
http://www.agric.nsw.gov.au/reader/sheep-
PARASITES (EXTERNAL) internal
http://www.agric.nsw.gov.au/reader/cattlehe
See also Itchmite, mange' alth
Diagnosis Diagnosis
Based on clinical findings and Clinical signs, faecal egg count, larval
demonstration and identification of parasite cultures and total worm count.
on skin scraping or skin biopsy.
Specimens required
Specimens required for diagnosis Live animal
i. Skin scrapings, submitted i. At least 30 g of faeces collected
unpreserved for parasitological from the rectum, submitted in a
examination. screw top jar filled to capacity for
ii. In cases where parasites cannot be egg count and larval culture. In flock
demonstrated in skin scrapings in or herd investigations, samples
the field, a skin biopsy sample from should be collected from 10 to 20
an affected area, submitted in animals showing evidence of
buffered formalin for histopathology. parasitism.
Specimens required for parasite WormTest is a convenient system for
identification collecting and submitting faecal
i. Flies, lice, fleas, ticks, spiders, etc. - samples to the laboratory.
submit preserved in alcohol/
glycerine (9 parts of 80 per cent ii. For F hepatica in cattle, serum
alcohol to 1 part glycerine) in a small sample chilled for ELISA serology.
leak proof container. Milk ELISA testing for liver fluke is
ii. Insect larvae - where live larvae are currently (2004) under validation; check
required for insecticide tests, with your veterinary laboratory for
consign about fifty in a ventilated availability.
container with about 3 cm damp
vermiculite in the bottom. Larvae Dead animal

VETERINARY LABORATORY MANUAL 80


Preferably:
i. The whole gastrointestinal tract Faecal egg counts are generally lower from
(double bagged in strong clear cattle than from sheep. If sheep are starved
plastic) submitted unopened, but for 24 hours, the count may be increased.
tied off, either chilled or frozen for Inappetence may cause the count to
parasitological examination. multiply 30 to 40 times. Diarrhoea
depresses the egg count.
Alternatively
i. Aliquot samples of 1 L from the Any statement of 'significant figures' for ova
stomach and small intestines, counts will be only a rough guide.
submitted in a 5% formalin solution Interpretation must be considered in
for parasitological examination. relation to the specific parasite responsible
ii. The abomasum and intestine, (as indicated by larval cultures) and the
submitted chilled or frozen for age, origin, state of nutrition and clinical
examination for immature parasites. history of the infected animals.

FAECAL EGG COUNTS See AgNote: WormTest for livestock and


(INTERPRETATION) guide to egg counts.
Faecal worm egg counts and differential
larval counts are a guide to the parasite Sheep
burden. The number of helminth ova An egg count of 500 eggs per gram (epg) is
passed per gram of faeces depends on generally considered high enough to
such factors as faecal consistency and require treatment in order to limit pasture
bulk, host resistance, stage of pregnancy, contamination and subclinical disease. The
effects of lactation and whether the worm following numbers of epg may indicate
burden consists of sexually mature clinical disease due to that parasite.
parasites.

Guide to faecal egg counts in sheep (indicating pathogenic burdens)

Species Eggs per gram of faeces


Young Sheep Older Sheep
Haemonchus contortus 2,000 2,000
Ostertagia spp. 500 500
Trichostrongylus spp. 500 1,000
Nematodirus spp. >200 500
Oesophagostomum columbianum 300 1,000
Chabertia ovina 500 1,000
Fasciola hepatica 100 >100
Paramphistomes >500
*Compiled from various reference sources: Cole VG (1986) Appendix 1, pp 233-239; Love SCJ,
Hutchinson GW (2003) Table 4, pp 329-333, Skerman KD, Hillard JJ (1966) p7.

The egg laying capacity of Ostertagia spp considered. This is of particular significance
and Nematodirus spp is poor and severe with Nematodirus, Ostertagia, Chabertia,
clinical signs may be seen before Fasciola hepatica and paramphistomes.
appreciable numbers of eggs are present in
the faeces. Dictyocaulus filaria is often associated with
mixed gastrointestinal infections on the
Low and medium egg counts will be more tablelands and slopes.
significant where the stocking rate is high,
when weather conditions are conducive to Cattle
epidemics (warmth, rain, humidity) and The clinical history and knowledge of the
where the biotic potential is high, e.g. seasonal pattern of worm parasites in
Haemonchus contortus. different areas of the State will assist in
interpretation of faecal egg counts.
Infections with one parasite only are rarely
seen and the additive effects of mixed In cattle greater than 18 months of age, the
infections will require assessment. The egg count gives little indication of the level
pathogenicity of immature stages not of the parasite burden.
indicated by egg count should always be

VETERINARY LABORATORY MANUAL 81


Guide to faecal egg counts in cattle (indicating possible pathogenic burdens in 6-18 month
old cattle)*

Species Eggs per gram of faeces


Ostertagia spp. 300->1,500
(<300 possibly significant)
Trichostrongylus axei 500-1,000
(medium infections)
Haemonchus placei 700->1,500
Cooperia spp (in 3-4 month old calves) 1,000-5,000
(10,000-30,000 in acute disease)
Oesophagostomum radiatum. >500
Bunostomum spp 500-800
(heavy infections)
Fasciola hepatica Any egg count is significant
Heavy infections may be indicated by >25 epg.
However, there is little relation between egg count and
fluke burden.
Paramphistomes >500
(heavy adult infection)
*Compiled from various reference sources: Cole VG (1986) Appendix 2, pp240-245; Love SCJ,
Hutchinson GW (2003) Table 5, pp 334-338 ; Skerman KD, Hillard JJ (1966) p 8; Smeal MG (1995)
pp 358.

Horses Haemonchus contortus. In young sheep


600 epg or more may represent a 500 worms (light), 3,000 (heavy) and in
pathogenic cyathostomin (small strongyle) adults 1,000 (light), 9,000 (heavy).
burden.
Ostertagia spp. 3,000 5,000 worms is a
Colic induced by verminous arteritis from heavy infection in young sheep; 5,000-
migrating Strongylus vulgaris larvae 10,000 worms in adult animals may
(without eggs in faeces), is not as common cause mortality. Usually there is a
as in previous times. concurrent infection with
Trichostrongylus spp. and other
Pigs intestinal worms.
Any positive count of Ascaris suum ova in
pigs up to 5 months of age is significant. Trichostrongylus axei. Heavy infections
Young pigs with heavy immature worm (>5,000 worms) may cause
burdens may not be passing any eggs in unthriftiness and gastritis.
the faeces. In infected pigs, eggs generally
appear after the age of 9 to 10 weeks. Small intestine
Trichostrongylus spp. In young poorly
TOTAL WORM COUNTS grown weaners, 5,000 worms may
(INTERPRETATION) cause mortality, while in older weaners,
Interpretation of total and differential worm 20,000 worms can be considered a
counts is not absolute, but depends on heavy infection. A similar number of
clinical history and several interacting worms in adult crossbred sheep can be
factors such as age, sex and nutritional responsible for ill thrift and scouring but
status of the host. Opinions vary on the probably 30,000 40,000 worms are
significance of counts for various worm needed to cause mortalities.
species.
Nematodirus spp. In young sheep
Sheep 1,000 2,000 worms is considered a light
Rumen and Reticulum burden, but heavier infections of 5,000-
Paramphistomes. The immature forms 10,000 worms may be an important
occur in the small intestine. Heavy cause of diarrhoea, ill thrift and
infections of adults, e.g. 5000 sometimes mortality. Usually observed
paramphistomes may be associated as a concurrent infection with
with ill health. Trichostrongylus spp. In the absence of
worm egg counts, the importance of
Abomasum immature stages of Nematodirus spp.
should be considered in the diagnosis
of clinical parasitism.

VETERINARY LABORATORY MANUAL 82


infections of adults, e.g. 5,000
Strongyloides spp. Not considered of paramphistomes, may be associated
pathogenic importance under field with ill health.
conditions except in young lambs when
infection is heavy (10,000-15,000). May Abomasum
be a problem under penned conditions. Haemonchus placei. In 6-month-old
calves, 5,000 adult worms may be
Cooperia spp. Heavy infections in pathogenic causing anaemia, sub-
young lambs may be of importance. mandibular oedema, and sometimes
diarrhoea. Less important in cattle than
Paramphistomes. Infections with > H contortus is in sheep.
5,000 immature stomach fluke cause
clinical disease. (80% of immatures Ostertagia ostertagi. An important
are in duodenum). pathogenic parasite of both young and
adult cattle as 30,000-50,000 adult
Moniezia spp. Questionable worms may cause diarrhoea, weight
importance. loss, anaemia and harsh coat. The
number of immature stages in the
Large intestine histotrophic form in the abomasal wall
Oesophagostomum spp. There may be should be considered when planning
a heavy infection with larval forms, or anthelmintic treatments against
very marked nodule formation in outbreaks of ostertagiosis. Following a
previously exposed older sheep due to single anthelmintic treatment which
Oesophagostomum columbianum, removes adult worms, the ensuing
when adults may be rare and egg count development of up to, 50,000-90,000
consequently low. Oesophagostomum immature worms could easily cause a
columbianum is usually restricted to second serious wave of adult infection
northern and western district pastoral (Type II ostertagiosis). Ostertagiosis
areas of NSW. Serious chronic effects should be considered in the differential
are caused by severe nodule formation. diagnosis of ill thrift and loss of
In young sheep 100 worms is a serious condition in adult cattle, including bulls.
infection and in adult animals 100-200
worms may be significant. Trichostrongylus axei. Usually found in
association with Ostertagia ostertagi,
Chabertia ovina. Both immature and but in lower numbers.
adult worms can cause ill effects. In
both young and adult sheep, 100-200 Small intestine
worms is a heavy infection. Cooperia spp. The most common small
intestinal worm. Always occurs in mixed
Trichuris ovis. Heavy infections have infections with Ostertagia ostertagi.
been observed in sheep during Heavy burdens of more than 200,000
prolonged drought periods. worms may occur in dairy calves, but
Inflammatory lesions in the caecum clinical signs are due primarily to
resulting from large numbers of ostertagiosis.
parasites probably cause scouring and
ill thrift. Trichostrongylus spp. Rarely found in
large numbers.
Liver
Fasciola hepatica. Mortalities from the Bunostomum phlebotomum. In young
acute disease may be associated with calves 500 worms may be significant.
700 or more immature liver fluke and This hookworm causes anaemia,
from the chronic disease with 50 or haemorrhagic enteritis and dark, foetid
more adult fluke. scour.
Lungs
Lung worms (Dictyocaulus filaria, Moniezia spp. Common in calves but
Muellerius capillaris) are relatively rare relatively unimportant.
in sheep. Greater than 50-100 worms
may be considered a serious burden. Nematodirus spp. Rarely found in large
numbers. Heavy infections may be
Cattle seen in dairy calves (10,000 worms).
Rumen and Reticulum Low numbers (2,000-3,000) may be
Paramphistomes. The immature forms found in young cattle in concurrent
occur in the small intestine. Heavy

VETERINARY LABORATORY MANUAL 83


infections with other intestinal trend to free-range pigs continues.
trichostrongyles. Heavy and pathogenic burdens of 50-
100 larvae.
Paramphistomes. Infections with
>10,000 immature stomach fluke cause References
clinical disease. (80% of immature
worms are in duodenum). Cole VG (1986). Animal Health in
Australia Volume 8, Helminth Parasites
Large intestine of Sheep and Cattle. Australian
Oesophagostomum radiatum. Found in Agricultural Health and Quarantine
calves 4-12 months old. A burden of Service, Department of Primary
500-800 worms is a light to medium Industries, AGPS, Canberra.
one, while 1,000 worms may produce
clinical signs of parasitism - anaemia, Love SCJ, Hutchinson GW (2003).
haemorrhagic enteritis, and a watery, Pathology and diagnosis of internal
mucoid scour. The prepatent stages of parasites in ruminants. In Gross
this worm in the intestinal wall may also Pathology of Ruminants, Proceedings
cause ill effects. 350, Post Graduate Foundation in
Veterinary Science, University of
Trichuris spp. Usually found only in low Sydney, Sydney;Chapter 16:309-338.
numbers and is of little pathogenic
importance. Skerman KD, Hillard JJ (1966). A
Handbook for Studies of Helminth
Liver Parasites of Ruminants. Near East
Fasciola hepatica. An important cause Animal Health Institute, Iran Unit,
of liver condemnations at abattoirs. UNDP, FAO, Rome.
Light infection up to 50; medium
infection 50-100 and heavy infection, Smeal MG (1995). Parasites of Cattle,
over 100. Veterinary Review No. 32, Post
Graduate Foundation in Veterinary
Lungs Science, University of Sydney, Sydney.
Dictyocaulus viviparus. Common in
dairy calves in cooler areas, but seldom PARVOVIRUS INFECTION IN PIGS
seen in beef cattle. Often seen in
conjunction with gastrointestinal Infection before 35 days of gestation
parasite burdens. causes foetal resorption and small litter
size, with returns to service. Infection from
Pigs 35 to 55 days of gestation results in foetal
Small intestine death and mummification, stillborn and
Ascaris suum. Very pathogenic. Any weak piglets. Infection after 55 days usually
number is significant in young pigs. has no effect, but occasionally causes
stillbirths if infection takes place before 70
Macracanthorhynchus spp. Not days gestation. Between 80 days and term,
common. Heavy infection is 200 worms. the virus does not cross the placenta.
Strongyloides spp. Heavy infections
may affect young suckers. Abortion is extremely rare in parvovirus
infection.
Large intestine
Oesophagostomum spp. Widespread Diagnosis
and common cause of scouring in pigs. Herd history, clinical findings,
histopathology and virology (demonstration
Trichuris spp. Heavy infections can of virus antigen or antibody in foetal tissue).
cause typhlitis in young pigs.
Specimens required
Lungs, liver and kidneys i. Whole mummified foetuses and
Metastrongylus spp. Prevalent stillborn foetuses submitted chilled
lungworm. Immature forms of Ascaris, for bacteriology, virology and
Stephanurus and Strongyloides may be histopathology.
found wandering in the lung.
Stephanurus dentatus (kidney worm) ii. Serology on individual samples from
very rare parasite in modern pig sows is of little value in diagnosing
husbandry, but still found in feral pigs in parvovirus infection. Parvovirus
some locations. May be re-introduced if serology should be used as a herd

