Bacterial Vaginosis Diagnosed by Analysis of First-Void-Urine Specimens

Bacterial vaginosis (BV) is traditionally diagnosed using vaginal samples. The aim of this study was to investigate whether BV can be diagnosed from first-void urine (FVU). Self-collected vaginal smears, vaginal swabs, and FVU were obtained from 176 women. BV was diagnosed by Nugent''s criteria. The FVU and vaginal swabs were analyzed by quantitative PCRs (qPCRs) for selected vaginal bacteria (Atopobium vaginae, Prevotella spp., Gardnerella vaginalis, bacterial vaginosis-associated bacterium 2, Eggerthella-like bacterium, “Leptotrichia amnionii,” Megasphaera type 1), and all had an area under the receiver operating characteristic (ROC) curve of >85%, suggesting good prediction of BV according to the Nugent score. All seven bacteria in FVU were significantly associated with BV in univariate analysis. An accurate diagnosis of BV from urine was obtained in this population by a combination of qPCRs for Megasphaera type 1 and Prevotella spp. The same two bacteria remained significantly associated with BV in a multivariate model after adjusting for the other five species. There was no statistically significant difference between the sensitivities and specificities of BV diagnosis by molecular methods performed on swabs and FVU samples. A linear regression analysis showed good agreement between bacterial loads from swabs and FVU, but Prevotella spp. could be detected in high numbers in a few FVU samples without being present in swabs. This method will allow diagnosis of BV in studies where only urine has been collected and where detection of BV is considered relevant.     

A DNA tool for early detection of vaginal dysbiosis in African women

A next-generation diagnostic tool for bacterial vaginosis, consisting of quantitative and/or qualitative molecular criteria, has not yet been identified. The optimal diagnostic tool should not only diagnose bacterial vaginosis in diverse populations, but should also detect early signs of transition to dysbiosis.We evaluated a tool based on log₁₀-transformed qPCR data for Lactobacillus crispatus, Lactobacillus iners, Lactobacillus jensenii, Lactobacillus gasseri, Lactobacillus vaginalis, Lactobacillus genus, Atopobium vaginae and Gardnerella vaginalis in vaginal specimens of 426 African women to detect dysbiosis and predict transition to dysbiosis.G. vaginalis (p = 0.204) and A. vaginae (p = 0.001) were more commonly present in women who evolved to an intermediate (Nugent 4–6) or bacterial vaginosis score (Nugent 7–10) compared to women who continued to have a normal Nugent score. The combination of G. vaginalis, A. vaginae and Lactobacillus genus counts performed best for diagnostic accuracy for bacterial vaginosis--sensitivity 93.4% and specificity 83.6%; and for predictive accuracy for bacterial vaginosis--sensitivity 79% and specificity 52%. L. crispatus combinations did not perform well.We conclude that a triple-G. vaginalis-A. vaginae-Lactobacillus genus-qPCR tool holds promise for research in sub-Saharan Africa or when developed as a next-generation clinical diagnostic modality for bacterial vaginosis, ideally engineered as a rapid assay.     

Strong correspondence in bacterial loads between the vagina and rectum of pregnant women

We sampled the vagina and rectum in 71 pregnant women and bacterial loads of Lactobacillus crispatus, L. jensenii, L. gasseri, L. iners, Gardnerella vaginalis and Atopobium vaginae were determined by culture and quantitative PCR (qPCR).Culture and qPCR results differed substantially with regard to the evaluation of vaginal and rectal occurrence of the six species tested. The vaginal-rectal prevalence of L. crispatus, L. jensenii, L. gasseri, L. iners, G. vaginalis and A. vaginae as established by culture vs. PCR was 32.3 vs. 91.5%, 32.3 vs. 77.4%, 28.1 vs. 91.5%, 12.6 vs. 68.5%, 12.6 vs. 74.6% and 5.6 vs. 69.0%, respectively.Using qPCR, a significant positive correlation was found between vaginal and rectal loads of L. crispatus (p < 0.0001), L. jensenii (p < 0.0001), L. gasseri (p = 0.005), L. iners (p = 0.003) and A. vaginae (p = 0.002).In summary, significant correlations between quantities of vaginal and rectal lactobacilli and of Atopobium vaginae were established by means of qPCR, indicating strong correspondence of vaginal and rectal microflora, not only in the occurrence of certain species in both niches, but also of cell densities per bacterial species.     

