Prevalence of Genotypes and Subtypes of Gardnerella vaginalis in South African Pregnant Women.

Background: Gardnerella vaginalis, a microorganism highly linked to bacterial vaginosis (BV), is understudied in terms of genotypic heterogeneity in South African populations. This study investigated the prevalence of G. vaginalis genotypes in BV-positive, BV-intermediate, and BV-negative South African pregnant women. Methods: The study population included n = 354 pregnant women recruited from a public hospital in Durban, South Africa. The women provided self-collected vaginal swabs for BV diagnosis by Nugent scoring. For the genotyping assays, the 16S rRNA and sialidase A genes from BV-negative, BV-intermediate, and BV-positive samples were amplified with G. vaginalis-specific primers. The16S rRNA amplicon was digested with TaqI to generate genotyping profiles, and subtypes were determined by correlating BamHI and HindIII digestion profiles. Phylogenetic analysis was performed on the 16S rRNA and sialidase A sequences. The data analysis was performed with R Statistical Computing software, version 3.6.2. Results: Two different genotypes, GT1 and GT2, were detected. The most prevalent genotype was GT1. Four subtypes (1, 2B, 2AB, and 2C) were shown to be present. The most prevalent subtype was 2B, followed by subtypes 1, 2C, and 2AB. The phylogenetic analysis of the 16S rRNA showed the presence of 5 clusters. The tree displayed clusters which contained sequences from the same BV group with different genotypes and subtypes. Clusters with sequences from across the BV groups carrying the same genotype and subtype were present. Diversity of the sialidase A across BV groups and genotypes was observed. Finally, the study did not find a significant association (p > 0.05) between reported symptoms of abnormal vaginal discharge and genotype harboured. Conclusion: This study provided the first report on the diversity of G. vaginalis in South African pregnant women. Diversity assessments of G. vaginalis with respect to genotypes and virulence factors may aid in a greater understanding of the pathogenesis of this microorganism.

Genetic Heterogeneity and Taxonomic Diversity among Gardnerella Species.

Gardnerella vaginalis has been considered a pivotal player in the progression of bacterial vaginosis (BV), a condition associated with serious health complications. However, G. vaginalis is also commonly found in asymptomatic or BV-negative women. This has generated interest in the question of whether genetic differences among isolates might distinguish pathogenic from commensal isolates. G. vaginalis was the only recognized species in its genus for four decades, but recently an emended description of G. vaginalis and descriptions of three new species - Gardnerella leopoldii, Gardnerella piotii, and Gardnerella swidsinskii - have been proposed. This review provides background on the heterogeneity and diversity within the genus Gardnerella, highlighting the main features that distinguish species and clades, and how these features may impact BV development.

Gardnerella vaginalis Clade Distribution Is Associated With Behavioral Practices and Nugent Score in Women Who Have Sex With Women.

BACKGROUND: Gardnerella vaginalis is detected in women with and without bacterial vaginosis (BV). Identification of 4 G. vaginalis clades raised the possibility that pathogenic and commensal clades exist. We investigated the association of behavioral practices and Nugent Score with G. vaginalis clade distribution in women who have sex with women (WSW). METHODS: Longitudinal self-collected vaginal specimens were analyzed using established G. vaginalis species-specific and clade-typing polymerase chain reaction assays. Logistic regression assessed factors associated with detection of G. vaginalis clades, and multinomial regression assessed factors associated with number of clades. RESULTS: Clades 1, 2, and 3 and multiclade communities (<2 clades) were associated with Nugent-BV. Clade 1 (odds ratio [OR], 3.36; 95% confidence interval [CI], 1.65-6.84) and multiclade communities (relative risk ratio [RRR], 9.51; 95% CI, 4.36-20.73) were also associated with Lactobacillus-deficient vaginal microbiota. Clade 4 was neither associated with Nugent-BV nor Lactobacillus-deficient microbiota (OR, 1.49; 95% CI, 0.67-3.33). Specific clades were associated with differing behavioral practices. Clade 1 was associated with increasing number of recent sexual partners and smoking, whereas clade 2 was associated with penile-vaginal sex and sharing of sex toys with female partners. CONCLUSIONS: Our results suggest that G. vaginalis clades have varying levels of pathogenicity in WSW, with acquisition occurring through sexual activity. These findings suggest that partner treatment may be an appropriate strategy to improve BV cure.

Resolution and Cooccurrence Patterns of Gardnerella leopoldii, G. swidsinskii, G. piotii, and G. vaginalis within the Vaginal Microbiome.