VETERINARY LABORATORY MANUAL 84


test or to assess the immune status Pepsinogen values should be interpreted
of individuals. on a herd rather than an individual basis.
False negatives may occur.
PASTEURELLOSIS
Level (IU/L) Interpretation
Infection by Pasteurella spp.and <5 No significant abomasal
Mannheimia haemolytica (syn: Pasteurella damage
haemolytica) can cause a septicaemic or 5-10 Minor damage*
pneumonic disease. It can also cause
10-15 Moderate damage
abortion and mastitis in sheep. In pigs,
> 15 Major abomasal damage
toxigenic strains of P. multocida are
associated with enzootic atrophic rhinitis
(see Atrophic rhinitis of swine).
* Levels of 5 or greater are considered
indicative of damage sufficient to cause
Diagnosis
production losses.
Clinical signs and necropsy findings.
Recovery of Pasteurella spp.
Histopathology. Toxigenicity testing of PERINATAL MORTALITIES IN
snout isolates from pigs. CATTLE
Bovine stillbirths and neonatal deaths are
Specimens required
often related to causes of abortion, and can
i. Sections of liver, lung, spleen, heart
be part of a foetal neonatal mortality
blood and any other organ showing
syndrome.
lesions, submitted chilled for
bacteriology.
The following conditions should be
ii. Sections of liver, lung, spleen and
considered, and specimens collected
kidney, submitted in buffered
according to the disease entity where
formalin for histopathology.
appropriate:
NB. In cases of abortion, see specimens
Dystocia
required for Abortion.
Congenital defects
PEPSINOGEN ESTIMATIONS FROM
SERUM OR PLASMA Genetic diseases
In adult cattle, estimations of serum or • Maple syrup urine disease (after first
plasma pepsinogen concentrations are feed)
considered a useful indicator of abomasal • Inherited congenital myoclonus (signs
damage and of value in the diagnosis of present at birth)
type II ostertagiosis (caused by the • Citrullinaemia (born normal; develops
synchronised emergence of histotrophic within 5 days of birth)
larvae). • Mannosidosis (progressive from birth)
• Generalised glycogenosis (progressive
Pepsinogen is produced in the gastric from birth)
mucosa as the inactive precursor of pepsin. • Cardiomyopathy and woolly haircoat
Abomasal damage results in increased syndrome (progressive from birth)
blood pepsinogen concentration. The assay
measures the presence of pepsinogen Environmental diseases
through the ability of the test serum or • Mineral deficiencies
plasma to breakdown a protein substrate to • Selenium deficiency (WMD; stillbirths
peptide fragments. Results are compared occasionally)
with a tryrosine standard, and expressed in
terms of IU/L of tyrosine. Infectious diseases
• Bacterial Leptospirosis,
Specimens required Campylobacter
i. Serum, heparinised or EDTA • Viral Pestivirus, Akabane, Palyam
plasma. • Chlamydial
o
Activity is stable for several days at 4 C for PERINATAL MORTALITIES IN
several days and several months at -20oC.
Haemolysis has little effect on activity.
SHEEP AND GOATS
Investigations should be based on a field
Interpretation of results necropsy with laboratory assistance to
confirm or make a specific diagnosis

VETERINARY LABORATORY MANUAL 85


Foetal infection which persists into and
In any detailed investigation, there must be through post natal life will have resulted
prior arrangement with the laboratory, from one of two circumstances:
regarding the number & nature of • Infection of a susceptible dam, probably
specimens to be examined. through contact with a virus carrier, in
early (usually 30 100 days) pregnancy.
Conditions to be considered should include: Immune tolerance to the virus is
• Abortion, including congenital infections induced; or,
• Dystocia • The dam herself is a persistently
• Starvation, mismothering or exposure viraemic and immuno-tolerant animal.
• Low birth weight from maternal
malnutrition during last 2-3 months of Diagnosis
pregnancy, with resultant inability of Clinical signs and pathology. Detection of
lamb to gain sufficient early feed. antigen or isolation of virus from blood clot
• Delayed colostrum production in ewes or tissue. Detection of antibody in
suffering nutritional stress in the last precolostral serum of deformed calves.
month of pregnancy
• Predation Specimens required
• Postparturient infections, including Live animal
necrobacillosis, pneumonia and i. 10 ml of unclotted blood (heparin
enteritis. blood is preferred), submitted chilled
• Nutritional factors, e.g. selenium for antigen detection. A clot (from 10
deficiency, maternal hypocalcaemia, ml of blood collected into a plain
maternal malnutrition (with foetal anoxia tube) may be used for antigen
from prolonged parturition), overfat detection but offers lower sensitivity.
ewes ii. Serum from clotted blood is
• Clinical conditions in the ewe likely to preferred for antibody detection.
result in a prolonged parturition (uterine iii. In cases of respiratory disease,
inertia) and resultant foetal hypoxia or nasal swabs in PBGS for virus
anoxia e.g. clover disease, foot isolation.
abscess, pregnancy toxaemia, poor Notes:
body condition due to other causes • In cases of embryonic mortality or
• abortion, serum samples from 10-
Diagnosis 15 cows may provide evidence of
Flock history, field investigation, supported herd infection and allow more
by the laboratory. selective investigation to confirm
pestivirus involvement.
Specimens required
Abortion and parturient death • A newborn calf with congenital
i. Specimens as required for Abortion deformities, the product of infection
in sheep. at 120-180 days gestation, may be
ii. seropositive. Some stillborn,
iii. Post-parturient death apparently-normal calves can also
iv. Whole lambs or kids submitted be seropositive. Other affected
chilled for bacteriological and [=viraemic] animals are usually
pathological examination. antibody negative. Demonstration
of an antibody titre in a live animal
usually indicates that the animal is
PESTIVIRUS INFECTION not persistently infected, assuming
Syn: Mucosal disease; mucosal disease it is old enough to have lost
virus infection; bovine viral diarrhoea virus maternal antibody (> 6 months).
However, around 5% of viraemic
Pestivirus infection is associated with animals may have antibody to
embryonic or foetal death, congenital pestivirus.
malformations, perinatal mortality, poor
growth rates, respiratory disease, non Dead animal
specific immunosuppression and disease i. In cases of mucosal disease,
with mortality in calfhood or early adult life. perinatal mortality, abortion or
Life long carriers of the virus including respiratory disease, demonstration
cases of mucosal disease [MD]) are a of virus antigen by ELISA on fresh
consequence of infection in early foetal life tissues provides a rapid diagnosis.
(usually before 90 days gestation). Preferred specimens are (in order):
spleen, lung, mesenteric lymph

VETERINARY LABORATORY MANUAL 86


node, bowel and parotid salivary
gland submitted chilled. One or Diagnosis
several of these should be Mainly on clinical examination.
submitted.
ii. Fresh lung, spleen, and kidney may Specimens required
be submitted chilled for virus i. Sections of liver and kidney,
isolation, especially from cases of submitted in buffered formalin for
respiratory disease where infections histopathology.
are transient and may not be ii. Plant material for identification.
detected by ELISA.
iii. Foetal fluids (pericardial, pleural or District Agronomists may be able to identify
peritoneal, in order of preference) suspect plant material.
should also be collected to test for
IgG and for pestivirus antibody of PIGLET ANAEMIA
the IgG level is elevated.
Iron deficiency causes a microcytic
Where pestivirus infection and/or mucosal hypochromic anaemia in sucking piglets.
disease has been confirmed, further testing
of maternal relatives and enquiry into the Diagnosis
history and management of the herd may History, clinical signs, haematology, gross
indicate how or when infection occurred, pathology.
and what steps can be taken to eliminate
the problem. Options available include Specimens required
removal of carrier families and Live animal
management to ensure the exposure of i. An EDTA blood sample for
breeders to carriers before joining. haematology.

PHALARIS POISONING Dead animal


i. The cadaver submitted chilled for
Can cause staggers or sudden death. bacteriology and histopathology to
investigate a differential diagnosis.
Diagnosis
Flock history, clinical findings, PINKEYE
histopathology.
See Ophthalmia'
Specimens required
i. In cases of staggers, specimens as PLANT POISONING
required for ataxia.
See Poisoning plant
ii. In cases of sudden death, there is
no specific diagnostic test available.
Specimens should be submitted to PNEUMONIA
exclude other possible causes of Diagnosis
death. Clinical and post mortem examination.
Bacteriological, virological, parasitological,
PHOSPHORUS DEFICIENCY histopathological examination.
See Hypophosphataemia'
Specimens required
Live animal
PHOTOSENSITIZATION Especially in feedlot or other intensively
This condition is characterised by the reared cattle.
occurrence of pruritus, oedema, frequently
with subsequent necrosis, jaundice being i. Nasal swabs in transport medium for
an inconstant feature. virus isolation.
ii. Paired (acute and convalescent)
Plants often causing photosensitization sera for virus serology (for IBR,
include Hypericum perforatum, medics Pestivirus, PI3 and RSV).
(Medicago spp.), clovers (Trifolium spp.),
and Tribulus spp. A number of other plants Dead or slaughter animals
are incriminated. Other causes of i. Portions of lung, pleural fluid and
photosensitization are phenothiazine, facial mediastinal lymph nodes, submitted
eczema, algae and conditions giving rise to chilled for microbiology.
liver damage. See also Protoporphyria ii. Portion of affected lung, trachea, etc
(Limousin calves). submitted chilled for virology.

VETERINARY LABORATORY MANUAL 87


iii. Sections of lung, submitted in NB The Specimen Advice Form
buffered formalin for histopathology should request a particular analysis.
iv. In pigs, see porcine ii. Approximately 500 g of suspect
pleuropneumonia and porcine material eg feed, soil for chemical
enzootic pneumonia (PEP'). analysis.
v. In goats where retrovirus infection is NB Analysis of this material is
suspected, serum sample from usually only requested after
affected animal submitted chilled for poisoning is confirmed and
virology. (see Caprine arthritis submitters wish to identify the
encephalitis'). source of the poison.
iii. Sections of liver, kidney and other
POISONING (CHEMICAL) organs showing lesions, submitted
in buffered formalin for
See also Arsenic, lead, organochlorine and histopathology.
organophosphate, sodium fluoroacetate, iv. Specimens as required to allow a
strychnine and urea poisoning differential diagnosis, e.g.
salmonellosis
NB Examination is not undertaken on
specimens likely to involve criminal
POISONING (PLANT)
prosecutions (e.g. in cases of suspected
malicious poisoning). In such cases, the See also Cyanide, nitrate and oxalate
police or other relevant regulatory authority poisoning and pyrrolizidine alkaloidosis
should be consulted before any action is
taken (see Conditions for acceptance of Diagnosis
specimens). History (sudden high morbidity/mortality,
evidence that suspected toxic plant has
Most chemical analyses are specific for at been eaten), clinical signs, necropsy
best a narrow group of chemicals eg findings,
organophosphates. As the analyses tend to
be expensive, clients should carefully If plant poisoning is suspected, the paddock
consider history, clinical findings and should be inspected to determine the
pathological findings so that only the most availability of possible toxic plants. A check
likely chemical is nominated when can then be made at necropsy to determine
completing a request for chemical analysis. if these plants are present in the ingesta.