Diagnostic accuracy of quantitative real-time PCR assay versus clinical and Gram stain identification of bacterial vaginosis

The purpose of this investigation was to determine the diagnostic accuracy of quantitative real-time polymerase chain reaction (PCR) assay in diagnosing bacterial vaginosis versus the standard methods, the Amsel criteria and the Nugent score. The Amsel criteria, the Nugent score, and results from the molecular tool were obtained independently from vaginal samples of 163 pregnant women who reported abnormal vaginal symptoms before 20 weeks gestation. To determine the performance of the molecular tool, we calculated the kappa value, sensitivity, specificity, and positive and negative predictive values. Either or both of the Amsel criteria (≥3 criteria) and the Nugent score (score ≥7) indicated that 25 women (15%) had bacterial vaginosis, and the remaining 138 women did not. DNA levels of Gardnerella vaginalis or Atopobium vaginae exceeded 10⁹ copies/mL or 10⁸ copies/mL, respectively, in 34 (21%) of the 163 samples. Complete agreement between both reference methods and high concentrations of G. vaginalis and A. vaginae was found in 94.5% of women (154/163 samples, kappa value = 0.81, 95% confidence interval 0.70–0.81). The nine samples with discordant results were categorized as intermediate flora by the Nugent score. The molecular tool predicted bacterial vaginosis with a sensitivity of 100%, a specificity of 93%, a positive predictive value of 73%, and a negative predictive value of 100%. The quantitative real-time PCR assay shows excellent agreement with the results of both reference methods for the diagnosis of bacterial vaginosis.     

Multiplex Detection of Bacteria Associated with Normal Microbiota and with Bacterial Vaginosis in Vaginal Swabs by Use of Oligonucleotide-Coupled Fluorescent Microspheres

