Schedule1 NMD Dbase Programme

Schedule 1 National Microbiological Database Programme
December 2012 Prelims
Prelims
December 2012
Table of Contents
Prelims ......................................................................................................................1 1. Administration and Participation .................................................................6 1.1 Species to which the National Microbiological Database Programme Applies ...............................................................................6 1.2 1.3 1.4 1.5 1.6 2. Participation...........................................................................................7 NMD Administration ..............................................................................7 Health of Personnel ...............................................................................8 Training of Samplers .............................................................................8 NMD analyses .......................................................................................8
NMD Sampling Requirements - Red Meat and Poultry ..............................9 2.1 2.2 2.3 2.4 2.5 2.6 2.7 2.8 2.9 2.10 2.11 2.12 Products sampled .................................................................................9 Number of samples per week ...............................................................9 Summary Table ...................................................................................13 Post Chill Carcass Sampling Programme ...........................................14 Salmonella Sampling Programme ......................................................14 Poultry Campylobacter Sampling Programme ....................................17 Sampling Location within the Process ................................................17 Carcass Sample Sites .........................................................................23 Analysis Required ...............................................................................37 Summary .............................................................................................38 Weekly Sampling Plan for Red Meat Random Rotational Sampling40 Poultry Broiler Sampling Requirements ..............................................46
3.
Sampling ......................................................................................................50 3.1 3.2 3.3 3.4 Manufacture of Diluents for Sampling and Analysis ...........................50 Templates Required for Swab Sampling ............................................52 Collection of Samples .........................................................................54 Sampling Method ................................................................................55
3.5 4.
Transportation of Samples to the Laboratory for All Species .............61
Laboratory Analysis ....................................................................................68 4.1 4.2 4.3 4.4 4.5 Dilutions Required ...............................................................................68 Duplicate Plating/Analytical Precision .................................................69 Preparation of Dilutions .......................................................................70 Preparation of Swab Samples for APC and E. coli Analysis ..............70 Preparation of Red Meat Separate Carcass Site Samples Swabbed for Salmonella Analysis .......................................................71 4.6 4.7 4.8 4.9 4.10 Preparation of Bulk Meat (Whole Tissue) Samples for APC, E. coli...72 Aerobic Plate Count, APC30 ................................................................72 Escherichia coli Petrifilm
TM
..................................................................76
Salmonella...........................................................................................77 Poultry Carcass Campylobacter Direct Plate Enumeration Method ...81
5.
Results and Formulas .................................................................................86 5.1 5.2 5.3 5.4 5.5 Result Calculation (Manual) Red Meat ...............................................86 Result Calculation (Manual) Poultry ....................................................88 Result Calculation (Automated) ..........................................................89 Limits of Detection ...............................................................................89 Reporting of Results ............................................................................91
6.
Targets Premises Analysis and Interpretation, Independent
Verification .............................................................................................................93 6.1 6.2 6.3 6.4 6.5 6.6 6.7 6.8 6.9 7. Premises Level Targets ......................................................................93 Bovine Species NMT: Escherichia coli Moving Window ....................93 Bobby Calf NMT: Escherichia coli Moving Window ...........................94 Ovine NMT 95 Percentile m Limits: APC ..........................................96 NMT: Documentation and Record Keeping .......................................97 Ranked List .........................................................................................98 Salmonella Performance Standards (SPS) .......................................100 Poultry Campylobacter Performance Target (CPT) ..........................102 Verification Requirements .................................................................108
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References .................................................................................................109
Interpretation
In this Schedule, unless the context otherwise requires,alert response means an immediate review of the process by an operator to identify and document factors that may have compromised hygienic processing, and if appropriate, take corrective and preventative action. batch of samples means all the samples collected in one sampling period. biosecurity (Campylobacter) means measures undertaken to prevent the introduction and spread of Campylobacter into grower flocks. birds means poultry broilers. bobby calf means bovine calf that is at least 4 days old, generally unweaned and less than 45 kilograms dressed carcass weight. bovine means beef, with 3 classes; bull, cow and prime. broiler chicken means a male or female chicken kept primarily for meat production, but does not include poussins. bulk meat means bulk packed meat, not including IW cuts. caprine means red meat species, goat. carcass post slaughter and dressing means a red meat carcass after completion of slaughter and dressing, after post-mortem inspection, prior to chiller entry at cold/warm boning premises or just prior to quartering at hot-boning premises. For the purposes of NMD post mortem inspection is considered to include removal of navel in bobby calves. cervine means deer, with 2 classes defined for NMD; fallow and other (wapiti, red and hybrid). domestic premises means premises that supply the domestic market only. dsBPW means double strength buffered peptone water. equine means red meat species which includes horse, mules and donkeys. extremes of temperature means, in relation to samples during collection and transport, less than 0 C and greater than 25 C. EU and US listed premises means premises that are listed for export to the EU and/or US markets or premises supplying red meat to markets specifying EU and/or US. farm means the location at which the poultry sheds are situated HACCP means Hazard Analysis Critical Control Points. initial item means is the first carcass, cut or bulk carton within the run to be sampled.
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initiation of analysis means a. In relation to APC and E.coli analyses, from the time the sample is suspended and ready for dilution; and b. In relation to Salmonella analysis, the commencement of incubation of the BP pre-enrichment. ILCP means Inter-Laboratory Comparison Programme. insulated container means a sample collection or transport container that is insulated. IW cut means an individually wrapped cut. MEGAREG means slang for US Pathogen Reduction HACCP Final Rule. MIMM 2005 means Meat Industry Microbiological Methods, Edition Four, March 2005. moving window means addition of 5 new samples to the bottom of the window to displace 5 samples from the top of the window. NMT means National Microbiological Target. non EU and non US premises means premises that do not export to EU and/or US, or any markets specifying EU and/or US. off-site laboratory means an independent consulting laboratory to which the operator of a premises has appointed responsibility to enter results directly to NMD. on-site laboratory means a laboratory facility located within the physical boundaries of a premises. ovine means species with 2 classes; sheep and lamb. porcine means red meat species pig. processing plant means poultry broiler slaughter house. processing season means bovine, caprine, cervine, ovine, ostrich and emu, poultry, 1 October to 30 September or from the commencement of a short season such as bobby calves, which may be before 1 October. primal cuts means a primal cut from red meat carcass. post chill carcass means carcass after chilling. post slaughter and dressing carcass means a carcass after post mortem inspection of carcasses. In the case of bobby calves after inspection, after the umbilicus/navel is removed and prior to any further trimming. PSW means a primary sampling window re Salmonella sampling programme. RMP means a Risk Management Programme. ssBPW means single strength buffered peptone water.
shed means building purpose built for sustenance of a flock of poultry broilers. SSW means secondary sampling window in relation to the Salmonella sampling programme. sub-contracted Laboratory means where a premises has a LAS approved laboratory but sub-contracts an off-site laboratory for specific analyses, the on-site laboratory may be subcontracted or seconded to train personnel to collect samples for the off-site laboratory. TNTC means too numerous to count. T means the time a poultry carcass takes to move from the point of selection on the chain to the first drop point. verifier means official assurance verifier which means a person accredited under section 103 of the Animal Products Act 1999 to undertake official assurance verification and includes animal product officers employed by MPI. VLT means very low throughput.
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December 2012 Schedule 1 National Microbiological Database Programme Administration
and Participation
1. Administration and Participation

1.1 Species to which the National Microbiological Database Programme Applies The species to which the NMD applies are bovine, ovine, bobby calf, caprine, cervine, ostrich, emu, poultry and porcine. The NMD operates on the same principles for each species with slight variations. A summary of the requirements for each species is listed below in Table 1. Note: If your premises is EU and/or US listed, whether or not you are exporting, you must comply with the NMD programme for all species processed that are covered by the NMD programme. If you are currently EU listed or US listed it is irrelevant whether the product is going to an EU or US market that week, NMD sampling/analysis must be conducted each processing week. Domestic and export premises; even if not processing, a report to NMD is required to confirm that there was no processing that week. Table 1: Domestic and export application of NMD programme. Species Bovine (excluding bobby calves) Ovine Bobby Calves Caprine Cervine (farmed) Domestic, non EU and non US Listed Premises Every processing week Every processing week Every processing week Every processing week Every processing week EU Listed Premises Every processing week Every processing week Every processing week Every processing week Every processing week US Listed Premises Every processing week Every processing week Every processing week Every processing week Currently no requirements for US. Domestic, non EU and non US or EU apply Every processing week Comply with the US Pathogen Reduction HACCP Final Rule. Must notify Director General of intentions Exporting to the US is prohibited
Ostrich and emu Porcine
Every processing week Every processing week
Every processing week Exporting to the EU is prohibited unless prior notification is obtained from the Director General Exporting to the EU is prohibited unless prior notification is obtained from the Director General Not required Not required
Poultry (broiler chickens)
Every processing week
Ducks, geese and guineas Horses, mules, and other equines
Not required Not required
Exporting to the US is prohibited Comply with the US Pathogen Reduction HACCP Final Rule. Must notify Director General of intentions.
and Participation
Table 1: Domestic and export application of NMD programme (continued). Species Domestic, non EU and non US Listed Premises Not required EU Listed Premises US Listed Premises
Wild game
Not required
Wild games, excluding feral goats and pigs, can be exported to US. Not required No negotiated entry requirements. Not required
Game estate
Not required
Not required
1.2
Participation All premises referred to in Table 1 must participate in the NMD programme as part of the New Zealand standard. Once a premises is participating it is mandatory to comply with all requirements of the NMD programme. Premises may voluntarily participate in additional NMD programmes but if submitting results to MPI must follow all requirements of the NMD. The NMD sampling plan applies to all premises irrespective of throughput. Allowances are made for premises classified as VLT; (see section 2.3 Table 2).
1.3
NMD Administration On commencing participation in the NMD an operator must submit completed demographic questionnaires to the NMD Administrator (these can be found on the MPI website). If any of the following details change, the operator must inform the NMD Administrator within 7 working days of the change occurring. a. name of premises; b. name of plant manager and contact details: phone, cell phone and email; c. address of premises, physical and postal;
d. name of NMD controller and contact details: phone, cell phone and email; e. name of LAS approved laboratory conducting NMD analysis and coordinating the NMD sampling programme; f. name of the data submitter and whether this is a person employed by the premises or the LAS approved laboratory conducting analysis; g. species required to be sampled for NMD; h. if species processed is VLT or standard throughput; i. any process details as outlined on demographics questionnaire;
and Participation
j.
location and reference number for each farm and reference number for each shed for which the operator processes poultry.
1.4
Health of Personnel Personnel handling product for the NMD programme must comply with the specifications and market access requirements for health of personnel, refer to Part 3 of the Animal Products (Specifications for Products Intended for Human Consumption) Notice 2004.
1.5
Training of Samplers All NMD sampling must be undertaken by persons trained and deemed competent for the species being sampled. Persons who have attended an LAS approved sampling course related to the species they will be sampling are collectively called Certified Trainers. Certified trainers are qualified to select personnel who are technically competent and are sufficiently familiar with the process associated with the species to be sampled, to be trained as Associate Trainers. Both Certified and Associated Trainers may train others to sample. Persons trained by Associate Trainers cannot train others. Samplers trained by Certified or Associate Trainers who are not qualified to train others are referred to as restricted samplers.
1.6
NMD analyses All sample analysis must be performed in LAS approved laboratories. Modification or substitution of methods is not permitted unless approved by LAS, and documented in LAS. Each NMD analysis must be performed according to the method listed in section 4 of the this Schedule. These analyses are based on those described in Meat Industry Microbiology Methods Edition Four (MIMM 2005) or later edition, general procedures. NMD analyses have been modified from those published in MIMM where appropriate, to reflect the specific requirements of the NMD programme. Methods as written in NMD procedures for application to the NMD programme take precedence over the MIMM 2005 or later edition. As required under LAS all quality assurance functions and quality control procedures for methods and media as per MIMM 2005 or later edition, Chapter 2 must be followed.
December 2012 Schedule 1 National Microbiological Database Programme NMD
Sampling Requirements Red Meat and Poultry
2. NMD Sampling Requirements - Red Meat and Poultry