Where NSW Department of Primary Specimens required


Industries Laboratories are not capable of i. Sections of liver, kidney and any
conducting the requested analysis, samples other organs showing lesions,
will be referred at cost to an outside submitted in buffered formalin for
provider selected on the basis of NATA histopathology.
accreditation and value. ii. If nervous signs are seen,
specimens as required for Nervous
Diagnosis disorders.
History, clinical signs, and in some cases,
laboratory examination. Confirmation of NB Examination of ingesta is usually
chemical poisoning depends upon the difficult and of limited value. It will only be
demonstration of the poison in body tissues performed for specific plants to which the
and organs. animals have had access, and provided
that specimens of those plants accompany
NB the specimens of ingesta.
• Irritant poisons will cause a
gastroenteritis. IDENTIFICATION OF SUSPECT
• When specific drugs (e.g. drenches, POISONOUS PLANTS
dips, feeds, additives) are suspected,
then material should be submitted as Advice in the first instance should be
recommended by the manufacturer. sought from the local District Agronomist.
• A limited range of analyses are When this is not possible, specimens
available on a routine basis. should be submitted as follows

Specimens required Collection of plant specimens


i. At least 100 g of stomach content, All specimens sent should consist of a
submitted chilled for toxicology. small branch or portion of the stem, 20 30
cm long, showing the leaves in position
together with flowers and/or fruits. Owing to

VETERINARY LABORATORY MANUAL 88


the tremendous number of plant species, it Specimens forwarded for identification
is difficult, and sometimes impossible to should be addressed to:
determine specimens from leaves alone.
The Director
For small plants and grasses, the (Attention: Botanical Inquiry Section)
whole of the plant should be sent, with National Herbarium
the exception of the terrestrial or Royal Botanic Gardens
ground orchids when only the parts SYDNEY 2000
above ground should be collected.
POLIOENCEPHALOMALACIA (PEM)
With small ferns and fern allies, the
rhizome (root like structure) is required, Syn: Cerebrocortical necrosis (CCN)
while in the tree ferns the scales or See also: Nervous disorders
hairs at the base of the stalk of the
frond are essential for identification. A sporadically occurring, nervous disease
principally of sheep and goats, less
Eucalypts can be difficult to identify commonly of cattle and occasionally pigs,
without samples of the buds, fruit and with characteristic brain lesions in
juvenile leaves, in addition to the adult advanced cases of cerebrocortical necrosis.
foliage. Information on the bark type, The disease in ruminants is considered due
the extent of rough bark when present, to either:
habit and the habitat are very important.
• production of thiaminases in the rumen
Labelling of plant specimens following changes in the ruminal flora,
Each plant specimens should be numbered particularly when indigestion occurs
and a second set of specimens with after sudden dietary change, or,
corresponding numbers retained. • ingestion of thiaminases in plants (e.g.
Nardoo, rock fern) or in chemicals (eg.
The data and place of collection should be amprolium in poultry feed/ manure).
given, as well details of the habit of growth,
height, flower colour, habitat, type of soil, Diagnosis
and, in the case of trees, a description of Clinical signs (typically progressive higher
the bark. CNS dysfunction with aimless wandering,
blindness, head pressing, recumbency,
Specimens from other states, which may be opisthotonus, convulsions and death).
unfamiliar to local botanists, particularly Gross pathology and histopathology of
require adequate specimen preparation, brain. Response to 6 10 mg/kg thiamine IV
precise locality details, and notes on the in live affected animals (preferably those
main features of the plants and their not yet recumbent).
habitats.
Differentiate from tetanus, focal
A list of identifying names or information symmetrical encephalomalacia/
corresponding to the numbers will be enterotoxaemia, lead poisoning, listeriosis,
reported. Specimens will only be returned salt poisoning, CAE (goats) and copper
on special request. deficiency induced leukomalacia.

Not more than 12 specimens will be named Specimens required


in one collection, except under special i. Whole brain, submitted in buffered
circumstances. formalin for histopathology.
ii. Specimens as required for diagnosis
The cooperation of inquirers is appreciated of other nervous disorders.
in supplying further information or material
when rare specimens, or specimens of POLYARTHRITIS
special interest, have been identified.
See Arthritis
Storage and despatch of plant
specimens POMPE'S DISEASE
Press and dry specimens between See Generalised glycogenosis
newspaper sheets.
PORCINE COLITIS
Fresh specimens should not be forwarded
unless particularly requested. Syn: Porcine intestinal spirochaetosis (IS),
spirochaetal diarrhoea, non-specific colitis

VETERINARY LABORATORY MANUAL 89


replacement of lost epithelium by low
Dd: Trichuriasis, swine dysentery, cuboidal enterocytes. It is possible that high
salmonellosis, yersiniosis, post-weaning energy grower diets may escape complete
colibacillosis, gastric ulcers, porcine digestion in the small intestine, to be
proliferative enteropathy (Lawsonia fermented in the large intestine, producing
intracellularis infection) abnormally high levels of SCFA. Many
cases reported previously may have been
Porcine colitis has been attributed to undiagnosed infections by B pilosicoli or
various non-Brachyspira hyodysenteriae possibly other weakly beta-haemolytic
spirochaetes and Campylobacter coli. spirochaetes, or Lawsonia intracellularis.

Porcine intestinal spirochaetosis Trichuriasis


(spirochaetal diarrhoea) Trichuriasis can produce an identical
Porcine intestinal spirochaetosis appears to macroscopic mucohaemorrhagic colitis and
be a distinct entity associated with infection typhlitis to that of swine dysentery. Disease
by Brachyspira (formerly Serpulina) outbreaks can be caused by massive
pilosicoli. It mainly affects weaners 2-6 infestations of immature T suis (no T suis
weeks post weaning but can affect growers, ova in faeces), which are not as easily
and leads to loose (’wet cement) faeces, detected at necropsy as the adult worms.
which may vary from watery to mucoid. The
disease is expressed as fluctuating and Dietary niacin deficiency
cyclical in a herd, persisting a few days, Dietary niacin deficiency produces severe,
resolving in 7-10 days, or becoming persistent diarrhoea, marked weight loss,
chronic. Lesions are often mild, patchy and and a thickening of the colonic mucosa.
focal superficial colitis and possibly typhlitis. Histological changes comprise hypertrophy
There may be no gross lesions, or mild of the colonic mucosa, associated with a
thickening and oedema of the colonic mixed leucocytic, mainly lymphocytic,
mucosa with localised mucosal infiltration of the lamina propria. Diagnosis
haemorrhages. In resolving lesions, conical is confirmed by assessment of the diet,
scales of fibrinonecrotic material may be which is usually home-mixed, with corn as
dislodged from the mucosal surface by the major component.
rinsing. Histological changes include
mucosal oedema, with shallow, scattered Diagnosis
mucosal erosions, crypt dilation, and Clinical signs (weaner ill-thrift associated
submucosal congestion; spirochaetes may with diarrhoea or mucoid faeces), post-
cover an intact epithelium as a false ‘brush mortem findings, bacteriology,
border’, and B pilosicoli may be ,parasitology, histopathology, ,
demonstrated in affected mucosa by PCR Confirmation of intestinal spirochaetosis
or culture. due to B pilosicoli; diagnosis of non-specific
colitis or niacin deficiency by dietary history
Dietary colitis syndrome (non-specific and elimination of infectious agents;
colitis) confirmation of T suis by parasitological
Dietary colitis syndrome is a recently examination; elimination of infectious
described condition of rapidly growing pigs, agents associated with other syndromes or
especially those 8-14 weeks of age. It is a diseases with similar presenting signs (L
diarrhoeal disease of unknown aetiology, in intracellularis; B hyodysenteriae,
which large-intestinal histological, and Salmonella spp, E coli, Yersinia spp);
sometimes macroscopic, lesions are seen. elimination of other possible causes of ill
Diarrhoea may progress from ‘wet cement’ thrift (e.g. gastric ulceration)
or ‘cow pat’ faeces to watery diarrhoea with
dehydration. Lesions include proprial Specimens required
leucocytosis, with variable epithelial i. Sections of affected intestine,
necrosis and erosion. It is likely that this submitted in buffered formalin for
syndrome has a variety of causes; in one histopathology.
associated with a pelleted diet, the only ii. Impression smears of affected
change detected is frothy large intestinal intestinal mucosa for bacteriology.
contents. Dietary change, poor quality oils iii. Fresh colon or caecum or faeces
and carbohydrates, ad-lib feeding and high (live animals) for detection of
density diets may be associated with this Brachyspira pilosicoli by PCR.
condition. Short chain fatty acids (SCFA) iv. Sections of affected intestine
can cause reversible changes to colonic (especially ileum and colon),
epithelium, comprising epithelial cell loss, submitted chilled for bacteriology
reduction of goblet cell numbers, and and PCR for differential diagnosis.

VETERINARY LABORATORY MANUAL 90


looked at in this light rather than on
NB. PCR tests are available for three an individual animal basis. For this
enteric pathogens: Brachyspira reason, it is recommended that
hyodysenteriae, Brachyspira pilosicoli and about 30-40 sera be submitted on
Lawsonia intracellularis. each occasion.
ii. Nasal swab material collected into
PORCINE ENTEROVIRUS sterile PBS for PCR (if required; may
ENCEPHALOMYELITIS be cost prohibitive)

Syn: Talfan disease Slaughter pigs:


i. Fresh lung samples submitted
A sporadic encephalomyelitis usually chilled or frozen for PCR. For PCR,
affecting pigs from 14 days to weaning, but samples taken from the affected site
also seen in older pigs. Low morbidity and and measuring 1 cm³ is sufficient.
mortality. Typically causes posterior paresis ii. Fixed affected lung for
and ataxia; may progress to all legs, with histopathology
recumbency and paddling. Tremors and iii. Culture is expensive and not
excitement may occur. routinely available. For samples
where an isolate is required (e.g. for
Diagnosis intensive investigation or research),
Histopathology, virus isolation. these must be fresh and reach the
laboratory within a few hrs of
Specimens required slaughter, and preferably come from
i. Brain fixed in neutral buffered well developed lesions.
formalin for histopathology.
ii. Sample of fresh, chilled brain for PORCINE PLEUROPNEUMONIA
virus isolation.
iii. Other samples to eliminate possible Actinobacillus pleuropneumoniae (formerly
differential diagnoses: e.g. bacterial Haemophilus pleuropneumoniae) causes a
meningoencephalitis; oedema fatal pleuropneumonia or non fatal chronic
disease; spinal cord abscess or respiratory disease. At least 7 serotypes
other lesions - including brain and (1, 2, 3, 5, 7, 11, 12) occur in Australia
small intestine for bacteriology; fixed isolates are currently being serotyped at the
spinal cord for histopathology. Animal Research Institute, Yeerongpilly,
Qld.
PORCINE ENZOOTIC PNEUMONIA
Diagnosis
Syn: Mycoplasmal pneumonia Gross pathology, bacteriology and
histopathology. Focal and diffuse gross
Diagnosis lesions of reddish black colour with or
Identification of Mycoplasma without pleurisy are suggestive. Concurrent
hyopneumoniae infection in a herd by infection with Pasteurella multocida is not
ELISA serology, or in individual animals by uncommon.
PCR on affected lung.
ELISA serology can provide a profile of the Specimens required
M. hyopneumoniae infection status of the i. Fresh affected lung and bronchial
herd. Blood samples may be collected from lymph node for culture.
the abattoir or from a subpopulation of ii. Fixed affected lung in buffered
animals in the piggery. Seroconversion formalin for histopathology.
after natural challenge can take 9 weeks,
so targeting older pigs for serological PORCINE INTESTINAL
profiling is more cost-efficient.
HAEMORRHAGE SYNDROME
Carriage in the live animal can be detected
by PCR from nasal swabs. However, nasal Syn; Intestinal volvulus, torsion of
excretion is variable, so a negative swab mesentery, whey bloat, colonic bloat,
PCR on a single occasion is of limited redgut, intestinal venous infarction.
diagnostic value.
Diagnosis
Specimens required History, clinical signs, post-mortem findings.
Live pigs:
i. Serum (or heparin plasma) samples Predisposing factors include transportation,
from affected pigs and their cohorts whey feeding. Clinical signs include
for ELISA. Since this is a herd abdominal distension and pain, and
disease, the results should be dyspnoea, but most cases are simply found

VETERINARY LABORATORY MANUAL 91


dead. Usually but not always associated Syn: Porcine intestinal adenomatosis (PIA),
with palpable torsion of the root of the proliferative haemorrhagic enteropathy (PHE),
mesentery that causes intestinal venous ileitis, regional ileitis (RI), necrotic enteritis
infarction. The proximal duodenum, which
receives its blood supply from the Diagnosis
gastroduodenal vessels, is unaffected.
Clinical signs, necropsy findings,
PORCINE MYOCARDITIS (PMC) histopathological examination of affected
intestine, and bacteriological examination,
Porcine myocarditis (PMC) is a recently
recognised condition of uncertain aetiology Lawsonia intracellularis is the aetiological
apparently affecting pigs in only two agent. It is an intracellular organism and
piggeries in NSW. . It is believed that the requires rat enterocyte cell lines for
syndrome is due to a viral infection maintenance. The uncomplicated proliferation
predominantly, if not exclusively, occurring of mucosa due to L. intracellularis is referred to
in utero. as PIA, while additional changes that may be
superimposed on this lesion may result in
PMC is a notifiable disease. Fees for tests regional ileitis (muscular hypertrophy), necrotic
undertaken to confirm or exclude a enteritis (coagulative necrosis) or PHE
diagnosis of porcine myocarditis are paid by (haemorrhage). It is likely that necrotic enteritis
NSW Department of Primary Industries. represents secondary bacterial infection
superimposed on a primary L.intracellularis
PMC causes an increase in stillbirths and lesion.
pre-weaning mortalities. There may be a
variable increase in the incidence of PIA and RI are associated with weaner ill thrift,
mummified foetuses. The gross but PIA can occur in all ages. PHE is
pathological changes consist of small pale associated with sudden death in finisher and
areas in the myocardium. There is often breeder pigs, while necrotic enteritis can cause
evidence of cardiac enlargement and an mortalities in grower pigs.
increase in the volume of body fluids,
consistent with congestive heart failure.
Specimens required
Histologically there is a non-suppurative
myocarditis
i. Sections of affected intestine
(especially ileum and colon),
Further information can be obtained from
submitted chilled for bacteriology for
the NSW Department of Primary Industries’
differential diagnosis.
website at
ii. Sections of affected intestine,
http://www.agric.nsw.gov.au/reader/an-
submitted in buffered formalin for
health/pmc-qa.htm.
histopathology
iii. Impression smears of affected
Diagnosis
intestinal mucosa for bacteriology
History, clinical signs, postmortem findings,
(MZN stain)
histopathology.
iv. Fresh intestinal contents or faeces
for PCR
Specimens required
iii. Chilled stillborn piglets for necropsy,
NB.
virology and histopathology.
PCR tests are available for three enteric
iv. Fresh heart, lung and body fluid
pathogens: Brachyspira hyodysenteriae,
collected separately and aseptically,
Brachyspira pilosicoli and Lawsonia
submitted chilled for virology.
intracellularis.
v. Sections of heart, liver, kidney, lung,
spleen and whole brain in buffered
formalin for histopathology. PORCINE STRESS SYNDROME
vi. Sections of liver, kidney, spleen, (PSS)
lung and heart blood submitted Syn: Pale soft exudative pork [PSE],
chilled for bacteriology. malignant hyperthermia

PORCINE PROLIFERATIVE An inherited neuromuscular disease of


ENTEROPATHY grower pigs, characterised by a
susceptibility to stress-induced collapse and
Porcine Proliferative Enteropathy death, particularly of Landrace and Pietrain
breeds.