Bacterial vaginosis (BV) is a recurrent condition that is associated with a range of negative outcomes, including the acquisition of human immunodeficiency virus and other sexually transmitted diseases, preterm births, and pelvic inflammatory disease. In contrast to the Lactobacillus-dominated normal vaginal microbiota, BV is characterized by a lack of lactobacilli and an abundance of anaerobic and gram-negative organisms, including Gardnerella vaginalis and Atopobium vaginae. To date, the laboratory diagnosis of BV has relied upon the fulfillment of criteria determined by microscopic observation of Gram-stained vaginal swabs. We describe a molecular-based method for the easy determination of the species profile within the vaginal microbiota based on the amplification of the chaperonin-60 genes of all bacteria present in the swab and hybridization of the amplicon to species-specific oligonucleotide-coupled fluorescent beads that are identified by flow cytometry with a Luminex instrument. We designed a nineplex Luminex array for characterization of the vaginal microbiota and applied it to the analysis of vaginal swabs from individuals from Africa and North America. Using the presence of A. vaginae or G. vaginalis, or both, as the defining criterion for BV, we found that the method was highly specific and sensitive for the diagnosis of BV using microscopy as a gold standard.Bacterial vaginosis (BV), which is defined by a reduction in the vaginal Lactobacillus populations and an increase in the number of microbial species present, including Gardnerella spp., Atopobium spp., and others, is increasingly recognized as an important risk factor for adverse reproductive health outcomes, such as miscarriage and premature birth, and sexually transmitted diseases, including human immunodeficiency virus infection (25). Despite intense investigation, the etiology and clinical course of BV have not been well defined, probably because the normal variation of the vaginal microbial communities between individuals and their dynamics over time are complex and poorly understood. Consequently, the accurate diagnosis of BV, as well as the elaboration of effective prevention and treatment strategies, remains a major challenge (26, 44).The current “gold standard” for the diagnosis of BV in the laboratory setting relies on microscopic profiling of microbial types in Gram-stained vaginal swab smears by using the criteria of Nugent et al. (29) or Ison and Hay (20). These scoring systems are based on the relative abundance of Lactobacillus morphotypes (large gram-positive rods) compared to the abundance of the bacterial morphotypes suggestive of BV (gram-negative or gram-variable coccobacilli and curved rods and gram-positive cocci). Although this method provides reliable information and is particularly well suited for use in resource-poor settings, it requires well-trained, highly experienced individuals to interpret the results and is simplistic in its reflection of the variety of organisms present in the microbial communities present in healthy individuals with a normal vaginal flora and patients with BV. Potential problems with the subjective interpretation of the Gram stain result, along with an increasing appreciation of the fact that the complexity of the vaginal microbiota can be missed by these classical methods, have resulted in a search for other methods for the diagnosis of BV that can also identify the different species present (5, 25).In-depth studies of the makeup of vaginal microbial communities are quite common, including studies that use the older culture-based approaches (31) and more recent molecular studies that involve the construction of a clone library on the basis of the 16S rRNA gene (30) or the chaperonin-60 (cpn60) (16) target. While the information generated by these studies is critical to obtaining a greater understanding of the variability and ecological dynamics of vaginal microbial communities, they remain far too labor-intensive at present for the screening of large numbers of samples or for the routine laboratory diagnosis of BV.Most recently described molecular approaches for the diagnosis of BV are based on quantitative PCR (qPCR) of the 16S rRNA gene or other targets in bacteria extracted from vaginal samples. For example, one recent study found that the simultaneous quantification of the Gardnerella cpn60 target and the Atopobium 16S rRNA gene target resulted in the sensitive and specific diagnosis of BV in comparison to the sensitivity and specificity of the diagnosis by use of the scoring system of Nugent et al. (29). The advantages of the cpn60 universal target (UT) as an alternative to the 16S rRNA gene for the identification of bacterial species are increasingly recognized and include the increased resolution available at the species level and its presence at a single copy per genome, improving the reliability of quantitation (7).Multiplexed, bead-based flow cytometric detection and quantification of bacterial targets by use of the Luminex platform have been described previously (3, 9, 14, 38). The main advantage of this approach is its ability to detect a large number of molecular targets simultaneously after a single PCR, making high-throughput analysis of multiple samples from large study groups or longitudinal studies technically feasible. In the context of BV, the use of this multiplex approach, coupled with highly discriminatory cpn60-specific probes, facilitates screening for multiple species of Lactobacillus and a number of BV-associated bacteria simultaneously, providing an unprecedented level of information about the variability of vaginal bacterial communities, defined as those associated with a normal vaginal flora and those associated with BV, in a single, rapid assay.The goal of the present study was the design of a flow cytometric bead array for the rapid profiling of microbial communities extracted from vaginal samples and its evaluation as a potential technique for the rapid diagnosis of BV in the laboratory setting.     

<Emphasis Type="Italic">Mycoplasma hominis</Emphasis> and <Emphasis Type="Italic">Gardnerella vaginalis</Emphasis> display a significant synergistic relationship in bacterial vaginosis

Gardnerella vaginalis plays an important role in bacterial vaginosis (BV,) while the role of genital Mollicutes is less obvious. The diagnosis of BV by use of the current Gram stain Nugent score is also suboptimal for defining the role of Mollicutes that lack a cell wall. Since bacterial load and diversity is an important prerequisite for BV, real-time quantitative polymerase chain reaction (qPCR) assays enable these to be assessed. The purpose of this study was to define the role of genital Mollicutes and potential patterns of synergy with G. vaginalis in women with BV. Vaginal swabs from 130 women categorised by Nugent score as BV (n?=?28), intermediate (n?=?22) and non-BV (n?=?80) were tested against four qPCR TaqMan assays targeting G. vaginalis, Mycoplasma hominis, M. genitalium, Ureaplasma urealyticum and U. parvum. Statistical analyses were used to compare bacterial prevalence and load between the three groups of women. Mycoplasma hominis and G. vaginalis co-infection was significantly more common in BV (60.7 %) compared to intermediate (36.4 %) and non-BV (8.8 %) Nugent scores (p?<?0.001). Significantly higher loads of M. hominis (p?=?0.001) and G. vaginalis (p?<?0.001) were detected in women with BV and the respective loads in M. hominis and G. vaginalis co-infections displayed a significant positive correlation (p?<?0.001; r?=?0.60). No significant associations were seen with the other Mollicutes. The findings strengthen the evidence of a role for M. hominis in BV and a potential synergy with G. vaginalis. This synergy could be an important trigger of the condition and sexual contact the conduit for the transmission of an otherwise commensal bacterium that could initiate it.     