2.1 Products sampled Non EU and non US listed premises, including domestic premises must sample the following: Bobby Calves and Caprine: all product types (fresh carcasses, primal cuts and bulk meat) processed under their RMP Bovine (excluding bobby calves), Ovine, Ostrich and Emu: fresh carcasses Poultry broiler: fresh carcasses Cervine: fresh carcasses and primal cuts Porcine: fresh carcasses.
EU and US listed premises must sample the following: Bovine (excluding bobby calves), Bobby Calves and Caprine: all products types (carcasses, primal cuts and bulk meat) processed under their RMP 2.2 Ovine, Ostrich and Emu: fresh carcasses only Cervine: fresh carcasses and primal cuts
Number of samples per week The number of samples is dependant on: whether the product is for the domestic market, non EU and non US markets or to be exported to EU and/or US; and the species processed; and whether the species to be sampled is classified as standard throughput or VLT; and the type of analyses required.
(Refer also to section 2.3 Summary, Table 2: Number of samples required per week). 2.2.1 VLT provisions for domestic only and non EU and/or non US Iisted premises
These VLT provisions do not apply to ovine and porcine species; and
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apply to premises that process product destined only for the local market and/or markets other than EU and US or any markets specifying EU and US; and
apply to bovine (excluding bobby calf) premises which process product destined only for local markets and/or markets other than EU and US or any other markets specifying EU and US, regardless of throughput.
Domestic only and non EU and/or non US listed red meat premises processing bovine as above must sample: one fresh carcass each week of processing, starting from the first week of operation and continuing for at least 16 processing weeks: where a VLT processing season is less than 16 weeks, the number of samples collected per product per week must be increased to ensure that 16 samples are collected within a season (for example an 8 week season would require collection of 2 samples per week for 8 weeks). VLT premises for bobby calf and/or caprine are premises which process product destined only for local markets and/or markets other than EU and US or any other markets specifying EU and US and have a throughput of less than 400 bobby calf/caprine animals in each processing week throughout the season. Domestic only and non EU and/or non US listed premises processing bobby calf and/or caprine qualifying as VLT premises must sample as follows: from the first week or part week of operation, including short seasonal operations or where processing is intermittent, sampling of one of each product type (fresh carcasses, primal cuts and bulk) must be conducted each processing week or part week for at least 16 weeks or, if the number of processing weeks in the season is less than 16 weeks, conducted each processing week or part week; or if at any time during the current season the weekly bobby calf and/or caprine throughput exceeds 400 animals in a processing week, sampling of five samples of each product type (fresh carcasses, primal cuts and bulk) each processing week is required. VLT premises for cervine species are defined as those that slaughter or cut/bone product from 100 cervine animals or less per week (this is based on throughput over a whole season). If at any time during the current season the weekly cervine throughput exceeds 100 cervine animals, cervine sampling must revert to standard throughput requirements. Premises qualifying as VLT for cervine species must sample as follows: one of each of the following product types; fresh carcasses and primal cuts each week of processing, starting at the first full week of operation within a season, and continuing for at least 16 processing weeks:
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where a VLT processing season is less that 16 weeks, the number of samples collected per product per week must be increased to ensure that 16 sampled are collected within a season (for example an 8 week season would require collecting on 2 samples per week for 8 weeks).
2.2.2 caprine
VLT for EU and US listed premises processing bobby calf, bovine, cervine and
VLT premises are defined as those that slaughter or cut/bone product from 100 bovine/cervine animals or less, and/or 400 bobby calf/caprine animals or less per week (this is based on throughput over a whole season). If at any time during the season the weekly throughput exceeds the VLT limit, samples must be collected at the standard rate for the remainder of the season. Premises qualifying as VLT for a given bobby calf, bovine, caprine or cervine species or product type of one of those species must sample as follows: at least one item of each product each week of processing, starting at the first full week of operation within a season, and continuing for at least 16 processing weeks: where a VLT processing season is less than 16 weeks, the number of samples collected per product per week shall be increased to ensure that 16 samples are collected within a season (for example an 8 week season would require collection of 2 samples per week for 8 weeks). 2.2.3 Discontinuing VLT sampling
These VLT provisions do not apply to porcine and ovine. The following are the only circumstances when VLT sampling may be discontinued: Bovine/bobby calf: These species have National Microbiological Targets (NMTs), see sections 6.2.1 NMT (bovine species): Escherichia coli moving window (m) and 6.2.4 Bobby calf NMT 80 percentile m limits: Escherichia coli re limits specified for E. coli. So, in addition to the above 16 week requirement, NMD sampling at the required rate may only be discontinued after E. coli moving window of 15 consecutive samples that does not elicit an m alert, is completed: VLT premises processing cervine and caprine may discontinue weekly testing of a product type after 16 samples of that product type have been collected. VLT premises processing bobby calf, bovine, caprine and cervine may continue to sample as per NMD requirements throughout the season if they wish. The NMD Administrator will continue to accept results.
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2.2.4
Recommencing VLT sampling
The VLT sampling requirements above/in section 2 are to be recommenced each season (which usually commences on 1 October) or whenever the process is changed such that new process conditions could affect microbiological outcomes. 2.2.5 VLT for porcine premises
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VLT porcine premises are defined as those that slaughter product from 10,000 pigs or fewer per annum. Porcine premises qualifying as VLT must sample at least one carcass each week or part week of processing for analyses required. 2.2.6 VLT for poultry premises
VLT poultry premises are defined as those that slaughter product from one million (1,000,000) birds or fewer per annum. Poultry premises qualifying as VLT for carcasses must sample at least three carcasses each week or part week of processing for analyses required.
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2.3
Summary Table Table 2: Number of samples required per week
Species
Product type
Technique
Domestic and non EU and non US listed 1 per week
EU and US listed standard throughput 5 per week
EU and US listed VLT
Bovine
Carcass
Multiple Swab Technique Multiple Swab Technique Whole Tissue Composite Sampling Multiple Swab Technique Multiple Swab Technique Multiple Swab Technique Multiple Swab Technique Multiple Swab Technique Whole Tissue Composite Sampling Multiple Swab Technique Multiple Swab Technique Multiple Swab Technique Multiple Swab Technique
1 per week
Primal Cuts
Not required
5 per week
1 per week
Bulk Meat Product
Not required
5 per week
1 per week
Post Chill Carcass Ovine Carcass
Not required
5 per week for 6 weeks 5 per week
1 per week for 6 weeks Not applicable
5 per week
Porcine
Carcass
5 per week; VLT 1 per week 5 per week VLT: 1 per week for 16 weeks 5 per week VLT: 1 per week for 16 weeks 5 per week VLT: 1 per week for 16 weeks Not required 5 per week 1 per week
Bobby Calf and Caprine
Carcass
Primal Cuts
5 per week
1 per week
Bulk Meat product
5 per week
1 per week
Post Chill Carcass Cervine Carcass
3 per week VLT: 1 per week 2 per week VLT: 1 per week 1 per week
Primal Cuts
2 per week
1 per week
Ostrich/Emu
Carcass
2 per week
Not applicable
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Table 2 continued: Number of samples required per week Species Product type Technique Domestic and non EU and non US listed EU and US listed standard throughput Not applicable EU and US listed VLT
Poultry
Carcass
Whole Carcass Rinse Method
3 per day VLT: 3 per week
Not applicable
2.4
Post Chill Carcass Sampling Programme Post chill carcass sampling is only required for bovine, bobby calf, and caprine species for EU and US listed premises. Post chill carcass sampling must be commenced on starting the NMD programme and then be undertaken annually at the start of the season. Post chill carcass sampling is required when the chiller operating conditions changes significantly (calibration). The season is classified as beginning at 1 October or the commencement of a short season such as bobby calves, which may be before 1 October. Post chill carcass sampling provides a microbiological profile of chillers at normal operating capacity for the particular species being tested. Post chill carcass sampling is required until 30 carcasses (at 5 carcasses per week for 6 consecutive processing weeks) have been monitored. If sampling takes more than 6 weeks it must be justified (for example, chiller capacity being lower than the normal for the species, multiple species and concurrent Salmonella sampling obligations exceeding laboratory capability for the sample collection and/or analysis). Post chill carcass sampling must be rotated on a weekly basis around the chillers, to a maximum of 6 chillers (5 samples per chiller) during each annual period. Premises that qualify as VLT for bovine, bobby calf and/or caprine species are required to have one post chill carcass sample per week for 6 weeks for each season.
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2.5
Salmonella Sampling Programme There is a performance standard for Salmonella sampling for all products (excluding chilled carcasses) from the following species: bovine
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bobby calf caprine cervine ostrich/emu porcine (see section 2.5.2) poultry
(Note: Salmonella testing is not required on ovine species). Salmonella sampling and performance standards are split into three groups: Table 3: Salmonella groups Group 1 Salmonella Bovine Bobby calf Caprine Cervine Group 2 Salmonella Ostrich/Emu Porcine Group 3 Salmonella Poultry
2.5.1
Group 1 (bovine, bobby calf, caprine, cervine) seasonal Salmonella
Seasonal Salmonella sampling is required for each product type processed. The processing season for bovine, caprine, and cervine is 1st October 30th September. For bobby calves the processing season is taken from the commencement of bobby calf processing during the calendar year 1st January 31st December. An initial primary sampling window (PSW) of 16 consecutive weeks, within a processing season must be conducted. Where a premises processing season is shorter than 16 weeks the PSW carries over to the subsequent processing season. A premises that has not detected Salmonella in a given product type of a given species in the preceding PSW must test product for the initial six consecutive processing weeks of the next season. (See Figure 1). This reduced sampling frequency is defined as a secondary sampling window (SSW). Where a premises doesnt complete either a PSW or SSW in their processing season, this then carries over to the new processing season. The carried over PSW or SSW does not displace the new processing season requirement. The new seasonal Salmonella sampling must commence on the completion of the carried over one. Examples: A bobby calf processing season, usually starting between March and August, might commence with PSW15; indicating a carry over from the previous season. In the first week of the new season the PSW of the previous season would be complete at PSW16 (providing no Salmonella detection occurs). The processor is then required to start a secondary sampling window (SSW1) in the following week.
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Where a SSW reads SSW2, SSW3, SSW4, SSW5 or SSW6 at the beginning of a new processing season this indicates an incomplete SSW from the previous season. The full SSW of the previous season must be completed and the new season SSW1 must be started in the next processing week (providing no Salmonella detection occurs). A premises that detects Salmonella in a given product type of a given species during a PSW or SSW must immediately begin another 16 week PSW for that species/product type. A sampling window terminates on detection of Salmonella and week one of a new PSW commences for that particular species/product type in the following processing week. Figure 1: Salmonella sampling windows succession
Salmonella: detected (+) not detected (-) An operator who demonstrates to a verifier that Salmonella was not detected in either a PSW or a SSW may, with approval of MPI VA, cease testing of that product for the remainder of the processing season. Unless the PSW or SSW was carried over from last season, where a SSW needs to be commenced for the current processing season following completion of the carried over seasonal window. Closures (seasonal or maintenance): An incomplete PSW or SSW carries over to the next processing week following completion of maintenance. Changes to process: A premises that is substantially modified in such a manner that it could affect expected microbiological outcomes of the process must immediately begin a 16 week PSW when processing recommences. Substantial modification includes: new premises under the same licence/registration number at the same or different location; and major changes in processing methods (for example, change from traditional to inverted dressing, installation of robotics, etc.) where changes in microbiological performance are possible or probable.
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2.5.2
Group 2 (Ostrich/Emu and Porcine) - Salmonella
An operator must conduct ostrich/emu and porcine Salmonella sampling each processing week. Porcine Salmonella sampling is a 27 month programme which commenced week beginning 5 October 2009 and will cease on the week beginning 2 January 2012. 2.5.3 Group 3 (Poultry) Salmonella
Poultry Salmonella sampling must be conducted by operators of; 2.6 Standard throughput premises; each processing day Very Low Throughput premises; on one processing day of each processing week.
Poultry Campylobacter Sampling Programme The Campylobacter sampling programme for poultry carcasses must be conducted by operators of; Standard throughput premises; each processing day Very Low Throughput premises; on one processing day of each processing week.
2.7 2.7.1
Sampling Location within the Process Red meat product selection
Refer to section 2.3 for products (carcasses post slaughter and dressing, post chill carcasses, primal cuts and bulk meat) required to be sampled for each particular red meat species NMD programme. 2.7.1.1 Carcasses post slaughter and dressing NMD sampling for post slaughter and dressing aims is conducted to gain a representative picture of microbiological contamination during the slaughter and dressing process. Procedures that result in a significant transfer of microbes to the carcass are completed before the post mortem inspection station. Post slaughter and dressing is defined, for the purposes of NMD, as after post mortem inspection and after final dressing procedures specified below. The NMD sampling for post-slaughter and dressing is as follows: NMD sampling must commence as soon as is practical, but no later than 30 minutes after post mortem inspection of carcasses. Carcasses detained by AsureQuality must not be sampled.
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Carcasses must be sampled BEFORE any procedure that may impact on the NMD carcass sampling sites. Processes following the post mortem inspection station may redistribute contamination over a carcass, add some contamination by handling from workers, or remove contamination due to particular process interventions. Such procedures include carcass stacking (contact), gross hot trimming, carcass washing, rail stimulation and worker contact during marshalling.
In the case of bobby calves, samples are to be collected immediately after inspection, after the navel is removed and prior to any further trimming.
Positioning of carcass for sampling: (i) Ovine, porcine, caprine and bobby calf carcasses can be moved to a dead rail for sampling, or another suitable position that does not mask the microbiological consequences of slaughter and dressing. (ii) Bovine and cervine carcass sampling must be conducted on a moving rail (as the carcass cannot be manually moved from the rail). This requires a greater level of training to ensure sample collection is consistently from the correct sites, the correct carcass side and with the correct sampling technique.
The specified carcass sampling sites require wet/dry swab sampling. The sampling sites will vary in location according to the species. The sites have been selected to represent the points where microbes are most likely to be transferred to the carcass during dressing procedures.
Record the time of sampling, run, shift, species, class and process type; inverted, traditional, gas de-pelted.
Rotation of chains on a weekly basis in premises with multiple slaughter chains is required.
2.7.1.2 Post chill carcasses Post chill carcasses of cold and warm boning premises require wet/dry swab sampling. Refer section 2.8.8. Post chill carcass sampling is not required for hot boning premises. Cold and warm boning premises must sample the post chill carcass: no later than 24 hours after the onset of chilling; or when the chilling cycle is completed in less than 24 hours, at the completion of the chilling cycle; and in either case in the chilling area itself prior to wrapping, transportation, freezing, boning or loadout.
Records must be taken of the: location of the chilled carcass at time of sampling; identifying the cooling floor, chiller, side chiller, quarter chiller or other,
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boning process (cold or warm), and number of hours chilled (timed from the onset of chilling).
2.7.1.3 Primal cuts and bulk meat product Primal cuts and bulk meat product the microflora comprises organisms from two sources; those which remain attached to the carcass after the initial transfer during slaughter and dressing; and those acquired by contact of the cut surfaces with other surfaces or materials (for example condensate, aerosols) in the processing environment (cross contamination). Primal cuts and bulk meat manufactured at a premise, but from carcasses supplied by another premises, shall be regarded as product of the boning premises. Boning premises must record the premises of a carcass origin and enter that information in the boning premises NMD report under the product description column of the NMD database. Only fresh product may be sampled for NMD (secondary processes and product that has either been chilled in the carton, frozen or thawed from frozen is not required to be sampled for NMD). Primal cuts (cold, warm or hot boned, bone-in or bone-out). Wet/dry swab samples must be taken from an outside (fell) surface of the cut, not an exposed, freshly cut internal surface. The sampling site selected on the primal cut should correspond to the equivalent carcass sampling site for the species concerned. Primal cut samples are to be taken immediately prior to vacuum packaging, wrapping or bulk meat packing into cartons. Bovine, bobby calf, caprine: hindquarter cuts or hindleg cuts (refer Table 4). Venison: hindquarter cut (rump/topside) and forequarter cut (shank/shoulder). Removal of the silver skin (de-sinewing) of cuts is more commonly carried out in venison further processing. Samplers must record whether the venison cuts sampled are de-sinewed or intact.
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Table 4: Primal cut descriptors Species Bobby Calf Bovine Hindleg Outside hindleg ATS Bottom Round Eye of Round Flat Inside Outside Less Eye of Round Outside Round Outside Smalls SAT Silverside Topside Hindleg Sinew Intact Hindquarter, rump Sinew Intact Hindquarter, topside Sinew Intact Hindquarter, outside Sinew Intact Forequarter, shank Sinew Intact Forequarter, shoulder Sinew Intact Forequarter, shoulder shank on Sinew Intact Forequarter, foreleg Sinew Intact Forequarter, knuckle Desinewed Hindquarter, rump Desinewed Hindquarter, topside Desinewed Hindquarter, outside Desinewed Forequarter, shank Desinewed Forequarter, shoulder Desinewed Forequarter, shoulder shank on Desinewed Forequarter, foreleg Desinewed Forequarter, knuckle No primal cut NMD required No primal cut NMD required No primal cut NMD required No primal cut NMD required Primal Cut Descriptor
Caprine Cervine
Ovine Porcine Poultry Ratite
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Figure 2: Beef carcass, location of cuts The bovine primal cut descriptors are highlighted in yellow:
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Bulk meat and trim. The micro organisms on the surface of bulk meat or trimmings will be distributed throughout the carton. Bulk meat will eventually be ground into a homogenous comminuted product and the original micro organisms will become evenly distributed throughout. Whole tissue samples from the original bulk meat carton are expected to give a good representation of the microflora. Boneless bulk meat product of 85-95 VL, not including individually wrapped (IW) product, is to be selected for sampling. Where a premises does not produce boneless bulk packed meat, cartons of trim in the 60-85VL range will be acceptable. Whole tissue samples are to be taken from bulk meat product immediately prior to closing and strapping the full carton before equilibration or freezing. Table 5: Bulk meat pack descriptors Species Bobby Calf Bulk Product Descriptor Trim Trunk Veal Inside Fores and Hinds Trim Shanks Trim Trunks No bulk NMD required No bulk NMD required No bulk NMD required No bulk NMD required No bulk NMD required
Bovine
Caprine Cervine Ovine Poultry Porcine Ratite
The time of sampling must be recorded (those conducting only further processing will enter this time on the NMD data entry sheet), species, class, description of boning process (cold, warm, hot) primal cut type, VL/CL grade percentage or estimate of and product description of bulk. Premises with multiple boning rooms must rotate sampling between boning rooms on a weekly basis.
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2.7.2
Poultry carcasses selection Whole poultry carcasses are to be selected in order for the whole carcass rinse method to be applied For premises where carcasses are mixed in bins at manual grading, immediately prior to grading (for example off the drain-table, after draining) For premises where carcasses are re-hung on the chain after immersion chilling, or are air chilled, the carcass must be selected immediately prior to the first drop point, or similar, at which the carcasses enter secondary processing or are bagged for retail.
Should the position at which the carcass is released from the chain not be readily accessible, the carcasses for sampling may be selected: At the last readily accessible position on the chain or from the drain-rack. In order to drain excess water, the carcasses must be hung prior to being sampled or bagged for shipment to the laboratory for the same time (T ) as the carcass would take to move from the point of selection to the first drop point. Note: This time (T ) will vary at each premises, but is representative of normal processing at that premises. Carcasses for sampling must be handled aseptically for removal from the drain table or main chain and for transfer into the sampling bag. Figure 3: Selection of poultry carcass Immersion chill Selection of carcass Pre-wash Drain table or re-hanging Air chill T
0 0 0
Drip-line
Bagging or Grading further processing
2.8
Carcass Sample Sites Individual samples are to be collected (ICMSF, 1986; MIRINZ, 1971) from the species specific sites described below and illustrated in Figure 4 (bovine), Figure 5 (ovine and caprine), Figure 6 (bobby calf), Figure 7 (cervine traditional), Figure 8 (cervine inverted),
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Figure 9 (porcine) and Figure 10 (ostrich/emu). Figure 11 illustrates sampling from the opposite sides of the carcass for APC/E. coli and Salmonella samples.
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2.8.1
Bovine
Three sites need to be swabbed on bovine. The three sites are as follows: Rump: (situated on the hindleg, refer Figure 4) The 100cm site is centred on the fascia overlying the semitendinosis muscle. The muscle is cut, packed and sold as the eye round. The centre of the sampling site is halfway along the muscle on its superficial lateral margin. Lateral to semimembranosis is the gluteobiceps muscle. This muscle is cut, packed, and sold as the outside flat. The 100cm template may overlap both muscles. Flank: The 100cm site is centred on the fascia overlying the cutaneous trunci muscle. The centre of the site is 10cm lateral to the umbilicus on a line drawn along the ninth rib from the spine and continued on to cut edge of the abdominal wall. Brisket: The 100cm site is centred on the fascia overlying the cranial ventral part of the cutaneous trunci muscle. The centre of the sampling area is 10cm off the central midline. The margins of the sampling site may overlap onto the fascia overlying the caudal margin of the pectoral profundis muscle. The lower edge of the sampling site is an imaginary line drawn transversely along the thoracic wall at the level of the point of the elbow.
2 2 2 1: 2
The naming of the bovine hindleg sample site as rump is required under our equivalence agreement with the US.
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Figure 4: Sampling sites for bovine carcass, 100cm
Rump site situated on the hindleg
Flank site
Brisket site
Photographs courtesy of David Walkinshaw, Taylor Preston Limited
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2.8.2 2.8.2.1
Ovine and Caprine Ovine: traditional and inverted
A single ovine carcass site needs to be swabbed. The site is as follows: Forequarter opening Y-cut: The Y-cut site is defined as the anterior aspect of the humeroradial joint close to the brachial vein, which is clearly visible at the site but a little erratic in its course. 2.8.2.2 Caprine Three carcass sites need to be swabbed for caprine. The three sites are as follows: Outside hindleg: The outside hindleg site is defined as an area one third up on a vertical line originating at the midpoint of a line between the ischial crest and stifle, and extending to a line horizontal to the cut end of the hock. Flap: The flap site is defined as an area ~50mm from the flap edge, midway between the flap joint and the xiphoid cartilage. Forequarter opening Y-cut: The Y-site is defined as the anterior aspect of the humeroradial joint close to the brachial vein, which is clearly visible at the site but a little erratic in its course.
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Figure 5: Ovine sampling site and caprine sampling sites, 5cm sampling area. Ovine carcasses (both traditional and inverted dressing) require sampling of the opening Y cut site only. Caprine carcasses require sampling of all three sites shown below.
Outside leg
Flap
Opening Y-cut
Photographs courtesy of Sandy Moorhead (AgResearch) and Monique Biss (MAF VA)
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2.8.3
Bobby calf
Three sites need to be swabbed on bobby calves. The three sites are: Fore rump: An area adjacent to the rectal canal, no greater than 50mm horizontally from the edge of the rectal canal, vertically with midpoint on a horizontal line dissecting the forward (lower) edge of the rectal canal. Flank: The site is defined as an area ~50mm from the flank edge, midway between the flank joint and the xiphoid cartilage. Foreleg: An area on the outside surface of the lateral head of the triceps brachii. The side (left or right of the animal) of the carcass APC/E. coli swabs are collected from must be recorded for bobby calf samples.
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Figure 6: Sampling sites for bobby calf carcasses, 25 cm sampling area
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2.8.4 2.8.4.1
Cervine Cervine traditional
Three sites need to be swabbed on traditional cervine. The three sites are as follows: Posterior hindleg: The hindleg sample site is centred longitudinally and laterally on an imaginary line drawn between the posterior aspect of the aitch bone and the archilles tendon. Sternum: The sternum site is immediately adjacent to the intersect of the abdominal cavity opening and brisket, but outside the area of standard brisket trim (by <10mm at its closest edge). Note: Tissue exposed by the standard trim must not be sampled. Inside foreleg: The inside foreleg site is defined as the anterior aspect of the humero-radial joint close to the brachial vein, which is clearly visible at the site but a little erratic in its course. 2.8.4.2 Cervine inverted
Three sites need to be swabbed on inverted cervine. The three sites are as follows: Inside hindleg: The sample site is on the medial side and proximal end of the hindleg immediately adjacent to the pelvic symphysis (midline). Brisket: The brisket sample site is centred longitudinally on the brisket, but immediately outside the area of standard brisket trim (by <10mm at its closest edge). Note: Tissue exposed by the standard trim must not be sampled. Inside foreleg: The inside foreleg site is defined as the anterior aspect of the humero-radial joint close to the brachial vein, which is clearly visible at the site but a little erratic in its course.
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Figure 7: Sampling sites for cervine carcasses (traditional), 25cm sampling area
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Figure 8: Sampling sites for cervine carcasses (inverted), 25cm sampling area
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2.8.5
Porcine
Three sites need to be swabbed on porcine carcasses. The three sites are as follows: Outside hindleg: Midway between the stifle and hip joint. Lower flap: 50mm from the abdominal incision midway between the two lower nipples. Outside shoulder: Above the forward top end of the shoulder blade. Figure 9: Sampling sites for porcine carcasses, 25cm sampling area Photos courtesy Land Meats
2
Porcine NMD site 1: Outside Hindleg
Porcine NMD site 2: lower flap between the lower two nipples
Porcine NMD site 3: outside shoulder site
Whole carcass view of NMD sites
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2.8.6
Ostrich and emu
LEG HUNG A single ostrich and emu carcass site needs to be swabbed. The site is as follows: Inside leg: Located longitudinally on the opening cut line and laterally where the edge of the remaining flap contacts the leg. It is recommended that this site be alternated between left right sides. Figure 10: Sampling sites for leg hung ostrich and emu carcasses, 25cm sampling area
2
Inside hindleg
Photo courtesy of Venison Packers Fielding Limited
WING HUNG No site identified at present as no NZ premises are using wing hung slaughter and dressing practice.
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2.8.7
Salmonella sampling
Where Salmonella sampling is being undertaken the same sites on the opposite side of the carcass from APC/E. coli samples are to be selected and sampled. In the case of adult cattle the carcass is physically split. Substitute a side from another carcass for sampling if the opposite side to that sampled for APC/E. coli is not available due to condemnation. Figure 11: Illustrates sampling from opposite sides of the carcass as required for APC/E. coli and Salmonella samples.
2.8.8
Post chill carcass sampling Where post chill sampling is being undertaken, select the same sites on the opposite side of the carcass from APC/E. coli sampling. For example, the carcasses post slaughter and dressing can be tagged and re-sampled on the opposite side of the carcass after chilling. Where Salmonella sampling of carcasses post slaughter and dressing coincides with post chill sampling choose another carcass for post chill sampling that has
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undergone the same chilling conditions adjacent to the carcass originally selected for carcass post slaughter and dressing sampling. Where matching is impractical (for example, pre-chill carcasses spread over several chillers) post-chill carcasses may be selected randomly from a single chiller, rotating weekly through all available chillers. NEVER sample a site that has been previously swabbed because the microbial load will have been removed/modified during the first swab sampling procedure. Note: post-chill carcass sampling can be conducted on the same day as fresh carcasses are sampled. 2.8.9 Poultry carcass sampling
The whole carcass is sampled for poultry. 2.9 Analysis Required The following analyses are required: Table 6: NMD analyses Product Bovine, bobby calf, caprine, cervine, ostrich and emu, and porcine carcasses post slaughter and dressing Ovine carcasses post slaughter and dressing Post chill carcasses Primal cuts Bulk meat Poultry whole carcass Analyses required Aerobic plate count E. coli Salmonella
Aerobic plate count Aerobic plate count Aerobic plate count Aerobic plate count E. coli E. coli E. coli Salmonella Salmonella Salmonella Campylobacter
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2.10
Summary
Table 7: Summary of NMD sampling requirements Product to be sampled Carcasses Post Slaughter and Dressing Bovine, bobby calf, caprine, cervine and porcine. Frequency Weekly Sites to be tested 3 sites, one side of carcass, alternating between the leading and trailing sides 3 sites, other side of carcass Salmonella (each seasonal sampling window, except porcinerefer 2.5.2) Carcasses post Slaughter and Dressing Ovine Weekly 1 site, one side of carcass, alternating between the leading and trailing sides. Carcasses Post Slaughter and Dressing Ostrich and emu Weekly 1 site, one side of carcass, alternating between the leading and trailing sides 1 site, other side of carcass Carcasses Post Chill Only required for bovine, bobby calf and caprine species Not required for hot boning premises Weekly, for a 6 consecutive week annual sampling window, at the start of the season 3 sites, one side of carcass, alternating between the leading and trailing sides APC/ E. coli Salmonella is not required Salmonella APC/ E. coli APC only. Salmonella is not required Analyses APC/E. coli
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Table 7: Summary of NMD sampling requirements (continued) Product to be sampled Primal Cuts Bovine, bobby calf, caprine, and cervine Weekly 1 site on required number of separate cuts (same site as on carcass) APC/ E. coli Diluent remaining after APC/E.coli test is composited for Salmonella Salmonella (each seasonal sampling window) Bulk Meat Bovine, bobby calf, and caprine. Weekly 10g from 5 locations within each carton composited, from which a 25g analysis sample is taken Salmonella (each seasonal sampling window) Poultry Whole Carcass Rinse Every processing day One processing day a week for VLT Whole carcass is rinsed with ssBPW Campylobacter for each carcass. Additionally - Salmonella for one carcass. APC/ E. coli Diluent remaining after APC/E. coli tests is composited for Salmonella Frequency Sites to be tested Analyses
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2.11
Weekly Sampling Plan for Red Meat Random Rotational Sampling
To ensure that a complete evaluation of the process is made, samples must be selected randomly such that every animal has an equal chance of selection. The sampling plan for red meat must include a randomly selected time each week to sample all products types (carcasses, cuts and bulk as per premises scope and as required for a particular species) for each species. When there are multiple red meat species processed at a premises, a single selected time period to sample all species can be used (same day, shift and run for all species) if you have enough samplers and laboratory personnel to process the samples. Alternatively, a different day of the week or time in the day may be randomly selected for the other species from the remaining days or part days. Red meat VLT premises must base their sampling plan on the farmed animal/cervine plan as above and randomly select a day, shift and run for each week or part week of processing. 2.11.1 Sample selection
First priority: to sample the required carcasses, cuts and bulk for each species each week. MPI can provide a random sample selection macro available to assist (contact the NMD Administrator). The order of selection; day, class, shift, run, time can be varied. The selection can be made in any order as long as all the parameters required by NMD random rotational sampling programme are met. 2.11.1.1 Day: random/rotational selection of day of week Ascertain the days in the week processing for each particular species is taking place. All processing or part processing days (including statutory holidays and weekends) must be eligible for selection. Randomly select an order of days from days available by randomly selecting one and rotation through the remainder (in any order). Random sampling only, without rotation through all available days is not permitted. All available processing days must be sampled from. For example if there are 5 days in the week available for processing over 5 weeks all days will be selected for sampling.
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Table 8: Random rotational of weekdays Week number Day 1 Wednesday 2 Monday 3 Friday 4 Tuesday 5 Thursday
Ovine species: Random/rotational sampling of sampling days is required for ovine species. However, where reasonable practical constraints exist, a specified day may be excluded from sampling without the scientific proof required under section 2.11.3 and on agreement with MPI VA. Where not all days in a processing week are available for sampling, random/rotational sampling must occur through the remaining days. 2.11.1.2 Class: random/rotational sampling of class Where there is more than one class random rotational sampling of classes is required. This is not dependent on throughput. The proportion of livestock classes processed at premises is irrelevant under the NMD programme and must not be used to deliberately bias the selection. Throughput only affects the number of samples collected if the premises is classified as Very Low Throughput. The class selected should be used for all product types (carcasses, cut, bulk) of that species that week. Ovine 1 class, lamb, is required to be sampled except on processing weeks where only sheep is being processed, in which case sheep must be sampled. Ostrich and emu 2 classes if both ostrich and emu are being processed. Cervine 2 classes; fallow and other - wapiti, red, hybrid and sika grouped as one class. Similar to tossing a coin for heads or tails randomly select the first class of fallow or other to be sampled every 2 weeks. Bovine 3 classes; bull, cow, prime. Randomly select one of the 3 classes for week 1, then randomly selected week 2 from the remaining two classes. The class remaining will be the one for week 3. Porcine 4 classes: suckers, porkers, baconers, choppers. Randomly select one of the 4 classes for week 1, randomly select week 2 from the remaining three classes, and week 3 from the remaining two classes and the class remaining will be the one for week 4. Variation to this may be required to ensure choppers are selected for one week of each month if choppers are being processed that month.