VETERINARY LABORATORY MANUAL 92


It is caused by a recessive mutation of the
HAL gene on chromosome 6 (susceptibility Specimens required
to PSS was initially determined by reaction See
to the anaesthetic halothane). Animals can http://www.dpi.nsw.gov.au/agriculture/vetm
be genotyped as stress-resistant (NN), anual/specimens-by-disease-
stress-carrier (Nn), or stress-positive (PSS- syndrome/diseases_of_livestock/dpi-samp-
positive) (nn). coll-prot.pdf
A large proportion of the stress-carrier and
PSS-positive animals produce carcases Diagnosis of the disease and
with inferior muscle quality. heterozygote detection
i. 20 to 25 hairs, with roots attached,
Diagnosis from the distal end of the tail.
Clinical history and signs, post mortem
findings (60-70% of pigs may show gross PYELONEPHRITIS (BOVINE)
muscle lesions). Laboratory examination of
necropsy specimens cannot assist the field Diagnosis
diagnosis. Clinical signs mainly of abnormal urine and
recovery of Corynebacterium renale from
A DNA test for PSS genotyping patented by urine or kidney tissue.
the University of Toronto is available at
commercial laboratories in North America. It Specimens required
is not routinely available in Australia. i. Sample of urine collected into a
sterile container, submitted chilled
for bacteriology.
PREGNANCY TOXAEMIA
ii. Sections of kidney lesion submitted
Diagnosis chilled for bacteriology.
Generally on clinical grounds and iii. Sections of kidney and other parts of
demonstration of ketones in urine. urogenital tract with lesions,
Differentiate from hypocalcaemia, metabolic submitted in buffered formalin for
or starvation ketosis, secondary ketosis due histopathology.
to anorexia from other causes and
polioencephalomalacia. PYOGENIC INFECTIONS
Specimens required Diagnosis
Samples are only to eliminate other Recovery of pyogenic organisms from
possible causes: abscesses and infected organs.

i. Sections of liver, kidney, adrenal Specimens required


gland and brain submitted in i. At least four (4) smears from
buffered formalin for histopathology. periphery of the lesion for
ii. At least 2 ml of serum, free of cells bacteriology.
and haemolysis, submitted chilled ii. Portions of affected tissue
for calcium, magnesium and beta containing lesions, submitted chilled
hydroxybutyrate (ketone) estimation. for bacteriology.
iii. Section of lesion, including the
NB Urine examination for ketones should margin, submitted in buffered
be carried out in the field. formalin for histopathology.

PSEUDOCOWPOX PYRROLIZIDINE ALKALOIDOSIS


See Teat lesions See also Poisoning, plant'
An intoxication of grazing sheep, cattle,
pigs and horses when hepatotoxic
PROTOPORPHYRIA
pyrrolizidine alkaloids are ingested from
(Limousin and Blonde d'Aquitaine cattle) plants of the genera Senecio, Crotalaria,
Heliotropium, Amsinckia, Echium and
An acute photosensitivity evident from birth others. In sheep, the condition may be
and characterised by alopecia, ulceration complicated by chronic copper poisoning.
and scarring of the pinnae, nares and
possibly along the midline. Diagnosis
History of access to hepatotoxic plants,
Diagnosis clinical and post mortem findings,
Clinical signs and history; confirmed by histopathology.
DNA analysis.

VETERINARY LABORATORY MANUAL 93


Specimens required RHODOCOCCUS EQUI PNEUMONIA
i. Sections of liver and kidney, IN HORSES
submitted in buffered formalin for
histopathology. Syn: Rattles
ii. In cases of suspected copper
poisoning, at least 30 g of liver and Diagnosis
kidney, in separate copper free jars, History, clinical signs, and post mortem
submitted chilled for copper findings. Recovery of Rhodococcus equi
estimations. from lungs and pulmonary lymph nodes.

Q FEVER Specimens required


i. Portions of affected lung and lymph
Q Fever is usually an asymptomatic nodes submitted chilled for
infection of animals caused by Coxiella bacteriology.
burnetii, but it can cause abortion in sheep ii. Swabs of lung abscesses in Amies
and goats and infections in man. Charcoal Transport medium,
submitted chilled for bacteriology.
Diagnosis iii. Portion of affected lung submitted
The complement fixation test is not a fixed in buffered formalin for
reliable test for Q Fever. At present there is histopathology.
no routine test available for diagnosis. The
CFT is used only for export certification. RHODOCOCCUS EQUI
Specimens required
LYMPHADENITIS IN PIGS
i. For abortions, those required for Characterised by oval or spherical lesions
abortion in sheep and goats. in lymph nodes which are encapsulated and
ii. For export certification, serum easily enucleated. It is often difficult to
sample forwarded chilled for CFT. differentiate from tuberculosis in swine.

RED GUT IN SHEEP Diagnosis


Recovery of Rhodococcus equi from
Syn: Intestinal volvulus, torsion of lesions, histopathology.
mesentery, colonic bloat, intestinal venous
infarction Specimens required
i. Affected lymph nodes, submitted
An acute haemorrhagic enterocolitis chilled for bacteriology.
occurring in sheep grazing some lucerne or ii. Affected lymph nodes, in buffered
clover pastures, or other fresh, young green formalin for histopathology.
feed. Some cases show severe abdominal
distension, with rapid death. Post mortem
findings can include distension of the small RINGWORM (DERMATOMYCOSIS)
and large intestine, severe haemorrhage See fungal infections
and congestion of intestinal mucosa and
blood stained intestinal contents. There ROCK FERN POISONING
may be rapid autolysis of the carcase.
Cheilanthes tenuifolia (rock fern) can cause
Diagnosis haemorrhages through out the body with
Post mortem findings, especially hepatic necrosis. Also associated with
displacement and/or torsion of the caecum staggers or ataxia in sheep.
and colon.
Diagnosis
Differential diagnosis from enterotoxaemia. History, clinical signs, post mortem findings,
histopathology.
NB. The ventral necropsy approach is
essential for the displacement and/or Specimens required
torsion to be seen. Live animal
i. 5 ml of blood in EDTA for
Specimens required haematology.
i. As for enterotoxaemia, together with ii. Blood films for haematology (refer to
ii. Sections of affected intestine, Haematology in first section of this
submitted in buffered formalin for manual).
histopathology.
Dead animal

VETERINARY LABORATORY MANUAL 94


i. Sections of liver, spleen and kidney
submitted in buffered formalin for Diagnosis
histopathology. Clinical signs, necropsy findings, recovery
ii. Specimens to differentiate from of Salmonella from heart blood, spleen,
other causes of haemorrhage and liver, bile, mesenteric lymph nodes and
septicaemia, based on post mortem intestinal contents in both septicaemic and
findings. acute forms. Serology.

Ataxic animal In the chronic form, the bacteria may be


Specimens as required for Ataxia. recovered from the intestinal lesions and
less frequently from other viscera.
ROTAVIRUS INFECTION
Specimens required
See also Scouring Live animal
i. Serum for Salmonella Typhimurium
Causes diarrhoea in young animals. and Salmonella Dublin SAT
serology in cattle and sheep.
Diagnosis ii. Faecal sample submitted chilled for
Demonstration of either virus particles in bacteriology.
faeces by electron microscopy or soluble
antigen by immunodiffusion, latex Dead animal:
agglutination or ELISA. i. Portion of liver, kidney, lung, bile,
spleen, mesenteric lymph nodes and
Specimens required small intestinal contents in separate
i. 20 g of faeces or intestinal contents, sterile containers, submitted chilled
submitted chilled for electron for bacteriology.
microscopy or antigen detection. ii. Sections of liver, kidney, lung,
ii. Specimens as required for the spleen and small and large intestine,
differential diagnosis of causes of submitted in buffered formalin for
scouring. histopathology.

RYEGRASS STAGGERS SALT POISONING IN PIGS


Syn; Perennial ryegrass staggers Diagnosis
History, clinical signs, histopathology.
Associated with fungal neurotoxins
produced by Acremonium loliae in the leaf, Specimens required
stem and seed of perennial ryegrass, which i. Brain, submitted in buffered formalin
are toxic to sheep and cattle. Outbreaks for histopathology.
more common in summer and autumn, and ii. Specimens as required for the
with close grazing of limited available feed. differential diagnosis of causes of
nervous disorders.
Diagnosis
History, clinical signs, histopathology.
SARCOSPORIDIOSIS
This syndrome can be diagnosed by clinical Infection with protozoa of the genus
signs in the field. Sarcocystis can cause anaemia, ill thrift and
myopathy in experimental infections in
Specimens required sheep and an acute febrile disease in
i. Brain, submitted in buffered formalin calves.
for histopathology.
Diagnosis
SALMONELLOSIS Histopathology.
Usually occurs as one of three major Specimens required
syndromes: i. Sections of cardiac and skeletal
• Peracute septicaemia seen mostly in muscle, submitted in buffered
young animals. formalin for histopathology.
• Acute enteric form most commonly
seen in adult animals. SCABBY MOUTH
• Chronic enteric form is common in
pigs, and occurs in cattle. There is See Contagious pustular dermatitis'
persistent diarrhoea, severe
emaciation and intermittent fever.

VETERINARY LABORATORY MANUAL 95


SCOURING tract, cardiac and skeletal muscle in
buffered formalin for histopathology.
Investigations into causes of scouring in iv. Specimens as required for specific
animals should take into consideration: diseases, e.g. Johne's disease,
• Age of affected animal9s) Pestivirus, arsenic poisoning, if
• Whether sudden or gradual in onset these are suspected.
• Condition of the animal v. Abomasum and small intestine
• Numbers affected submitted chilled for total worm
• Intensity of grazing or housing count.

Diagnosis SELENIUM DEFICIENCY


History, clinical examination, bacteriology,
parasitology, histopathology, virology. See Muscular degeneration, nutritional'

Specimens required SEMEN EXAMINATION


Depending on the age of the animal and the In investigations of infertility, semen
results of the clinical examination, examination should be considered after a
specimens should be forwarded as required careful clinical examination. Laboratory
for the following diseases: examination can be considered:
• Colibacillosis, especially in neonates • When the field veterinarian is confident
• Rotavirus infection in young animals the semen sample is typical of the
• Cryptosporidiosis in young animals semen to be obtained, and
• Salmonellosis • There is an obvious deficiency with
• Pestivirus (Mucosal disease) in cattle, regard to density, motility, wave motion,
especially < 2 yrs or
• Parasitism • There is evidence of inflammatory
• Johne's disease in cattle, sheep and products in the sample, i.e. polymorphs,
goats clotted semen.
• Yersiniosis
• Poisoning, plant or chemical Veterinarians should conduct the initial
• Proliferative enteropathy in pigs examination in the field.
• Swine dysentery in pigs
• Enteritis in pigs, calves, dogs Specimens required
associated with Campylobacter jejuni i. For morphology, air dried semen
• Copper or selenium deficiency in smears.
ruminants ii. For live/dead sperm examination,
• Coccidiosis smears stained with freshly
• Hypomagnesaemia in milk fed calves prepared nigrosin eosin.
iii. For bacteriology, semen collected
In general, the following specimens should aseptically and submitted in sterile
be submitted. containers.