Analysis of vaginal acetic acid in patients undergoing treatment for bacterial vaginosis

A "gold standard" method for the diagnosis of bacterial vaginosis (BV) is lacking. The clinical criteria described by the Amsel technique are subjective and difficult to quantify. Alternatively, the reading of Gram-stained vaginal smears by scoring techniques such as those that use the Nugent or Hay-Ison scoring systems is again subjective, requires expert personnel to perform the reading, and is infrequently used clinically. Recently, a new diagnostic device, the Osmetech Microbial Analyzer--Bacterial Vaginosis (OMA-BV), which determines a patient's BV status on the basis of measurement of the amount of acetic acid present in a vaginal swab specimen, was approved by the Food and Drug Administration. The present study uses the conducting polymer gas-sensing technology of OMA-BV to measure the concentration of acetic acid in the headspace above vaginal swab specimens from patients undergoing treatment for BV with metronidazole. In 97.8% of the cases the level of acetic acid detected fell sharply during the treatment period, crossing from above to below the diagnostic threshold of 900 ppm. The diagnosis obtained on the basis of the level of vaginal acetic acid was compared with the diagnoses obtained by use of the Amsel criteria and the Nugent scoring system both at the time of initial entry into the study and at the repeat samplings on days 7 and 14. The results obtained with OMA-BV showed overall agreements compared with the results of the Amsel and Nugent tests of 98 and 94%, respectively, for the 34 patients monitored through the treatment process. This provides further evidence that the measurement of vaginal acetic acid by headspace analysis with conducting polymer sensors is a valid alternative to present tests for the diagnosis of BV.     

Trimethylamine content in vaginal secretion and its relation to bacterial vaginosis

The presence of a fishy odor emanating from women who present with a malodorous vaginal discharge is well known. The odor is due to bacterial reduction of trimethylamine oxide to trimethylamine (TMA) in vaginal secretion. The release of TMA from specimens of vaginal fluid following the addition of alkali is often used in making a clinical diagnosis of bacterial vaginosis (BV). We now report a sensitive gas chromatographic method for analysis and quantification of TMA in vaginal fluid in which weighed samples were used. In addition, a proper diagnosis of BV was obtained using Gram-stained smears of the vaginal fluid according to the method of Nugent et al. (R. P. Nugent et al., J Clin Microbiol 1991;29:297-301). We also diagnosed BV according to Hallén et al. (A. Hallén et al. Genitourin Med 1987;63:386-9). TMA was present in all women with a Nugent score between 7 and 10 and in almost all women diagnosed with BV according to the method of Hallén et al. TMA was not found or was only found in very low concentrations in vaginal fluid from women with Nugent scores of 0 to 3. TMA was also found in four women with a negative sniff test. It seems that high levels of TMA in samples of vaginal fluid are typical for BV regardless of the scoring method used for diagnosis. However, low levels of TMA, <5 microg/g vaginal fluid, do not always correlate with BV.     