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Table 9: Random rotation of classes Week number Ostrich/Emu Cervine Bovine Porcine 1 Ostrich Fallow Cow Suckers 2 Emu other Prime Porkers 3 ostrich other Bull Choppers 4 Emu Fallow Bull Baconers 5 Emu other Cow Choppers 6 ostrich Fallow Prime Suckers
The italicised class indicates where the rotation starts for each species. 2.11.1.3 Shift: random/rotational sampling of shift Samples must be collected from all processing shifts. When multiple shifts are running for a species or a product type of a particular species rotate through all shifts. Ovine species: Rotational sampling through all processing shifts is required for ovine species. However, where reasonable practical constraints exist, a specified shift may be excluded from sampling without the scientific proof required under section 2.11.3 and on agreement with MPI VA. Sampling must be carried out on the other shift on the selected day of that week or from the same shift on another randomly selected day of that week. Bovine, bobby calf, caprine cervine, ostrich, emu and porcine where there are two shifts: alternate between day and night shift such that over two weeks both shifts are sampled. Table 10: Alternating shifts Week number Shift 1 Day 2 Night 3 Day 4 Night 5 Night 6 Day
2.11.1.4 Random selection of run It is important that the choice of run from which samples are to be collected is randomly selected. Every carcass, primal cut or bulk carton has to have an equal chance of being selected during the chosen run. 2.11.1.5 Random selection of time Select the initial item by randomly selecting the time in the run. No element of routine timetable practises is acceptable. Only the initial item needs to be randomly selected. Subsequent, but not consecutive, items can be selected in sequence after the previous item has been sampled and returned to the rail or the conveyor. Selection of items will be at time intervals equivalent to the time required to sample. Selecting 5 primal cuts at once from the conveyor belt and placing them on a table to swab at
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the same time is NOT ACCEPTABLE, except where there is insufficient product type remaining in the week to make up the samples. Another acceptable method is selecting the first carcass randomly from carcass ticket numbers in that run. The time may be chosen by the 24 hour clock system or minutes into the run with a sampling error of 10 minutes either side. For example, 15:33 10 minutes (the range is between 15:23 15:43). If time segments are used the time segments are to be no greater than 20 minutes. No time either side of the time segments is permitted. It is NOT 20 minutes plus or minus 10 minutes at each end, allowing 40 minutes altogether. If the carcass, cut or bulk sample available at that time is unsuitable (e.g. carcass goes on to detain rail), then select the next available appropriate carcass, cut or bulk. The time recorded at time of sampling (am/pm, 24 hour clock, carcass ticket number or minutes into a run) must be in a format that can be interpreted/translated to 24 hour clock units in the final results submitted to NMD. Table 11: Random/rotational selection of day, random selection of run and time Week number Day Run Time (using ticket number) Time (24 hour clock) Time (minutes into run) 2.11.2 1 Wednesday 4 4045 2 Thursday 3 3125 3 Friday 1 0215 4 Monday 4 3701 5 Tuesday 2 2245 6 Tuesday 2 1523
15:33
13:01
08:22
14:45
11:55
10:37
15
78
61
23
42
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Departure from the originally selected sampling period
Any changes from the originally selected sampling period must be justified. If cuts and bulk cannot be sampled in the same run or day selected for carcasses, randomly reselect for cuts and bulk. If the production schedule cause days available to sample for a species to alter from what was expected, randomly reselect from remaining times available for that week. The most important feature is to complete the required sampling and analyses for each species each week.
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Regarding random run and time selection, it is vital there is adequate communication between the samplers and the process personnel on the day. The slaughter floor should notify the sampler if they suspect processing may finish early or that there may be other interruptions that may have an impact on sampling. The sampler must then randomly reselect from remaining time available that day or reselect the day and time from remaining days available that week. Where the random time selection is near the end of the run sampling may commence 10 minutes prior to the randomly selected time. Any remaining samples must be collected at the beginning of the next run unless not available (refer below for further options to complete sampling). The key task is to collect the carcass, cut and bulk samples required for each species each week. Where there is insufficient product in the selected sampling period the following remedial actions can be taken to complete the sampling requirements. In order of priority, collect samples from: incomplete cartons; across multiple runs within a shift; multiple product types: bulk 95CL, 60CL, 80CL within a class; different classes; across multiple shifts; across multiple days.
Any of the changes described above (for example, alterations to shift, day, class run and or time) must be documented along with reasons for the departure from the original randomly selected parameter. 2.11.3 Modifications to the random sampling plan
Modifications to the random sampling plan at individual premises for all species, except ovine species, may be approved by MPI. Examples of modifications include: 1. Use of dayshift sampling only instead of day and night shift sampling. 2. Choosing only some particular days of the week for sampling instead of all available processing days. Sound statistical analysis will be required to show that microbiological outcomes will not be compromised by any modifications. It is recommended that where there are shifts that data is collected for shift comparisons and that verification of no significant differences between shifts is completed before progressing to statistical comparison between days of the week.
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It must also be shown that by electing to sample from only one shift and/or particular day of the week that this is not bias the coverage of all possible processing scenarios. The statistical analysis and conclusions must be fully documented by the premises and approved by MPI VA. MPI VA may choose to confirm the statistical conclusions drawn with MPI before granting approval. Once approved the premises must notify the NMD Administrator of the modified sampling plan and provide a copy of the approved justification. Ovine species: Modification of random sampling plans is prohibited except on occasion where practical constraints prevent the required sampling. Practical constraints such as courier schedules must be identified and documented by the premises and agreed with MPI VA. Premises are required to document and maintain records of the departure from sampling a particular shift (day or night shift) and of the particular days of the week nominated for ovine sampling. These records must be available for MPI VA to view at any time. The NMD Administrator must be notified of the modified ovine sampling plan. 2.11.4 Practical constraints and technical failures
Due to the premises production schedules, breakdowns, sample transport problems, or other, meeting the random sampling plan and number of weekly samples required may not always be possible. Should a technical problem occur randomly reselect from the remaining available times that week (This applies to red meat standard throughput and VLT premises). If no other times are available in that sampling week, two sets of samples are not required in the following week. When expected sample results are non-existent due to technical failure or production schedule constraint, the factors involved must be recorded by the premises and supplied to the NMD Administrator promptly. The NMD Administrator will note this in the sample ledger for future reference. If the practical constraint is routine, for example no sampling on Friday afternoons because of courier problems, a detailed justification must be documented. This must show that all alternatives have been researched and are impossible to implement. This must be approved by MPI VA who may choose to discuss this with MPI before granting approval. Once approved the premises must notify the NMD Administrator of the approved derogation and supply a copy of the supporting documents.
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2.12 2.12.1
Poultry Broiler Sampling Requirements Number of samples
A set number of poultry broiler samples must be taken: Standard throughput: select 3 carcasses each processing day: all 3 carcasses must be analysed for Campylobacter, randomly select 1 of the 3 carcasses to be analysed for Salmonella in addition to Campylobacter, VLT; 3 carcasses must be selected on one* processing day per week; all 3 carcasses must be analysed for Campylobacter, randomly select one of the 3 carcasses to be analysed for Salmonella.
The operator must ensure 3 carcasses are collected each processing day for standard throughput and 3 carcasses are collected on one processing day each processing week for VLT. The operator must communicate production schedules and any variation to production schedules to the laboratory at the earliest opportunity. * Operators of a VLT premises may apply to the NMD Administrator to modify the collection of 3 carcasses in one week to be conducted over more than one processing day (where there is more than one processing day available in a particular processing week). The alternative sample selection programme must be fully documented by the operator and approved by MPI VA prior to application. 2.12.2 Selection of sampling days
Standard throughput: Sampling must be conducted every processing day. VLT: 3 samples from one single randomly selected day of the processing days available that week. The table below gives two examples of how this requirement can be met.
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Table 12: VLT sampling days Monday Tuesday Wednesday Thursday Friday Saturday Sunday Total Samples
Randomly select 1 sampling day from processing days available 5 processing days, Monday Friday: Exclude Exclude 3 Exclude Exclude Np Np 3
1 processing day 3 Np Np Np Np Np Np 3
2.12.3
Shift; random/rotational sampling of shift
If shifts are undertaken then take samples from alternate shifts. Table 13: Standard throughput shifts Processing day Shift 1 day 2 Night 3 day 4 night 5 day 6 Night
Table 14: VLT Shifts Week number Shift 2.12.4 1 day 2 Night 3 day 4 night 5 day 6 Night
Random selection of time within a processing day All processing times during that processing day must be available for selection. You must not alter the randomly selected time to allow for convenient times to sample or according to flocks, sheds or cuts or any other preference.
Random selection of time From unique carcass identification numbers across the entire days processing, or From all times processing is underway that day (in minutes) The randomly selected time (in minutes) must be recorded prior to sampling. If it is not practicable to conduct the test on the specified minute a window of up to 10 minutes either side of the originally selected random time is permitted in which to conduct the sampling. The reason for departure from the originally selected time must be recorded by the sampler.
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The actual sample time must be recorded in a format (am/pm, 24 hour clock, unique carcass number) that can be interpreted/translated to 24 hour clock units in the final results submitted to NMD.
Standard throughput: Each of the three carcasses needs to be independently randomly selected from all the times available in each processing day. Where premises are operating day and night shifts they must collect all three samples from one shift each day as per Table 13 above. VLT: The initial carcass must be randomly selected. Collect the remaining carcasses at time intervals equivalent to the time required to bag or rinse each carcass. An alternative is to independently randomly select all 3 carcasses across the processing day as per standard throughput. Carcasses for the day must not be collected all at once from the line and then bagged or rinsed, unless there are no other opportunities to complete collection of the 3 carcasses. 2.12.5 Changes to processing schedules
If processing schedules change in a manner that means that the original random sampling plan can not be followed any remaining samples that day (standard throughput) or week (VLT) must be randomly selected from the remaining available processing time. For VLT premises if there is doubt on the number of days (or which single day that week) processing is to be undertaken, ensure that sampling is undertaken on the first processing day. This variation to random selection of sampling day must be recorded. For standard throughput and VLT; if less than the required samples are taken each missed sample will be recorded on the database as non-compliant for the poultry Campylobacter performance target (CPT). 2.12.6 Poultry Campylobacter Performance Target (CPT) programme sampling
requirements The required number of samples must be taken, as specified in 2.12.1. There are no practical constraints allowed for within the CPT Programme. If the required numbers of samples are not taken, the default will be to register the missed sample result/s as a result greater than the CPT moving window limit. Where there are technical failures such as samples not delivered to the laboratory within the correct time, or a laboratory error during analysis, these will be recorded on the database as technical failures. Where premises processing schedules change suddenly, there are breakdowns, sample transport problems or laboratory errors, variations to the above random sample selection protocols are permitted to allow sampling to be undertaken from the remaining processing that day or week to make up the required numbers. Such variations must be recorded.
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The operator may wish to undertake extra sampling routinely to allow for practical constraints and technical failures. Where extra sampling is undertaken, the operator must identify the original NMD selection and the extra sample/s selected. The Campylobacter enumeration results entered in the NMD database will commence using the original NMD samples in the first instance followed with the extra sample/s if necessary, to give a total of 3 results over 1 processing day for standard throughput and 3 results over one processing day per week for VLT.
December 2012 Sampling
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3. Sampling
3.1 Manufacture of Diluents for Sampling and Analysis A component of the equipment required for sampling will be sterile diluents for moistening swabs. Diluent aliquots may already be added to vials that swabs will be directly placed into at time of sampling. Sterile diluents are used for: 1. Moistening APC/E. coli carcass and primal cut swabs. 2. Moistening APC/E. coli/Salmonella primal cut swabs. 3. Suspension of APC/E. coli bulk meat samples. 4. In dilution series for APC and E. coli analyses. 5. Rinsing whole poultry carcasses. Whether sterile diluents have been manufactured by the laboratory or an approved supplier the expiry dates must be recorded and no sterile diluent outside of expiry date used for sampling. Peptone diluent 0.1% peptone with 0.85% NaCI. Note: for bobby calves where the chemical intervention, acidified sodium chlorite (ASC), has been applied prior to sampling, the addition of Sodium thiosulphate to dechlorinate the sampled is required. Table 15: Peptone diluent manufacture 0.1% Peptone Ingredients Peptone Sodium chloride (NaCI) Distilled water Gram/litre 1.0 8.5 1000 ml
Reference: MIMM 2005 or later edition, 5.7.2 (0.1% peptone manufacture) which applies to all species and 11.1.5 (dechlorination of water sample using sodium thiosulphate) which applies to bobby calf samples post ASC intervention only.
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Table 16: The volume of peptone diluent required Species Sterile one diluent volume peptone Zero (swab, whole tissue immersion) dilution Ovine, caprine Bobby calf, bovine, cervine, ostrich, emu, porcine Bulk meat sample suspension Buffered Peptone Water (BPW) Single strength buffered peptone water (ssBPW) Sterile ssBPW is used for 1. Moistening swabs for collection of carcass samples solely for Salmonella analysis. 2. Carcass composite Salmonella analysis making up the volume required for the sample pre-enrichment broth. 3. Poultry whole carcass rinse diluent for poultry broiler carcasses sampled for Campylobacter and Salmonella testing. 400ml of ssBPW per carcass is required for all poultry rinses. For poultry whole carcass rinse samples and bobby calf Salmonella samples where chlorinated antibacterial agents are used the addition of sodium thiosulphate is required. The proportion of sodium thiosulphate to add to ssBPW is listed in Table 17. Where antibacterial agents other than chlorine based ones are used suitable non-antimicrobial neutralising additives must be determined. Table 17: ssBPW manufacture ssBPW Ingredients Peptone Sodium chloride (NaCl) Disodium phosphate (Na2HPO4) Potassium phosphate (KH2PO4) Sodium thiosulphate (Na2S2O3) where chlorinated antibacterial agents are used to decontaminate carcasses (poultry and bobby calves only). Distilled water Final pH 7.2+/-0.2 at 25C Reference: MIMM 2005 or later edition, Chapter 7.7.4 gram/litre 10.0 5.0 3.5 1.5 1.0ml of a 3% (Na2S2O3) solution 1000ml 10ml 15ml 225ml
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Double strength buffered peptone water (dsBPW). Sterile dsBPW is used for: 1. For primal cut swab sample composite Salmonella analysis; addition of equal volume of dsBPW with remaining peptone diluent to constitute as ssBPW pre-enrichment broth. 2. For bulk meat sample composite Salmonella analysis addition of equal volume of dsBPW with remaining peptone diluent to constitute an ssBPW pre-enrichment broth. Table 18: dsBPW manufacture dsBPW Ingredients Peptone Sodium chloride (NaCl) Disodium phosphate (Na2HPO4) Potassium phosphate (KH2PO4) Distilled water Final pH 7.2+/-0.2 at 25C Note: this is a formulation specific to NMD requirements. 3.2 Templates Required for Swab Sampling Sample collection for carcasses and primal cuts requires the use of templates to delineate the site to be swabbed. 3.2.1 Template materials Gram/litre 20.0 10.0 7.0 3.0 1000ml
Materials from which templates can be constructed: 1. Hoops (circular or square) made of 3mm stainless steel rod with handle that can be hooked over a finger. 2. Flexible material (plastic or cardboard). 3. Flat rigid sheet metal. Unsuitable for 100cm templates due to the large area and curved/irregular nature of the surface to be sampled. The size of all templates used for sampling must be verified in laboratory calibration records.
2
53
Examples of template designs:
Table 19: Suitable template designs Area to be sampled 5cm Ovine, caprine
2
25cm Bobby calf, cervine, ostrich and emu, and porcine 56.4mm 50.0mm x 50.0mm Suitable
100cm Bovine
Area measurements
Circular diameter/mm Square Side/mm
25.2mm 22.4mm x 22.4mm Suitable
112.8mm 100.0mm x 100.0mm Suitable
Hoops (circular or square) made of 3mm stainless steel rod with handle Flexible material plastic or cardboard Flat rigid sheet metal 3.2.2
Suitable Suitable
Suitable Suitable
Suitable not suitable
Sterilisation of templates
It is preferable to use pre-sterilised (autoclaved) multiple templates. Otherwise for metal hoops and templates after each sample sterilise these for the next sample by: 70% ethanol for flame sterilisation; or Ethanol/iso-propanol based disinfectant wipes.
54
3.2.3
Sterile cotton tipped swabs
Table 20: Swabs required for sampling SPECIES Ovine, caprine Bobby calf, cervine, ostrich/emu, porcine 4 2 wet, 2 dry Bovine Poultry
Number of swabs required per carcass site or cut site
2 1 wet, 1 dry
6 3 wet, 3 dry
No swabs Whole carcass rinse method
Swabs must be sterilised before use. A pack of about 30 for use with each sampling batch is a practical number. Any remaining swabs must be re-sterilised before re-use in another sampling batch. Note: Commercial swabs supplied in individual plastic tubes and returned after sampling to those individual plastic tubes can be used however certain conditions apply. All tubes must be labelled. The suspension buffer required for each site sampled must be used to rinse out the inside of all the tubes of swabs associated with that site in order to remove any bacteria that may have been smeared onto the inner walls by the swabs during sampling and transport. 3.3 Collection of Samples Trained sample collectors must be used for sample collection. Refer to Part 1 clause 4 and Schedule 1 section 1.5 of this Notice. 3.3.1 Equipment required for sampling
For swabbed sites the following requirement is required: 1. Templates size required for species. 2. Sterile vials containing 4 9 plastic or glass beads with or without diluent. If the vials do have diluent ensure the diluent volume is correct for the species to be sampled. 3. Diluent for swab moistening as required for the type of sampling. 4. Sterile cotton tipped swabs. 5. Flaming alcohol or wipes if required for sterilising templates.
55
For bulk meat: For bulk meat the following equipment is required: 1. Knives (sharpened with appropriate equipment or by experts on site) or scalpels and blades. 2. Tweezers if required. 3. Plastic sample bags with twist ties or equivalent for example, whirl pac bags. 4. Gloves (vinyl or latex as required) or plastic bags (these must be clean. Sterilisation not required) to cover hands. 5. Flaming 70% ethanol or ethanol/iso-propanol containing disinfectant wipes for sterilising the knife, or scalpel blade, and tweezers. For whole rinse poultry carcasses: For whole rinse poultry carcasses plastic sample bags with twist ties or equivalent for example, poultry rinse bags with a tear tie are required. General: In general the following equipment is required: 1. Appropriate headwear, overalls/coats and trousers, boots, aprons for entering an edible area. 2. Pens and paper to record sampling details and indelible pen to label vials as required. 3. Spare sterile templates, vials, swabs and/or gloves/plastic bags. 4. Insulated containers with ice packs (slicker pads or bags of shaved ice). 5. Ladders or stools for access to sampling sites where necessary. 3.4 3.4.1 Sampling Method Wet/dry swabbing technique
The following is the wet/dry sampling technique: 1. Fill out the laboratory sample log and label all vials (with indelible pen or coded adhesive labels) to correspond with date, time, sample sites, product type, class and species. 2. 3. 4. Loosen the caps of vials, but do not open. Document the time that sampling begins. Unwrap a sterile template or alcohol flame/wipe a hoop or metal template. Ensure that the template does not become contaminated. Hold the template in one hand. 5. Carefully extract the required number of swabs from their sterile wrap.
56
6.
Hold the shaft/s of half the number of swabs required per site between the fingers and thumbs of one hand and moisten in the diluent for 5 seconds. Leave the other half of the sterile swabs dry and hold either separately like chopsticks in the same hand or the hand in which the template is being gripped.
7.
Press the sterile template onto the surface to be sampled. Hold firmly. Care is required not to smear the template over the sampling area.
8.
Take the wet swab/s and rub them over the area defined by the template. Rub the wet swab/s vertically, then horizontally, then diagonally across the entire surface. The sampler is required to swab in all three directions to cover the surface area adequately but not necessarily in the order stated. Rub in all three directions, one way following the other. Rub for at least the specified times in Table 21.
Table 21: Time required to rub wet swabs Ovine, caprine 2 5 cm 10 seconds swabbing time Bobby calf, cervine, ostrich/emu, porcine 2 25 cm 20 seconds swabbing time Bovine 2 100 cm 20 seconds swabbing time
It is important that the swab/s are rotated slowly and continually, by rolling the shafts between the fingers and the thumb (take care not to drop or contaminate the dry swab in the process). Use as much pressure as possible, but avoid breaking the shaft. Do not hold a swab on such an oblique angle that the shaft contacts the sample site or template. Only the bud should make contact with the sample, not the shaft. If a swab becomes contaminated by the contact with foreign non-target surfaces discard the contaminated swabs and resample another carcass or primal cut. 9. Hold the wet swabs to one side, and repeat swabbing with dry swab/s until the area being sampled is dry. 10. Insert the wet and dry swabs into a labelled transport vial. The vial may or may not contain the diluent zero dilution volume required for number of swabs/species. Break the swabs by pressing the buds against the inside of the vial near the rim and press down on the lid to aid in snapping the shafts from the buds. Care is required with aseptic technique and dexterity to prevent fingers contaminating the swabs, inside of lid or inside of vial. Discard and resample from a new carcass or cut if any contamination event occurs at this point. 11. Close the vials and place in an insulated container avoiding direct contact with the ice pack. 12. Disinfect templates between sample sites and carcasses/cuts, or use separate presterilised templates for each sample.
57
3.4.2
Whole tissue composite sampling technique for bulk meat
The following is the whole tissue composite sampling technique for bulk meat: 1. Fill out the laboratory sample log and label all plastic sample bags with twist ties or equivalent for example, whirl pac bags (with indelible pen or coded adhesive labels) to correspond with date, time, sample sites, product type class and species. 2. 3. 4. 5. Document the time that sampling begins. Select a carton after packing and before strapping. Open the carton to be tested and carefully peel back the plastic liner. Select 5 whole tissue samples, each of approximately 10g in volume, from the 4 diametrically opposite corners of the full carton and 1 from the centre of the full carton. No extra whole tissue samples are needed when Salmonella testing is required. Figure 12: The position of the 5 sites to take whole tissues samples from in a bulk meat carton
6.
Using a glove or a plastic bag inverted over your hand or sterile equipment such as meat hooks, tweezers or knives, reach into one of the 5 sites in the carton identified in figure 12. Cut out a whole tissue sample by trimming a thin layer from an original external carcass surface if possible. Each whole tissue sample weighing approximately 10 gram to result in at least 50 gram collected per carton.
7. Deposit the sample aseptically into a sterile sample bag. 8. Repeat at the 4 remaining sites in the carton depositing samples into the same sample bag. 9. Seal the sample bag with tape or twist tie. 10. Place the sample bag into an insulated container avoiding direct contact with ice packs.
58
11. Re-sterilise knife with alcohol flame/wipes (and tweezers if you are using these) or change scalpel blade between cartons. 12. Make sure all implements used (knives, tweezers, gloves or plastic bags, ties) are not left inside the carton after completion of sampling. 3.4.3 Poultry: whole carcass rinse
Each poultry whole carcass rinse sample will be analysed for Campylobacter. In addition one whole carcass rinse sample of the set taken on the day of sampling will be analysed for Salmonella as specified in section 2.12.1. 1. Fill out the laboratory sample log and label all plastic sample bags (with indelible pen or coded adhesive labels) to correspond with date and time of sampling. 2. Sampling periods: a. Standard throughput; 3 samples per processing day are required. Each of these samples will be taken at separate randomly selected times. There must be three discrete sampling times across that days processing. For each of the three sampling periods the date and time must be recorded. b. VLT: Select the first carcass of 3 for sampling (or bagging) from the chain. Record the time of selection of the first carcass. Collect the remaining carcasses in this single sampling period, ensuring that carcasses are not taken consecutively from the chain. 3. If the carcass is collected prior to the first drop point suspend the carcass for T to drain excess water from the carcass. This procedure will mimic the usual handling and draining of carcasses at the premises. 4. Invert the bag over the hand to form a glove. Take care not to contaminate the exposed internal surface of the bag. 5. Open the sample bag aseptically. 6. Pick up the drained bird/carcass by the legs (or air-chilled bird from the chain) using the bagged hand and reverse the bag over the carcass. 7. Bottles (400ml) of sterile ssBPW for rinsing the carcass for Campylobacter, E coli and Salmonella analyses will have been prepared as per section 3.1 Table 17 and must be pre-cooled to 4C for use at 4C at time of sampling. 8. The carcass can be rinsed on selection or transported to the laboratory for rinsing in which case the carcass must be transported to the laboratory in an insulated container with ice packs, but not in direct contact with ice packs, and not frozen; arriving at the laboratory within 15-30 minutes of carcass selection. The temperature of the carcass on arrival at the laboratory does not need to be measured.
59
3.4.3.1 Rinse procedure 1. Rinsing involves pouring 400ml of ssBPW diluent over the carcass in the bag, expelling most of the air from the bag then securing the bag tightly by hand or with a twist tie. 2. While securely holding the bag, rinse the outside of the carcass, including the wings, legs, and the inside cavity by gently rocking the carcass in an arching motion, transferring the weight of the carcass from one hand to the other. The hands can simultaneously be used to massage the carcass surface, particularly around the wings and the legs. 3. Time require for the poultry whole carcass rinse procedure is two minutes (120 seconds). Commence by rocking the carcass permitting the rinsate to pass across the external surfaces. The rinsate will appear yellowy in colour with suspended fats. Periodically position the bird and clear any fat or skin covering the vent and tilt such that the rinsate can enter the cavity. Swirl to rinse the entire internal cavity. The rinsate on exit from the cavity should be a pink to red colour. Continue the rocking motion to pass the rinsate over external surfaces and complete an internal cavity rinse at least once more during the two minutes. After the two minutes a rinsate with a reddish tint and thicker with suspended fats and cells should be visible. Table 22: Time required Poultry carcass whole rinse 120 seconds (2 minutes) of rocking for 400ml rinsate
4. To transport the rinse sample to a laboratory place the sample bag/vial into a second bag, secure the opening and place in an insulated container with ice packs for transport to the laboratory. Avoid direct contact of the rinse sample with the ice packs.
60
Figure 13: Poultry whole carcass rinsing
Arrival of poultry broiler sample; 400ml of ssBPW to be added.
Rocking the bird
Yellowish rinsate after the rocking procedure, tipped into a corner
Opening the vent entry into the cavity.
Rinsate after passage through cavity
Final rinsate view after 2 minutes.
61
5. Isolating the carcass rinse for analysis Reset the bag with the carcass on a flat surface. Open the bag and remove the carcass aseptically using a gloved hand. Expel the air (to ease packing) and secure the top of the stomacher bag to prevent contamination and/or spillage of the rinse sample; OR Prior to removal of the carcass aseptically transfer at least 70ml of the rinse sample to a sterile vial for transportation of the laboratory. Cutting a corner of the bag off is one practical means to achieve this. 70ml will provide enough rinse sample for analysis plus back up. 6. Disposition of carcasses Carcasses rinsed in the processing room may be returned to the immersion chiller. Carcasses subjected to air chilling shall be rinsed with potable water prior to return to the chain. 3.5 Carcasses rinsed in the laboratory must not be returned to the processing room.
Transportation of Samples to the Laboratory for All Species In order to protect samples from extremes of temperature and minimise changes to the microflora during collection and transport, all samples must be stored and transported in an insulated container. For NMD purposes extremes of temperature are defined as <0C and >25C. All samples must be subjected to sufficient frozen coolant during sample collection and transportation to keep the samples cold, but not frozen, regardless of whether the sample is to be immediately analysed, transported off-site or held for a period of time.
62
3.5.1
Red meat species samples
Premises may choose to have samples transported to an on-site lab for some analyses and off-site for others according to schedules, laboratory scope of testing and other factors. Figure 14: Red meat species sampling timetable
ssss
3.5.2
Transportation temperature and times for red meat samples Sample collection temperatures The temperature within the insulated container used at the time of sampling should be maintained between 0C and 5C (not frozen), and must not exceed 10C.2 Storage temperatures A refrigerator operating in the 0C 5C range must be used to store samples and diluents. Transportation temperatures The temperature of samples within the insulated container used for transport should be maintained between 0C and 5C (not frozen), and must not exceed 10C in transit. Controls - Transport sample temperature shall be measured/monitored (using a control vial/sample to prevent contamination) at a site within the transportation container that will most likely reflect the temperature of the samples;
Four Small Studies for NMD, AgResearch 2001
63
for swab samples stored transported dry, immediately adjacent to the swabs;
o o
for swab samples stored/transported in liquid medium, within the medium; for whole tissue samples, immediately adjacent to tissue samples.
Frozen - Samples that are received frozen shall be rejected by the laboratory and replacement samples collected.
High Temperature - Samples received that exceed 10C must be rejected and replacement samples sought.
Sample receipt - The laboratory receiving the samples must: o o record in the sample register the sample temperature on receipt. verify that analysis can be initiated within 24h, or 30h under exceptional circumstances, from time of collection of the first sample. Reference: ISO 17604:2003. Extensions to a maximum of 30 hours are permitted where practical constraints dictate, but not routinely. Extensions must be justified with reasons recorded. o verify that the sampler recorded as collecting the samples was a currently listed Certified, Associate or restricted sampler.
Verification of temperatures the laboratory must use data loggers to verify temperatures of insulated containers/samples from time of sampling to receipt by laboratory. Verification of temperatures is required every 6 months.
Premises must implement procedures to ensure that requirements for all analyses of samples, including Salmonella analysis, are complied with within the time periods specified. Laboratories, including off-site laboratories, are responsible for training on-plant personnel seconded to collect samples. Off-site laboratories are responsible for ensuring that transport companies, for example, on-site personnel, couriers are aware of the transport and storage temperature and time requirements. Off-site and on-site laboratories are responsible to ensure night shifts are informed of storage temperatures, transport to laboratory and time requirements. 3.5.3 Collection and transport of whole tissue samples
All bulk meat whole tissue samples; cold boned, hot or warm boned must be placed in an insulated container with sufficient frozen coolant at time of sampling to maintain a temperature between 0C and 5C (not frozen), and not exceeding 10C . Hot/warm boned whole tissue samples must be rapidly cooled to <5C but not frozen. Trials must be conducted to demonstrate;
64
temperature falls rapidly to <5C, samples do not freeze, that transport arrangements can still be met, and that E. coli NZRM 916 when inoculated into the whole tissue samples can be recovered.
All bulk meat whole tissue samples requiring storage before transport and/or analysis must be stored in a refrigerator calibrated to 0 - 5C. All bulk meat whole tissue samples must be transported in an insulated container maintaining between 0C and 5C (not frozen), and not exceeding 10C temperature in transit. The laboratory must conduct six monthly verification of temperature ranges by datalogging the temperature of bulk meat samples from time of sampling until commencement of analysis. 3.5.4 Transport of red meat Salmonella samples
Salmonella samples include: carcass Salmonella swabs, APC primal cut swabs, the bulk meat sample itself, primal cut and bulk original peptone APC/E. coli suspensions, and the final BPW suspensions. All Salmonella samples must be in an insulated container maintained between 0C and 5C (not frozen), and not exceeding 10C in transit, and/or stored in a refrigerator operating between 0C and 5C until commencement of analysis. Following receipt of samples by the laboratory initiation of Salmonella analysis is defined as the commencement of incubation of the BPW pre-enrichment broth. This may mean cooling of these original suspensions during APC/E. coli analysis and use of pre-chilled dsBPW to <5C to make up the final Salmonella sample suspension.
65
3.5.5
Poultry samples
Figure 15: Poultry sampling timetable