Live animal NB. Semen collected using an artificial


i. In cases of suspected enteric or vagina is not suitable for bacteriology.
systemic infections at least 30 g of
faeces, submitted chilled for SERUM ENZYMOLOGY
bacteriology or virology. NSW Department of Primary Industries
ii. At least 30 g of faeces from 10 to 20 refers all clinical chemistry testing to a
animals in the flock, submitted specialist laboratory. Test results are
chilled for parasitology. incorporated into the laboratory report
iii. Blood or serum samples as required which includes normal ranges supplied by
for virology. the testing laboratory.

Dead animal: Serum enzymology is usually undertaken


i. Sections of small and large intestine, as part of a package of tests eg liver profile.
especially areas showing lesions, renal profile, ruminant metabolic profile.
submitted chilled for bacteriology. The most frequently requested serum
ii. Sections of liver, kidney, lung and enzymes are:
spleen submitted chilled for
bacteriology in cases of suspected • Aspartate aminotransferase (AST)
enteric or systemic infection. also known as Glutamic oxaloacetic
iii. Sections of liver, kidney, spleen, transaminase (GOT)
selected sites in the gastrointestinal

VETERINARY LABORATORY MANUAL 96


• Creatine kinase (CK), also known as NB If blood cannot be sent to the
creatine phosphokinase (CPK) laboratory overnight, it should be allowed to
• Gamma glutamyltransferase (GGT) clot and the serum poured off; it should be
• Glutamate dehydrogenase (GLDH) frozen if the delay will be greater than 24
hrs. Approximately 2ml of serum per
Specimens required animal should be provided. It must be free
i. 10 ml of blood, clotted in vacuum of haemolysis.
tubes.

Note that the stability of these enzymes is variable, as is their half life in the blood:

Stability in serum sample Half life in blood after insult


AST Intermediate Weeks
CK Unstable Days
GGT Very stable -
GLDH Intermediate Weeks

Applications and Interpretation of Results in Sheep and Cattle


Enzyme Major source Suggested normal serum levels (IU/L)
AST Liver, muscle < 100
CK Muscle, CNS < 300
GGT Liver < 80 sheep
< 50 cattle
GLDH Liver < 20 sheep
< 50 cattle

Typical disease syndromes with elevation of serum enzymes


Enzyme Disease process
AST Sustained necrosis of hepatic and/or muscular tissue (skeletal or cardiac)
CK Sustained necrosis of muscular tissue (eg. WMD) and/or CNS (eg. PE)
GGT Bile duct epithelium damage, cholestasis incl cases with biliary hyperplasia (eg.
chronic fascioliasis; pyrrolizidine alkaloidosis) or acute hepatic necrosis
GLDH Hepatocellular damage (rises rapidly in acute hepatic necrosis)

SODIUM FLUOROACETATE (1080) submitted in buffered formalin for


POISONING histopathology.
Causes neurological signs in canines and
cardiac signs in ruminants. Continued SPORADIC BOVINE ENCEPHALOMYELITIS
exposure to low doses can cause (SBE)
myocardial degeneration in cattle and See also Chlamydial infections'
sheep.
Diagnosis
Diagnosis History, clinical signs, histopathology,
History, elimination of other causes, serology.
detection of poisoned material, e.g. carrot in
the stomach contents; chemical analysis Specimens required
confirms fluoroacetate in stomach contents. i. Paired serum samples from
individually identified and observed
NSW Department of Primary Industries animals collected in the clinical and
does not analyse for fluoroacetate. convalescent phase (2 to 3 weeks
Specimens will be sent to a specialist apart), submitted chilled or frozen for
laboratory on request. chlamydial serology.
ii. Portions of liver and spleen collected
Specimens required aseptically, submitted chilled for
i. Approximately 100ml of stomach bacteriology.
contents.. iii. Brain and sections of liver, spleen,
ii. In cases of suspected poisoning in kidney and other organs with
sheep and cattle, sections of serositis, submitted in buffered
myocardium, liver and lung formalin for histopathology.

VETERINARY LABORATORY MANUAL 97


iv. Heparinised or citrated blood from Clinical signs, confirmation by detection of
acute cases for chlamydial isolation strychnine in ingesta by chemical analysis.
and FAT.
NSW Department of Primary Industries
ST GEORGE DISEASE does not analyse for strychnine. Specimens
Caused by ingestion of desert riceflower will be sent to a specialist laboratory on
(Pimelea simplex). request.

Diagnosis Specimens required


History of access to P simplex, clinical i. Approximately 100ml of stomach
signs. contents.

Specimens required SWAINSONA POISONING


i. Sections of liver, kidney, spleen,
See also Poisoning, plant'
heart and lung, submitted in
buffered formalin for histopathology.
Diagnosis
History, clinical signs, histopathology.
STRANGLES IN HORSES
Strangles is a notifiable diseases in NSW. Specimens required
Submission of appropriate specimens i. Brain and spinal cord, submitted in
accompanied by a Specimen Submission buffered formalin for histopathology.
Form requesting exclusion of strangles is ii. Sections of liver, kidney and lymph
suitable notification. There are no nodes, submitted in buffered
laboratory fees for tests taken to exclude formalin for histopathology.
strangles.
SWINE DYSENTERY
Diagnosis
Clinical findings, isolation of Streptococcus Syn: Brachyspira hyodysenteriae
equi from lesions. infectionDD: Lawsonia intracellularis
(proliferative enteritis/ileitis), Brachyspira
Specimens required pilosicoli (intestinal spirochaetosis;
i. Pus samples or swabs taken from spirochaetal colitis), Non-specific (dietary)
lesions and upper respiratory tract, colitis; less commonly: Salmonella,
submitted chilled in Amies charcoal Whipworm (Trichuriasis), Gastric ulcers.
transport medium for bacteriology.
Brachyspira hyodysenteriae (formerly
STREPTOCOCCUS SUIS INFECTION IN Serpulina hyodysenteriae) is a relatively
PIGS common pathogen with a severe impact on
Streptococcus suis is associated with production. Swine dysentery can occur in
meningitis, septicaemia, polyarthritis, any age (suckers - breeders) but is most
endocarditis and bronchopneumonia in common in 15-70 kg grower pigs (5-20
young pigs. At least 28 serotypes have weeks). It may be less common in weaners
been identified. Disease in Australian pigs if medication for respiratory diseases is
has been associated with serotypes 1,2,3,7 used.
and 9.
Clinical expression is normally low
Diagnosis mortality/ high morbidity, and affected pigs
Isolation of S suis from affected tissues can vary from the classical
(brain, joints, lung, liver, kidney, spleen). mucohaemorrhagic diarrhoea - various
amounts blood and mucus, or faeces
Specimens required varying from soft, grey to light brown to
i. Fresh chilled portions of brain and/or dark. Severe cases can show watery
lung, liver, kidney, spleen faeces, with blood and mucus, and stained
ii. Sections of brain, liver, kidney in hindquarters. Swine dysentery may be
buffered formalin for histopathology difficult to detect in straw-based shelters. A
less severe, chronic form of loose, dark
Serotyping of S suis isolates is not routinely faeces ("black scour") is associated with
available. intermittent medication.

History is typically one of depressed growth


STRYCHNINE POISONING rate, and partial loss of appetite that further
Diagnosis decreases growth. There is gradual spread
in group, and increased incidence of "slab

VETERINARY LABORATORY MANUAL 98


sided" pigs (hollow in flanks). Abattoir
inspection may reveal an index case Can be caused by infection by paravaccinia
through evidence of an accumulation of virus (pseudocowpox), bovine herpes
blood-tinged and jelly-like material in the mammilitis virus, bacterial infections,
large intestinal wall and between the coils, chapping and photosensitisation.
linked to a history of ‘ordinary’ growth rate.
Pathology affects proximal large intestine, Diagnosis
causing a fibrinohaemorrhagic colitis and Clinical signs, demonstration of virus by
typhlitis. This is often severe and extensive, electron microscopy, bacteriology.
with erosions, fresh blood and mucus on
necropsy. Epithelial hyperplasia is a Specimens required
feature. i. Scabs with underlying tissue, and
necrotic debris chilled for virology
Diagnosis (electron microscopy examination)
Gross and histopathology. Smears of and bacteriology.
affected intestinal mucosa. Identification of ii. Biopsy of lesion in buffered formalin
Brachyspira hyodysenteriae infection in for histopathology
individual animals by PCR on affected
intestine or faeces. Culture of affected TETANUS
intestine or faeces.
Diagnosis
Specimens required History, clinical signs, , elimination of other
Live pigs causes including polioencephalomalacia,
i. Faeces for PCR focal symmetrical encephalomalacia,
ii. Faeces for culture if isolation is strychnine poisoning.
required (e.g. for antibiotic
sensitivity). It is preferable to advise Specimens required
the laboratory so special selective There is no routine diagnostic test for
media can be available. tetanus. Specimens should be forwarded to
eliminate other causes.
NB: PCR are available for Lawsonia
intracellularis and Brachyspira pilosicoli. TICK FEVER OF CATTLE
Serology is available to exclude Lawsonia,
Includes Anaplasmosis, babesiosis
if clinical signs have been evident for over
one week.
Tick fevers are notifiable diseases in
NSW. Submission of appropriate
Dead pigs specimens accompanied by a Specimen
i. Fresh, chilled portion of affected gut
Submission Form requesting exclusion
for smears, PCR and possible
of tick fever is suitable notification.
culture.
There are no laboratory fees for tests
ii. Fixed affected intestinal section in
taken to exclude tick fever.
buffered formalin for histopathology
Diagnosis
TAPEWORM INFESTATION IN Clinical signs, demonstration of organisms
SHEEP in blood or organ smears.
Occasionally suspected of causing ill thrift
In the Tick Quarantine Area, tick fever
and scouring in sheep under poor
should be suspected in animals showing
nutritional conditions.
haemoglobinuria, elevated temperature,
jaundice and anaemia. Babesia bovis
Diagnosis
(argentina) infections often cause nervous
Response to treatment, must be
symptoms, followed by coma and death.
differentiated from other causes of
scouring.
Specimens required
Live animal
Specimens required
i. Thick blood films air dried, avoiding
exposure to heat or direct sunlight,
The condition should be diagnosed in the
for parasitology.
field.
ii. Conventional unstained thin blood
smears for haematology. (see
TEAT LESIONS Specimens for Haematology).
Includes Pseudocowpox, herpes mammilitis
virus'

VETERINARY LABORATORY MANUAL 99


iii. Blood collected into EDTA, risk to laboratory workers. In instances
submitted chilled for parasitology where clinical disease requires confirmation
and haematology. in the dead animal, fixed sections in
iv. At least 2 ml of serum, submitted buffered formalin of small intestine, liver,
chilled for serology. kidney, heart, muscle, spleen, and brain
v. Urine sample, submitted chilled for should be submitted for histopathology.
bacteriology and parasitology.
TOXOPLASMOSIS IN SHEEP AND
Dead animal GOATS
i. Blood films, air dried for
parasitology. Abortion and perinatal loss in both sheep
ii. Impression smears from the kidney, and goats is caused by the asexual
heart, liver and brain for proliferative stages of Toxoplasma gondii,
parasitology. with formation of tissue cysts.
iii. Brain squash preparation from the
grey matter of the cerebral cortex, Infection of the dam can be by sporulated T
prepared by crushing a section of gondii oocysts from cats, or by T gondii
the cerebral grey matter the size of a cysts in infected intermediate-host tissues
match head between 2 slides and (eg infected placenta).
spreading lightly.
iv. Sections of kidney, heart, liver, brain Diagnosis
and spleen, submitted chilled for Typical lesions in placenta and foetal brain
bacteriology and parasitology. (necrogranulomas in brain). Serology by
v. Sections of kidney, heart, liver, Latex test.
spleen and brain, submitted in
buffered formalin for Specimens required
histopathology. i. Those required for abortion in sheep
(see 'Abortion in Sheep').
TOXAEMIC JAUNDICE
NB Foetal membranes must be submitted
See Copper poisoning if toxoplasmosis is to be diagnosed.