Gardnerella vaginalis is a hallmark of vaginal dysbiosis, but it is found in the microbiomes of women with and without vaginal symptoms and those who do not have microbiologically defined dysbiosis. G. vaginalis encompasses diverse taxa differing in attributes that are potentially important for virulence, and there is evidence that clades or subgroups within the species are differentially associated with clinical outcomes. The G. vaginalis species description was recently emended, and three new species within the genus were defined (G. leopoldii, G. swidsinskii, and G. piotii). 16S rRNA sequences for the four Gardnerella species are all >98.5% identical, and no signature sequences differentiate them. We demonstrated that Gardnerella species can be resolved using partial chaperonin 60 (cpn60) sequences, with pairwise percent identities of 87.1 to 97.8% among the type strains. Pairwise cooccurrence patterns of Gardnerella spp. in the vaginal microbiomes of 413 reproductive aged Canadian women were investigated, and several significant cooccurrences of species were identified. Abundance of G. vaginalis and G. swidsinskii was associated with vaginal symptoms of abnormal odor and discharge. cpn60 barcode sequencing can provide a rapid assessment of the relative abundance of Gardnerella spp. in microbiome samples, providing a powerful method of elucidating associations between these diverse organisms and clinical outcomes. Researchers should consider using cpn60 instead of 16S RNA for better resolution of these important organisms.

In Silico Analysis of Gardnerella Genomospecies Detected in the Setting of Bacterial Vaginosis.

BACKGROUND: Gardnerella vaginalis is implicated as one of the causative agents of bacterial vaginosis, but it can also be isolated from the vagina of healthy women. Previous efforts to study G. vaginalis identified 4 to 6 clades, but average nucleotide identity analysis indicates that G. vaginalis may be multiple species. Recently, Gardnerella was determined to be 13 genomospecies, with Gardnerella piottii, Gardnerella leopoldii, and Gardnerella swidsinkii delineated as separate species. METHODS: We accessed 103 publicly available genomes annotated as G. vaginalis. We performed comprehensive taxonomic and phylogenomic analysis to quantify the number of species called G. vaginalis, the similarity of their core genes, and their burden of their accessory genes. We additionally analyzed publicly available metatranscriptomic data sets of bacterial vaginosis to determine whether the newly delineated genomospecies are present, and to identify putative conserved features of Gardnerella pathogenesis. RESULTS: Gardnerella could be classified into 8 to 14 genomospecies depending on the in silico classification tools used. Consensus classification identified 9 different Gardnerella genomospecies, here annotated as GS01 through GS09. The genomospecies could be readily distinguished by the phylogeny of their shared genes and burden of accessory genes. All of the new genomospecies were identified in metatranscriptomes of bacterial vaginosis. CONCLUSIONS: Multiple Gardnerella genomospecies operating in isolation or in concert with one another may be responsible for bacterial vaginosis. These results have important implications for future efforts to understand the evolution of the Gardnerella genomospecies, host-pathogen interactions of the genomospecies during bacterial vaginosis, diagnostic assay development for bacterial vaginosis, and metagenomic investigations of the vaginal microbiota.

Comparative analysis of virulence factors & biotypes of Gardnerella vaginalis isolated from the genital tract of women with & without bacterial vaginosis.

BACKGROUND & OBJECTIVES: : Bacterial vaginosis (BV) involves the presence of a thick vaginal multispecies biofilm, where Gardnerella vaginalis is the predominant species. The reason for an increase in the number of G. vaginalis which are usually present as normal flora of the female genital tract in cases of BV, is not known. Hence, the objective of the present study was to compare the biotypes and virulence factors of G. vaginalis isolated from the genital tract of women with and without BV. METHODS: : High vaginal swabs collected from 811 women of reproductive age were cultured. G. vaginalis isolates were biotyped and tested for adherence to vaginal epithelial cells, biofilm formation, agglutination of human red blood cells (RBCs), protease production, phospholipase production and surface hydrophobicity. RESULTS: : Of the isolates from women with BV, 83.3 per cent (60/72) showed good adherence, 78.4 per cent (58/74) produced biofilm, 82.9 per cent (63/76) produced phospholipase, 67.1 per cent (51/76) produced protease, 77.3 per cent (58/75) were positive for surface hydrophobicity and 61.6 per cent (45/73) were positive for haemagglutination of human RBC. In case of G. vaginalis from non-BV women, 25 per cent (15/60) isolates showed good adherence, 18.4 per cent (9/49) biofilm production, 35 per cent (21/60) phospholipase, 36.6 per cent (22/60) protease, 41.7 per cent (25/60) surface hydrophobicity and 10.1 per cent (6/59) agglutination of human RBCs. Maximum number of isolates belonged to biotypes 6, 2 and 3. Biotype 3 was more associated with non-BV rather than BV; biotype 6, 2 and 1 were more associated with cases of BV. Maximum virulence factors were expressed by biotypes 6, 2 and 1. INTERPRETATION & CONCLUSIONS: : Virulence factors were more expressed by G. vaginalis isolates obtained from women with BV rather than from non-BV. Biotypes 6, 2 and 1 were more associated with cases of BV and expressed maximum virulence factors.

