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Bacterial vaginosis: an update

on diagnosis and treatment
Expert Rev. Anti Infect. Ther. 7(9), 1109–1124 (2009)

Hans Verstraelen† and
Rita Verhelst
†
Author for correspondence
Department of Obstetrics &
Gynaecology, Ghent University
Hospital, De Pintelaan 185,
B-9000 Ghent, Belgium
Tel.: +32 9332 3796
Fax: +32 9332 3831
hans.verstraelen@ugent.be
Bacterial vaginosis is the most common cause of vaginal complaints. Bacterial vaginosis is further
associated with a sizeable burden of infectious complications. Diagnosis relies on standardized
clinical criteria or on scoring bacterial cell morphotypes on a Gram-stained vaginal smear. A few
point-of-care tests have not gained footage in clinical practice, but molecular diagnosis is now
pending. Treatment remains cumbersome and clinicians are currently rather poorly armed to
treat bacterial vaginosis in the long run. As an adjuvant to standard treatment with antibiotics,
alternative treatments with antiseptics and disinfectants, vaginal-acidifying and -buffering
agents, and probiotics hold some promise for long-term prevention.
Keywords : bacterial vaginosis • diagnosis • therapy • vaginal microflora • vaginitis

Bacterial vaginosis (BV) is a condition character-
ized by the partial loss of the indigenous vaginal
lactobacilli on the one hand, and massive poly-
microbial anaerobic overgrowth of the vaginal
mucosa on the other. The etiopathogenesis of
this infestation remains largely elusive, although
a limited number of risk factors have consistently
been associated with BV, including black ethnic-
ity, sexual intercourse and vaginal douching.
Although BV often remains asymptomatic, it still
is, along with vulvovaginal candidiasis, the most
common cause of vaginitis, and hence among the
most common reason for women to seek medical
help. However, in recent years BV has emerged as
a global issue of concern due to the vast infectious
disease burden that results from the diminished
colonization resistance with BV. Infections related
to BV may broadly be categorized as opportunis-
tic infections with BV-associated bacteria and as
infections due to sexually transmitted agents. In
the first category, ascending genital tract infection
in the setting of BV implicates postabortion and
postpartum endometritis, pelvic inflammatory
disease (PID) and, in pregnancy, late fetal loss and
spontaneous preterm birth. In the second cate-
gory, where it has rather recently been recognized
as a state of diminished colonization resistance,
BV renders women particularly vulnerable to the
acquisition of Trichomonas vaginalis, Neisseria
gonorrhoeae, Chlamydia trachomatis, HSV‑2 and
HIV‑1. Moreover, it has been documented that
BV propagates viral replication and vaginal shed-
ding of the HIV‑1 and HSV‑2 viruses, thereby
further enhancing the spread of these sexually
transmitted diseases. Tackling the HIV burden
through eradicating BV and restoring the vaginal
microflora is therefore now considered one of the
most promising answers to the HIV epidemic.
Diagnosis of BV
Signs & symptoms
Bacterial vaginosis is confined to an asymptom-
atic state in at least half of the cases. Symptomatic
BV, on the other hand, is most typically accom-
panied by foul-smelling, profuse vaginal dis-
charge in the absence of any appreciable signs of
inflammation. Symptoms of vaginitis are overall,
however, rather nonspecific and therefore clini-
cal diagnosis will at best give an indication of the
presence of vaginitis, and warrants microscopic
investigation to determine an infectious cause.
In case of BV, such targeted diagnosis can basi-
cally be made according to the so-called clini-
cal criteria or according to what is designated as
the microbiological criteria, although both gold-
standard approaches actually encompass vaginal
fluid microscopy.
Clinical diagnosis
In 1983, Amsel et al. launched clinical diagnostic
criteria [1] for BV, which have proved particularly
useful in clinical practice and hence are still in
use today. The clinical diagnosis of BV is made
if three of the four following signs are present:
• An adherent and homogenous grayish-white
vaginal discharge;
• A vaginal pH exceeding a value of 4.5;

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 Review Verstraelen & Verhelst
• The presence of so-called clue cells – vaginal epithelial cells
with such a heavy coating of bacteria that the peripheral borders
are obscured – on saline wet mount;
• A fishy or amine odor after the addition of a 10% potassium
hydroxide solution (positive whiff or sniff test).
The Amsel’s criteria, among others, however, have been criticized
because two of the four criteria, in particular the appearance of
the discharge and the appraisal of the odor, are rather subjective
and hence may lead to misdiagnosis. By contrast, a pH greater
than 4.5 is considered the most sensitive criterion, whereas the
presence of clue cells has been considered the single most specific
predictor of BV [2,3] . However, even today, diagnosis through
Amsel’s criteria still remains the best option for in-office testing
for BV by the clinician.
Gram-stain-based diagnosis
As an alternative, Gram-stain-based microbiological diagnosis of
BV has been proposed. To perform a Gram stain, vaginal fluid or
discharge is collected on a glass slide, allowed to air-dry, stained
in the laboratory and examined under an oil immersion for the
presence of specific bacteria. This diagnostic method has several
advantages, including a permanent record, a high frequency of
interpretable results, low cost, and ease of transport and stor-
age [2] . In addition, Gram-stained vaginal smears can be evaluated
repeatedly or independently by more than one assessor, thereby
increasing diagnostic reliability.
The most widely performed method is the Gram-stain-
based scoring system developed by Nugent et al. [3] . Basically,
the Nugent scoring system accounts for three bacterial cell
morphotypes – that is, Lactobacillus morphotypes (large
Gram-positive rods), Gardnerella and Bacteroides morphot-
ypes (small Gram-variable or Gram-negative rods) and curved
Gram-variable rods (typically Mobiluncus spp.) Based on the
abundance of each of these morphotypes per oil immersion
field, each of the three morphotypes is then quantitated from
0 to 4+, and the summary score is obtained by adding up the
morphotypes-specific scores equates the overall Nugent score:  vaginal pH by inserting one finger into the vagina. The glove has
(Mobiluncus score)
Gardnerella score + [4 - (Lactobacillus score)] + [
2
The criterion for BV is a score of 7 or higher. A score of 4–6
corresponds to so-called intermediate vaginal microflora, and a
score of 0–3 is considered to represent normal vaginal microflora.
Overall, the Nugent scoring system for Gram-stained vaginal
smears has shown high intracenter and intercenter reliability,
as well as high intraobserver and interobserver reproducibil-
ity. In addition, validity studies have repeatedly documented a
high degree of accuracy. However, practitioners are not usually
familiar with performing in-office Gram-stain-based diagnosis,
and hence point-of-care testing will usually rely on clinical cri-
teria according to Amsel, possibly followed by laboratory-based
Gram-stain confirmation. For research purposes, however a
defined preference for Gram-stain-based diagnosis of BV seems
to exist.
1110
Some concern remains, however, over the performance of the
Nugent scoring system. First, it has been acknowledged that
Nugent’s criteria are widely applied in the absence of standard-
ized pre-analytical and analytical conditions, which may impinge
on Gram-stain diagnosis. Forsum et al. emphasized the need for
quality specifications in this respect [4] , as different sampling
devices and procedures, different ways of spreading the vaginal
specimen on the glass slide leading to differences in homogeneity
of the sample and in the thickness of the smear, different fixation
methods and time, and differences in the area of the high-power
oil immersion field at magnification ×1000 all may affect Gram-
stain interpretation [4–6] . Second, Gram-stain interpretation with
regard to Nugent’s criteria per se is also a matter of concern, since
no definite criteria have been proposed to distinguish between the
three basic morphotypes handled in the Nugent scoring system.