TOXOPLASMOSIS IN CATS Serological titres of 4 or more in the Latex


test are considered positive.
The cat is the definitive host of the
coccidian parasite Toxoplasma gondii, with
sexual stages in the gut resulting in TOXOPLASMOSIS IN OTHER
oocysts shed in faeces. Clinical disease is ANIMALS
not common in cats. Except initially, oocyst The asexual proliferative stages of
excretion is sporadic, and in small numbers. Toxoplasma gondii with formation of tissue
cysts, cause disease in a wide range of
The asexual proliferative stages of intermediate animal hosts (including
Toxoplasma gondii with formation of tissue humans) that are infected by
cysts, cause disease in a wide range of
• Consuming sporulated T gondii
intermediate animal hosts (including
oocysts shed by cats,
humans) that are infected by
• Consuming tissues of other
• Consuming sporulated T gondii
intermediate hosts containing T
oocysts shed by cats,
gondii cysts, or
• Consuming tissues of other
• Transplacentally.
intermediate hosts containing T
gondii cysts, or
Diagnosis
• Transplacentally. Clinical signs, histopathology.
Oocysts of Toxoplasma gondii are highly Specimens required
infectious to humans and animals. i. Sections of liver, kidney, heart,
spleen, muscle and brain, submitted
There is no routine serological test in buffered formalin for
available. Serological tests are not reliable histopathology.
in cats.
There are no routine serological tests
Faecal examination will only be performed available for animals other than sheep and
in special circumstances after prior goats.
arrangements have been made with the
laboratory. Such examinations pose a high

VETERINARY LABORATORY MANUAL 100


TRANSMISSIBLE SPONGIFORM Diagnosis
ENCEPHALOPATHY (TSE) Herd history of infertility and abortion,
demonstration of T foetus.
Syn: Prion disease, bovine spongiform
encephalopathy (BSE), scrapie in sheep Specimens required
and goats Abortion
i. Specimens as required for abortion
The National TSE Surveillance Program in cattle. See 'Abortion in Cattle'.
(NTSESP)
(http://www.animalhealthaustralia.com.au/a Herd infertility
ahc/programs/adsp/tsefap/tsefap_home.cf i. From cows, vaginal or uterine
m) is a national program jointly funded by exudates of vaginal mucus from
government and industry to demonstrate selected females taken 10 to 20
Australia’s ongoing freedom from BSE and days after service submitted in
scrapie, and to provide early detection of selective transport medium for T.
those diseases should they occur. The foetus.
NTSESP provides payments for producers, ii. From bulls, preputial scrapings
private veterinarians and government submitted unchilled in selective
agencies for the submission of specimens transport medium for T foetus
for laboratory testing from adult cattle and (InPouch TF). See
sheep with progressive neurological http://www.dpi.nsw.gov.au/agricultur
disease. e/vetmanual/specimens-by-
discipline/bacteriology/02-in-pouch-
NTSESP submissions must be guide.pdf
accompanied by:
• A completed NTSESP Clinical History NB Selective transport medium for T.
and Post-mortem Report Form. foetus (InPouch TF) together with
• A completed Specimen Submission instructions for the collection and despatch
Form of specimens will be forwarded from the
laboratory upon request. See
Diagnosis http://www.dpi.nsw.gov.au/agriculture/vetm
Clinical signs of progressive neurological anual/specimens-by-
disease, histopathology. Immunochemistry discipline/bacteriology/02-bull-vd-coll.pdf
is used to specifically identify accumulated
abnormal prion protein (PrP), usually in the TUBERCULOSIS
brain and cord.
Diagnosis
Specimens required Clinical examination, tuberculin test,
i. Brain and cord in buffered formalin necropsy findings, recognition and/or
for histopathology. recovery of pathogenic mycobacteria from
ii. Fresh, chilled cervical spinal cord* tissues, histopathology.
for detection of prion protein.
Tuberculosis is a notifiable disease in
*It is prudent to routinely collect at least a NSW. Submission of appropriate
sample of fresh, unfixed cervical spinal from specimens accompanied by a Specimen
all cases of progressive neurological Submission Form requesting exclusion
disease in any animal species, for testing of tuberculosis is suitable notification.
for PrP if required. There are no laboratory fees for tests
taken to exclude tuberculosis.
TRICHOMONIASIS OF CATTLE When required, specimens for culture of
A contagious venereal disease of cattle mycobacteria will be submitted to the
causing infertility, abortion and pyometron, Australian Reference Laboratory in Western
caused by the protozoon Tritrichomonas Australia.
foetus.
Specimens required
Trichomoniasis is a notifiable disease in Suspect TB lesions
NSW. Submission of appropriate i. Portions of lesions unpreserved in
specimens accompanied by a Specimen sterile sealed jars, submitted chilled
Submission Form requesting exclusion for bacteriology.
of trichomoniasis is suitable notification. If considerable delays (> 3 d) will occur
There are no laboratory fees for tests before culture, freeze fresh tissues (and
taken to exclude trichomoniasis. prevent from thawing during transport).

VETERINARY LABORATORY MANUAL 101


ii. Portions of lesions, submitted in ii. A biopsy from the affected area, in
buffered formalin for histopathology. buffered formalin for histopathology.

Reactor cattle ULCERS, GASTRIC IN PIGS


i. In the case of tuberculin reactor
cattle, the following lymph nodes Haemorrhage from an ulcerated of the pars
should be collected as aseptically as oesophagea with haemorrhage is an
possible in separate sterile important cause of anaemia and sudden
containers, and submitted chilled for death in grower and finisher pigs. Ad lib
culture: feeding, stress and finely ground grains
increase the prevalence of lesions.
Essential
• L & R medial retropharyngeal LN (also Diagnosis
known as suprapharyngeal LN) Gross pathology is usually sufficient to
• L & R bronchial LN confirm the diagnosis.
• Cranial and caudal mediastinal LN
• L & R Supramammary LN (in cows) UREA POISONING
Diagnosis
Desirable History of access to urea, clinical signs of
• L & R lateral retropharyngeal LN (also ataxia, respiratory distress, coma.
known as atlantal LN)
• L & R mandibular LN Specimens required
• L & R parotid LN There are no routine laboratory procedures
• Mesenteric LN available to diagnose urea poisoning.
• L & R internal iliac LN
URINE EXAMINATION
It is important that the property of origin be
clearly identified, either by name or tail tag, Urine examinations should be performed in
or tattoo in the case of pigs. the field using the commercially available
dip stick kits. The results obtained should
NB. Suspect tuberculosis specimens must be interpreted according to the instructions
be packed and despatched separately from with the kit.
other specimens.
The field veterinarian should then consider
As organisms responsible for tuberculosis diagnostic possibilities according to the
in livestock are pathogenic for man, care results of the urine examination and make a
should be taken in collecting samples. field diagnosis or send the appropriate
specimens.
Specimens should be forwarded in such
a manner that they will offer no risk of Proteinuria
infection to persons handling the • following violent exercise or stress
containers in transit or on arrival at the • nephritis
laboratory. • haematuria from any cause
• nephrosis, especially if due to chemical
NB. Tissues are cultured for 12 weeks poisoning
before being classed as negative. Positive • inflammation of the postrenal urinary
cultures may appear at approx 8 weeks tract
(possibly earlier in lesioned tissues with • chronic passive congestion of the
massive infection), and can be confirmed kidney due to a number of conditions
either as Mycobacterium bovis or not including cardiac and hepatic diseases
Mycobacterium bovis by monoclonal dot and fevers.
blot in a matter of days, providing there are
sufficient colonies for testing. Glycosuria
Large quantities of ketone bodies may
ULCERATIVE SPIROCHAETOSIS IN depress the colour development and result
PIGS in a false negative reaction.
• diabetes
Diagnosis • brain lesions including tumours,
Clinical signs, bacteriology, histopathology. haemorrhage, encephalitis,
• enterotoxaemia in sheep, also
Specimens required sometimes in listeriosis and botulism
i. Smears from skin lesions.
Acetonuria (Ketonuria)

VETERINARY LABORATORY MANUAL 102


Occurs in any clinical condition associated •
with deficient carbohydrate metabolism, VIBRIOSIS
including: See Campylobacteriosis'
• ketosis and pregnancy toxaemia in
ruminants VITAMIN DEFICIENCY
• starvation in pregnancy, lactation,
Diagnosis
anorexia
Nutritional history, clinical findings,
• diabetes mellitus response to treatment, biochemistry.
• acidosis
• prolonged vomiting and diarrhoea NSW Department of Primary Industries
• febrile and cachexic diseases laboratories do not analyse for vitamin A or
• after ether or chloroform anaesthesia vitamin E. Specimens will be tested at a
specialist laboratory when these tests are
Haematuria, haemoglobinuria requested.
• neoplasia of the urinary tract
• urolithiasis Specimens required
• infectious diseases, including As vitamins A and E are light sensitive and
babesiosis, leptospirosis degraded in tissues after collection, all
• chronic copper poisoning samples for analysis should be protected
• postparturient haemoglobinuria from light by being wrapped in foil or brown
• sulphonamide treatment paper and kept chilled during transport to
• plant poisoning, including ranunculas, the laboratory.
turnips, privet, broom
• azoturia (myoglobinuria) i. 5 ml of serum, heparin plasma or
• acute nephritis, nephrosis, renal EDTA plasma
infarcts.
NB Ensure you obtain at least a full
Bilirubinuria vacutainer of blood before separating off
• hepatocellular disease including the serum/plasma. If inadequate volumes
infections and toxins are presented, pools may be prepared
where possible.
• bile duct obstructions
• jaundice due to haemolysis, after liver
ii. 20 g liver
damage occurs.
Interpretation of vitamin A and E concentrations in blood and liver (cattle, sheep, pigs)
Species Sample Units Deficient Deficient
Vitamin A Vitamin E
Sheep Serum or µmol/L < 0.9 < 2.3
plasma
Cattle Serum or µmol/L < 0.9 < 4.6
plasma
Sheep Liver µmol/kg < 700 < 4.6
Cattle Liver µmol/kg < 11.6
Pig (piglet) Liver µmol/kg < 11 <7
Pig (adult) Liver µmol/kg < 40 <7

VOMITING AND WASTING DISEASE OF • Enterocolitis and diarrhoea


PIGS • Systemic abscesses and mortality
See Haemagglutinating encephalomyelitis • Placentitis, abortion and perinatal
virus (HEV) infection' mortality
• Pneumonia
WHITE MUSCLE DISEASE • Mastitis
See Muscular degeneration nutritional • Epididymo-orchitis

YERSINIOSIS In addition, specific entities have been


Yersiniosis occurs in a variety of domestic associated with Y pestis in domestic
animals. Disease is usually associated with carnivores (plague) and Y ruckeri in fish
Yersinia . pseudotuberculosis or Yersinia (enteric redmouth).
enterocolitica. However, Y intermedia, Y
kristenseni and Y frederickseni occasionally Syndromes associated with yersiniosis Y
cause disease. Yersiniosis can be manifest pseudotuberculosis and Y enterocolitica in
as one or more of the following conditions: domestic animals:

VETERINARY LABORATORY MANUAL 103


Cattle Y. enterocolitica
Y. pseudotuberculosis: • Acute septicaemia, enteritis,
• Enterocolitis, diarrhoea, ‘flood mud splenomegaly, diarrhoea, mortality
scours’, mortality • Chronic systemic abscessation, chronic
• Placentitis, abortion, perinatal mortality enteritis, diarrhoea, ill thrift
• Pneumonia • Systemic myositis (turkeys)
• Mastitis
Diagnosis
Y. enterocolitica Clinical signs, bacteriology, histopathology.
• Abortion
Recovery of Yersinia spp from the
Sheep alimentary tract and internal organs using
Y. pseudotuberculosis routine and selective media.
• Enterocolitis, diarrhoea, ill thrift,
mortality NB Many Yersinia spp inhabit the intestinal
• Systemic abscessation, ‘pyaemic tract of clinically healthy animals and a
hepatitis’ diagnosis of enteric yersiniosis is
• Placentitis, abortion, perinatal mortality confirmed by finding typical
• Epididymo orchitis histological lesions in sections of
the intestine and internal tissues.
Y. enterocolitica
• Enterocolitis, diarrhoea, ill thrift Recovery of Y pestis is important in
view of the zoonotic potential of this
Goats bacterium.
Y. pseudotuberculosis
Y ruckeri is of regulatory importance in
• Enterocolitis, mortality
fish.
• Placentitis, abortion, perinatal mortality,
endometritis, infertility
Specimens required
• Systemic abscessation i. Samples of faeces, intestinal
• Mastitis contents (particularly ileal contents)
and fresh tissue submitted chilled for
Y. enterocolitica bacteriology.
• Enterocolitis, mesenteric lymphadenitis, ii. Portions of tissue fixed in neutral
diarrhoea, ill thrift, mortality buffered formalin for histopathology.
Samples of fixed internal organs
Deer with lesions should be taken if
Y. pseudotuberculosis and Y. enterocolitica systemic disease is suspected.
• Enterocolitis, diarrhoea, mortality Segments from all levels of the
• Systemic abscessation intestinal tract, particularly ileum and
caecum, as well as mesenteric
Pigs lymph nodes should be taken in
Y. pseudotuberculosis cases of enterocolitis.
• Mild enterocolitis, mild diarrhoea iii. Samples suitable for diagnosis of
• Mild systemic abscessation other causes of diarrhoea, pyaemia,
abortion, pneumonia, mastitis and
Y. enterocolitica epididymo-orchitis should be
• Mild enterocolitis, mild diarrhoea collected as appropriate to the
syndrome under investigation.
Poultry (domestic fowl, turkeys, ducks,
geese)