Select whole carcass in process room day shift or night shift. Record time of collection of first carcass.*
*First carcass in each discrete consignment of samples dispatched to the laboratory for that processing day.
Rinse carcass immediately in process room.
Transport carcass in an insulated container to the laboratory within 15 30 minutes of sampling.
Transport sample to on-site or off-site laboratory.
Rinse carcass immediately.
Insulated, arriving at or below 10C and stored at <5C.
Commence analysis immediately or store at <5C.
Initiate analysis within 24 hours after first carcass selected.
The maximum period may be extended to 30 hours, not routinely, with reasons recorded
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3.5.6
Preparation of poultry samples for transportation
See also 3.4.3 The process for the preparation of poultry samples for transportation is as follows: 1. Tightly seal the sample bag, transportation vial, or bag containing the carcass to be sampled; and 2. All samples must be immediately placed into an insulated container with frozen coolant ensuring the sample bags or transport vials are not in direct contact with the frozen coolant. 3.5.7 Transport and temperature requirements for poultry whole carcass samples
The carcass rinse sample or the carcass itself must be transported to the laboratory in an insulated container with ice packs, but not in direct contact with ice packs, and not frozen. 1. Whole carcasses not rinsed immediately in the process room, but transported to the laboratory for rinsing must be transported in insulated containers with sufficient frozen coolant to cool the sample. The temperature of the carcass on arrival at the laboratory does not need to be measured. 2. Whole carcass for rinsing in the laboratory must be preferably delivered to the laboratory within 15 minutes (no later than 30 minutes) of selection of the whole carcass from the processing chain and immediately rinsed. 3. Rinse samples transported to the laboratory must be transported in insulated containers with frozen coolant. The temperature of rinse samples within the insulated container used for transport should be maintained between 0C and 5C (not frozen), and must not exceed 10C. The temperature of the diluent, not just the airspace in the container must be maintained in this temperature range. 4. All samples are to be delivered to the laboratory as soon as possible after collection. 5. The time of collection of the whole carcass sample must be recorded. 6. The temperature of the rinse sample on arrival at the laboratory must be recorded. 7. Samples that are frozen shall be rejected by the laboratory and replacement samples collected from the same days processing if possible. 8. Premises must be able to verify that analysis can be initiated within 24h from time of collection for the first sample. This may be extended to 30h under exceptional circumstances, not routinely, with reasons recorded. 9. Laboratories, including off-site laboratories, are responsible for training on-plant personnel seconded to collect samples. Off-site laboratories are responsible for
67
ensuring that transport companies, e.g. on-site personnel, couriers are aware of the transport and storage temperature and time requirements. 10. Off-site and on-site laboratories are responsible to ensure night shifts are informed of storage temperatures, transport to laboratory and time requirements.
December 2012 Schedule 1 National Microbiological Database Programme Laboratory
68
Analysis
4. Laboratory Analysis
4.1 Dilutions Required Table 23: Dilutions required Analyses APC30 E. coli Campylobacter Red Meat zero to 10 (dilution series of 5) zero to 10 (dilution series of 4)
-3 -4
Poultry
0 0 -2
10 to 10 (dilution series of 3) 10 and 10 plus higher dilutions if required.