Albeit in two international workshops the interobserver reliability
in scoring Gram-stained vaginal smears was generally good [4,5] ,
specific problems occurred; in particular there seems to be dis-
agreement between researchers as to which morphotypes should
be considered Gram-positive rods and hence which morphotypes
are scored as lactobacilli, the differentiation between cocci and
small rods varies among investigators and, importantly, small
bacteria morphotypes such as Gardnerella (and Prevotella) may
vary in size from round to more elongated – as they may vary
in Gram-staining aspect – which may lead to confusion in their
categorization [4,5] .
Commercial point-of-care tests for the diagnosis of BV
Several rapid point-of-care diagnostic tests for BV have been
developed and commercialized, although none of these tests
are being widely used. Here we provide a brief, nonexhaustive
overview of tests that may be of use in clinical practice or in
­epidemiological settings.
Self-test pH glove
A glove with an integrated pH indicator paper was developed in
Germany in the early 1990s by which women can monitor their
been broadly applied in population-based screening programs in
] Germany for the prevention of preterm birth, by which women
were instructed to consult their physician if the pH was 4.7 or
more [7] . As mentioned earlier, an increased vaginal pH is a very
sensitive, although nonspecific, indicator of BV, and hence still
warrants further assessment as outlined previously.
Testing for the presence of trimethylamine (electronic sensor
array or electronic nose)
The volatile organic amino acids responsible for the characteristic
odor in BV have been used as a target for BV diagnosis. This was
initially accomplished through gas–liquid chromatography [8] – a
laborious procedure. This knowledge was then translated to the
development of an electronic sensor array by which vaginal fluid
is passed over an application-specific array of conducting polymer
sensors, each of which has specific interactions with different
volatile organic species based upon their size, shape and functional
Expert Rev. Anti Infect. Ther. 7(9), (2009)

group [9] . In a large diagnostic study, Hay et al. obtained a sensi-
tivity of 81.45% and specificity of 76.1% with the electronic nose
compared with Amsel’s criteria, and a sensitivity and specificity of
82.9 and 77.3% compared with Gram-stain diagnosis [10] .
Testing for the presence of trimethylamine in combination with
vaginal pH assessment
This point-of-care test, the FemExam® (CooperSurgical, Inc, CT,
USA) test card, is based on determining pH and trimethylamine
levels in vaginal fluid for the diagnosis of BV; however, the test
did not compare favorably with Amsel’s criteria or with Nugent
criteria in published studies [11–13] .
Testing for sialidase activity
The BVBlue ® (Gryphus Diagnostics, AL, USA) system is a
chromo­genic diagnostic test based on the presence of elevated
sialidase enzyme activity in vaginal fluid samples. This point-of-
care test has consistently shown good sensitivity, specificity, and
positive and negative predictive values when weighted against
both Amsel’s criteria and Nugent criteria [14–16] .
Testing for proline aminopeptidase activity
Promising results in detecting proline aminopeptidase activity (Pip
Activity TestCard™, Quidel Corp., CA, USA and CooperSurgical
CT, USA) of anaerobes, especially Gardnerella vaginalis, in vagi-
nal discharge for the diagnosis of BV were reported by Schoon­
maker et al. in 1991 [17] . Two ­subsequent studies obtained a high
­d iagnostic accuracy with the test for proline ­a minopeptidase
­activity [18,19] .
DNA probe for G. vaginalis rRNA
The Affirm™ VP III (BD Diagnostic Systems, NJ, USA) G. vagi-
nalis DNA hybridization assay is a DNA hybridization test that is
positive only for concentrations of G. vaginalis in excess of 2 × 105
bacterial cells per ml of vaginal fluid [20] and should therefore be
positive most often in women with BV and rarely in women with
normal vaginal microflora. The Affirm system can also detect
the presence of Candida spp. and Trichomonas vaginalis in the
same specimen, making it a quite attractive tool for evaluating
women with vaginal discharge. The procedure requires between
30 and 45 min to complete. The test can be performed in an
office setting but this is not very efficient and it is usually better
done in a laboratory setting. Nonetheless, the rapid turnaround
time permits return of results to clinicians within 24 h. Promising
results have been reported with this approach in two populations,
one in pregnant women and one in otherwise healthy women of
childbearing age [12,20] . Both studies concluded that the probe
can be used as a supplement to the Amsel and Nugent methods.
Molecular diagnosis of BV
Following the recent surge of molecular analysis-based studies of
the vaginal microflora, several prospects for the molecular diag-
nosis of BV have emerged. This approach might overcome at least
some of the aforementioned problems with regard to the reliability
and reproducibility of Gram-stain diagnosis.
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Bacterial vaginosis Review
Until the first study in 2002 that used broad-range PCR to
characterize the vaginal microflora [21] , essentially all our knowl-
edge of the vaginal microflora has been obtained from isolat-
ing organisms by culture and subsequently identifying them by
pheno­t ypic means. This approach has long been the mainstay for
studies on the human vaginal ecosystem. However, cultivation
of microbes as a means to characterize microbial communities in
a natural ecosystem has major shortcomings, as it is recognized
that many microbes in different ecosystems cannot be cultivated
using standard culture techniques [22] .
Since the beginning of this decade, several research groups have
applied PCR-based culture-independent methods to study the
bacterial microflora of the human vagina and showed – by means
of cloning of the 16S rRNA gene or the chaperonin-60 gene, by
species-specific PCR (sPCR), by denaturing gradient gel electro-
phoresis of the 16S rRNA gene, by terminal restriction fragment
length polymorphism of the 16S rRNA gene or by FISH – that
previously unrecognized difficult-to-culture organisms are part
of the vaginal microflora [23,24] . These broad-range bacterial PCR
studies identified key organisms related to BV and opened the
door for the detection of these bacteria by either conventional
sPCR or quantitative real-time (QRT)-PCR.
In the past, several sPCR assays have been developed for sensi-
tive detection of vaginal bacteria that are either characteristic for
a normal microflora or are BV related. Several assays for the detec-
tion of G. vaginalis, targeting, respectively, the 16S–23S rRNA
spacer region [25–27] or the 16S rRNA gene [28] were designed.
In addition, the fastidious anaerobic Mobiluncus spp. [25,29] and
Mycoplasma spp. [27] were interesting species for specific detection,
especially the latter, as this species is not detected by Gram stain-
ing. Recently, several research groups applied sPCR for Atopobium
vaginae, since this species was found to be more specific for the
detection of BV than G. vaginalis [30–33] .
Although these PCR assays proved to be highly sensitive for
BV detection, none of these individual assays gained a footing
in the diagnosis of BV. Attempts were made to combine several
sPCRs in order to obtain more predictive assays. Obata-Yasuoka
et al. proposed, as a diagnostic test for BV, a multiplex PCR assay
using primers specific to 16S rRNA genes of Mobiluncus mulieris
and Mobiluncus curtisii, the nanH gene of Bacteroides fragilis,
and an internal spacer region of the rDNA of G. vaginalis [25] .