VETERINARY LABORATORY MANUAL 104


DISEASES OF POULTRY laboratory for advice before
proceeding.
Many poultry diseases can be diagnosed in
the field. Before specimens are submitted, STORAGE AND DESPATCH OF
the field veterinarian should obtain a SPECIMENS
complete history, examine the flock and Live birds should be submitted in pet
affected birds, and perform several transport boxes or the like, with adequate
necropsies. air holes and absorbent lining.
If a diagnosis cannot be made or if The birds must not be overcrowded. The
laboratory confirmation is required, then birds must reach the laboratory overnight
appropriate specimens should be submitted (summer) or within 24 hours (winter),
with a full history and description of clinical unless arrangements have been made for
and necropsy findings. Appropriate tests for them to receive water and food in transit.
specific diseases should be requested.
Sick birds must be able to withstand
STANDARD CONTAINERS AND transport. If they are not likely to survive,
EQUIPMENT they should be killed humanely and
submitted chilled in an insulated container
See Specimens (General). with an icebrick or ice in a sealed container.
They must not be frozen.
COLLECTION OF SPECIMENS
Specimens required They must reach the laboratory within 24
i. For the investigation of most hours, preferably overnight.
diseases, submit at least 5 affected
live or freshly dead birds which are, Fresh tissues and serum should also be
or were, showing typical signs. transported chilled, but not frozen, and
ii. Tissues from necropsied birds, fixed must reach the laboratory within 24 to 48
in formalin and/or fresh, are also hours, preferably overnight.
acceptable.
iii. For serology, 1 to 5 ml of blood Formalin fixed tissues must be in leak proof
should be collected from each of 10 containers and packaged securely to
to 25 birds. The jugular and wing prevent breakage in transit.
veins are the best vessels from
which to collect blood. Blood should If chlamydiosis or tuberculosis is
be collected into suitable containers suspected, particular care must be taken
and allowed to clot. If the clotted to avoid infection of couriers and laboratory
blood will not reach the laboratory staff. Live birds must not be sent. Dead
within 48 hours, the serum should birds should be sealed inside two plastic
be poured off into a suitable clean bags and submitted chilled inside a strong
container before submission to the outer container.
laboratory. The blood must not be The Specimen Submission Form should be
allowed to haemolyse. attached to the outside of the outer
iv. For bacterial or fungal culture, whole container and should prominently warn that
birds or fresh tissues are required. In chlamydiosis or tuberculosis is suspected.
general, swabs (with or without
transport media) are not suitable, SEROLOGICAL TESTS AVAILABLE
due to loss of viability of fragile FOR POULTRY
organisms and/or overgrowth of
contaminants. For the culture of Serological tests are currently available at
Mycoplasma spp. or Haemophilus EMAI for the following diseases:
paragallinarum, live birds must be • Avian encephalomyelitis (AE)
submitted. • Avian Influenza (AI)
v. For virology, whole birds, fresh or • Big Liver and Spleen Disease (BLS)
frozen tissues are acceptable. For • Egg Drop Syndrome 76 (EDS)
virus isolation particularly for NDV (Haemagglutinating avian adenovirus;
and AI- swabs in PBSG (which is adenovirus 127)
available in 5-ml plastic screw-top • Fowl adenovirus (FAV)
containers from the RVL Menangle • Haemorrhagic enteritis of turkeys (HE;
/EMAI Virology Laboratory. If THE)
unfamiliar with the correct • Infectious Bursal Disease (IBD)
specimens, please contact the • Marek's Disease (MD)

VETERINARY LABORATORY MANUAL 105


• Mycoplasma gallisepticum infection Caution Chlamydiosis is a zoonotic
(Mg) disease, causing serious illness and
• Mycoplasma synoviae infection (Ms) sometimes death in humans.
• Newcastle Disease (ND)
• Reticuloendotheliosis (REV) Diagnosis
• Salmonella pullorum (Sp) Gross pathology, examination of tissue
impression smears for acid fast organisms,
Most serological tests are flock tests. antigen detection in tissues by
Positive results indicate that the flock has immunofluorescence (IFAT),
been exposed at some time to the histopathology, serology (pigeons only),
infectious agent concerned, either by isolation.
natural infection (subclinical infection or
clinical disease) or due to vaccination. To Specimens required
confirm recent infection, paired serums are i. Whole dead birds, for pathology,
required to demonstrate flock acid fast and FAT stains.
seroconversion or rising flock titres. The ii. Liver, spleen submitted fresh, chilled
tests are of limited value for individual birds. for acid-fast stain and IFAT on fresh
smears (made at testing laboratory).
Where (i) or (ii) are not feasible, air dried
AVIAN ENCEPHALOMYELITIS (AE)
impression smears from the serosal (not
Diagnosis cut) surface of liver and spleen, and from
Young chickens and turkeys conjunctiva. Submit duplicate smears, for
Clinical signs (neurological disease), acid fast stain and IFAT.
histopathology, serology.
iii. Liver, spleen, conjunctiva and other
Laying chickens and turkeys representative tissues in buffered
Clinical signs (egg production drop), formalin, for histopathology.
serology. iv. Sera, for serology. Ideally, paired
sera should be submitted to
Serology is also used in monitoring efficacy demonstrate rising titres.
of vaccination.
NB Serology is unreliable for most avian
Specimens required species other than pigeons.
i. Whole live birds, or,
ii. Brain, spinal cord, skeletal muscle, CHOLERA
myocardium, proventriculus, gizzard See Pasteurellosis'
in buffered formalin, for
histopathology. COCCIDIOSIS (AVIAN)
iii. Fresh or frozen sera, collected early
Diagnosis
during the problem (within 7 days of
Gross lesions, wet intestinal smears,
onset), and again during
histopathology.
convalescence (3-4 weeks later).
Unless the birds are vaccinated, the
Specimens required
titre range could indicate recent
i. Whole birds, or:
exposure and the paired sera may
ii. Fresh intestinal scrapings (ie
not be required in all circumstances.
mucosa plus contents), for
microscopic examination for oocysts
BIG LIVER AND SPLEEN DISEASE and schizonts.
Diagnosis iii. Portions of affected gut in buffered
Gross lesions, histopathology, serology. formalin, for histopathology.

Specimens required CORYZA


i. Whole birds, or: (Haemophilus paragallinarum infection)
ii. Liver and spleen in buffered
formalin, for histopathology. Diagnosis
iii. Sera, for serology. Clinical signs, gross lesions, bacteriology.

CHLAMYDIOSIS (AVIAN) Specimens required


Syn: Psittacosis; ornithosis i. Whole live birds, for pathology and
bacteriology. (Do not submit dead
birds, tissues or swabs for
bacteriology, as Haemophilus spp
quickly lose viability)

VETERINARY LABORATORY MANUAL 106


INFECTIOUS LARYNGOTRACHEITIS (ILT)
EGG DROP SYNDROME (EDS 76) Diagnosis
A drop in egg production due to EDS in Gross pathology, histopathology, antigen
layer flocks is usually associated with the detection in tissues, virus isolation.
appearance of shell-less eggs. A drop in
egg production without shell-less eggs can Specimens required
be due to a variety of causes. These i. Whole birds, or,
include nutritional, management, ii. Fresh or frozen tracheas (including
environmental and disease factors. Many larynx) in sterile jars, for antigen
causes can be identified in the field and ELISA and virus isolation.
specimens should only be submitted to iii. Tracheas (including larynx) in
confirm the diagnosis of disease caused by buffered formalin, for histopathology.
infectious agents. Because of relatively poor sensitivity
of the antigen ELISA test, 5-10
Diagnosis tracheas are required.
Clinical signs, serology.
LEUKOSIS
Specimens required
i. Fresh or frozen sera, collected early Diagnosis
during the problem (within 7 days of Gross pathology, histopathology.
onset of egg drop), and again during
convalescence (3 4 weeks later). If Specimens required
the flock is not vaccinated, the range i. Whole birds, for gross and
of titres in conjunction with clinical histopathology.
signs would lead to diagnosis, ii. The following tissues fixed in
without the need for paired sera. buffered formalin: liver, spleen,
kidney, heart, ovary/testicle,
FOWL POX proventriculus, gizzard, sciatic
nerves, brain, bursa (if present),
Diagnosis
tumours (if present).
Gross pathology, histopathology, electron
microscopy.
MAREK'S DISEASE
Specimens required Diagnosis
i. Whole birds, for gross and Clinical signs, gross pathology,
histopathology. histopathology.
ii. Lesions fixed in buffered formalin.
iii. Fresh tissues for electron Specimens required
microscopy. i. Whole birds, for gross and
histopathology.
INFECTIOUS BRONCHITIS (IB) ii. The following tissues fixed in
Diagnosis buffered formalin: liver, spleen,
Histopathology, virus isolation. kidney, heart, ovary/testicle,
proventriculus, gizzard, pancreas,
Specimens required sciatic nerves, brain, bursa (if
i. Whole birds, or: present), tumours (if present).
ii. Trachea, lung, kidney, oviduct in
buffered formalin, for histopathology. MYCOPLASMOSIS (AVIAN)
iii. Fresh trachea, lung, kidney for virus
isolation (and serotyping). (Mycoplasma gallisepticum, M synoviae
infections)
INFECTIOUS BURSAL DISEASE (IBD)
Diagnosis
Diagnosis Gross pathology, histopathology and
Gross pathology, histopathology for clinical bacteriology for clinical disease.
or subclinical disease. Serology to monitor Bacteriology for subclinical disease.
exposure or vaccination efficacy. Serology to monitor exposure.
Specimens required Specimens required
i. Whole birds, or, i. Whole live birds, for pathology and
ii. Bursa of Fabricius in buffered bacteriology. (Do not submit dead
formalin, for histopathology. birds, tissues or swabs for
iii. Fresh or frozen sera, for serology. bacteriology, as mycoplasmas
quickly lose viability).

VETERINARY LABORATORY MANUAL 107


ii. Fresh sera, for serology. Preferably ii. To check reactors to the field test,
off the clot and free of red blood submit fresh sera clearly identified to
cells. individual birds.
iii. To confirm reactors, submit live or
NB Do not submit frozen sera, as false freshly humanely killed birds (clearly
positives may occur. identified) for bacteriology.

PASTEURELLOSIS (AVIAN) RETICULOENDOTHELIOSIS (RE)


(Pasteurella multocida, P anatipestifer Diagnosis
infections) Gross pathology, histopathology, serology.

Diagnosis Specimens required


Clinical signs, gross pathology, i. Whole birds, for gross and
bacteriology. histopathology.
ii. The following tissues fixed in
Specimens required buffered formalin: liver, spleen,
i. Whole birds, for gross pathology and kidney, heart, ovary/testicle,
bacteriology, or, proventriculus, gizzard, sciatic
ii. Fresh tissues (liver, heart, lung and nerves, brain, bursa (if present),
(from ducks) brain), for bacteriology. tumours (if present).
iii. Sera, for serology.
PULLORUM DISEASE
TUBERCULOSIS (AVIAN)
(Salmonella pullorum infection)
Diagnosis
Diagnosis Necropsy, histopathology, isolation of
Serology, bacteriology for detection of mycobacteria.
carriers.
Specimens required
Specimens required i. Fresh lesions in sterile sealed jars,
i. For an initial flock test, submit fresh for bacteriology.
(not frozen) sera from every bird, ii. Portions of lesions in buffered
individually identified. formalin for histopathology.

VETERINARY LABORATORY MANUAL 108


DISEASES OF CAGE BIRDS, COLLECTION OF SPECIMENS
AVIARY BIRDS AND RACING Specimens required
PIGEONS i. At least two affected live or freshly
dead birds which are typical of the
flock problem.
Isolated mortalities or sporadic disease
problems in cage and aviary birds and racing If chlamydiosis is suspected, do not
pigeons are NOT examined at Regional send live birds.
Veterinary Laboratories, unless a notifiable or
exotic disease is suspected. However, ongoing ii. Tissues from necropsied birds, fixed
problems involving a number of birds in larger in formalin. Small birds may, after
aviaries or pigeon lofts may be investigated, necropsy, be submitted in toto in 10
depending on the circumstances. Before times their volume of buffered
submitting specimens, please phone the RVL formalin.
to discuss the case. You should have
conducted a thorough investigation of the
problem and be able to provide a detailed
STORAGE AND DESPATCH OF
history, clinical and necropsy findings. SPECIMENS
As for Diseases of Poultry.
STANDARD CONTAINERS AND
EQUIPMENT
See Specimens (General).