Molecular analysis of the relationship between specific vaginal bacteria and bacterial vaginosis metronidazole therapy failure

Bacterial vaginosis frequently persists, even after treatment. The role of some strains of bacteria associated with bacterial vaginosis treatment failure remains poorly defined. The aim of our study was to define the risk of bacterial vaginosis treatment failure, including pre-treatment detection of specific vaginal bacteria. Bacterial vaginosis is present when the Nugent score is ≥7 and the modified Amsel criteria is positive. Women with bacterial vaginosis were treated with intravaginal metronidazole gel nightly for 5 nights. The 454 pyrosequencing method was used to detect bacteria in vaginal fluid. By univariate analysis, a history of bacterial vaginosis, intrauterine device use and the presence of Facklamia, Corynebacterium and Veillonella were significantly associated with bacterial vaginosis treatment failure. Lactobacillus crispatus, Lactobacillus pentosus and Megasphaera were significantly associated with curing bacterial vaginosis. After logistic regression analysis and detection of these bacteria for test-of-cure, we found that women who had a history of bacterial vaginosis had a higher incidence of bacterial vaginosis treatment failure, whereas women with L. crispatus had a lower incidence of treatment failure. Post-treatment sexual activity was not associated with the treatment effect. Our data suggested that treatment failure may be not caused by drug resistance. Rather, it has a closer relationship with the failed restoration of lactobacilli.     

Self-collected vaginal swabs for the quantitative real-time polymerase chain reaction assay of Atopobium vaginae and Gardnerella vaginalis and the diagnosis of bacterial vaginosis

The aim of this study was to assess the feasibility of using self-collected vaginal specimens for the quantitative real-time polymerase chain reaction (qPCR) assays of bacterial vaginosis (BV)-associated bacteria versus practitioner-collected swabs. A cross-sectional study included 190 pregnant women enrolled before 20 weeks’ gestation from September 2008 to November 2009. Self- and practitioner-collected swabs were taken during the same prenatal visit for each woman, qPCR assays performed for each, and the results compared. The quantification of the human albumin gene was used as an internal control to ensure sampling quality and accurate comparisons. The level of agreement of the qPCR assays for each microorganism was calculated with the Spearman product moment correlation coefficient and the kappa statistic. In all, 370 vaginal samples (185 self- and 185 practitioner-collected swabs) had a narrow range of values for the number of albumin gene copies and a significant correlation coefficient (Spearman’s rho = 0.532; p < 0.001). The agreement between both sampling methods was excellent (Spearman’s rho was 0.748 for Atopobium vaginae, 0.918 for Lactobacillus species, 0.940 for Gardnerella vaginalis; p < 0.001), especially for high concentrations of A. vaginae (≥10⁸ copies/mL; kappa value = 0.973; p < 0.001) and G. vaginalis (≥10⁹ copies/mL; kappa value = 0.903; p < 0.001). This study demonstrates the validity and reliability of self- versus practitioner-collected swabs for the molecular quantification of Lactobacillus species, G. vaginalis, and A. vaginae.     

Reciprocal Interference between Lactobacillus spp. and Gardnerella vaginalis on Initial Adherence to Epithelial Cells

Bacterial vaginosis (BV) is the most common vaginal disorder in women of child-bearing age. It is widely accepted that the microbial switch from normal microflora to the flora commonly associated with BV is characterized by a decrease in vaginal colonization by specific Lactobacillus species together with an increase of G. vaginalis and other anaerobes. However, the order of events leading to the development of BV remains poorly characterized and it is unclear whether the decrease in lactobacilli is a cause or a consequence of the increase in the population density of anaerobes. Our goal was to characterize the interaction between two Gardnerella vaginalis strains, one of which was isolated from a healthy woman (strain 5-1) and the other from a woman diagnosed with BV (strain 101), and vaginal lactobacilli on the adherence to cervical epithelial cells. In order to simulate the transition from vaginal health to BV, the lactobacilli were cultured with the epithelial cells first, and then the G. vaginalis strain was introduced. We quantified the inhibition of G. vaginalis adherence by the lactobacilli and displacement of adherent lactobacilli by G. vaginalis. Our results confirmed that pathogenic G vaginalis 101 had a higher capacity for adhesion to the cervical epithelial cells than strain 5-1. Interestingly, strain 101 displaced L. crispatus but not L. iners whereas strain 5-1 had less of an effect and did not affect the two species differently. Furthermore, L. iners actually enhanced adhesion of strain 101 but not of strain 5-1. These results suggest that BV-causing G. vaginalis and L. iners do not interfere with one another, which may help to explain previous reports that women who are colonized with L. iners are more likely to develop BV.     