The diagnostic sensitivity, specificity, positive predictive value
and negative predictive value of multiplex PCR in comparison
with Gram-stain examination were 78.4, 95.6, 82.9 and 94.2%,
respectively. Verhelst et al. used a combination of the sPCRs for
G. vaginalis and A. vaginae for diagnosing BV [34] . The simul­
taneous presence of A. vaginae and G. vaginalis in a vaginal swab
specimen as detected by species-specific PCR had an accuracy
of 90% (95%  CI: 86–92%), a sensitivity of 82% (95%  CI:
59–94%) and a specificity of 90% (95% CI: 87–92%) in assess-
ing BV. This preliminary predictive model further showed a nega-
tive predictive value as high as 99% (95% CI: 98–100%); but
a positive predictive value of merely 26% (95% CI: 17–39%)
in assessing BV, the latter resulting from the poor discrimina-
tive value of qualitative differentiation between normal and BV
1111
 Review Verstraelen & Verhelst
microflora, considering the common presence of G. vaginalis in
low numbers with normal microflora. The most thorough sPCR
approach was reported by Fredricks et al., who targeted 17 dif-
ferent vaginal bacteria that were previously found to be either
highly specific for BV or novel [32,35,36] . The study of 264 vagi-
nal samples obtained from 81 subjects with BV and 183 subjects
without BV in two clinics in Seattle (WA, USA), revealed that
while Lactobacillus crispatus was inversely associated with BV,
three novel bacteria from the Clostridiales order (BVAB 1–3),
as well as Atopobium, an Eggerthella-like bacterium, Sneathia/
Leptotrichia, Megasphaera types 1 and 2, and a bacterium from
the TM7 division were highly specific for BV. Moreover, detecting
the combination of one of the Clostridiales bacteria (BVAB2) or
Megasphaera type 1 produced the highest sensitivity and specific-
ity for sPCR diagnosis of BV so far reported (sensitivity 99% and
specificity 89%).
Although conventional sPCR merely allows assessing the pres-
ence or absence of a particular target, it revealed the presence of
noncultivable species, some of which allowed the diagnosis of BV
with much higher specificity than G. vaginalis detection. Of note
is that sPCR studies reported the traditional BV-associated spe-
cies Mobiluncus [25,29] and Mycoplasma [27,37,38] at approximately
the same incidence as culture-based studies, while broad-range
bacterial PCR studies failed to detect these species. Most impor-
tantly, sPCR studies that applied combinations of key organisms
suggested that PCR-based amplification might potentially be used
for the molecular diagnosis of BV and resulted in the application
of QRT-PCR, allowing us to also take into account the inoculum
of bacteria.
The first efforts to investigate the applicability of QRT-PCR
for the diagnosis of BV came from Zarrifard et al. who deter-
mined the feasibility of using this technique to detect and quan-
tify Lactobacillus spp., G. vaginalis and Mycoplasma hominis in
the genital tract of 21 women using stored vaginal samples [27] .
The results show that samples from women with BV who were
clinically diagnosed had significantly higher numbers of G. vagi-
nalis, but significantly lower numbers of lactobacilli. Moreover,
there was a noticeable pattern where low numbers of lactobacilli
were found in samples with high numbers of G. vaginalis and,
conversely, low numbers of G.  vaginalis organisms were seen
in samples that had high numbers of lactobacilli. In a follow-
ing study [39] , this research group compared Nugent score with
Amsel criteria and quantitative bacterial PCR for diagnosing BV
in 203 cervicovaginal lavage samples from women with Nugent
scores of 7–10 (BV group) and 203 samples from women with
BV Nugent scores of 0–3 (no-BV group). Although there was
significant overlap in the log10 Lactobacillus counts between the
two groups, their data demonstrate that quantitative bacterial
PCR for G. vaginalis, M. hominis and lactobacilli significantly
correlates with the Nugent Gram-stain method to diagnose BV.
In addition, they were able to identify cut-off points for G. vagi-
nalis and M. hominis that differentiated the BV group from the
no-BV group. Utilizing all three log10 bacterial counts (G. vagi-
nalis, M. hominis and lactobacilli), the sensitivity and specificity
of the PCR assay were 83 and 78%, respectively, in comparison
1112
with Nugent score, while the sensitivity and specificity of the
Amsel criteria compared with the Nugent score were 37 and
99%, respectively.
Several research groups also applied an A. vaginae QRT-PCR
assay to study the role of this species in BV [40–43] . Bradshaw et al.
and Ferris et al. studied the presence of A. vaginae before and
after treatment with metronidazole [40,41] . Bradshaw found higher
recurrence rates in women in whom both A. vaginae and G. vagi-
nalis were detected pretreatment, and Ferris’ QRT-PCR assay
indicated that high pretreatment A. vaginae concentrations are
predictive of adverse treatment outcomes for BV patients. Tabrizi
et al. studied the occurrence of G. vaginalis and A. vaginae in
44 women who were virgins and found that 45% had G. vaginalis
and 7% had A. vaginae [42] . De Backer et al. confirmed by QRT-
PCR that the presence of A. vaginae seems to be a diagnostically
more valuable marker for BV than the presence of G. vaginalis [44] .
Zozaya-Hinchliffe et al. assessed the prevalence and abundance of
uncultivated Megasphaera-like bacteria in the vaginal niche using
QRT-PCR [45] . Megasphaera type 1 concentrations were higher in
subjects with BV (up to five orders of magnitude) than subjects
without BV, and this bacterium was significantly associated with
BV (p = 0.0072), as was Megasphaera type 2 (p = 0.0366).
Recently, a few research groups applied broader sets of QRT-
PCR assays for the diagnosis of BV. Menard et al. used a series of
species-specific primers for QRT-PCR targeting Lactobacillus spe-
cies, G. vaginalis, M curtisii, Mobiluncus mulieris, Ureaplasma urea-
lyticum, A. vaginae, Candida albicans and M. hominis [46] . They
were able to document that the presence of A. vaginae at a level of
108 copies/ml or more and of G. vaginalis at a level of 109 copies/ml
or more was highly predictive for the diagnosis of BV, with a sen-
sitivity of 95%, a specificity of 99%, a negative predictive value
of 99% and a positive predictive value of 95% [46] . Interestingly,
Menard et al. also showed that, according to this criterion, 57%
of the samples rated as intermediate microflora on Gram stain
actually involved BV [46] . Fredricks et al. used eight QRT-PCR
assays targeting both easily cultivated vaginal bacteria (G. vagi-
nalis and L. crispatus) and fastidious bacteria (BVAB1, BVAB2,
BVAB3, Leptotrichia/Sneathia, Atopobium and Megasphaera-like
species) to determine how concentrations of vaginal bacteria
change in women with BV by comparing women who were cured
with women with persistent BV 1 month following vaginal met-
ronidazole treatment [47] . Successful antibiotic therapy resulted
in 3- to 4-log reductions in median bacterial loads of BVAB1,
BVAB2 and BVAB3, a Megasphaera-like bacterium, Atopobium
species, Leptotrichia/Sneathia species and G. vaginalis. By contrast,
median post-­treatment bacterial levels did not change significantly
in subjects with persistent BV except for a decline in BVAB3 levels.
Fredricks et al. therefore concluded that the presence or absence of
BV is reflected by vaginal concentrations of BV-associated bacteria
such as BVAB1, BVAB2, Leptotrichia/Sneathia species, Atopobium
species, G. vaginalis and a Megasphaera-like bacterium, and hence
that these bacteria may be suitable markers of disease and treat-
ment response [47] . In conclusion, there are now some prospects
for the molecular diagnosis of BV. The study by Menard et al.
that assessed the utility of quantitative loads of G. vaginalis and
Expert Rev. Anti Infect. Ther. 7(9), (2009)

A. vaginae as a diagnostic tool for BV clearly seems to provide
a sound basis, thereby possibly overcoming the aforementioned
problems with regard to the reliability and reproducibility with
Gram-stain diagnosis [46] . Indeed, it might be hypothesized that
BV is characterized by a pathological core of G. vaginalis and A.
vaginae in a biofilm configuration with a unique virulence profile
regardless of the variability of associated species [48] . However, as
recently reiterated by Kalra et al., distinct BV profiles consisting
of differing species might predispose to different health outcomes
[49] . As a consequence, the broader set of QRT-PCR assays applied
by Menard et al. or by Fredricks et al. will be required for subtype-
specific BV profiles [46,47] . In addition, the molecular diagnosis
of the normal, lactobacilli-dominated vaginal microflora may be
particularly interesting in the identification of women at risk of
developing BV and associated adverse health outcomes [44,50,51] .