-1
Note: zero is the undiluted sample. 4.1.1 High end dilutions
All dilutions must be plated in duplicate until the premises data indicates at what dilution a colony count of greater than the maximum allowable is never, or rarely exceeded. The maximum allowable counts are: 300 CFU for spread plates; 150 CFU for spread plate half plates; 250 CFU for Petrifilm 150 CFU for Petrifilm
TM
APC; E. coli.;
TM
150 CFU for mCCDA direct plate Campylobacter When a valid count exceeds (>300, >250 or >150 on the highest dilution) the dilution series shall be immediately extended until such time as the premises demonstrates that the higher counts are not a consistent trend. It is very important to have a valid actual count of higher results for statistical purposes. A TNTC count can be any value greater than the valid count value of 300, 250 or 150 on the highest dilution tested and is thus more variable and unknown than a not detected count. A TNTC result can range from as low as a 2 log10 value up to any higher value (infinite). It could be 5 log10 or 10 log10. If a dilution series is not great enough this will lead to TNTC results and such high counts cannot be included in the database, which, in turn, leads to a poor representation of the true range of results. When insufficient dilutions are prepared and counts are above the maximum allowable count on the lowest dilution the laboratory must report to NMD a too numerous to count result which will be classed as a technical failure. A TNTC must be regarded as detection and/or higher than expected count when assessing compliance with premises performance criteria under HACCP and RMPs.
69
Analysis
Note that the area sampled will have an effect on the number of dilutions required. More bacteria are expected to be recovered from a larger sample area therefore more dilutions will be required. As regulatory limits will be applied to poultry broiler carcass Campylobacter enumeration results laboratories need to ensure that a suitable series of dilutions is plated to minimise the chance of TNTC results. If a TNTC result occurs it will be recorded as exceeding the CPT limit. Refer sections 4.10 and 6.9. When a TNTC count occurs the test does not need to be repeated. Upon receipt of a TNTC result the laboratory must extend the dilution series, in anticipation of higher counts, for at least 3 subsequent poultry processing days (standard throughput) or 3 subsequent processing weeks (VLT) following the TNTC result notification. 4.1.2 Low end dilutions
TM
All cases Undiluted (zero dilution) plates or Petrifilm
plates that contain less than the
lowest counting range of 30, 25, or 15 are to be counted and reported. Table 24: Summary of acceptable counting ranges Dilutions Spreadplate APC Spreadplate APC plate mCCDA direct plate 0-150 15-150 Petrifilm
TM
APC
Petrifilm E. coli
TM
10
-1 -2
0 -n
0-300 30-300
0-250 25-250
0-150 15-150
10 , 10 ,..10 4.2
Duplicate Plating/Analytical Precision Plating must be performed in duplicate, i.e. two agar plate, both sides of a plate or two Petrifilm
TM
inoculated from each dilution until laboratory precision data (for all analysts
performing testing) indicates otherwise. Duplicate plating (APC30 and E. coli) must be undertaken to give statistical confidence to the results obtained. Use the Analytical Precision calculation (for a single operator) in chapter 5 Appendix 1.7 of MIMM to determine a Z score which is then compared to a limit of 1.96. Record all duplicate plate counts that fall within the acceptable counting ranges as specified in section 4.1. If a laboratory demonstrates that it meets the Analytical Precision criteria outlined in MIMM 2005 or later edition, it may use singlet plating for all dilutions other than the undiluted (zero) dilution sample. For ILCP analyses related to NMD APC and E. coli tests use the NMD procedures described in this publication. Although successful precision analysis may have eliminated the need for duplicates on all dilutions for routine NMD analyses, duplicates are required for all dilutions for the corresponding ILCP tests.
70
Analysis
4.3 4.3.1
Preparation of Dilutions Introduction
When conducting a quantative bacterial analysis, sample suspensions must be diluted to an appropriate level that will result in 30 300 colony forming units (CFU) on the plate, 25 250 CFU if using APC Petrifilm mCCDA direct plate. Dilutions for viable bacterial counts are aseptically prepared in ten fold steps. All dilutions are performed using 0.1% peptone + 0.85% NaCI diluent. Reference: ISO 17604:2003. The dilutions volume used must be sufficient to inoculate duplicate plate count agar plates, duplicate Petrifilm , enough volume for the composite Salmonella pre-enrichment if required and leaving enough volume in case of error. Two dilution volumes are recommended: a. 9ml in 25ml universal bottles or vials, with 1ml transfer volume. b. 4.5ml in microcentrifuge or microdilution tubes, with 0.5ml transfer volume. The use of pre-poured dilution volume is allowed only if a laboratory can provide evidence that they do not lose volume during autoclaving (as defined in MIRINZ Bulletin 35 or MIMM 2005 or later edition, Section 2.5.6), and that they have an appropriate QA programme in place for ongoing verification. Always use a fresh pipette or pipette tip for each transfer of diluted sample. Ensure that dilutions are as homogenous as possible at all stages by: Carefully following the instructions for the use of micropipettors; Mixing each dilution thoroughly prior to the next dilution step to ensure that the bacteria are randomly distributed; 4.3.2 Definition of zero dilutions for calculation purposes
TM TM TM
or 15 150 CFU if using half plates, E. coli Petrifilm
or
The swab suspension (swabs immersed in the initial volume of either 10ml or 15ml) is an undiluted sample referred to as the zero dilution. The swab suspension is entered into the calculations as the 10 dilution. The whole tissue suspension is dilution but in order to maintain consistency with plate labelling, it is entered into the calculation as the 10 dilution (undiluted) and accounted for in the final numeric multiplier. 4.4 Preparation of Swab Samples for APC and E. coli Analysis Add to each vial with swabs.
0 0
71
Analysis
Table 25: Swab suspension volumes required Bovine, bobby calf, cervine, ostrich/emu, porcine Ovine and caprine
15ml sterile 0.1% peptone diluent to 10ml sterile 0.1% peptone diluent to each each vial vial Make certain there are 4 9 sterile glass or plastic beads in each vial. Close the vial and shake vigorously to loosen the cotton bud of the swab for 2 minutes (ensure consistency). Shaking can be performed by hand or by using a combination of initial shaking by hand and a vortex mixer in 3 x 10 second bursts. Note that shaking by hand is usually sufficient and enables several vials to be processed at once. The cotton buds should be well broken up by this treatment. The cotton swabs must be left in the diluent until all the dilution and plate inoculation procedures are completed. Do not remove the swabs from the vials at any stage. 4.5 Preparation of Red Meat Separate Carcass Site Samples Swabbed for Salmonella
Analysis Note: Salmonella testing to support official assurances for beef exported to Sweden and Finland or to any other country with the same requirements, eg Iceland, must use the required sampling and method specified for routine official test 2.4.2. Table 26: Preparing Salmonella sample suspensions Species Bovine 5 carcasses
6 swabs x 3 sites x 5 carcasses = 90 swabs 250 ml
Bobby calf, Porcine 5 carcasses