Prevalence of bacterial vaginosis in antenatal women

Bacterial vaginosis is an established risk factor in pregnant women for premature rupture of membranes and preterm delivery. This study was carried out to find out the prevalence of Bacterial Vaginosis (BV) in antenatal women with vaginal discharge and the effect of treatment with Metronidazole gel on pregnancy outcome. One hundred and fifty symptomatic and fifty asymptomatic women in second trimester of pregnancy in the age group of 20-30 years were included in the study. Gram stained smears of vaginal discharge were examined for evidence of BV with a scoring system by Nugent et al and was found to be positive in 38.5% in symptomatic antenatal women. Intravaginal metronidazole gel application was found to be an effective therapeutic option. Incidence of preterm labour was more in untreated cases.     

Comparison of Nucleic Acid Amplification Assays with BD Affirm VPIII for Diagnosis of Vaginitis in Symptomatic Women

A commercially available, nonamplified, nucleic acid probe-based test system (BD Affirm VPIII) was compared with nucleic acid amplification (NAA)-based assays for determining the etiology of vaginitis in a cohort of 323 symptomatic women. First, a semiquantitative, multiplexed PCR assay (BV-PCR) and the Affirm VPIII Gardnerellavaginalis test were compared with a unified bacterial-vaginosis (BV) reference standard incorporating both Nugent Gram stain scores and Amsel clinical criteria. In the evaluable population of 305 patients, BV-PCR was 96.9% (191/197) sensitive and 92.6% specific (100/108) for BV, while Affirm VPIII was 90.1% sensitive (179/197) and 67.6% specific (73/108). Second, a multiplexed PCR assay detecting Candida albicans and Candida glabrata (CAN-PCR) was compared with the Affirm VPIII Candida test using a reference standard for vulvovaginal candidiasis (VVC) of yeast culture plus exclusion of alternate vaginitis etiologies. In the population evaluated (n = 102), CAN-PCR was 97.7% sensitive (42/43) and 93.2% specific (55/59) and Affirm VP III was 58.1% sensitive (25/43) and 100% specific (59/59) for VVC. Finally, the results of a commercial NAA test (GenProbe Aptima Trichomonas vaginalis assay; ATV) for T. vaginalis were compared with the Affirm VPIII Trichomonas vaginalis test. In the absence of an independent reference standard for trichomonal vaginitis (TV), a positive result in either assay was deemed to represent true infection. In the evaluable cohort of 388 patients, the sensitivity of ATV was 98.1% (53/54) versus 46.3% (25/54) for Affirm VPIII. The diagnostic accuracy of the combined NAA-based test construct was approximately 20 to 25% higher than that of the Affirm VPIII when modeled in populations with various prevalences of infectious vaginitis.     

Development and validation of a semiquantitative, multitarget PCR assay for diagnosis of bacterial vaginosis

Quantitative PCR assays were developed for 4 organisms reported previously to be useful positive indicators for the diagnosis of bacterial vaginosis (BV)--Atopobium vaginae, Bacterial Vaginosis-Associated Bacterium 2 (BVAB-2), Gardnerella vaginalis, and Megasphaera-1--and a single organism (Lactobacillus crispatus) that has been implicated as a negative indicator for BV. Vaginal samples (n = 169), classified as positive (n = 108) or negative (n = 61) for BV based on a combination of the Nugent Gram stain score and Amsel clinical criteria, were analyzed for the presence and quantity of each of the marker organisms, and the results were used to construct a semiquantitative, multiplex PCR assay for BV based on detection of 3 positive indicator organisms (A. vaginae, BVAB-2, and Megasphaera-1) and classification of samples using a combinatorial scoring system. The prototype BV PCR assay was then used to analyze the 169-member developmental sample set and, in a prospective, blinded manner, an additional 227 BV-classified vaginal samples (110 BV-positive samples and 117 BV-negative samples). The BV PCR assay demonstrated a sensitivity of 96.7% (202/209), a specificity of 92.2% (153/166), a positive predictive value of 94.0%, and a negative predictive value of 95.6%, with 21 samples (5.3%) classified as indeterminate for BV. This assay provides a reproducible and objective means of evaluating critical components of the vaginal microflora in women with signs and symptoms of vaginitis and is comparable in diagnostic accuracy to the conventional gold standard for diagnosis of BV.     