These ambiguities can only be resolved by large, prospective
cohort studies in which microbiological profiles obtained through
molecular techniques are correlated with known reproductive and
infectious health outcomes.
Treatment of BV
Despite numerous therapeutic research efforts, treatment of BV
remains cumbersome and clinicians are currently rather poorly
armed to treat BV properly in the long run. Moreover, appraisal
of the available evidence on treatment options for BV is hindered
by methodological concerns.
Rationale for treatment of BV
Treatment of BV is primarily targeted at resolving or alleviating
the presenting symptoms, most commonly profuse, foul-smelling
vaginal discharge. In the absence of treatment BV may resolve
spontaneously; however, it often recurs over an extended period
of time. As recurrence of BV is common, treatment of BV is
ideally also targeted at prophylaxis of BV recurrences. On a sec-
ondary level, BV treatment aims to prevent infectious complica-
tions associated with BV, and hence also involves treatment of
asymptomatic BV.
Definition of cure
Assessment of therapeutic efficacy of a given treatment depends
on the clinical context in which such an evaluation is to be made.
In clinical practice, cure of BV will therefore typically be consid-
ered a fact if a patient reports the resolution of the symptoms for
which therapy was instigated on the occasion of a control visit.
Contrary to this symptomatic approach, a more objective manner
of establishing cure is to make a reassessment by use of the Amsel’s
criteria [1] or Nugent’s criteria [3] .
This approach has not been properly defined, however, and
hence some ambiguity prevails. If cure is evaluated through
Amsel’s criteria, cure could be accepted as long as no more than
two signs defined by Amsel et al. [1] persist following therapy, since
it takes at least three out of four signs to build the diagnosis. Many
researchers agree however that so-called clinical cure is reflected
by the resolution of all Amsel’s signs. Similarly, so-called micro­
biological cure as assessed through Nugent’s criteria [3] has not
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Bacterial vaginosis Review
been unequivocally defined: does microbiological cure indicate
that the condition of BV has resolved, that is, a Nugent score
below 7, or does it entail the reconversion to a normal vaginal
microflora, that is, a Nugent score of less than 4? Both defini-
tions have been applied throughout the literature. The distinc-
tion between both situations is particularly important, as the
former definition, reconversion to a vaginal microflora with a
Nugent score of less than 7, may implicate intermediate microflora
(Nugent score 4–6) as the normalized state, while intermediate
microflora are significantly more likely to shift to BV than normal
microflora defined by a Nugent score of less than 3.
Another as yet undefined aspect with regard to the assessment
of cure relates to the time frame relative to therapy. Many stud-
ies have reported cure rates through the assessment of patients
after treatment cessation or at 1-week follow-up. Only a small
number of studies typically report on cure as defined through
Amsel’s or Nugent’s criteria at 3–4 weeks of follow-up. As more
than 50% of woman may have persistence or recurrence within
2 months of antibiotic treatment [52] , assessment of cure at such
extended follow-up might be an even better indicator of therapeu-
tic efficacy, although very few studies have actually adopted this
approach. At present, the sole guideline on the assessment of cure
following treatment of BV comes from a draft guidance docu-
ment that was launched by the US FDA [53] . According to this
guidance, cure is defined as the absence of all four Amsel’s signs
and a Nugent score of less than 4 at a test-of-cure visit 21–30 days
after the first day of treatment. This stringent approach is now
increasingly gaining attention as a standard to define cure of BV
by researchers in this field.
Treatment modalities
Antibiotics have served as the mainstay of BV treatment over the
past six decades. Several alternative approaches also deserve atten-
tion, however, and can grossly be categorized as treatment with
antiseptics and disinfectants, with vaginal acidifying or buffering
agents, and with probiotics, which have been used as a standalone
therapy or in conjunction with antibiotics.
Standard treatment with antibiotics
Soon after the advent of antibiotics and as early as in the 1950s,
attempts were made to identify the optimal antibiotic treatment
for BV. A number of antibiotics passed the revue in the following
decades, although many of the alleged early therapeutic successes
in this era could not be repeated later [54] .
Eventually, Pheifer et al. reported, in their 1978 New England
Journal of Medicine paper, on an oral regimen of 500 mg met-
ronidazole twice daily for 7 days that eradicated Haemophilus
vaginalis in all 81 patients treated, with resolution of symptoms
and signs in 80 patients [54] . A total of 10 years later, Greaves
et al. found an oral regimen of 300 mg clindamycin twice daily
to be equally safe and effective as the metronidazole regimen [55] .
Since then, metronidazole and clindamycin have been the drugs
of choice in the treatment of BV, as also recommended by the
most recent sexually transmitted disease treatment guidelines by
the US CDC (Box 1) [56] .
1113
 Review Verstraelen & Verhelst
So how strong is the evidence in support of the CDC guide-
lines and, hence, how effective are the recommended regimens?
There are over 100 published clinical trials on treatment of BV
with antibiotics, although this bulky material has never been
subjected to a meta-ana­lysis, presumably due to the notorious
methodological heterogeneity across studies as indicated ear-
lier. The only evidence-based approach comes from a narrative
systematic review that is regularly updated within the frame-
work of the BMJ Clinical Evidence series [52] , which at present
provides a systematic appraisal of the evidence of interventions
for BV published before July 2006, of which the main findings
on strength of evidence are summarized in Table 1. From this
evidence-based review, it was concluded that all CDC recom-
mended regimens are likely to beneficial in the treatment of BV in
terms of short-term benefit, with no obvious differences between
the different recommended regimens, albeit the overall quality
of evidence is actually rather low [52] . In addition, few studies
have assessed long-term cure rates, but it is generally accepted
that recurrence of BV even after a proper treatment course is
high, with some 30–50% of women experiencing a BV relapse
within 2–3 months.
Therefore, even with the drugs-of-choice, it appears as if anti­
biotic treatment largely fails to prevent the relapse of BV and
hence clinicians are currently rather poorly armed to properly
treat BV in the long run. Much as a result of the deploring long-
term cure rates with standard antibiotic treatment of BV, several
alternative antibiotics and regimens have recently been evaluated
for their clinical efficacy in eradicating BV.
Of particular relevance here is the renewed interest in 5-nitro-
imidazole derivates other than metronidazole, such as secnidazole
[57,58] , ornidazole [57,59–61] and tinidazole [59,62–78] . Overall, the
metronidazole relatives, secnidazole, ornidazole and tinidazole
seem to be as effective as metronidazole itself.
Other alternative antibiotics and regimens have also recently
been considered. In a recent large randomized, controlled trial the
search for alternative antibiotic regimens was directed towards an
extended-release metronidazole regimen and to azithromycin as
yet another candidate in BV treatment. In this rigorously designed
Box 1. Treatment guidelines for bacterial vaginosis according to
the CDC.
Recommended regimens (CDC, 2006)
• Metronidazole 500 mg orally twice daily for 7 days, or
• Metronidazole gel (0.75%), one full applicator (5 g) intravaginally, once daily for 5 days, or
• Clindamycin cream (2%), one full applicator (5 g) intravaginally at bedtime for 7 days
Alternative regimens (CDC, 2006)
• Clindamycin 300 mg orally twice daily for 7 days, or
• Clindamycin ovules 100 mg intravaginally once at bedtime for 3 days
Recommended regimens for pregnant women (CDC, 2006)
• Metronidazole 500 mg orally twice daily for 7 days, or
• Metronidazole 250 mg orally three times daily for 7 days, or
• Clindamycin 300 mg orally twice daily for 7 days
Data from [145].