Cervine 3 carcasses
Ostrich/Emu 2 carcasses
4 swabs x 1 site x 2 carcasses = 8 swabs 30 ml
Caprine 5 carcasses
Total number of swabs Volume of ssBPW
VLT (Very low throughput) premises with only one carcass sample (no composite) Species
Total number of swabs Volume of ssBPW
Bovine VLT
6 swabs x 3 sites x 1 carcass = 18 swabs 50 ml
Bobby calf, Porcine VLT

Cervine VLT
4 swabs x 3 sites x 1 carcass = 12 swabs
Ostrich/Emu VLT
4 swabs x 1 site x 1 carcass = 4 swabs 15 ml
Caprine VLT
30 ml
Add an appropriate number of glass beads to the suspension, close the bottle and shake vigorously for 2 minutes to remove the bacteria from the swabs into the diluent.
72
Analysis
4.6
Preparation of Bulk Meat (Whole Tissue) Samples for APC, E. coli Note: Salmonella testing to support official assurances for beef exported to Sweden and Finland or to any other country with the same requirements, for example, Iceland, must use the required sampling and method specified for routine official test 2.4.2 Place the whole tissue samples (5 X ~10g) onto a sterile chopping board and finely dice (size reduce) with a sterile knife or other suitable aseptic means. Re-sterilise or use separate chopping boards and knives etc for each sample. Aseptically weigh 25g of diced tissue directly into a stomacher bag. To each bag add 225ml of sterile diluent (0.1% peptone + 0.85% NaCl). Stomach for 2 minutes.
4.7
Aerobic Plate Count, APC30
Applicable to red meat NMD programmes. Not applicable to poultry NMD programmes. The following APC methods are approved for the red meat NMD programmes: 1. Spread Plate method (Routine Official test 2.1.2) 2. Petrifilm
TM
Aerobic Plate Count method (Routine official test 2.1.3)
3. Spiral Plater method (Routine official test 2.1.4) 4.7.1 General considerations for plating
The following are general considerations for plating: 1. Pre-poured plate count agar plates must be dried before use. Plates may be dried using one of the following methods: i. ii. iii. incubation for 24h at 30-37C inverted with lids in place; inverting with lids ajar in a laminar flow cabinet for 20-30 minutes; inverting with lids ajar in a still air incubator at 30-37C for 1 to 2h;
iv. inverting with lids ajar in a forced air incubator at 30-37C for 30 minutes. 2. The swab samples in diluent with beads must be shaken for 2 minutes, bulk meat samples 25g/225ml peptone diluent must be stomached for 2 minutes before commencing analysis. 3. The same pipette/tip may be used when working up a dilution series from the most dilute level to the most concentrated level. 4. The required incubation temperature for the APC is 30C +/- 1C (ISO 17604:2003).
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Analysis
5. Initiation of analysis for APC is defined as the time when the sample is suspended and ready for serial dilution. 6. The incubation time is 48 hours. Where plates cannot be counted at 48h plates may be: I. incubated for up to but not longer than 72h II. removed from the incubator and stored for no longer than 48h in a laboratory refrigerator set at not greater than 5C. The extra storage time of the plates must be recorded on the result sheet. 7. Count only plates with between 30 to 300 colonies, 25 to 250 colonies for Petrifilm APC, 15 to 150 colonies for plates and Petrifilm See table 22: Acceptable counting ranges. 8. If colonies are indeterminate at 48h estimate a count, re-incubate for a further 18-24h and recount. The total incubation time must be recorded on the result sheet. 9. Express the result as the number of colony forming units (CFU)/cm or /g of sample. When there are nil counts on both of the duplicate zero dilution plates, express the result as not detected. 4.7.2 APC30 by spread plate method
2 TM TM
E. coli plates (ISO 17604:2003).
(Reference MIMM 2005, Chapter 6 or later edition). Carry out all quality control procedures for the media and methods using Pseudomonas aeruginosa NZRM 981 or Lactobacillus viridescens NZRM 3313 as the positive control. 4.7.2.1 Method The following is the APC30 by spread plate method: 1. Dispense 0.1ml (100l) volumes of the appropriate dilutions (100 non-diluted, 10 , 10 , 10 , 10 ) onto duplicate, dried plate count agar plates, or a single plate. 2. Start with the highest (most dilute) level of dilution. Mix the dilutions thoroughly before dispensing. The same pipette/tip may be used for all inoculations. 3. Spread the inoculum over the surface of the agar as evenly and as quickly as possible using a sterile spreader (bent glass rods or disposable plastic spreaders). In order to prevent cross-contamination, sterilise the spreader between plates both within and between dilutions. 4. Allow the inoculum to soak into the agar surface. This should occur within 15 minutes. If liquid does not soak in (i.e. the plates were not dried sufficiently), the viable bacteria present in the inoculum may start to replicate and spread, possibly resulting in an inaccurate count.
-3 -4 -1 -2
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Analysis
5. Invert the plates before incubation so that the agar is in the upper half of the petri dish. Inversion prevents condensation dropping onto the surface of the agar reducing contamination and preventing the spread of motile micro-organisms. Stack if necessary, and incubate at 301C for not less than 48h. 6. Count all colonies. Record all counts on the undiluted plates and only those between 30-300 on full plates or 15-150 on half plates. 7. Express the results as the number of colony forming units CFU/cm or /g of sample. 8. If colonies are not detected on both of the zero dilution plates express the results as not detected. 9. Where counts are greater than 300 on full plates or 150 on half plates on the highest dilution plate or half plate record the result as to numerous to count (TNTC). The TNTC must be regarded as a detection and /or higher than expected count when assessing compliance with premises performance criteria under HACCP and RMPs.
2
4.7.3
APC30: Petrifilm
TM
Aerobic Count Plate Method
(Reference MIMM 2005, Chapter 6 or later edition). 4.7.3.1 Method The following is the APC30 Petrifilm
TM
Aerobic Count Plate Method:
1. Carry out all quality control procedures for the media and methods using Pseudomonas aeruginosa NZRM 981 or Lactobacillus viridescens NZRM 3313 as the positive control. 2. Place the Petrifilm
TM
Aerobic Count plate onto a flat surface, label and lift the top film.
Do not allow the plate to curl. 3. Inoculate and process each Petrifilm inoculate several at the same time. 4. Lift the top film, and carefully dispense 1ml of the appropriate dilution onto the agar. 5. Slowly roll the top clear plastic film down onto the inoculum to prevent entrapment of air bubbles. 6. Distribute the sample evenly using the spreader provided (ridge side down). Petrifilm
TM TM
plate individually. Under no circumstances
Aerobic Count plates do not have a foam dam thats why the spreader is used ridge side down. Apply a gentle even pressure, but do not twist or slide the spreader. Carefully lift the spreader clear or the Petrifilm 7. Move the completed Petrifilm set.
TM TM
plate.
plate to one side and allow one (1) minute for the agar to
75
Analysis
8. In the meantime inoculate and process the next Petrifilm 9. Incubate all Petrifilm
TM
TM
plate.
plates for not less than 48h at 30C +/- 1C clear side up and in
stacks of no more than 20. 10. Count only plates with 25-250 colonies, the exception being the undiluted (zero dilution) plate if counts are less than 25. 11. Express the results as the number of colony forming units CFU/cm or /g of sample. 12. If colonies are not detected on both of the zero dilution plates express the result as not detected. 13. Where counts are greater than 250 on the highest dilution plate record the result as too numerous to count (TNTC). A TNTC must be regarded as a detection and/or higher than expected count when assessing compliance with premises performance criteria under HACCP and RMPs.
2
4.7.4
APC30: Sprial Plater Method
(Reference MIMM 2005, Chapter 6 or later edition). 4.7.4.1 Considerations The quality of pre-poured agar is paramount to the successful use of the Spiral Plater. Plates must be prepared according to the manufacturers guidelines supplied with the Spiral Plater. The plate surface must be smooth, the depth uniform and the surface uniformly dry but not dehydrated. Pre-poured agar plates must be dried before use. 4.7.4.2 Method The following is the Spiral Plater Method: 1. Carry out all quality control procedures for the media and methods using Pseudomonas aeruginosa NZRM 981 or Lactobacillus viridescens NZRM 3313 as a positive control. 2. Follow all instructions for use in the manual supplies with the spiral plater. 3. After inoculation, leave the plates on the bench right-way-up for 10 minutes to allow the inoculum to soak into the agar. 4. Invert the plates, stack if necessary, and incubate at 30C +/- 1C for no less than 48h. 5. Obtain a count either manually according to the manual supplied with the spiral plater or automatically using a computerised plate scanner.
76
Analysis
6. Do not count spiral plates with an irregular distribution of colonies caused by dispensing errors. 7. Express the result as the number of colony forming units; CFU/cm or /g of sample. 8. If colonies are not detected on the plate express the result as not detected. 9. Where counts out of range on the highest dilution plate record the result as too numerous to count (TNTC). A TNTC must be regarded as a detection and/or higher than expected count when assessing compliance with premises performance criteria under HACCP and RMPs. 4.8 Escherichia coli Petrifilm
TM 2
Note that red meat samples will be in a peptone diluent suspension. The Petrifilm
TM
E. coli method is the only method of E. coli analysis approved for the red
meat NMD Programme. Reference MIMM 2005, chapter 8.4 or later edition. 4.8.1 Method
1. Carry out all quality control procedures for the media and methods using Escherichia coli NZRM 916 as the positive control and Klebsiella pneumoniae NZRM 482 or Enterobacter aerogenes NZRM 798, as a negative control. 2. Initiation of analysis is defined as when the sample has been suspended as is ready for serial dilution. 3. Place a Petrifilm plate to curl. 4. Mix the dilutions thoroughly before dispensing. The same pipette tip may be used for all inoculations if you commence with the highest (most dilute) dilution. 5. Lift the top film and carefully dispense 1ml of the appropriate dilution into the circular well containing the agar. Note: Do not inoculate several at the same time. It is imperative to inoculate and lower the top film before proceeding to the next. 6. Slowly roll the top clear plastic film down on to the inoculum to prevent entrapment of air bubbles. 7. If necessary use the spreader provided flat side down, as Petrifilm
TM TM
E. coli plate on a flat surface and hold the plate flat. Do not allow the
E. coli plates have
a rim around the well, to distribute the sample evenly over the well. Apply a gentle even pressure to the spreader, but do not twist or slide the spreader. Carefully lift the spreader clear of the plate. 8. Move the completed plate to one side and allow one (1) minute for the agar to set. 9. In the meantime inoculate and process the next plate.
77
Analysis
10. Incubate all plates for 18-24h at 35 +/- 1C or 37 +/- 1C (36 +/- 2C is not acceptable) clear side up and in stacks of no more than 20. 11. Where plates cannot be counted at 24h the plates may be removed from the incubator and stored for no longer the 72h in a laboratory refrigerator set at not greater than 5C, or freezer. Storage time/temperature must be recorded on the result sheet. 12. If colonies are indeterminate at 24h, count as best as possible, re-incubate for a further 18-24h and recount. Record the incubation time on the result sheet. 13. Count as E. coli all blue colonies with or without gas bubbles (3M approved). i. Do not count colonies that have grown on the foam dam as the may not have been subjected to the selective influence of the medium thus might not be E. coli. ii. Count only plates with between 15 and 150 counts, the exception being if the plate from the non-diluted (zero) dilution sample has less than 15 colonies. iii. High numbers of E. coli will turn the medium blue and high numbers of Enterobacteriaceae will turn the medium red. Both situations make accurate enumeration improbable. Should this occur the highest dilution plated the samples must be recorded as (TNTC) and the operator immediately notified. 14. Express the results as the number of E. coli CFU/cm or /g or /ml of sample. If blue colonies with or without gas are NOT detected express the results as not detected. 15. Where counts are greater than 150 on the higher dilution plate record the result as too numerous to count (TNTC). A TNTC must be regarded as a detection and/or higher than expected count when assessing compliance with premises performance criteria under HACCP and RMPs. 4.9 Salmonella This method is applicable to all NMD Programmes; red meat and poultry. Note: Salmonella testing to support official assurances for beef exported to Sweden and Finland or to any other country with the same requirements, for example, Iceland, must use the required sampling and method specified for routine official test 2.4.2 4.9.1 Sample preparation
2
Reference MIMM 2005, Chapter 7.7. 1. Carry out all quality control procedures for the media and methods using Salmonella menston NZRM 383 as the positive control and E. coli NZRM 916 as the negative control. 2. Sample preparation. Samples are derived either from separate carcass swabs taken from a specific site sampled for Salmonella analysis only or from the primal cut swab, bulk meat or whole carcass (poultry) suspension.
78
Analysis
4.9.1.1 Red meat carcasses with a separate site swabbed for Salmonella analysis See section 4.5 Table 26. 4.9.1.2 Poultry carcass rinse samples and red meat primal cut and bulk meat swab suspensions for Salmonella analysis Poultry carcass rinse samples. On receipt of the poultry carcass rinse sample transfer 30ml to a separate sterile vial for use as the BPW suspension described in section 4.9.2. Red meat. See also Section 2.10 Table 7 for red meat species requiring Salmonella sampling/analysis for cuts and or bulk. For each of the primal cut initial swab suspensions take a specified volume of the diluent and composite into one vial then add an equal volume of dsBPW to the composite sample. Note: For VLT premises there will be only one cut and bulk sample each week. Thus there will be no composite sample. Table 27: Primal cut Salmonella pre-enrichment composite sample from swab suspensions Bovine 100cm
2
Bobby calf 25cm
Cervine 25cm
Caprine 5cm
5 samples of 8ml = 40ml + 40ml dsBPW = 80ml in total
VLT (very low throughput) premises with only one sample (no composite) 1 sample of 8ml = 8ml + 8ml dsBPW = 16ml in total 1 sample of 8ml = 8ml + 8ml dsBPW = 16ml in total 1 sample of 8ml = 8ml + 8ml dsBPW = 16ml in total 1 sample of 1ml = 1ml + 1ml dsBPW = 2ml in total
Bulk meat bovine, bobby calf, caprine species Salmonella enrichment composite sample from five 25g bulk meat stomached suspensions: 5 x 1ml suspension diluent = 5ml + 5ml dsBPW = 10ml in total. For samples of 60-80VL trim add 0.05ml (50l) of Tween 80 or 0.03ml (30l) of Terigol 7 to the enrichment volume. For VLT premises where there is only one bulk meat sample: To 1ml of suspension dilution add 1ml dsBPW = 2ml in total. For samples of 60-80VL trim add 0.01ml (10l) of Tween 80 or 0.006ml (6l) of Terigol 7 to the enrichment volume. Bulk meat samples are not required for cervine, ostrich and emu, ovine or poultry.
79
Analysis
4.9.1.3 Transport or retention of Salmonella samples for later analysis For red meat carcass sample where the Salmonella sample sites/swabs are separate: Whether or not the swabs are suspended in ssBPW they must be transported to the laboratory under normal transport requirements; maintaining temperatures between 0 C and 5C (not frozen), and not exceeding 10C. For primal cut swabs and bulk meat ex APC and E. coli testing: These samples suspensions are then composited and mixed with dsBPW to form the composite BPW suspension for Salmonella analysis. These BPW suspensions must be transported or stored maintaining temperatures between 0 C and 5C (not frozen), and not exceeding 10C. The original suspensions also need to be kept below 10C during APC/E. coli analysis. All Salmonella analysis must commence within 24 hours of the original sample collection. Under special circumstances this may be extended to 30 hours, with reasons recorded. 4.9.2 Method
1. BPW suspensions must not be stored or left on the bench for long periods (such as over the weekend) before incubation commences. Incubation must commence as soon as possible after the BPW suspension is prepared. Initiation of Salmonella analysis is defined as from the time the BPW suspension is placed in the incubator, the start of the BPW pre-enrichment step. 2. Incubate the pre-enrichment samples at 35+/-1C or 37+/-1C (36+/-2C is not acceptable) for 18-24 hours. Reference: Mills and Barea, (2003) AgResearch Client Report (CR) 907. After initiation of analysis, refrigeration of this pre-enrichment broth and/or the RVS selective enrichment broth is not permitted at any time. All steps in the analysis must be completed within the time temperature bounds described in this method. 3. Transfer 0.1ml of the BPW enrichment culture to 10ml of RVS selective enrichment broth pre-warmed to 42C. 4. Incubate the RVS selective enrichment broth for 24 hours +/- 2 hours at 42+/-0.2C in a water bath (preferable) or incubator certified as capable of control to this specific temperature range. The temperature is critical for maximum recovery of Salmonella. 5. Transfer a loop-full (10l) of the RVS culture to one plate of BGM agar and one plate of XLD agar selective plating media. Plates must be labelled with agar type (BGM or XLD) as the agar colour can change during incubation making the two types of media indistinguishable. Streak to obtain single, well isolated colonies.
80
Analysis
6. Incubate both plates for 18-24 hours at 35+/-1C or 37+/-1C (36+/-2C is not acceptable). 7. Examine the plates for typical Salmonella colonies (refer to MIMM 2005 or later edition, chapter 7.7, Table 1). BGM agar: Pink surrounded by bright red medium. XLD agar: Red with a black centre. H2S negative serotypes have red colonies without a black centre. If swarming bacterial growth covers the plates refer to MIMM 2005 or later edition, Chapter 7.7.5 for further instruction. 4.9.3 Confirmation of presumptive positive Salmonella
Select five or more single, well isolated, typical colonies from the selective plating media. Use biochemical tests and/or commercial kits as listed in MIMM 2005 or later edition, Chapter 7. Perform the agglutination reaction as per manufactures instructions. The five colonies may be tested sequentially and if any one of these colonies is positive the sample is deemed positive. The remaining colonies need not be tested. 4.9.3.1 Obtaining pure cultures of Salmonella for serotypical analysis Streak colony from either selective media onto MacConkey agar. Incubate for 24h at 35+/1C or 37+/-1C (36+/-2C is not acceptable). Subculture typical yellow colonies to plate count agar and incubate overnight at 35+/-1C or 37+/-1C (36+/-2C is not acceptable). Refer to MIMM 2005 or later edition, Chapter 7.7.7 and Table 4 or 7.7.7. 4.9.3.2 Final confirmation and serotyping Colonies may be confirmed in-house using Poly O and Poly H anti-sera. If either is positive, submit purified colonies to ESR. Where presumptive Salmonella has been confirmed as in 4.9.3 by latex agglutination purified colonies must be submitted directly to ESR Kenepuru Science Centre, Porirua for final confirmation and serotyping. 4.9.3.3 Reporting results Where samples are composited for Salmonella testing express the result as Salmonella detected in a composite sample or not detected in a composite sample. For poultry whole carcass rinse samples report the results as not detected/carcass.
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Analysis
4.10
Poultry Carcass Campylobacter Direct Plate Enumeration Method
This method is applicable to the NMD poultry broiler programme for carcasses selected for Campylobacter testing rinsed with 400ml of ssBPW. If chemical decontamination washes were used during processing, ensure that nonantimicrobial neutralising additives tailored to the decontamination concerned were added to the ssBPW used for rinsing. Refer section 3.1. 4.10.1 Plating Media
Selective enrichment medium: Modified charcoal cefoperazone desoxycholate agar (mCCDA). Basal medium: (commercially available) Lab-lemco powder Peptone Sodium chloride Bacteriological charcoal Casien hydrolysate Sodium desoxycholate Ferrous sulphate Sodium pyruvate Agar Distilled water pH 7.4+/-0.2 Suspend 22.75g of dehydrated mCCDA base in 500ml or distilled water, mix well and boil to dissolve the agar. Sterilise by autoclaving at 121C for 15 minutes. mCCDA Selective Supplement (commercially available) Vial quantities suitable for 500ml basal medium are: Cefoperazone 16mg Amphotericin B 5mg Complete mCCDA medium Cool autoclaved basal medium to about 50C. Aseptically add 1 vial of each selective supplement to 500ml of mCCDA base, mix thoroughly and pour into Petri plates. Plates can be stored for up to two weeks in sealed containers at 4C. Prior to use, air dry plates (do NOT use an laminar flow cabinet), either by leaving unopened on the bench overnight, or when plates are used on the day of preparation, in an incubator at 42C. 10.0g/l 10.0g/l 5.0g/l 4.0g/l 3.0g/l 1.0g/l 0.25g/l 0.25g/l 12.0g/l 1000ml
82
Analysis
4.10.2
Direct Plating
Taking 2ml of rinsate from the rinse bag or sample container apply over six labelled, appropriately dried, mCCDA plates. All of the 2ml must be dispensed onto the six plates. The exact amount per plate of the 2ml does not need to be measured as all six plates will be counted. Spread each aliquot on the agar surface with a sterile spreader. Allow the 6 plates to remain upright at room temperature to permit the sample to soak in before inverting. Plate out an additional 0.1ml of the rinsate onto each of an additional two mCCDA plates. If higher numbers are expected set up a further 10 fold dilution series in ssBPW and plate 0.1ml of each dilution (in duplicate) onto mCCDA plates as described above. Note that a suitable dilution series must be set up to minimise the occurrence of TNTC results.
DILUTION SERIES 2ml rinsate over 6 plates
Zero dilution 2ml rinsate plated over all 6 plates
0.1ml rinsate plated onto each plate
0.1ml
0.1ml
- 1 dilution 1:10 ssBPW dilution of rinsate, 0.1 plated onto each plate
0.1ml
0.1ml
- 2 dilution 1:10 ssBPW dilution of above, 0.1 plated onto each plate
0.1ml
0.1ml
Incubate in a microaerobic atmosphere (5% O2, 10% CO2, 85% N2). Ensure an appropriate number of sachets for the jar/container are used. If too many sachets are used a moist environment and condensate on plates will occur, which could influence colony growth and cause colony spreading. Too many, or too few sachets, will not create the correct gaseous mixture in the container. Incubate for 48 hours +/- 2 hours at a single temperature of 42+/- 0.5C. The microaerobic atmosphere must be maintained at all stages of incubation. 4.10.3 Presumptive Count
Select plates from dilutions of the 2ml set of 6 plates, or 0.1ml duplicate plates with not more than 150 colonies on any plate. Counts of over 150 on any plate are considered TNTC as Campylobacter colonies are so variable in size. Examine these plates for characteristic thermotolerant Campylobacter spp colonies: flat grey and moistened, variable size for 1mm 5mm in diameter, from pinpoint colonies to roundish
83
Analysis
or flattened out colonies with irregular spreading margins. Count and record the numbers of characteristic thermotolerant Campylobacter spp. colonies on this set of plates. Other organisms that may be present on mCCDA plates include creamy coloured yeast colonies, which can easily be distinguished by 10x magnification, Arcobacter and Pseudomonads.
Photograph of Campylobacter jejuni colonies and some creamy white yeast colonies on a mCCDA plate, courtesy of Tegel Foods Limited, Christchurch. 4.10.4 Confirmation
Two methods of confirmation are permitted. Laboratories may elect which they will use routinely, but must use their elected method on a regular basis to comply with their accreditation requirements. Confirmatory testing should be conducted on fresh cultures from mCCDA agar plates immediately following incubation and presumptive count. 4.10.4.1 Oxidase/Latex confirmation Select five characteristic colonies ensuring that any variation in colony size and shape is included in the selection (or all colonies if there are less than 5 colonies). a. Screen all 5 colonies for oxidase activity. Record result. b. On the first colony that is oxidase positive carry out a latex agglutination test for Campylobacter. The latex agglutination test confirms three Campylobacter species: C.
84
Analysis
jejuni, C. coli, and C. lari. If the latex test is positive assume all 5 are Campylobacter positive. If not conduct the latex test on the remaining oxidase positive colonies. c. PCR may be used as an alternative to the latex agglutination assay confirmatory test.
d. If none of the five characteristic colonies selected confirm as Campylobacter, select a further five atypical colonies and repeat steps (a-c). If there are no atypical colonies present no further confirmation is required. e. Small colonies may not contain enough cells for the oxidase test and latex test. It is recommended that small colonies are subcultured to blood agar prior to confirmation. Validation: Once a week choose a set of 5 characteristic Campylobacter colonies from mCCDA agar plates that are oxidase positive and test all 5 with latex to validate the presumption that an oxidase positive test will be latex positive. Table 28: Example of confirmatory results by Oxidase/Latex Typical colonies selected 1 2 3 4 5 Oxidase positive positive positive positive positive Latex Positive Presume remaining oxidase positive are latex positive Result per colony Campylobacter Campylobacter Campylobacter Campylobacter Campylobacter
Where the first oxidase positive colony selected of 5 records a latex negative result: Typical colonies selected 1 2 3 4 5 Oxidase negative positive positive positive negative Latex not conducted Negative Positive Positive not conducted Result per colony Negative Negative Campylobacter Campylobacter Negative
4.10.4.2 Confirmation and species identification as per MIMM sections 7.3.7 and 7.3.8 Select five characteristic colonies as per 4.10.4.1 (or all colonies if there are less than five colonies). a. For each colony carry out gram stain, motility, oxidase, hippurate hydrolysis and antibiotic sensitivity using Nalidixic acid and Cephalothin as per MIMM sections 7.3.7 and 7.3.8. b. Record as positive colonies that are gram stain negative with small gull shaped rods, exhibiting wet mount motility, are oxidase positive and where the hippurate hydrolysis result and antibiotic sensitivity confirm positive as C. jejuni, C. coli or C. lari.
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Analysis
c.
If none of the five characteristic colonies selected confirm as Campylobacter, select a further five atypical colonies and repeat steps (a-b). If there are no atypical colonies present no further confirmation is required.
Table 29: Example of confirmatory results by MIMM Colonies Selected 1 2 3 4 5 4.10.5 Reporting results MIMM 7.3.7 and 7.3.8 confirmation and species identification to C. jejuni, C. coli or C. lari C. jejuni Negative C. coli C. jejuni C. jejuni Result per colony Campylobacter Negative Campylobacter Campylobacter Campylobacter
Final count: If no thermotolerant Campylobacter spp colonies were confirmed report as not detected. Refer to section 5.4, Table 30. Where thermotolerant Campylobacter spp colonies were confirmed the counts obtained must be calculated according to section 5.2.2. Report result as per the other NMD poultry analysis (Salmonella).
December 2012 Schedule 1 National Microbiological Database Programme Results
86
and Formulas
5. Results and Formulas