Validation of the use of Pap-stained vaginal smears for diagnosis of bacterial vaginosis

Papanicolaou-stained cervicovaginal smears (Pap smears) are used to screen for cervical cancer. Since there is a lack of consensus in published reports respecting the efficacy of Pap-stained smears in BV diagnostics, there is a need to validate their use for diagnosis of BV. Slides from the international BV00 workshop were Pap stained and independently analyzed by four investigators under a phase-contrast microscope. All workshop slides--whether Pap-stained, Gram-stained or rehydrated air-dried smears--were scored according to the same Nugent classification. The diagnostic accuracy of Pap smears for diagnosis of BV had a sensitivity of 0.85 and a specificity of 0.92, with a positive and negative predictive value of 0.84 and 0.93, respectively. The interobserver weighted kappa index was 0.86 for Pap-stained smears compared to 0.81 for Gram-stained smears, and 0.70 for rehydrated air-dried smears using the mean Nugent score as the criterion standard. Provided that the samples are taken from equivalent locations (the vaginal fornix) and analyzed according to the same scoring criteria, there is no discernable difference in the diagnostic accuracy of the three smear-staining methods. The Pap-stained vaginal smears can be used as a wholly adequate alternative to Gram-stained smears for BV diagnosis.     

Adherence of Human Vaginal Lactobacilli to Vaginal Epithelial Cells and Interaction with Uropathogens

Three strains of Lactobacillus, identified as Lactobacillus acidophilus, Lactobacillus gasseri, and Lactobacillus jensenii, were selected from among 70 isolates from the vaginas of healthy premenopausal women for properties relevant to mucosal colonization or antagonism. All three self-aggregated and adhered to epithelial vaginal cells, displacing well-known vaginal pathogens, such as G. vaginalis, and inhibiting the growth in vitro of Escherichia coli and Streptococcus agalactiae. The surface components involved in self-aggregation appeared to be proteins for L. gasseri and lipoproteins for L. acidophilus and L. jensenii, as judged by susceptibility to treatment with appropriate degrading enzymes. The factors responsible for adherence to epithelial vaginal cells seemed to be glycoproteins (L. acidophilus and L. gasseri) and carbohydrate (L. jensenii). The receptors of the vaginal cells were glycolipids, which presumably were the targets of the competition observed between the lactobacilli and the pathogenic microbes.     

BVBlue test for diagnosis of bacterial vaginosis

Bacterial vaginosis (BV) is a disorder of the vaginal ecosystem characterized by a shift in the vaginal flora from the normally predominant Lactobacillus to one dominated by sialidase enzyme-producing mixed flora. It is the most common cause of abnormal vaginal discharge in adult women. The BVBlue system (Gryphus Diagnostics, L.L.C.) is a chromogenic diagnostic test based on the presence of elevated sialidase enzyme in vaginal fluid samples. BVBlue was compared to the standard method for diagnosing BV (Amsel criteria and Nugent score). Fifty-seven nonmenstruating women of > or =16 years of age who presented for a pelvic examination were recruited. Demographic features were collected via a self-administered questionnaire. The Amsel criteria were assessed based on three of four of the following characteristics of vaginal discharge: consistency, odor, pH, and presence of clue cells on Gram stain. BVBlue was compared to the Gram stain and Amsel criteria. The sensitivity, specificity, positive predictive value, and negative predictive value for BVBlue versus the Gram stain and Amsel criteria were 91.7, 97.8, 91.7, and 97.8% and 50.0, 100, 100, and 88.2%, respectively. A significantly greater proportion of patients with a vaginal pH of >4.5, a positive amine test, or with clue cells on vaginal Gram smear were found to have a positive BVBlue test (P < 0.001). Women previously treated for BV were 2.98 times more likely to have another episode of BV. BVBlue is a useful point-of-care diagnostic tool to provide a presumptive diagnosis of BV, especially in situations where microscopic capabilities are unavailable.     