1114
clinical trial, Schwebke and Desmond randomized 568 women
with symptomatic BV to one of four arms; that is, a regimen of
extended-release metronidazole (750 mg orally) once per day for
7 days was compared with a regimen of extended-release met-
ronidazole (750 mg orally) once per day for 14 days, with both
metronidazole regimens additionally being evaluated with and
without azithromycin 1 g orally on days 1 and 3 as an adjuvant
antibiotic [79] . As compared with the standard 7‑day regimen, the
extended 14‑day administration regimen for metronidazole was
associated with an increased cure rate at 7 days post-treatment
(82.5 vs 73.8%), although not at 21 days post-treatment (65.9
and 75.6% respectively). Furthermore, there was no therapeutic
effect at all associated with the administration of azithromycin
in addition to metronidazole [79] .
Antibiotic resistance with BV treatments
There are few reports on antimicrobial resistance of BV-associated
anaerobes, except for a vast number of studies on metronida-
zole susceptibility of G. vaginalis, with widely varying rates of
metronidazole resistance having been reported. A single study
that was addressed in two reports is of particular interest here
[80,81] . In this randomized, clinical trial, 119 nonpregnant women
diagnosed according to Amsel’s criteria and with a Nugent score
of 4 or more were randomized to receive either intravaginal met-
ronidazole gel for 5 days or intravaginal clindamycin ovules for
3 days. Quantitative vaginal cultures were then performed at three
follow-up occasions over 90 days and isolated anaerobes were
assessed for metronidazole and clindamycin susceptibility, respec-
tively. Emerging resistance to metronidazole following therapy
was a rare phenomenon, occurring with merely 0.3% of Gram-
negatives evaluated for susceptibility. By contrast, approximately
half of the anaerobic isolates tested in the clindamycin arm rapidly
developed antibiotic resistance to clindamycin, primarily involv-
ing Prevotella spp [80,81] . It was concluded from this study that
emerging clindamycin resistance of vaginal anaerobic bacteria
is a matter of concern, especially since clindamycin and other
­macrolides are widely applied in obstetrics and gynecology.
The recent discovery of several difficult-to-culture bacteria
with BV through molecular techniques
has, however, shed a new light on the
resistant nature of BV in response to treat-
ment with metronidazole. In particular, a
considerable number of strains of the BV
index species A. vaginae have been found to
elicit pronounced metronidazole resistance
in subsequent studies [40,82–84] , while the
limited number of strains evaluated thus far
consistently showed proper susceptibility to
clindamycin [84] .
The picture has become even more
intricate, as BV has recently been found to
involve the development of a dense bacte-
rial biofilm consisting of G. vaginalis and
A. vaginae [48] . Swidsinski et  al. indeed
documented that the biofilm persisted but
Expert Rev. Anti Infect. Ther. 7(9), (2009)

Table 1. Evidence-based appraisal of the CDC treatment guidelines for bacterial vaginosis.
Comparison Effect difference
Oral metronidazole or clindamycin No RCTs
vs placebo
Oral metronidazole vs oral clindamycin Equally effective at 7 days
Intravaginal metronidazole or clindamycin Equally effective at
vs placebo 4–8 weeks
Intravaginal metronidazole No RCTs
vs intravaginal clindamycin
Intravaginal metronidazole or clindamycin Equally effective
vs oral metronidazole or clindamycin at 4 weeks
RCT: Randomized, controlled trial.
temporarily switched to a metabolically latent state with stan-
dard treatment with oral metronidazole and then rapidly regained
activity following treatment c­ essation [85] .
So when accounting for the complexity of the bacterial com-
munity dynamics with BV, it may be questioned whether antibi-
otic susceptibility testing in vitro is actually of any value to the
study of BV.
BV recurrence
It remains unclear at present whether BV recurrence reflects resis-
tance, recurrence and/or reinfection [86] . The surge of molecular
studies is likely to clarify this issue in the nearby future. Recent
studies indicated, for instance, that the presence of A. vaginae with
BV is an indicator of treatment failure [41] , as well as indicator of
BV recurrence [87] . It has been further postulated that following
the resolution of BV, Lactobacillus iners is the Lactobacillus species
most likely to replenish the vagina in appreciable amounts, which
in turn may render patients more vulnerable to a new episode of
BV, considering the rather moderate colonization resistance offered
by L. iners [49] . Albeit based on very small numbers of observa-
tions, the prevailing hypothesis is that recuperation of a stable
Lactobacillus microbiota following treatment is crucial to the long-
term outcome of BV treatment, and that the abundance of L. iners
with BV and following BV cure may compromise this evolution.
Alternative treatment with antiseptics & disinfectants
Antiseptics have been applied for over half a century in the treat-
ment of vaginal infections. Essentially, antiseptics are applied
from the same perspective as antibiotics, which is, the eradication
of the vaginal microflora with BV, following which recolonization
with indigenous lactobacilli is expected. Antiseptics generally
have a very broad spectrum as they act nonspecifically on bacteria
through mechanisms such as bacterial cell membrane disruption.
In accordance, there are very few reports on antimicrobial resis-
tance with these agents. Antiseptics are also generally regarded
as safe for mucosal application when administered in appropriate
www.expert-reviews.com
Bacterial vaginosis Review
Cure rate Level of evidence Ref.
No RCTs No RCTs
94 vs 96% (Greaves [1988]) Low-quality evidence [146]
95 vs 93% (Aubert [1994]) [147]
Clindamycin: Moderate-quality evidence
82 vs 35% (Joesoef [1999]) [148]
Metronidazole:
71 vs 50% (Joesoef [1999]) [148]
No RCTs No RCTs
Paavonen (2000): Very low-quality evidence [149]
Clindamycin cream (82%)
Metronidazole gel (71%)
Oral metronidazole (78%)
concentrations. Hence, antiseptics may be unduly overlooked
in contemporary gynecological practice, as they may provide a
­valuable alternative without systemic exposure to antibiotics.
We are currently performing a systematic review on the efficacy
and safety of antiseptics and disinfectants in the treatment of BV.
To this purpose, we conducted a systematic literature search on
antiseptics and disinfectants listed in the Anatomical Therapeutic
Chemical (ATC) Classification System under the code D08A
(‘antiseptics and disinfectants’), with the exceptions of hydrogen
peroxide, which is listed in the ATC classification system under
the code S02AA for ‘anti-infectives’ and benzydamine which is
listed under ATC code G02CC03
������������������������������������
for ‘anti-inflammatory prod-
ucts for vaginal administration’. Through this thorough search we
identified a total of 12 ­clinical trials that are further considered
for evidence-based appraisal.
Antiseptics that have been administered to women with BV as
vaginal suppositories, bioadhesive gel formulations and occasion-
ally loaded on pessaries in these clinical trials include benzyda-
mine [88] , chlorhexidine [89,90] , dequalinium chloride [91] , poly-
hexamethylene biguanide [92,93] , povidone iodine [65,88,91,94–96]
and hydrogen peroxide [97,98] .
Most of these studies were actually rather well-designed, how-
ever, most were single-blinded and none was placebo-controlled.
Although possibly prone to publication bias, most studies docu-
mented cure rates that were at least as effective as the antibiotics they
were weighted against. However, as each particular antiseptic, except
for povidone iodine, has been studied only once or twice, no firm
conclusions on these antimicrobial agents can be drawn, although
as a whole, this approach definitely warrants further scrutiny as it
may offer a superior means – without involving systemic exposure
to antibiotics – of treatment and prevention of BV recurrence.