5.1 Result Calculation (Manual) Red Meat Calculate the number of colony forming units (CFU) per cm or per g using the following formulae. Note: 1. That the swab suspension is not a dilution and is entered into calculations as the 10 dilution or zero dilution (non-diluted). 2. Although the whole tissue suspension is a 1:10 dilution, in order to maintain consistency in plate labelling it is entered into calculations as the 10 dilution or zero dilution. 3. The dilution series values for the calculations below are to be mathematically expressed as 10 , 10 , 10 , 10 , 10 Refer to section 4.1. 5.1.1 Aerobic plate count (spread plate) Swab samples 100cm area and 15ml suspension volume CFU / sq. cm = Count 1 + Count 2 2
2 2 0 -1 -2 -3 -4 0 0 2
1 Dilution
1 0.1ml
15ml 100cm
2
Swab samples 25cm area and 15ml suspension volume CFU / sq. cm = Count 1 + Count 2 2
2
1 Dilution
1 0.1ml
15ml 25cm
2
Swab samples 5cm area and 10ml suspension volume CFU / sq. cm = Count 1 + Count 2 2 x 1 Dilution x 1 0.1ml x 10ml 5 cm
2
Whole tissue samples 25g sample + 225ml diluent = 250ml total volume CFU / g = Count 1 + Count 2 2 x 1 Dilution x 1 0.1ml x 250ml 25g
87
and Formulas
5.1.2
Aerobic plate count (Petrifilm ) Swab samples 100cm area and 15ml suspension volume CFU / sq. cm = Count 1 + Count 2 2
2 2
TM
1 Dilution
1 1ml
15ml 100cm
2
2
1 Dilution
1 1ml
15ml 25cm
2
Swab samples 5cm area and 10ml suspension volume CFU / sq. cm = Count 1 + Count 2 2 x 1 Dilution x 1 1ml x 10ml 5cm
2
Whole tissue samples 25g sample + 225ml diluent + 250ml total volume CFU / g 5.1.3 = Count 1 + Count 2 2 x 1 Dilution x 1 1ml x 250ml 25g
Aerobic plate count (spiral plater)

2
Calculate the number colony forming units per cm or per g using the procedures outlined in the spiral platers operating instructions. The typical volume for the inoculum is 50l = 0.05ml or 100l = 0.1ml. Swab samples 100cm area and 15ml suspension volume CFU / sq. cm = Count 1 + Count 2 2
2 2
1 Dilution
1 Inoculum ml
15ml 100cm
2
2
1 Dilution
1 Inoculum ml
15ml 25cm
2
Swab samples 5cm area and 10ml suspension volume CFU / sq. cm = Count 1 + Count 2 2 x 1 Dilution x 1 Inoculum ml x 10ml 5cm
2
88
and Formulas
Whole tissue samples 25g sample + 225ml diluent = 250ml total volume CFU / g = 5.1.4 Count 1 + Count 2 2 Escherichia coli (Petrifilm ) Swab samples 100cm area and 15ml suspension volume CFU / sq. cm = Count 1 + Count 2 2
2 2 TM
1 Dilution
1 Inoculum ml
250ml 25g
1 Dilution
1 1 ml
15ml 100cm
2
2
1 Dilution
1 1 ml
15ml 25cm
2
Swab samples 5cm area and 10ml suspension volume CFU / sq. cm = Count 1 + Count 2 2 x 1 Dilution x 1 1 ml x 10ml 5cm
2
Whole tissue samples 25g sample + 225ml diluent = 25ml total volume CFU / g = 5.2 Count 1 + Count 2 2 Result Calculation (Manual) Poultry Calculate the number of colony forming units (CFU) per carcass of rinse sample using the following formulae. 5.2.1 Campylobacter enumeration x 1 Dilution x 1 1 ml x 250ml 25g
For counts obtained from 2ml spread over six (6) plates: CFU/carcass = *(number colonies confirmed as Campylobacter/n) x count characteristic Campylobacter morphology colonies (plate 1 + plate 2 + plate 3 + plate 4 + plate 5 + plate 6) x 400ml/2ml = number of Campylobacter organisms/poultry carcass sample. For duplicate plates of higher dilutions: CFU/carcass = *(number colonies confirmed as Campylobacter/n) x count characteristic Campylobacter morphology colonies (plate 1 + plate 2)/2 x 400ml/0.1ml x 1/dilution = number of Campylobacter organisms/poultry carcass sample. n = number or characteristic colonies examined, usually 5 unless there are less than 5 characteristic colonies altogether.
89
and Formulas
* Usually five/five if the first colony of five is confirmed as positive. Will be a proportion of five if the remaining colonies are required to be confirmed e.g. three/five. 5.3 Result Calculation (Automated) Laboratories and operators must use web based data entry or other that would generate NMD data for submission to NMD in an equivalent manner. The web based data entry takes raw counts and dilution data and automatically performs all calculations, provides descriptive statistics, and graphical profiles, and provides an ongoing database for storage of information. 5.3.1 Procedure
Enter premises demographics and sample descriptors such as shift, day, time of day, etc., by selecting check boxes, etc., within the spreadsheet. Enter data into web based data entry or Microsoft Excel spreadsheet. Follow instructions supplied with the software. Authorise results: a. Laboratory Reconcile data entered into the spreadsheet against the laboratory workbook, and sign off by the LAS signatory. b. Premises Where premises personnel independent of the laboratory enter authorised laboratory results into the spreadsheet, the results for submission to the NMD must be signed off by authorised, technically competent premises personnel. 5.4 Limits of Detection Not detected results are considered to be results less than the lowest limit of detection (LLD), which is Count plate 1 = 0, Count plate 2 = 1 (or vice versa) on a zero dilution pair of duplicates. Note: Because CFU/cm results (arithmetic) are transformed to logarithmic Log CFU/cm results for NMD reporting all results above the LLD, even though they may be negative logarithmic values, are to be considered as detections/counts. For example the LLD value of 0.075 CFU/ cm for a Petrifilm
2 TM 2 2 2 2
E. coli 100 cm sample site converts to 1.12 Log CFU/ cm
which represents a count, a detection, even though it is a negative log value. The values used to represent LLD and not detected results in the NMD database are listed in the table below.
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and Formulas
Table 30: Limits of detection Lowest Limit of Detection Value APC CFU/cm or /g Spreadplate 0.1ml 5cm
2 2 2 2
Not Detected E. coli Log10 value Log10 value
APC Log10 value
E. coli 2 CFU/cm or /g or /ml Not applicable
10 3 0.75 50
1.00 0.48 -0.12 1.70
Not applicable
-0.31 -0.53 -1.13 -0.31
25cm
100cm
Bulk Meat (25g) Petrifilm 5cm

2 2 2 TM
1ml 1 0.3 0.075 5 0.00 -0.52 -1.12 0.70 1 0.3 0.075 5 0.00 -0.52 -1.12 0.70 -0.31 -0.53 -1.13 -0.31
25cm
100cm
Bulk Meat (25g) Spiral plater 50l 5cm

2 2 2
20 6 1.5 100 CFU/carcass 200
1.30 0.78 0.18 2.00 Log10 value per carcass 2.30
Not applicable
Not applicable
-0.31 -0.53 -1.13 -0.31
25cm
100cm
Bulk Meat (25g) Campylobacter direct plating 2ml
Not detected Log10 value per carcass 2.00
If the number of colonies on the highest dilution plate exceeds the maximum allowable count; for example>300 APC spreadplate,>150 Petrifilm
TM
E. coli enter that result as TNTC.
Such a result is classified as a too numerous to count (TNTC) technical failure. Notwithstanding omission from the database, the result must be regarded as a detection and/or higher than expected count when assessing compliance with premises performance criteria under HACCP and RMPs. The TNTC result must still be used at premises level for assessing compliance with inhouse targets and HACCP performance criteria. For mCCDA Campylobacter where the colonies on the highest dilution plate exceed the maximum allowable count the result must be classified as TNTC, reported and will default to a greater than 3.78 Log10 CFU/carcass result of 3.79 Log10 CFU/carcass on the database. See section 6.8.1.
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and Formulas
5.5 5.5.1
Reporting of Results General requirements
The following are general requirements for process and sample descriptors to be used for reporting results: All reports to the NMD must clearly indicate the premises from which samples were taken and any required process descriptor. All reports to the NMD must have complete sample description information according to the product. Red Meat: species, APC method used, sampling week, class, shift, time, boning process, cut product description, cervine cuts (skinned or not skinned), bulk meat VL/CL grade, bulk product description, chiller identification, chilling period of time. Poultry: sample date, shift, sample time, method of drainage, carcass rinse location, 1% Tween 80 addition, rinse diluent temperature, carcass to laboratory time (when carcass is transported from the premises to be rinsed in the laboratory), time analysis initiated, cut number, average age of birds, farm reference number, shed number. Results are to be submitted to NMD in the required format, in Log10cfu/cm or /g, /ml or carcass units, or detected/not detected for pathogen presence/absence tests. If colonies are not detected express the results as the not detected assigned value e.g. -1.13, -0.53, -0.31. Results must be entered directly by web based data entry to MPI. Data provided manually will not be accepted unless agreed by the NMD Administrator. Entering a subset of results from more substantial sampling programme is allowed. However, in order to avoid biasing the NMD, the samples from which results would be entered must be selected prior to collection of the samples, as per random sample selection, not after the results had been obtained. 5.5.2 Reporting for the database to premises and use of data
2
The NMD administration must: Post on the MPI web site, summary reports to the NMD National Microbiological performance profiles on a quarterly basis. Use the RMP identifier when reporting the ranked list to the operator.
Quarters for reporting are defined as follows: 1 Quarter: January 1 March 31

st
92
and Formulas
2 Quarter: April 1 June 30 3 Quarter: July 1 September 30 4 Quarter: October 1 December 31 Provide reports as soon as possible after completion of a quarter, preferably within one (1) month. hold results from individual premises in the strictest confidence from other contributors. Individual premises data will only be available to the operator who provided it, and any LAS approved laboratory supplying data on behalf of the operator.
th rd
nd
December 2012 Schedule 1 National Microbiological Database Programme Targets
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Premises Analysis and Interpretation, Independent Verification
6. Targets Premises Analysis and Interpretation, Independent Verification

6.1 Premises Level Targets All premises shall set premises level microbiological targets (all species) and Salmonella performance standards (excluding ovine species), and present NMD data in a manner that facilitates early identification and visualization of loss of process control. 6.2 Bovine Species NMT: Escherichia coli Moving Window The NMT consist of two levels of target administered on an ongoing basis by the operator: Moving window (m) based on the percentage of samples that exceed the 80 percentile. For bovine E. coli the 80 percentile approaches not detected. Thus E. coli detected/not detected (presence/absence) is used as the limit. For m-alerts a carcass is considered a single entity. A detection of E. coli on any carcass site renders the carcass as a whole as detected. An upper limit (M) based on the 98 performance percentile (M) of E. coli numbers, as specified in the US Pathogen Reduction/HACCP Final Rule (MegaReg), which is 100cfu/cm . This is converted to logarithmic notation for NMT purposes. 100cfu/cm = 2.00 log10cfu/cm . Results from each carcass site are considered separately. The 3 carcass site results are not composited. 6.2.1 NMT (bovine species): Escherichia coli moving window (m)
2 2 2 th th th
A five (5) sample increment moving window. Addition of 5 new samples to the bottom of the window displaces 5 samples from the top. An overall window size of 15 samples (3 weeks). NMT shall be implemented for each product: carcasses (post-slaughter and dressing), primal cuts, and bulk meats. In this case, the carcass is considered a single entity. If any of the three sites of a carcass are positive, the carcass as a whole is considered positive. A premises may detect E. coli in four (4) samples of product type within the 15 sample (3 week) window, without response. Detection of E. coli in five (5) samples or more shall elicit an Alert response. An Alert response shall consist of immediate review of the process by the operator to identify and document factors, that may have compromised hygienic processing, and if appropriate, take corrective and preventative action.
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MPI VA need not be notified. However, the company response to an Alert shall be verified during routine MPI VA audits. 6.2.2 NMT (bovine species): Escherichia coli upper limit (M)
A NMT must be implemented for each bovine product: carcass post slaughter and dressing, primal cuts, and bulk meats. This NMT must be implemented for each site on the bovine carcass; thus there are three (3) sets of five (5) results for carcasses each week, five (5) rump (hindleg), five (5) flank and five (5) brisket sets of results to consider. For M-alerts results from each carcass site sample are considered separately. The 3 carcass site results are not composited. Premises may detect E. coli in numbers greater than 2.00 log10cfu/cm in one (1) carcass site, cut or bulk sample for bovine product type in any given sampling week. Detection of E. coli in numbers greater than 2.00 log10cfu/cm in two (2) carcass sites, cut or bulk sample, in a sampling week must elicit an Alert response. Table 31: Carcass example (units 2.00 log10cfu/cm ) Carcass Site Carcass 1 Carcass 2 Carcass 3 Carcass 4 Carcass 5 M-Alert Rump (hindleg) >2.00 >2.00 <2.00 <2.00 <2.00 Yes Flank <2.00 <2.00 <2.00 <2.00 <2.00 No Brisket <2.00 <2.00 <2.00 >2.00 <2.00 No
2 2 2
Bovine carcasses must be listed with M-alert status if one or more carcass sites generate a M-alert. An Alert response must include immediate review of the process by the premises to identify and document factors that may have compromised hygienic processing, and, if appropriate, take corrective and preventative action. The premises MPI VA verifier must immediately be notified, and must prior to company review of the process, assess and approve the review procedures according to the principles outlined in Section 5.10 (Processing Conformance) or IS-8 (ref. 5.1 & 5.2). 6.3 6.3.1 Bobby Calf NMT: Escherichia coli Moving Window Bobby calf product types
2
For m-alerts a carcass is considered a single entity. The arithmetic (cfu/cm ) results from the three separate carcass sites samples are composited by averaging and the expressing this arithmetic average as a single log10cfu/cm results per carcass.
2
95
For M-alerts results from each carcass site sample are considered separately. The 3 carcass site results are not composited. Primal cut and bulk meat results are taken as individual sample results in log10cfu/cm or /g units. 6.3.2 Bobby calf NMT 80 percentile m limits: Escherichia coli
th 2
Analysis of data from a 15 sample moving window, (15 carcass, 15 cuts and 15 bulk results) canvassing 3 weeks is required. The addition of the 5 latest samples to the window displaces the oldest dated 5 samples. The m-alert limit is bases on the 80 percentile E. coli values of NZ industry 25cm bobby calf data from July 2002 to December 2002. Carcasses post slaughter and dressing Primal Cuts Bulk meat
th th 2
1.23 log10cfu/cm (whole carcass average) 0.32 log10cfu/cm 2.08 log10cfu/g

2
A premises may detect E. coli >80 percentile limit in four (4) samples of a product type within a 15 (3 week) sample window without response. Detection of E. coli counts exceeding the 80 percentile in five (5) or more samples within a 15 sample (3 week) window for any product type shall elicit a m-alert response. The m-alert response requires an immediate review of the process by the company to identify and document factors that may have compromised hygienic dressing. Corrective and preventative actions shall be taken if required. MPI VA need not be notified. The company must record its responses to the m-alert, as this will be reviewed during routine MPI VA audits. 6.3.3 Bobby calf NMT M limits: Escherichia coli
th
M-alerts apply to each carcass site of five (5) carcass samples, and five (5) cut and bulk sample results (one weeks results). There are three (3) sets of five (5) results for carcasses each week, five (5) fore rump, five (5) flank and five (5) fore leg sets of results to consider. The bobby calf M-alert is based on the 98 percentile of E coli results from NZ industry 25cm bobby calf data from July 2002 to December 2002. The M value is set at not less than the FSIS M of 100 cfu/cm = 2.00 log10cfu/cm . Carcasses post slaughter and dressing Primal Cuts Bulk meat 2.11 log10cfu/cm 2.00 log10cfu/cm 2.98 log10cfu/g
2 2 2 2 2 th
Premises may detect E. coli in numbers greater than M in not greater than one (1) sample for a bobby calf product type in any given sampling week (5 samples).
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Detection of E. coli in numbers greater than M: in two (2) or more samples for any bobby calf product type in sampling week (5 samples) shall elicit an M-alert response.
Table 32: Carcass example (units log10 cfu/cm ) Carcass Site Carcass 1 Carcass 2 Carcass 3 Carcass 4 Carcass 5 M-Alert Fore Rump >2.11 <2.11 <2.11 <2.11 >2.11 Yes Flank >2.11 <2.11 >2.11 >2.11 <2.11 Yes Fore Leg >2.11 <2.11 <2.11 <2.11 <2.11 No
Bobby calf carcasses must be listed with M-alert status if one or more carcass sites generate an M-alert. An M-alert response must include immediate notification to the MPI VA and immediate review of the process by the premises to identify and document factors that may have compromised hygienic processing. When factors are identified corrective and preventative action is to be implemented. MPI VA must, prior to company review of the process, assess and approve the review of procedures according to the principles outlined in Section 5.10 (Processing Conformance) of IS-8 (ref, 5.1 & 5.2). 6.4 Ovine NMT 95 Percentile m Limits: APC The ovine species APC NMT requires analysis of APC data from a fifteen (15) sample moving window; single Y-cut site, five (5) carcasses, three (3) week period. Addition of the 5 most recent samples to the window displaces the 5 least recent samples. The ovine APC m-alert is based on the 95 percentile of APC results from NZ industry ovine fresh lamb carcass Y-cut site 5cm APC data over two years of NMD data from the beginning of January 2002 to the end of December 2003. The 95 percentile ovine APC malert value over this range of results is 4.65 log10 CFU/cm . Detection of an APC value greater than m (4.65 log10 CFU/cm ) in three (3) or more individual samples in a 15 sample (3 week) moving window is classified as an m-alert. In contrast to the NMT for other species, the moving window resets on eliciting an m-alert. Non-conforming samples (3 or more APC values greater than m) do not carry over into subsequent sampling windows, i.e. they contribute towards just one m-alert. Responses to ovine m-alerts: 1. First m-alert. Premises shall:
2 2 th 2 th th
97
review the process, identity contributing factors, propose preventative measures, and reset window.
2. Second m-alert in the moving window immediately after the first m-alert Premises must: inform MPI VA (premises level), who will review the process and the effectiveness of the implementation of the preventative measures or changes proposed in response to the first m-alert, and reset window.
3. Third m-alert in the moving window immediately after the second m-alert Premises must: inform MPI, who will carry out an independent regulatory review, carry out a full review of HACCP plans, implement process changes as required, reset window.
Actions taken, including any sanctions, will be subject to ongoing verification of compliance. 6.5 NMT: Documentation and Record Keeping All operators must document the system by which they monitor performance against the NMT, and perform any required process reviews, according to the requirements of section 5.10 (Process Conformance) of IS-8. Documentation must include: authorities; review procedures on notification of an Alert; corrective actions; preventative actions.
The verifier must validate documentation as per the requirements for implementation of IS-8, and shall assess and approve the documented process review procedures prior to implementation. All operators shall record the results of review procedures and consequent corrective and preventative actions implemented when exceeding a premises level target or on notification of an Alert.
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All documents shall be reviewed during routine MPI VA verification audits. 6.6 6.6.1 Ranked List Bovine and bobby calf
For bovine and bobby calf the percentage of APC results greater than the published national 80 percentile (Table 33) will be calculated for each premises each report quarter. Bovine and bobby calf E. coli results will be compared against the values specified by the bovine and bobby calf Escherichia coli NMT targets (sections 6.2 and 6.4) each quarter. Note that for bovine E. coli the 80 percentile is taken as not detected and the percentage detected is calculated for ranking purposes. Table 33: Bovine and bobby calf APC national 80 percentile NMT limits BOVINE Bovine Sites/Cuts/Bulk Carcass (rump hindleg) Carcass (flank) Carcass (brisket) Primal cuts Bulk meat
3 th th th
BOBBY CALF Bobby Calf Sites/Cuts/Bulk