In vitro activity of secnidazole against Atopobium vaginae,an anaerobic pathogen involved in bacterial vaginosis

Bacterial vaginosis is a polymicrobial syndrome. The most important marker for bacterial vaginosis is the presence of Gardnerella vaginalis and Atopobium vaginae. In this study, the in vitro susceptibilities to metronidazole and secnidazole of 16 strains of A. vaginae were tested with the agar dilution method. We observed an MIC range for metronidazole of 4–64 mg/L (MIC₅₀, 8 mg/L; MIC₉₀, 32 mg/L) and an MIC range for secnidazole of 4–128 mg/L (MIC₅₀, 16 mg/L; MIC₉₀, 64 mg/L). According to these findings, we can conclude that the activity of secnidazole is similar to that of metronidazole.     

Evaluation and subsequent optimizations of the quantitative AmpliSens Florocenosis/Bacterial vaginosis‐FRT multiplex real‐time PCR assay for diagnosis of bacterial vaginosis

Traditional microscopy‐based methods for diagnosis of bacterial vaginosis (BV) are underutilized in many settings, and molecular techniques may provide opportunities for rapid, objective, and accurate BV diagnosis. This study evaluated the quantitative AmpliSens Florocenosis/Bacterial vaginosis‐FRT multiplex real‐time PCR (Florocenosis–BV) assay. Vaginal samples from a previous study including unselected female subjects (n = 163) and using Amsel criteria and 454 pyrosequencing for BV diagnosis were examined with the Florocenosis–BV test and additionally tested for the presence and quantity of Gardnerella vaginalis clades 3 and 4. The Florocenosis–BV assay demonstrated 100% and 98% sensitivity compared with the Amsel criteria and 454 pyrosequencing, respectively, with 91% specificity. The modified Florocenosis–BV assay (detecting also G. vaginalis clades 3 and 4) resulted in 100% sensitivity vs the Amsel criteria and 454 pyrosequencing with specificity of 86% and 88%, respectively. Further optimizations of thresholds for the quantitative parameters used in the kit resulted in 99–100% accuracy vs Amsel criteria and 454 pyrosequencing for selected parameters. The Florocenosis–BV assay is an objective, accurate, sensitive, and specific method for BV diagnosis; however, the performance of the test can be further improved with some minor optimizations.     

Effectiveness of Lactobacillus-containing vaginal tablets in the treatment of symptomatic bacterial vaginosis

The purpose of this study was to determine the effectiveness of Lactobacillus -containing vaginal tablets in the treatment of bacterial vaginosis (BV) and in the restoration of a healthy vaginal flora. Thirty-nine women with BV were enrolled in a double-blind, placebo-controlled clinical trial. Patients received either one Lactobacillus -containing tablet or placebo daily for 7 days. Clinical criteria, vaginal Gram stain scores and symptoms were compared with those at the initial visit and those at completion of therapy and 2 weeks later. After completion of therapy, all of the patients in the Lactobacillus -treated group ( n  = 18) were free of BV, showing a normal (83%) or intermediate (17%) vaginal flora, as compared with only two patients free of BV with intermediate flora (12%) from among the 16 placebo-treated women (p   <0.001). Two weeks after completion of therapy, treatment was successful (score <7) in 61% of Lactobacillus -treated patients as compared with 19% of those in the placebo group (p   <0.05). In the treatment group, the total number of symptomatic patients and the intensity of their symptoms, in particular vaginal malodour, were significantly reduced at both follow-up visits. The data indicate that intravaginal administration of exogenous selected strains of lactobacilli can restore a normal vaginal microbiota and be used in treating bacterial vaginosis.