GENITAL TRACT BACTERIA LINKED TO PRETERM PRE-MATURE

RUPTURE OF MEMBRANE (PPROM) AND THEIR ANTIBIOTIC

SUSCEPTIBILITY PATTERN

AMAH HC1, UCHEGBU U1, AMAH CC2, NDUDIM – DIKE J1, NWOSU DC
EGEDE N3 and ULONEME GC4
1. Department of Medical Laboratory Science, Faculty of Health
Science, Imo State University Owerri, Imo State, Nigeria
2. Orange Specialists Medical Laboratory, No 10 Oparaugo Street
Owerri, Imo State
3. Department of Gynaecology and Obstetrics, Federal Medical
Centre, Owerri, Imo State.
4. Department of Anatomy and Neurobiology, Faculty of Medicine,
Imo State University, Owerri Imo State, Nigeria.

ABSTRACT
BACKGROUND: Genital tract infection has been associated with
increased risks for preterm premature rupture of the membranes
(PPROM). This study was carried out to ascertain the role of genital tract
infection in the aetiology of PPROM as well as the antibiotic susceptibility
profile of the incriminating pathogens.
METHODS: A total of 102 pregnant women presented with PPROM
between 24 weeks and 37 weeks of gestation and 102 control cases
were enrolled while attending prenatal clinic of the federal medical
centre, Owerri. The sociodemographic characteristics of the study
population was taken and microbial flora were isolated using standard
bacteriological methods. Disc susceptibility test was performed according
to NCCLS methods.
RESULTS: Pathogens were isolated in 85 patients, giving a recovery
rate of 83.3%. The common pathogens include Escherichia coli (23.5%),
Staphylococcus aureus (20.6%) Streptococcus spp (16.7%) and Candida
albican (13.7%). Levofloxacin was the most effective antibiotics against
all the isolated pathogens while ampicillin-cloxacillin was the least active.
CONCLUSION: The timely detection and administration of ceftriaxone
erythromycin, cefuroxime or augmentin were suggested for conservative
management of PPROM.
GENITAL TRACT BACTERIA LINKED TO PREGNANT WOMAN
WITH PPROM AND THEIR ANTIBIOTICS SUSCEPTIBILITY
PATTERN

Introduction:
Preterm premature rupture of membrane (PPROM) is the rupture of the
fetal membranes before the onset of labour before 37 weeks of
gestation. It occurs in approximately 3 percent of pregnancies and leads
to one third of preterm births. It increases the risk of prematurity and
leads to a number of other perinatal and neonatal complications
including a 1 to 2 percent and risks of fetal deaths. The most serious
outcome of preterm premature rupture of membranes is often
associated with adverse maternal and infants outcomes related to
infection. Studies `indicates that one of the causes is genitals tract
infection, has been associated with increased risks for premature rupture
of the membranes (gravett et al., 2002).

Fetal membranes are made of an outer four to six layered chorion

attached to a collagen rich connective tissue and an inner single cell
amnion (kitzimiller et al., 2004). Hypothesis is that several organisms
implicated in bacterial vaginosis secretes proteases that degrade
collagen and weaken the fetal membranes leading to PPROM
inflammatory markers like IL-1, IL-6 and tumor necrosis factor during
infections resulting to increase in metalloproteinase causing collagen
degradation, decreased tensile power of membrane and eventual
rupture.

Patients with PPROM may present with leakage of vaginal fluid or

bleeding but without contractions. Diagnosis of PPROM is made through
history from the patient and by a sterile speculum vaginal examination.
Pooling of liquor in the posterior vaginal fornix or leakage of it from the
cervical ostium confirms the diagnosis. Ferning of liquor is observed on
the microscope or change of nitrazine paper to blue because of the
alkalinity of the amniotic fluid is supportive of the diagnosis of PPROM
(parry et al., 2008).
The aim of the study was to estimate the incidence rate, identification
and antibiotics susceptibility pattern of incriminating microbial pathogens
of pregnant women with PPROM, as there was no record of this research
in this locality.
In PPROM, the management involves administration of antibiotics that
reduces the risk of perinatal infection and increase the latency period
while steroids reduce perinatal morbility and mortality (Mercer et al.,
2005).

MATERIAL AND METHODS

A prospective cross-sectional study was undertaken involving pregnant
women admitted to Federal medical centre, Owerri Imo State, southeast
Nigeria, between April 1, 2013 to April 1, 2015. Federal medical centre
Owerri is a tertiary hospital that serves as a referral centre for many
cases of PPROM. The PPROM group comprised women (102) between
24 weeks and less than 37 weeks of pregnancy presenting to the pre-
natal and labour ward of the hospital with a confirmed diagnosis of
PPROM. The non-PPROM control groups were invited to enroll in the
study while attending prenatal clinic of the hospital. The women in the
PPROM and non-PPROM groups were matched for age (+ 2 years),
parity and gestational age (+ 2 weeks).