Alternative or adjuvant treatment with acidifying agents
A basic component of the healthy vaginal ecosystem is the main-
tenance of an acidic vaginal environment at an average pH of
4 ± 0.5 – inhospitable to most bacteria and viruses – through
1115
 Review Verstraelen & Verhelst
the enzymatic conversion of epithelial cell-derived glycogen pri-
marily into lactate [99,100] . Alkalinization of the vaginal milieu
as induced, for example, by menses or sperm leads to decreased
epithelial adherence of the lactobacilli and gives a free rein to the
overgrowth of typical BV-associated microorganisms. From this
perspective, it has been postulated that actively acidifying the
vagina with naturally occurring acids like lactate or buffering
the vagina against alkalic exposures may enhance lactobacillary
colonization and prevent anaerobic overgrowth. Hence, vaginal
acidifying or so-called buffering agents might serve as either a
therapeutic or preventive means to BV.
In exploratory studies not involving BV, a tampon lubricated
with a lactate-buffered gel was not found effective in one study [101] ,
whereas in another study an intelligent tampon comprising a poly-
meric delivery system that upon absorption of menstrual fluid
gradually releases lactic acid and citric acid ­convincingly leveled
off the pH increase associated with menses [102] .
However, in recent clinical trials, the acidifying approach did
not unequivocally prove effective. In a placebo-controlled ran-
domized study by Holley et al., a 0.92% acetic acid-based gel
applied twice daily for 7 days was not superior to a placebo gel
in the cure of vaginosis [103] . In another study, the acid-buffering
ACIDFORM gel was also significantly less effective than a 10%
metronidazole gel [104] . Intravaginal vitamin C has been proposed
as yet another potentially beneficial acidifying agent [105] .
Two studies evaluated the therapeutic efficacy of gels that con-
tain polycarbophil, a weak polyacid that it is able to stick on the
vaginal epithelial cells until they turnover (up to 3 to 5 days) and
that is assumed to buffer the vaginal secretions. A noncontrolled
study found polycarbophil to be moderately effective in treating
BV, despite the lack of the alleged pH-controlling effect [106] ,
whereas in a randomized, double-blind, placebo-controlled trial,
a polycarbophil–carbopolol-based vaginal gel was found to be
very effective at 1 week following a 5-week treatment course [107] .
Interestingly, a couple of studies documented promising results
with vaginal acidifiers for the long-term treatment of recurrent
BV. Almost two decades ago, Andersch et al. treated 42 women
with recurrent BV with a lactate gel for 7 days and then random-
ized these study participants to receive either prophylactic lactate
gel 3 days monthly for 6 months or a placebo gel [108] . Patients
treated prophylactically with lactate gel for 6 months had – albeit
not defined by conventional criteria – an 83% cure rate as com-
pared with 16% of the women in the control group, although it
must be acknowledged that this comparison might be biased,
as 1-week cure rates (following a 6-month treatment course) in
the active treatment group were compared with 6-month treat-
ment rates in the placebo group. In turn, Wilson et al. enrolled
61 women with recurrent BV, of whom 49 were followed for a
mean duration of 18 months [109] . Following standard antibiotic
treatment, patients were instructed to use a commercial vaginal
gel containing 0.94% glacial acetic acid either nightly or after trig-
ger factors such as menses and coitus. BV recurrences were treated
with standard antibiotic treatment. Although there was no control
group, the authors concluded that the use of the ‘maintenance
acetic acid vaginal gel’ was associated with a significant decrease in
1116
the number of women having any further relapses, as well as in the
rate of recurrences among the remainder, the overall recurrence
rate being reduced from 4.4 to 0.6 recurrences per woman/year
[109] . Clearly, more rigorously designed trials are needed to evalu-
ate the potential role of vaginal acidifying and buffering agents in
the treatment and particularly in the ­prevention of recurrent BV.
Alternative or adjuvant treatment with probiotics
Probiotics have been defined by the United Nations Food and
Agriculture Organization and the WHO in 2001 as live micro-
organisms, which when administered in adequate amounts confer
a health benefit to the host [110] . Somewhere down the line, the
presence of lactobacilli in fermented dairy products and yoghurt
in particular has led to the idea that yoghurt could also offer
an alternative treatment for vaginal infections. Eating yoghurt
or inserting yoghurt into the vagina, for instance, by soaking a
tampon in it, has therefore been a popular alternative treatment
for vaginal infections over the past decades. Only one clinical trial
has actually studied oral yoghurt therapy – in an open, ­crossover
design – for the prophylaxis of recurrent BV [111] . The latter study
suffered, however, from such a huge attrition bias – merely 25%
of study participants completing the study protocol – that any
conclusion is prevented from being drawn. In another two trials,
the effect of intravaginal administration of fermented dairy prod-
ucts on BV was assessed. Fredricsson et al. used a fermented milk
product and found that at 4 weeks, only one out of 13 patients
had been successfully treated with fermented milk, comparing
unfavorably with 13 out of 15 patients being successfully treated
with metronidazole [112] . Neri et al. enrolled 84 women with BV
diagnosed during early pregnancy and obtained a cure rate of
87.5% in the intravaginal Lactobacillus acidophilus yoghurt group
as compared with a cure rate of 37.5% in the group who applied
a vaginal tampon soaked in 5% acetic acid [113] .
The yoghurt approach is questionable, however, taking into
consideration the dissimilarity of lactobacilli pertaining to food
products and the vaginal Lactobacillus species. Yoghurt and other
fermented milk products rely on the mandatory addition of two
lactic acid bacteria, Streptococcus salivarius subsp. thermophilus
and Lactobacillus delbrueckii subsp. bulgaricus, to initiate the fer-
mentation process in milk. These elementary bacteria are often
cocultured with other lactic acid bacteria, including L. acidophi-
lus, Lactobacillus casei and Bifidobacterium species, but not with
typical vaginal Lactobacillus species.
Hence, it was not until more recently that a limited number of
well-documented Lactobacillus strains have been subjected to study
of their colonization potential upon oral or vaginal administration,
such as the L. crispatus strain CTV-05 [114] , the Lactobacillus rham-
nosus strain GR-1 [115] and the Lactobacillus  reuteri strain
RC-14  [116] . As a matter of fact, 100 years after the probiotic
principle was launched by Metchnikoff, treatment of BV with
pro­biotics is regarded as the most promising therapeutic perspec-
tive in this particular area. A number of potentially probiotic
strains are currently under study, that is, vaginal Lactobacillus are
carefully selected based on their properties relating to mucosal
colonization and microbial antagonism, including adhesion to
Expert Rev. Anti Infect. Ther. 7(9), (2009)

vaginal epithelial cells, hydrogen peroxide production, bacteriocin
production, co-aggregation with pathogens, and overall inhibitory
or antagonistic activity towards BV-associated microorganisms.
In order to assess the current evidence on the safety and efficacy
of probiotics in the treatment of BV, we conducted a systematic
review in collaboration with the Cochrane Collaboration [117] .
We performed a thorough search for published and unpublished
trials conducted or published before 2008 on any probiotic used
alone or in conjunction with antibiotics in treating BV diagnosed
according to Nugent’s or Amsel’s criteria. The primary outcome
measure set forward in the study protocol was BV cure accord-
ing to Nugent’s criteria at 21–30 days post-treatment. Overall,
16 studies published between 1992 and 2006 were identified, of
which four met the criteria for inclusion in the systematic review.
