Abstract:
Bacterial vaginosis (BV) is a highly prevalent condition and the most common cause
of abnormal vaginal discharge. Despite the high prevalence of BV and associations
with other infections such as HIV-1 and obstetric and gynecological morbidity
associated with it, the etiology of BV is still not clearly understood. It has been
hypothesized that some unknown influences cause a significant decline in hydrogen
peroxide producing lactobacilli allowing the overgrowth of anaerobic bacteria. The
organisms implicated with BV have mainly been isolated by culture methods. The
present study was done in two phases where the first was a pilot study done in
California with the aim of determining the novel and predominant culturable and
unculturable microorganisms associated with BV. To achieve this, micro-array and
shot-gun sequencing techniques were used. Simpler techniques were also developed
in phase one part of the study. The second phase was done in Kenya with the aim to
determine the prevalence of BV among women infected with HIV and determine the
association of the condition with bacteriophages and the specific microorganisms
identified in phase one study. This was achieved using multiplex and simplex PCR
with organism’s specific primers. The pilot study in California identified two main
organisms that had a relationship with BV presence or absence. Oenococcus oeni
were present only in BV negative samples while Bifidobacterium species were
significantly associated with BV positive samples (p=0.0472, Fisher’s exact test).
Bacteriophage type A2 was detected in 2 of 14 (14%) BV negative samples.
Unculturable bacteria were over 90% of the total bacteria identified by sequencing.
In Kenya, BV prevalence showed a declining trend while CD4 cells count increased
with visit count. Oenococcus oeni was not detected in any sample. Bifidobacteria
xviii
were present in 39/250 (16%) of BV positive and in 19/250 (8%) BV negative
samples. Though higher in BV positive the difference was not statistically
significant (p>0.05; Chi-square). Bacteriophages Bradley types A1, A2, B1 and B2
were detected in BV positive and negative samples. Bacteriophage Bradley type B2
was detected at significantly higher rates in BV positive samples than in BV negative
samples (p<0.05; Fisher’s exact test). The use of different methods; shot-gun
sequencing versus specific PCR may not have contributed to the differences of
Oenococcus detection since, both methods are known to have high sensitivities with
detection limits of 1 pg μl−1 chromosomal DNA having been reported. Identification
of Oenococcus and unculturable bacteria confirms that shot gun sequencing is an
appropriate technique for identification of novel and unculturable organisms
associated with BV. In addition, detection of BV associated organisms by PCR
provides an effective screening method for BV. Bifidobacterium species are variably
associated with presence or absence of BV and phylogenetic analysis shows some
are close to BV related organisms while some are not. This indicates the likelihood
that there exists variant species of Bifidobacteria. This study therefore concludes that
novel and unculturable bacteria are the largest population of microorganisms in the
vagina of women with and without BV.