The women beyond 24 weeks of gestation diagnosed to have PPROM

with Diabetes mellitus, preclampsia, polyhydroamniosis, IUD, abruption,
placenta previa, more than one fetus, known congenital malformation,
cervical incompetence were exclude from the study. Also women were
excluded from PPROM when they had had PPROM more than 24 hours
prior to presentation, received antibiotic treatment within 7 days of
presentation, PPROM with temperature up to 380C or active vaginal
bleeding. Ethical approval was obtained from the ethical committee of
the hospital and written informed consent was obtained from all the
participants.
Endocervical swabs were taken aseptically from all recruited women. All
women were evaluated for rupture of membrane with the aid of a
detailed history taking, a physical examination and a sterile speculum
examination. A diagnosis of membrane rupture was made at the initial
examination using standard clinical assessment criteria if two of the
following three clinical signs were present: a visual pooling of fluid in the
posterior fornix, a positive nitrazine test, or microscopic evidence of
ferning (caughey et al., 2008).

The specimen were inoculated on blood, chocolate and Mac Conkey agar
plates. All plates were incubated for 48 hours aerobically with the
exception of chocolate agar that was incubated in a candle jar.
Thereafter, a wet mount and gram staining study was done for each
specimen. Emergent colonies were identified according to standard
bacteriology methods. Disc susceptibility test was performed according
to NCCLS method.

The results were analyzed using the statistical software of STATA 8.0 in
the analysis of statistical differences was used test person CHI square
and test based on an analysis of variance. The level of statistical
significance was P<0.05.

Table 1:
Sociodemographic characteristics of the study population
characteristics

Age , y PPROM Non-PPROM X2 P value

n=102 (n=102)
16-20 3 3 0.00 1.000
21-25 10 11
26-30 31 32
31-35 42 41
36-40 13 13
41-45 3 1

Parity 0.03 0.862

0 21 20
1 28 30
2 28 27
3 6 3
4 14 14
25 5 8

Gestational age, week 0.00 1.000

28-39 35 34
31-33 17 22
34-36 50 46
Level of education 0.50 0.477
No formal education 2 0
Primary education 3 1
Secondary Education 31 41

Marital Status 0.50 0.477

Single 2 0
Married 101 103

Table 1:
Microbial flora isolated from the genital treat of participating
women

Organisms PPROM NON-PPROM Pvalue

(n – 102) (N-102)
Staphylococcus aureus 21(20.6) 3(2.8) <0.001
Escherichia coli 24(23.5) 1(1.0) <0.001
Streptococcus spp 17(16.7) 2(2.0) <0.001
 Gardnerella vaginalis 8(7.8) 1(1.0) <0.001
Candida albican 14(13.7) 2(2.0) <0.001
Proteus mirabilis 3(2.9) 0 <0.150
Klebsiella pneumonia 2(2.0) 0 <0.130
Negative specimen 17(16.7) 93(91.2) <0.001

Total number of positive

Isolate 85(83.3) 9(8.8) <0.001

Table: 3

SENSITIVITY TO ANTIBIOTICS DRUGS AMONG ISOLATED

BACTERIA IN THE PPROM GROUP
Antibiotic Streptococcus Staphylococcus Escheriachia Proteus Klebsiella
spp aureus coli mirabilis Pneumonia
N=17 N=21 N=24 N=3 N=2

Gentamicin 13(76.5) 14(66.7) 20(83.3) 2(66.7) 1(50.0)

Ceftriaxone 15(88.2) 18(85.7) 22(91.7) 2(66.7) 1(50.0)
Erythromycin 12(70.6) 13(61.9) 9(37.5) 0 0
Ciprofloxacin 14(82.4) 16(76.2) 22(91.7) 2(66.7) 1(50.0)
Levofloxacin 16(94.1) 19(90.5) 23(95.8) 3(100) 2(100.0)
Cefuroxime 11(64.7) 12(57.1) 13(54.2) 1(33.3) 1(50.0)
Ampicilin- 7(76.5) 8(38.1) 6(02.5) 1(33.3) 0
cloxacillin
Amoxiclav 13(76.5) 15(71.4) 4(16.7) 1(33.3) 1(50.0)