In the study by Parent et al., microbiological cure on day 28
was documented in seven out of eight women in the arm who
received one Gynoflor® tablet (10 million viable ‘L. acidophilus
bacteria’ and 0.03 mg estriol per tablet) daily for 6 days com-
pared with one out of seven women in the placebo arm (OR: 0.02;
95% CI: 0.00–0.47%) [118] . It may be added here that the so-called
L. acidophilus labeled by the Gynoflor manufacturer is actually
a L. crispatus strain, based on a product ana­lysis we performed
[Verhelst R, Verstraelen H, Unpublished data] . In the study by Eriksson
et al. 225 subjects were initially enrolled in the open part of the
trial and all were treated with 100 mg clindamycin ovules inserted
vaginally once daily for 3 days and then randomized to one of two
arms [119] . During the first menstrual period following clindamy-
cin treatment, patients received either lactobacilli-impregnated
tampons (1 × 108 freeze-dried L. rhamnosus, Lactobacillus gas-
seri and Lactobacillus fermentum cells per tampon) or placebo
tampons, and participants had to use at least five tampons to be
included in the efficacy ana­lysis. After the second menstruation,
during which the women used their normal catamenial protec-
tion, and according to an intention-to-treat ana­lysis, 75 out of 108
women in the treatment group and 80 of the 109 women in the
placebo arm presented with normalized Nugent scores (OR: 0.82;
95% CI: 0.46–1.49%). The authors explained these results by the
fact that the Lactobacillus strains were not properly released from
the tampons. In a first study by Anukam et al., patients were ran-
domized in a double-blind manner and given oral metronidazole
500 mg twice daily for 7 days, plus either oral L. rhamnosus GR-1
and L. reuteri RC-14 (1 × 109 cells per capsule) twice daily for
30 days starting on day 1 of metronidazole treatment or identical-
looking placebo capsules [120] . In the per-protocol ana­lysis, BV
cure at day 30 according to Nugent’s criteria, occurred with 43
of the 49 women in the antibiotic/probiotic treatment arm and
with 23 of the 57 women in the antibiotic/placebo arm (OR: 0.09;
95% CI: 0.03–0.26%). In a second randomized, controlled trial
by Anukam et al., 40 women diagnosed with BV were randomized
to receive either two dried capsules containing L. rhamnosus GR-1
and L. reuteri RC-14 each night for 5 days, or 0.75% metroni-
dazole gel, applied vaginally twice a day [121] . Follow-up at days
6, 15 and 30 showed cure of BV in significantly more probiotic-
treated subjects (16/20, 17/20 and 18/20, respectively) compared
with metronidazole treatment (9/20, 9/20 and 11/20, respectively;
www.expert-reviews.com
Bacterial vaginosis Review
p = 0.016 at day 6, 0.002 at day 15 and 0.056 at day 30). Hence, in
this study the probiotic regimen proved superior to metronidazole
gel on 1- and 2-week cure rates, but marginally missed significance
on 3-week cure rates (OR: 0.27; 95% CI: 0.07–1.10%), which
was the primary outcome of the Cochrane Review.
In brief, two of the four studies, one involving vaginally admin-
istered lactobacilli combined with estriol [118] and one study
involving orally administered lactobacilli [120] , documented a
highly beneficial effect on the resolution of BV at 21–30 days
post-treatment, whereas in the other two studies involving the
administration of intravaginal lactobacilli, the probiotic treatment
was not significantly better compared with placebo [119] or com-
pared with intravaginal metronidazole gel [121] . Therefore, based
on a limited number of studies that were selected based on their
level of evidence, it must be concluded that there is insufficient
evidence available as yet to recommend the use of probiotics in
addition to antibiotics in the treatment of BV, although the results
from some studies were particularly promising.
Several, similarly promising studies have been published since we
performed the systematic literature search within the framework of
the Cochrane Review thereby including studies published before
2008. Larsson et al. randomized 100 women with BV in two groups
of 50 and the participants were given a 7-day course of daily 2%
vaginal clindamycin cream directly followed by vaginal gelatine
capsules containing 108–9 freeze-dried lactobacilli (a L. gasseri and
a L. rhamnosus strain) or placebo capsules of identical appearance,
sufficient for 10 days or until menstruation commenced [122] . After
each menstruation, treatment with vaginal lactobacilli capsules or
placebo was repeated during 10 days for three cycles. Thus, the
treatment regime included one treatment course with clindamycin
followed by four lactobacilli/placebo courses; one within the same
menstrual cycle and the other during the next three consecutive
cycles. The initial intent-to-treat ana­lysis for the 1-month cure rate
was 64% in the lactobacilli group and 78% in the placebo group
(p = 0.8). The 76 cured women were followed for six menstrual
cycles or until relapse within that timespan. At the end of the
study, 64.9% (24 out of 37) of the lactobacilli-treated women were
still BV-free compared with 46.2% (18 out of 39) of the placebo-
treated women (p = 0.042). Marcone et al. randomized 84 patients
with BV in a single-blinded manner [123] . All patients received oral
metronidazole 500 mg twice a day for 7 days, then followed in
the treatment group by one vaginal tablet containing freeze-dried
L. rhamnosus once a week at bedtime for 2 months starting 1 week
after the last antibiotic administration. Follow-up was performed at
days 30, 90 and 180, with cure rates in the treatment group of 88,
88 and 83% and in the control group of 81, 71 and 67% at 1, 3 and
6 months, respectively (p = 0.4, p= 0.05 and p = 0.07). In a rather
small but well-designed double-blinded randomized, controlled
trial, Mastromarino et al. randomized 39 women with BV to receive
either a vaginal probiotic tablet (containing at least 109 viable lacto-
bacilli, in particular a Lactobacillus brevis strain, a Lactobacillus������
 sali-
varius subsp. salicinius strain, and a Lactobacillus plantarum strain)
or an identical placebo for 7 days [124] . The 2-week cure rates were
61% (11 out of 18) in the active treatment group as compared with
19% (three out of 16) in the placebo group (p = 0.017).
1117
 Review Verstraelen & Verhelst
Treatment of recurrent BV
As apparent from the aforementioned information, the main chal-
lenge with BV remains the quest for long-term cure and hence
the prevention of BV recurrence. Standard antibiotic treatment
regimens are associated with high relapse rates, and we have very
few options for dealing with relapsing BV. As outlined, several
alternative treatment options, including long-term treatment with
vaginal acidifying agents and probiotics, may offer promise for
overcoming this longstanding frustration.
The most rigorously studied approach to the prevention of recur-
rent BV, however, certainly comes from a US multicenter trial in
which women were initially treated during the open-labeled phase
with a 10‑day course of 0.75% metronidazole gel and in which
asymptomatic responders were then randomized to receive twice-
weekly 0.75% metronidazole vaginal gel or placebo for 16 weeks
and off therapy for 12 weeks [125] . During suppressive therapy,
recurrent BV occurred in 13 women (25.5%) receiving metro-
nidazole and in 26 (59.1%) receiving placebo (RR: 0.43; 95%
CI: 0.25–0.73%; p = 0.001). During the entire 28-week follow-
up, recurrence occurred in 26 (51.0%) on treatment compared
with 33 (75.0%) on placebo (RR: 0.68; 95% CI: 0.49–0.93%;
p = 0.02). Adverse effects were uncommon, although second-
ary vaginal candidiasis occurred significantly more often in
­metronidazole-treated women.
In a recent study, Reichman et al. performed a retrospective ana­
lysis of a modified long-term suppressive triple-phase treatment
scheme in which patients with recurrent BV were treated con-
secutively with 7 days of oral nitroimidazole, followed by 21 days
of intravaginal boric acid 600 mg/day and if in remission then
treated with metronidazole gel twice weekly for 16 weeks  [126] .
Cure after nitroimidazole and boric acid therapy ranged from
88 to 92%, 7 and 12 weeks after the initial visit, respectively.