2
Bovine APC NMT 1.77 log10 CFU/cm 2 1.69 log10 CFU/cm 2 0.95 log10 CFU/cm 2 2.01 log10 CFU/cm 3.53 log10 CFU/g
Bobby Calf APC NMT 2.03 log10 CFU/cm 2 2.85 log10 CFU/cm 2 2.13 log10 CFU/cm 2 2.82 log10 CFU/cm 4.07 log10 CFU/g
2
Carcass (fore-rump) Carcass (flank) Carcass (foreleg) Primal cuts Bulk meat
6.6.2
Ovine
Each quarter ranked lists will be published based on fresh carcass APC results for the opening Y cut site. Premises will be ranked in descending order of the percentage of results greater than the national profile ovine carcass opening Y cut APC 80 percentile all data to date value. Premises with percentage APC exceeding the 80 percentile that fall into the following category greater than 2 x the average, and prevalence statistically (Chi-square) of P <0.05
th th
will be bolded for microbiological performance information purposes only. Alerts as described in section 6.6.4 will not be elicited.
Derived from 100cm2 NMD data for the year to 31 December 2007. All bobby calf 25cm2 data from July 2002 to December 2002.
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6.6.3
Caprine, cervine, ostrich and emu and porcine

th
For cervine, caprine, ostrich and emu, and porcine the national profiles 80 percentile values for APC and E. coli all data to date will be used to compare premises data against to compile the quarterly ranked lists. 6.6.4 Ranking and alerts
th
Premises will be ranked in descending order of APC/E. coli prevalence based on the 80 percentiles/NMT for the particular species. The average prevalence will be calculated. Premises with percentage APC/E. coli exceeding the 80 percentile that fall into the following two categories shall elicit an Alert:
th
greater than 2 x the average, and prevalence statistically (Chi-square) of P <0.05 less than 0.5 x the average, and prevalence statistically (Chi-square) of P<0.05
Premises that elicit an upper or lower Alert shall be highlighted in bold on a ranked list issued by MPI on a quarterly basis. 6.6.4.1 Upper alert Notified premises must review the process to identify and document factors that resulted in a prevalence significantly higher than the industry average. The review is likely to result in modification of process control programmes programmes (pre-requisite and HACCP). 6.6.4.2 Lower alert A lower alert highlights a very low percentage prevalence. It is accepted that this result may be a reflection of consistent application of good hygienic dressing practices by the premises rather than any issue with implementation of the NMD sampling programme. Notified premises must review the NMD sampling programme; in particular sample collection, transportation (time/temperature), and laboratory procedures, to identify and correct any deficiencies that may impact on microbiological results. Where there are successive recurring lower alerts and the following points are met, the MPI verifier may decide that further reviews are no longer required: There is a history of compliance with NMD microbiological sampling requirements; and Review of both past approved laboratory audits (refer NMD Notice clause 10, sub clause (5)), and NMD verifications completed by the MPI verifier are found to have been acceptable; If after a period time outside the lower alert category the premises ranked list includes a lower alert then the review process must be reinstated.
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6.6.5
Poultry quarterly report ranked list
Poultry premises are ranked in descending order of their Campylobacter averages over the last completed quarter. Poultry premises are bolded high and low where the premises average is greater or less than +/- 0.5 log10 CFU/carcass of the average of all poultry results for that quarter. 6.7 6.7.1 Salmonella Performance Standards (SPS) Group 1 and porcine Group 2 Salmonella performance standard
Absence of Salmonella is the desired objective, and the presence of these organisms must not be overlooked, if and when detected. The contributing factors and source of Salmonella must be investigated. Upon detection of Salmonella the operator must undertake the following actions addressing product, livestock and personnel: Immediately inform the MPI VA verifier of the detection. Ensure the laboratory has submitted purified cultures of isolates detected to ESR Enteric Reference Laboratory at National Centre for Biosecurity and Infectious Disease (NCBID), Wallaceville for serotypic confirmation. Ensure records from the original product sampled are traced back to the catchment area of the stock being processed. Consult the laboratory(s) providing an animal health service in the stock catchment area, and inquire as to any increase in reported cases of animal salmonellosis. Determine if symptoms of salmonellosis have been reported by any personnel.
In addition to the above actions related to product, livestock and personnel the operator must undertake the following process responses which escalate according to the frequency of Salmonella detections in the season. Monitoring and control of this pathogen is also of interest to overseas authorities. The appropriateness of country listings of an operator will be reconsidered by MPI VA when there are ongoing detections of Salmonella at the premises. 6.7.1.1 First detection On the first detection for the season in a PSW or SSW the operator must: Review the entire process. Identify contributing factors. Implement corrective and preventative measures.
101
6.7.1.2 Second detection On the second detection in a subsequent PSW or first PSW following a first detection in a SSW the operator must: Under take a further review as per the first detection. Review the effectiveness of the corrective and preventative actions implemented in the first detection. Ensure any new or existing preventative and corrective actions are implemented. Inform the MPI VA primary verifier, who will review the process and the effectiveness of the operators actions in response to the first and second detections. 6.7.1.3 Third detection On the third detection in a subsequent third PSW or second PSW if the first detection was a SSW the operator must: Undertake a further review as per the first detection. Carry out a full review of HACCP plans. Reassess the effectiveness of corrective and preventative actions implemented in the first and second detections. Implement further corrective and preventative actions to address previous or newly identified contributing factors. The premises shall submit a Salmonella Management Plan, describing process/HACCP reviews and the measures implemented to reduce the prevalence of pathogens, to MPI VA. The MPI VA verifier must: Review operator corrective and preventative actions; and Review the operators Salmonella Management Plan and submit to MPI VA Specialist Advisor. Actions taken, including any sanctions, will be subject to ongoing verification of compliance.
6.7.2
Group 3 Salmonella performance standard
The Poultry Salmonella performance standard is based on the US Performance Standard. Operators are recommended to monitor the prevalence of Salmonella according to the US Performance Standard, PR/HACCP Salmonella Performance Standards described in Table 2 Federal Register/Volume 61, No. 144 page 38867.
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The US Standard states that Salmonella may be detected in no more than 12 of 51 consecutive poultry samples. It is recommended that on breaching the US Performance Standard operators immediately review the process and livestock Salmonella status to identify and document factors that resulted in breach of the performance standard. This may lead to modification of process control programmes (pre-requisite and HACCP). 6.8 Poultry Campylobacter Performance Target (CPT) The poultry Campylobacter performance target (CPT) consists of two regulatory limits requiring Campylobacter analysis of poultry broiler carcass rinse samples over set processing periods as follows: Standard processing premises, a total of three samples must be taken per processing day. Each of the three samples must be collected at a separate randomly selected sampling time per processing day. A processing period is five days processing equalling a total of 15 samples. Very low throughput (VLT) premises, a total of three samples on a single randomly selected day of one processing week must be randomly selected over available processing times. A processing period is one processing week equalling a total of three samples. Table 34: Table of CPT sampling requirements Sampling period A moving window of three processing periods. Standard throughput 45 samples over 15 processing days. Very low throughput (VLT) 9 samples over 3 weeks.
The moving window is defined as three processing periods. The addition of the samples of the latest processing period displaces the samples of the oldest processing period. Two CPT regulatory limits are used to determine compliance over each moving window: Number of samples with a result of greater than 6000 CFU/carcass, 3.78 log10CFU/carcass and Number of positive samples; those samples with a result of 2.30 log10CFU/carcass or higher representing Campylobacter detection.
5
A not detected Campylobacter result will be recorded as 2.00 log10CFU/carcass on the
NMD database.
103
Transitional arrangements: The new requirements in this Schedule will be implemented from Monday 7 January 2012. The first processing period under the new system will start on the first day of processing on or after that date. All premises will be reset to compliant on that date.
6.8.1
Recording of sample descriptors and default results
Each sample must have its sample descriptors recorded on the NMD database; sample time, farm reference number, shed number, cut number and average age of birds in that cut. Failure to sample If less than the required number of samples have been collected during a processing period the missed samples will each default to a greater than 3.78 log10 CFU/carcass result recorded as 3.79 Log10 CFU/carcass on the database. Technical Failures Samples which have been collected, but where a technical failure (TF) has not permitted a result the sample descriptors must be entered as proof of sampling with TF recorded in the result field. Entering the sampling descriptors of samples taken ensures that a 3.79 default result is not generated. Too numerous to count results Too numerous to count (TNTC) results must be reported. Each TNTC result will default to a greater than the 3.78 log10 CFU/carcass result; recorded as 3.79 Log10 CFU/carcass on the database.
6.8.2
CPT non-compliance
There are two classes of CPT non-compliance (1) Enumeration Failure (EF): An EF will be generated upon detection of a value greater than 6000 CFU per carcass (3.78 log10CFU/carcass) in: a. Standard throughput: premises: seven (7) or more out of 45 individual carcass samples taken from a 3 successive processing period moving window; OR b. VLT premises: two (2) or more out of 9 individual carcass samples taken from a 3 successive processing period moving window. (2) Detection Failure (DF)
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A DFwill be generated upon a result of 2.30 log10CFU/carcass or greater in: a. Standard throughput: premises: 30 or more out of 45 individual carcass samples taken from a 3 successive processing period moving window; OR b. VLT premises: Six (6) or more out of 9 individual carcass samples taken from a 3 successive processing period moving window. If the premises has an EF, a DF or both for a moving window it is counted as one non-compliant window. Responses to CPT escalate according to the number of consecutive non-compliant moving windows. To clear the non-compliance a moving window without an EF and without a DF is required. The database then resets to zero to show that the premises is compliant. Note that a noncompliance will be recorded in the database as soon as the EF or DF becomes evident (which may be before the results from all samples for that moving window have been entered). This enables corrective actions to be initiated at the earliest opportunity. 6.8.3 Required responses to CPT non-compliance
The premises NMD controller must check the NMD results at least once every processing period to determine whether or not the premises is CPT compliant. The NMD Controller must notify the operator and the MPI-assigned verifier within 24 hours of determining each non-compliant moving window. Responses escalate with each consecutive non-compliant moving window. With each noncompliant moving window the investigations, corrective actions undertaken and further actions planned to restore control must be recorded by the NMD Controller in the NMD ledger. The following responses must be undertaken. Consecutive noncompliant moving windows 1 Within 1 week of a noncompliant window being reported in NMD: the NMD Controller must: o notify the Operator and the , MPI-assigned premises verifier, and o 2 indicate that this has been done in the NMD ledger, and Response Required
the Operator must initiate corrective actions to restore control.

nd
As soon as a 2
consecutive noncompliant window is reported in NMD:
the NMD Controller must notify the Operator, and the Operator must document the investigations done, corrective actions taken to date and further actions planned to restore
105
control, and the Operator must copy this information to the MPI-assigned premises verifier, and the NMD Controller must indicate that this has been done in the NMD ledger. As soon as possible the MPI- assigned premises verifier must: review the actions and if necessary visit the premises or ask for additional information to ensure that the actions are appropriate, and indicate that this has been done in the NMD alert screen, and report any concerns to nominated MPI managers/technical people. If the MPI-assigned premises verifier or an MPI manager/technical person is not satisfied that the actions are appropriate they may notify this to an appropriate MPI Director who may require an immediate response as per 7 consecutive non-compliant windows. 3 As for non-compliance 2 with information updated on the NMD ledger by both the NMD controller and the MPI-assigned premises verifier. 4 As for non-compliance 3 with information updated on the NMD ledger by both the NMD controller and the MPI-assigned premises verifier. 5 As for non-compliance 4 with information updated with information updated on the NMD ledger by both the NMD controller and the MPIassigned premises verifier. 6 As for non-compliance 5 with information updated on the NMD ledger by both the NMD controller and the MPI-assigned premises verifier. The operator must document any product disposition options they could implement in order to minimise the risk of contaminated product reaching the consumer. The product disposition options must be provided to the MPI-assigned premises verifier.
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Responses to be undertaken continued: Non-compliant moving window 7 As for non-compliance 6 with information updated on the NMD ledger by both the NMD controller and the MPI-assigned premises verifier. The MPI-assigned premises verifier must: provide all information received to date to a nominated MPI Campylobacter expert. The MPI Campylobacter expert must: review the actions taken and the results to date then recommend to an MPI Director whether or not to initiate a Campylobacter Response Team (CRT) visit to the non-compliant premises and which experts should be in the team. The MPI Director must: sign-off the decision to initiate the CRT visit, and nominate a CRT Leader and any other relevant experts to form the team, or sign a statement declining the recommendation with associated justification. If authorised by the MPI Director, the CRT must visit the premises at the first available opportunity to: review all actions to date and recommend to the Operator other corrective actions likely to bring the premises into compliance, and where necessary, require corrective actions; where necessary, recommend the application of sanctions as per non-compliance level 8 under Section 89 of the Animal Products Act 1999 to protect the consumer. The Response Team Leader must: provide a report to the Operator summarising the visit findings, a required action plan and recommendations. copy the report to the MPI-assigned premises verifier, and the MPI Director who approved the visit. The Operator must: Expected response
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comply with the required action plan unless an alternative is agreed and signed off by the Response Team Leader and copied to the premises verifier, and
consider the recommendations.
The MPI-assigned premises verifier must: monitor the actions taken and report any concerns to the CRT Leader.
If actions are not taken as agreed then non-compliance 8 response is required. 8 MPI must apply sanctions under Section 89 of the Animal Products Act to protect the consumer. The sanctions may include, but are not limited to one or more of the following: Revisit(s) by the CRT and further required actions / recommendations by the CRT Increased verification Full-time supervision of processing Introduction of further interventions Product disposition Further sampling and research initiatives and Premises closure.
When any of the above sanctions are applied they must remain in place until revoked by an Animal Products Officer After the premises has a compliant moving window or At the direction of an MPI Director.
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6.9
Verification Requirements MPI VA/the verifier must verify that premises comply with the NMD Programme according to the Performance-Based Verification procedures to be issued by MPI. In addition, on two occasions in each processing season (Oct 1 to Sept 30), NZFSA VA/the verifier shall verify the collection, packaging and reporting of a complete NMD sample set against the NMD specification, by: observing the rotational and/or random selection of sampling day, shift, run, time and carcass/cut/carton to be sampled. observing the collection of samples, including documentation of sample descriptors and chronological information. apply a MPI carton seal to a set of NMD samples to permit a check with the receiving laboratory to verify samples are received in the correct condition/temperature and that analyses are commenced in the required time from time of sampling. observing the storage of samples prior to packaging, and subsequent packaging of samples for transport. requesting and observing the laboratory records and reports for samples transported under the MPI carton seal. observe and verify chiller exit sampling. confirm submission of current demographics to NMD Administrator. confirm commencement of appropriate Salmonella sampling programme (16 week PSW, 6 week SSW, or continuation of PSW requirements for the season). confirm reporting of Salmonella sampling programme results and completion of PSW or SSW for the season. confirm implementation of Campylobacter sampling programme where required.
December 2012 References
109
7. References
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Issued under section 167 of the Animal Products Act 1999. Date of notification in Gazette: This notice is administered in Ministry for Primary Industries.

Schedule1 NMD Dbase Programme

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Schedule 1 National Microbiological Database Programme

December 2012 Prelims

Schedule 1 National Microbiological Database Programme

Schedule 1 National Microbiological Database Programme

December 2012 Prelims

Transportation of Samples to the Laboratory for All Species .............61

Salmonella...........................................................................................77 Poultry Carcass Campylobacter Direct Plate Enumeration Method ...81

Targets Premises Analysis and Interpretation, Independent

Schedule 1 National Microbiological Database Programme

December 2012 Prelims

Schedule 1 National Microbiological Database Programme

December 2012 Prelims

Schedule 1 National Microbiological Database Programme

December 2012 Prelims

December 2012 Schedule 1 National Microbiological Database Programme Administration

1. Administration and Participation

Ostrich and emu Porcine

Every processing week Every processing week

Poultry (broiler chickens)

Every processing week

Ducks, geese and guineas Horses, mules, and other equines

Not required Not required

December 2012 Schedule 1 National Microbiological Database Programme Administration

December 2012 Schedule 1 National Microbiological Database Programme Administration

December 2012 Schedule 1 National Microbiological Database Programme NMD

Sampling Requirements Red Meat and Poultry

2. NMD Sampling Requirements - Red Meat and Poultry

December 2012 Schedule 1 National Microbiological Database Programme NMD

Sampling Requirements Red Meat and Poultry

December 2012 Schedule 1 National Microbiological Database Programme NMD

Sampling Requirements Red Meat and Poultry

December 2012 Schedule 1 National Microbiological Database Programme NMD

Sampling Requirements Red Meat and Poultry

Recommencing VLT sampling

December 2012 Schedule 1 National Microbiological Database Programme NMD

Sampling Requirements Red Meat and Poultry

Summary Table Table 2: Number of samples required per week

Domestic and non EU and non US listed 1 per week

EU and US listed standard throughput 5 per week

EU and US listed VLT

Bulk Meat Product

Post Chill Carcass Ovine Carcass

5 per week for 6 weeks 5 per week

1 per week for 6 weeks Not applicable

Bobby Calf and Caprine

Bulk Meat product

Post Chill Carcass Cervine Carcass

5 per week for 6 weeks 3 per week

1 per week for 6 weeks 1 per week

December 2012 Schedule 1 National Microbiological Database Programme NMD

Sampling Requirements Red Meat and Poultry

Whole Carcass Rinse Method

3 per day VLT: 3 per week

December 2012 Schedule 1 National Microbiological Database Programme NMD

Sampling Requirements Red Meat and Poultry

Group 1 (bovine, bobby calf, caprine, cervine) seasonal Salmonella

December 2012 Schedule 1 National Microbiological Database Programme NMD

Sampling Requirements Red Meat and Poultry

December 2012 Schedule 1 National Microbiological Database Programme NMD

Sampling Requirements Red Meat and Poultry

Group 2 (Ostrich/Emu and Porcine) - Salmonella

Sampling Location within the Process Red meat product selection