Table: 4
SENSITIVITY TO ANTIBIOTICS DRUGS AMONG ISOLATED
BACTERIAL IN THE NON-PPROM GROUP
Antibiotic Streptococcus spp Staphylococcus aureus Escheriachia coli
N=2 N=3 N=1
Gentamicin 2(100.0) 2(66.7) 1(100.0)
Ceftriaxone 2(100.0) 2(66.7) 1(100.0)
Erythomycin 2(100.0) 3(100.0) 1(100.0)
Ciprofloxacin 2(100.0) 3(100.0) 1(100.0)
Levofloxacin 2(100.0) 3(100.0) 1(100.0)
Cefuroxime 1(50.0) 1(33.3) 0
Ampicillin- 0 0 0
cloxacilin
Amoxiclav 1(50.0) 2(66.7) 0

3. RESULTS
In total, 204 women participated in the study. The demographic
characteristics of the participants are shown in Table 1. Both groups
were apparently homogenous (P>0.05). The mean age of the women in
the PPROM group was 30.2 + 3.2 years and the mean age of the non-
PPROM group was 30.4 + 2.3 years. Most women in the study
population had a parity of 0 – 2. The mean gestation age of the fetuses
in the PPROM and non-PPROM groups were 32.5 + 1.5 weeks and 33.1
+ 1.8 weeks respectively.

Bacteria were recovered from 85(83.3%) of the specimens taken from

women with PPROM (P<0.001) (table 2). Escherichia coli were the
bacteria isolated most frequently in the PPROM group; while the
common next organism isolated was Staphylococcus aureus (30.6%).
These bacteria identified were frequently significant in the PPROM than
the non-PPROM group (Table 2).

In the PPROM, Levofloxacin was the most effective antibiotic: 63

(94.0%) of the bacterial isolates cultured from the 67 bacterial isolates,
isolated were sensitive to levofloxacin (Table 3). Ampicillin-cloxacillin
was the least effective of the antibiotics tested: only 22(32.8%) of the
bacterial isolates were sensitive to the antibiotic (Table 3). The
sensitivity of isolates from the non-PPROM group is shown in table 4.

DISCUSSION:
Despite many years of research the etiology of preterm delivery remains
to be unraveled. Several studies contain the complexity of reasons both
from the mothers side or the blastocyst and depending on the external
environment (Hiller et al, 2005).

In studies by Das CR et al., (1996) showed that infection was 2-3 times
more common in patients with rupture of membranes before 37 weeks
of gestation than when foetal membranes ruptured at term. In this
study, we found a significant correlation between PPROM and
pathogenic micro-organisms. The following bacterial pathogens were
significantly isolated: Staphylococcus aureus, Escherichia coli,
Streptococcus spp, proteus mirabilis, klebsiella pneumonia and
Gardnerella vaginalis.
According to sherman et al., 2007 the colonization of Streptococcus
agalactiae or Enterobactecteriacae alone do not induce preterm labor
and PPROM only lowering the percentage of lactobacilli or co-infection
with other micro-organisms of high virulence (such as klebsiella spp and
pseudomonas aeruginosa) increase the risk of premature contractions.
In our research has confirmed that infection with Klebscella spp, etiology
was positively correlated with PPROM, as klebsiella spp was not isolated
from the non-PPROM group. According to Krychowska – cwkla et al.,
(2011), ureaplasm urealyticum, Mycoplasma hominis, Bacteriodes,
Chlamydia trachomatis and Neisseria Gonorrrhoea are implicated in
premature spontaneous birth. These bacterial agents may contribute to
the negative specimens with no bacterial growth, as they are not
cultivated by the conventional methods used in our study.
The rationale for prophylactic treatment of PPROM with antibiotics is
that infection appears to be both a cause and consequence of PPROM
and can lead to premature delivery (Lewis et al., 1995). The
administration of antibiotics in cases of PPROM forms part of the current
standard care due to strong evidence that antibiotics prolong latency
period in short-term neonatal morbidity by eradication of intrauterine
infection and decreasing inflammatory responses (Kenyons et al., 2003).
Ampicilin-cloxacillin is one of the antibiotic habitually implicated in self-
medication in Nigeria, which might explain why only 22(32.8%) of the
isolates were sensitive to ampicillin-cloxacillin.

Due to the teratogenic and fetotoxic concerns of quinolone, Berkovitch

et al (2004) the use of effective ciprofloxacin and Levofloxacin should be
discouraged. In conclusion, based on the outcome of the antibiotic
sensitivity test, the use of ceftriaxone, Erythomycin and Cefuroxime and
augmentin suggested for the conservative management of PPROM.

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