Cumulative cure at 12, 16 and 28 weeks from the initial visit was
87, 78 and 65%, respectively. A failure rate of 50% was docu-
mented by 36 weeks of follow-up. Although very encouraging,
this approach requires validation in a prospective randomized,
controlled study.
Partner treatment prevention of BV
Of all risk factors explored thus far, the majority of epidemio-
logic studies have identified sexual behaviour as the primary risk
factor to the occurrence of BV. The nature of this association
remains fraught, however, considering that a number of observa-
tions somewhat paradoxically point at a very consistent correla-
tion with sexual contact, yet seem to contradict at least in part a
traditional mode of sexual transmission.
Still, as the epidemiological profile of BV mirrors that of estab-
lished sexually transmitted infections, the putative role of partner
treatment with antibiotics and condom use have been scrutinized
with regard to the treatment and prevention of BV.
To date, six randomized controlled trials [74,127–131] have been
directed towards the effectiveness of male partner treatment in
the treatment of BV. Five out of the six studies failed to document
any benefit from partner treatment with antibiotics [74,127–129,131] .
It may be acknowledged here that most of these studies suffer
1118
from multiple methodological shortcomings [132] . Moreover, the
antibiotic regimens applied to male partners of women diagnosed
with BV, mostly single doses or short courses with metronidazole
or tinidazole, are also poorly effective in women with BV and are
therefore not recommended by the CDC [56] . In only one of the
six randomized, controlled trials, a CDC-recommended regimen
for women was administered to the spouses of women with BV,
consisting of a 7‑day course of oral clindamycin, although again
without any noticeable effect [131] . Finally, while partner treatment
might not be effective in treating BV, what one really wants to
know is whether male treatment might prevent the recurrence of
BV among their female partners, presuming that women might
get reinfected from a male reservoir. This was addressed in two
studies with a 3 month follow-up [129,131] – when a recurrence rate
of at least 50% among women is expected [52] – although both
studies failed to document any benefit of male sexual partner
treatment on 3-month cure rates among their female partners. It
may be concluded that the evidence suggests that there is no ben-
efit in treating the sexual partner of women with BV with the drug
regimens tested, although it may be argued that no evidence of
effect does not equate to evidence of no effect [132] . It may further
be acknowledged that when assuming a male-to-female route of
transmission, the true effect of male treatment on the incidence
of BV could only be evaluated in a study in which male carriers
would be treated prophylactically.
With regard to condom use as a means of preventing BV, six
cross-sectional studies were equivocal [133–138] , whereas longi­
tudinal and cohort studies on the other hand are more in line with
each other towards a beneficial effect of condom use in relation
to BV acquisition [139–142] , although overall the observed effects
tend to be very moderate, with an average relative risk reduction
associated with condom use in a recent meta-ana­lysis estimated
to be merely 20% [143] . Interestingly, the two most recent studies
also addressed recurrent incident BV [142,144] . Hutchinson et al.
found a very strong overall protective effect of consistent condom
use on the occurrence of both incident and recurrent incident
BV in a 3-year follow-up study (adjusted OR: 0.37; 95% CI:
0.20–0.70%) [142] . Conversly, Yotebieng et al. found that con-
sistent condom use in a 6‑month follow-up study was protective
against incident BV, although not against recurrent incident BV
[144] . Hence, the evidence on consistent condom use as a protective
means for BV overall seems to suggest a rather moderate effect
on the ­prevention of BV.
Expert commentary & five-year view
Diagnosis of BV in the nearby future will definitely continue to
rely on standardized clinical criteria that can be assessed rapidly
as a point-of-care test by the attending physician. It is even com-
mendable that this approach would be uniformly implemented,
as vaginitis is too often treated empirically. Diagnosis is ideally
confirmed by ana­lysis of a Gram-stained vaginal smear, although
there is a defined need to standardize the procedures and cri-
teria involved. Computer-based cell morphotype recognition is
being developed and may help tremendously in the scoring of
Gram-stained vaginal smears. Furthermore, following the recent
Expert Rev. Anti Infect. Ther. 7(9), (2009)
 Bacterial vaginosis Review

surge of molecular ana­lysis of the vaginal bacterial biota, molecu-
lar diagnosis of BV is now pending. Species-specific QRT-PCR
techniques are being patented and may become commercially
available as diagnostic assays. In addition, highly sensitive high-
throughput ana­lysis through deep pyrosequencing will further
enlighten our knowledge of bacterial diversity with BV. This in
turn may lead to the recognition of distinct microflora patterns
representing specific subtypes of BV that may be associated with
specific health outcomes. Finally, molecular diagnosis will also
document that part of what is now categorized as intermediate
microflora actually involves BV biota. As for the treatment of
BV, there are very few perspectives, if any, for shotgun therapy
and singular treatment courses. Therefore, the focus will shift to
long-term treatment regimens aiming at restoring and maintain-
ing a normal vaginal microflora, thereby in itself preventing the
recurrence of BV. Further research may explore the potential of
disinfectants and vaginal acidifying and buffering agents; how-
ever, the expectations of the scientific community are primarily
directed to the development of prebiotics, probiotics and synbiot-
ics. Results obtained thus far with the administration of probiotic
bacilli following an antibiotic course have not proven unequivo-
cally successful, but at least we have a proof-of-concept for this
approach through a couple of randomized clinical trials. Probiotic
preparations will be sophisticated further by adding prebiotics
and other substances that enhance the colonization and growth of
the lactobacilli administered. Finally, therapeutic research in this
particular area will be guided by our recently acquired knowledge
of BV as a biofilm condition.
Financial & competing interests disclosure
The authors have no relevant affiliations or financial involvement with any
organization or entity with a financial interest in or financial conflict with
the subject matter or materials discussed in the manuscript. This includes
employment, consultancies, honoraria, stock ownership or options, expert
testimony, grants or patents received or pending, or royalties.
No writing assistance was utilized in the production of this manuscript.

Key issues
• Bacterial vaginosis (BV) is a condition characterized by a loss of the indigenous vaginal lactobacilli and massive anaerobic overgrowth of
the vaginal mucosa with a mix of anaerobes.
• BV is a highly common infestation among women of childbearing age and is associated with a vast disease burden of adverse obstetric
outcome, pelvic inflammatory disease, and the acquisition and secondary spread of HIV‑1 and other sexually transmitted diseases.
• In-office or point-of-care diagnosis of BV relies on a set of clinical criteria (discharge, amine odor, increased vaginal pH and the
pathognomonic presence of epithelial cells on microscopy loaded with BV-associated bacteria).
• Laboratory diagnosis typically involves the quantitation of bacterial cell morphotypes on a Gram-stained vaginal smear.
• Therapy of BV typically involves treatment with oral or intravaginal metronidazole or clindamycin and is associated with particularly high
relapse rates.
• Alternative treatment options – including maintenance treatment following an antibiotic course – are increasingly explored and involve
treatment with antiseptics and disinfectants, vaginal acidifying or buffering agents, and probiotics.

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•• Second paper from the same group on the
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Affiliations
• Hans Verstraelen, MD, MPH, PhD
Department of Obstetrics & Gynaecology,
Ghent University Hospital, De Pintelaan
185, B-9000 Ghent, Belgium
Tel.: +32 9332 3796
Fax: +32 9332 3831
hans.verstraelen@ugent.be
• Rita Verhelst, PhD
Laboratory Bacteriology Research,
Department of Clinical Chemistry,
Microbiology and Immunology, Ghent
University Hospital, De Pintelaan 185,
B-9000 Ghent, Belgium
Tel.: +32 498 081 982
Fax: +32 9332 3659
rita.verhelst@ugent.be
Expert Rev. Anti Infect. Ther. 7(9), (2009)