VETERINARY LABORATORY MANUAL 109


DISEASES OF BEES i. Preferably 30 adult bees submitted
live at ambient temperature for
microbiology. Bees should be
Bee diseases are investigated at the Regional submitted in aerated containers with
Veterinary Laboratories. Diseases seen a small amount of a candied mixture
include American Foul Brood, European Foul of sugar and honey.
Brood and many viral diseases affecting both ii. If only dead specimens are
adults and larvae. obtainable, submit chilled for
microbiology.
Diagnosis
Clinical signs and demonstration of causal
LARVAL DISEASES
agent. i. Four (4) air dried larval smears.
ii. A brood comb sample containing
Specimens required diseased larvae, submitted chilled
for microbiology.
ADULT BEE DISEASES

VETERINARY LABORATORY MANUAL 110


DISEASES OF FISH STANDARD CONTAINERS AND
EQUIPMENT
The Special Veterinary Officer (Fish Water sample containers
Health), located at the Regional Veterinary The types of containers recommended for
Laboratory, Wollongbar can be contacted collection of water samples include:
for advice on the investigation of fish health • Polyethylene or glass for ammonia,
problems.(Phone: 066 240261) nitrate/nitrite, pH, cyanide and algae;
• Glass stoppered glass for oxygen, filled
Expertise in the diagnosis of fish diseases
to exclude air;
is currently available at other Regional
• Nitric acid washed polyethylene for
Veterinary Laboratories, and the assistance
metals;
of officers at these locations can also be
sought in the first instance. • Hexane washed glass for pesticides;
• Sterile 200 ml glass bottles for
bacteriology, chilled at 4oC, and
submitted within 24 hours of collection.
Fixatives

ml
BOUIN'S FIXATIVE
Saturated aqueous picric acid 75
Formalin (Commercial solution; 38% formaldehyde w/v) 25
Acetic acid 5

DAVIDSON'S SOLUTION
Glycerine 10
Formalin (38% formaldehyde) 20
95% Alcohol 30
3.5% NaCl solution (or Filtered seawater) 30
Glacial acetic acid 10

Fixation time: 24 hours (minimum)

HISTORY AND DIAGNOSIS estuarine or ocean environments are


important because of their
History taking differences in salinity. A general
i. A comprehensive history assists the description of the land use in the
investigation. Estimates of morbidity local drainage basin or watershed
and mortality, based on stocking may be appropriate.
density on farms or the abundance
of the species in natural waterways iii. Describe the condition of the water
should be provided. Include in terms of colour, turbidity, odour
measurements of length and weight, and flow. Is the water level high,
with an estimate of the age or stage low or normal? Are algae, organic
of growth. Larger fish are more likely debris, froth or oil floating at the
to suffer oxygen deprivation, surface? Does the ecosystem
whereas smaller fish tend to be appear "healthy"?
more susceptible to toxic insults. If
other inhabitants of the aquatic iv. Record the temperature of the water
environment are affected, poor at several sites and depths. Submit
water quality or exposure to toxins the results of any onsite water
should be suspected. quality tests performed. These
measurements, especially O2 and
ii. A description of the locality of the pH, could be taken at various sites
fish kill should include the type of (inlet, middle and outlet) and depths
habitat, whether natural or man (surface, mid level and bottom) in
made, river, stream, lake, pond, the body of water.
dam, tank, raceway, cage or
aquarium. On fish farms the design v. Details of composition of diet, history
and construction of facilities and the of dietary changes, source or
source of water should be related to manufacturer of feed, rate of
the number and distribution of ponds feeding, storage conditions and
affected. Freshwater, brackish, duration of storage could allow

VETERINARY LABORATORY MANUAL 111


nutritional deficiencies or toxicities to be displayed. Skin colour may
be identified. change.

vi. List any treatments, including • Superficial lesions affecting


chemicals and antibiotics, which the skin, fins, gills, eyes,
have been administered to the fish. mouth or anus may be
On many occasions malachite green visualised in the live fish. The
or formalin will have been used; results of any clinical
although these treatments may have examination should be
eliminated the causative agent, detailed.
rendering subsequent examinations
negative, they may have caused iii. Any lesions noted at autopsy or
further direct or indirect injury to the during handling should be recorded.
fish and these potential If several fish are available for
complications should be considered. examination, an onsite autopsy
What dosage regime (time of should be performed on at least
treatment, duration of treatment and one. Collect fresh and preserved
concentration or dose rate of specimens from any autopsies and
chemical) has been applied? submit these along with any live
affected or freshly dead fish which
Diagnosis are available.
History, clinical signs, field assessment of
water quality*, field evaluation of COLLECTION, STORAGE AND
environmental and ectoparasitic causes of DESPATCH OF SPECIMENS
disease**, histopathology, parasitology,
bacteriology, virology, toxicology. Specimens required
i. Live moribund or affected fish
*Water quality assessment is best (preferred specimens).
performed in the field using commercial test Fish to be submitted are placed in a
kits for ammonia, nitrite/nitrate, acidity, strong plastic bag inside another plastic
hardness and copper. bag, together with a half volume of
water from the source environment. If
**Microscopic examination of wet likely to be in transit for any length of
preparations of gills, skin and fins should time, the plastic bag should be inflated
allow common parasites to be identified. with oxygen before twisting, double
folding and sealing with strong rubber
Diagnostic hints bands. The plastic bag is then placed
i. Mortality patterns can be fitted to the inside an esky and sent to the
time of year, day or season. For laboratory.
example, deaths due to oxygen
deprivation are most likely to be Fish should never be fed before
clustered in the early hours of the transport; this increases metabolic rate
morning when water oxygen levels which increases oxygen consumption
are depressed. Some diseases, and may lead to fouling of the water.
such as infestations with the gill Sick fish should also be protected from
protozoan Chilodonella in many changes in temperature.
species or epizootic haematopoietic
necrosis (EHN) virus in redfin perch, Despatch of several survivors and/or
are recurrent seasonal problems. several healthy control fish is often
beneficial in reaching a diagnosis, as
ii. The signs exhibited by diseased fish this allows the pathologist to compare
are readily observed in glass sided affected and unaffected/recovered
aquaria, but may be difficult to animals in greater detail.
detect in open water, unless fish are
surfacing. Diseased fish may ii. Dead fish (rarely of much diagnostic
change the depth which they value, unless they are examined on
normally occupy in the water column site)
or their spatial relation to inlet or Dead fish are virtually useless after
outlet points to a pond. Feeding transport, even if chilled, because
patterns may be altered. decomposition in fish occurs rapidly
Behavioural abnormalities such as and many ectoparasites will detach
gasping at the surface of the water, from the host soon after death. If only
flashing or turning upside down may

VETERINARY LABORATORY MANUAL 112


dead fish are available for submission containers. A pool of fresh chilled brain,
to the laboratory, submit: liver, kidney and spleen is often suitable
• Fixed tissues for histopathology for isolation of viruses. Swabs are less
(must be fixed on site) suitable.
• Frozen tissues for
toxicology/virology. If only dead fish v. Smears of blood and body fluids (for
are submitted, bacteriology is haematology and bacteriology).
precluded (culture must be Many examinations for ectoparasites
undertaken within 1 hour of death are best carried out in the field using
to be of value). Frozen fish are wet preparations. Rapid blood stains
unsuitable for histopathology. can also be used for smears from fish.

iii. Tissues in fixative (submitted in In special cases and after consultation


addition to live fish). specimens can be forwarded to the
This ensures that serviceable Australian Fish Health Reference
specimens will be available should Laboratory, Geelong, Victoria.
there be transportation delays that
result in death or deterioration of other SOME DISEASES OF FISH,
specimens. Small fish may be CRUSTACEANS AND SHELLFISH
immersed in fixative whole, with a
ALREADY ENCOUNTERED IN NSW
section of abdomen removed to allow
penetration of fixative to internal Fish Kills
organs. A range of tissues could be Attributed to:
taken from larger fish at autopsy, • Anoxia
concentrating especially on those • Environmental toxins e.g.
tissues which exhibit gross pathology. superphosphate
• Infectious diseases e.g. Epizootic
In addition to internal organs such as Haematopoietic Necrosis (EHN) in
heart, liver, kidney, spleen and redfin perch.
intestine, consider submitting brain,
skin, muscle and gonad. As autolysis is Skin Lesions in Fish
rapid in the gills, sections of the gill Often multifactorial; may involve nutritional,
arches should be placed in fixative at parasitic, bacterial and viral pathogens.
the start of the examination. Histopathology of lesions has been of
value. Causative agents include:
Ten percent neutral buffered formalin or • Bacterial infection with Aeromonas
5% formol saline are suitable fixatives hydrophila in hatcheries, seen in
for fish tissues. Bouin's fixative (see stressed adult fish.
‘Fixatives’ above) can be used if the • Yellow spot
specimens are small, for rapid • Red spot in estuarine fish on the North
penetration and optimal preservation. Coast
The tendency of some ectoparasites to • Goldfish ulcer disease due to
detach from gills and skin, even in the Aeromonas salmonicida or Vibrio
presence of formalin, may be overcome infections. The former is an important
by the use of Davidson's solution (see pathogen of Salmonids
‘Fixatives’ above).
Shellfish Diseases
iv. Fresh tissues (for bacteriological
• Discolouration in Abalone
examinations, virology and
toxicology).
Crustacean Diseases
Tissues from large fish (which have
• Protozoal myositis with ataxia in
been freshly killed) may be removed
Yabbies
and transported chilled in sterile

VETERINARY LABORATORY MANUAL 113


FEEDS AND PASTURE ANALYSIS
NSW Department of Primary Industries TESTS AVAILABLE
provides an independent Feed Quality Service Individual and package tests are available
to sheep, beef and dairy producers. A limited from the following list:
range of analyses is available for pig and • Dry Matter
poultry feeds. The Feed Quality Service • Nitrogen (crude protein)
accepts most types of feeds for analysis e.g. • Digestibility
grain, hay, pasture, silage, mixed rations, crop • Organic matter
residues, browse and leaves. • Metabolizable energy
• Acid detergent fibre
Charges are made for evaluating feed • Neutral detergent fibre
samples, in accordance with Departmental
• Lignin
policy. The charges are based on which
• Soluble carbohydrate
analyses are requested. For information on
analyses and charges, contact the Customer • Fat
Service Unit on 02 6763 1187. • Nutrient minerals (B, Ca, Cu, Fe, K, Mg,
Mn, Na, P, S, Zn)
SAMPLE COLLECTION • pH (silage only)
A standard procedure for the collection of • Ammonia-Nitrogen (silage only)
samples should result in a representative
sample and minimal sample deterioration. Other specialised tests are available.

Detailed instructions for the collection of SUBMISSION OF SAMPLES


samples are available from Livestock All samples must be accompanied by a
Advisory Officers or directly from the Feed Feed Quality Service submission form The
Quality Customer Service Unit on 02 6763 form is available from the Feed Quality
1187. Customer Service Unit and can be sent to
you by fax, e-mail or by post. NO SAMPLE
The main points to be considered are: WILL BE TESTED WITHOUT A
• Samples collected must be CORRECTLY COMPLETED SUBMISSION
representative of the feed and not FORM.
taken from one bale or bag.
• Sampling Hay: 10-15 core or grab Each sample should be sealed in a unique,
samples from at least 10 small or 5 strong paper or plastic bag (not freezer
large bales selected at random need to bags etc.) with as much air squeezed out
be sampled. Mix the sample thoroughly as possible. DO NOT SEND SAMPLES IN
and send approximately 500g in a GLASS.
paper bag to the laboratory.
• Sampling Silage: 10-20 grab samples The outside of the bag should be labelled
from fresh silage or from at least 10 with:
small or 5 large selected at random • Your name
need to be sampled. Mix the sample • Sample type (hay, pasture, silage etc)
thoroughly and freeze approximately • A unique sample number or name
500g in a plastic bag and send to the
laboratory. Wet samples such as silage, fresh crop
• Sampling pasture: Sample at random (green leaf) and fresh pasture must be
15-20 locations across the paddock. sealed in a plastic bag (air tight) and frozen
The sample must reflect the pasture immediately. These samples should then
consumed by the animal. Mix the be packed in an esky and sent to the Feed
sample thoroughly and prepare Quality Service.
approximately 500g by either:
microwave the sample on high for no NB. DO NOT DESPATCH FROZEN
longer then 30 seconds, or dry at 600C SAMPLES ON THURSDAYS OR FRIDAYS
for 24hrs, or freeze the sample and
send to the laboratory. All samples should be sent to:
• Grain and feed mixes: 10-15 grab
samples from different locations or Feed Quality Service
bags within the supply. Mix thoroughly NSW Department of Primary Industries
and send approximately 500g to the RMB 944
laboratory. Calala Lane
Tamworth NSW 2340

VETERINARY LABORATORY MANUAL 114


NOTIFICATION TO LABORATORY CHARGES FOR FEED QUALITY
The Feed Quality Service should be notified SERVICES
by telephone (02) 6763 1187 when samples DO NOT SEND ANY MONEY with the
are sent for analysis. The work load can sample. An invoice will be issued with the
then be anticipated and a potential turn laboratory report.
around time can be given to the officer
involved. Any special conditions needed for Discounts for large number of samples may
sample transport can be discussed at the be available. Contact the Customer
time of notification; e.g. silage and other Service Unit (02 6763 1187) for more
fresh material (as noted above) needs to be information.
cooled or frozen.

VETERINARY LABORATORY MANUAL 115


WATER TESTING
The quality of water from bores, wells, creeks, Further information on water testing is
rivers and farm dams can vary widely. available from the NSW Department of Primary
Common problems include hardness, iron and Industries website at
salinity. Water quality can impact on plant http://www.agric.nsw.gov.au/reader/das-
growth and animal production, and can affect laboratory/das-farm-water.htm
irrigation equipment.
Water for human consumption
NSW Department of Primary Industries' NSW Department of Primary Industries does
Environmental Laboratories provide a water not undertake testing of water for human
testing service for farmers and graziers consumption. Contact either the Health
wishing to determine the suitability of their Inspector at the Local Council, or the NSW
water for agricultural and domestic Department of Health's Water Laboratory,
applications. Sampling kits are available from Lidcombe regarding any testing for human
all Departmental offices. Standard laboratory consumption:
reports provide detailed information on pH,
salinity, chloride, alkalinity, turbidity, hardness, NSW Health
saturation index, sodium absorption ratio and Division of Analytical Laboratories
electrical conductivity. Weerona Road
LIDCOMBE NSW 2141
Testing for nutrients, pesticide residues and
blue-green algae is also available.

VETERINARY LABORATORY MANUAL 116

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