Unveiling the role of Gardnerella vaginalis in polymicrobial Bacterial Vaginosis biofilms: the impact of other vaginal pathogens living as neighbors.

Bacterial vaginosis (BV) is characterized by a highly structured polymicrobial biofilm, which is strongly adhered to the vaginal epithelium and primarily consists of the bacterium Gardnerella vaginalis. However, despite the presence of other BV-associated bacteria, little is known regarding the impact of other species on BV development. To gain insight into BV progress, we analyzed the ecological interactions between G. vaginalis and 15 BV-associated microorganisms using a dual-species biofilm model. Bacterial populations were quantified using a validated peptide nucleic acid fluorescence in situ hybridization approach. Furthermore, biofilm structure was analyzed by confocal laser scanning microscopy. In addition, bacterial coaggregation ability was determined as well as the expression of key virulence genes. Remarkably, our results revealed distinct biofilm structures between each bacterial consortium, leading to at least three unique dual-species biofilm morphotypes. Furthermore, our transcriptomic findings seem to indicate that Enterococcus faecalis and Actinomyces neuii had a higher impact on the enhancement of G. vaginalis virulence, while the other tested species had a lower or no impact on G. vaginalis virulence. This study casts a new light on how BV-associated species can modulate the virulence aspects of G. vaginalis, contributing to a better understanding of the development of BV-associated biofilms.

Emended description of Gardnerella vaginalis and description of Gardnerella leopoldii sp. nov., Gardnerella piotii sp. nov. and Gardnerella swidsinskii sp. nov., with delineation of 13 genomic species within the genus Gardnerella.

Whole genome sequence analysis (digital DNA-DNA hybridization and average nucleotide identity) was carried out for 81 sequenced full genomes of the genus Gardnerella, including ten determined in this study, and indicated the existence of 13 genomic species, of which five consist of only one strain and of which only five contain more than four sequenced genomes. Furthermore, a collection of ten Gardnerella strains, representing the emended species G. vaginalis and the newly described species Gardnerella leopoldii, Gardnerella piotii and Gardnerella swidsinskii, was studied. Matrix-assisted laser desorption ionization time-of-flight MS analysis of the protein signatures identified specific peaks that can be used to differentiate these four species. Only strains of G. vaginalis produce ß-galactosidase. We emend the description of G. vaginalis (type strain ATCC 14018T=LMG 7832T=CCUG 3717T) and describe the novel species Gardnerella leopoldii sp. nov. (UGent 06.41T=LMG 30814T=CCUG 72425T), Gardnerella piotii sp. nov. (UGent 18.01T=LMG 30818T=CCUG 72427T) and Gardnerella swidsinskii sp. nov. (GS 9838-1T=LMG 30812T=CCUG 72429T).

Increased richness and diversity of the vaginal microbiota and spontaneous preterm birth.

BACKGROUND: The bacterial community present in the female lower genital tract plays an important role in maternal and neonatal health. Imbalances in this microbiota have been associated with negative reproductive outcomes, such as spontaneous preterm birth (sPTB), but the mechanisms underlying the association between a disturbed microbiota and sPTB remain poorly understood. An intrauterine infection ascending from the vagina is thought to be an important contributor to the onset of preterm labour. Our objective was to characterize the vaginal microbiota of pregnant women who had sPTB (n = 46) and compare to those of pregnant women who delivered at term (n = 170). Vaginal swabs were collected from women at 11-16 weeks of gestational age. Microbiota profiles were created by PCR amplification and pyrosequencing of the cpn60 universal target region. RESULTS: Profiles clustered into seven community state types: I (Lactobacillus crispatus dominated), II (Lactobacillus gasseri dominated), III (Lactobacillus iners dominated), IVA (Gardnerella vaginalis subgroup B or mix of species), IVC (G. vaginalis subgroup A dominated), IVD (G. vaginalis subgroup C dominated) and V (Lactobacillus jensenii dominated). The microbiota of women who experienced preterm birth (< 37 weeks gestation) had higher richness and diversity and higher Mollicutes prevalence when compared to those of women who delivered at term. The two groups did not cluster according to CST, likely because CST assignment is driven in most cases by the dominance of one particular species, overwhelming the contributions of more rare taxa. In conclusion, we did not identify a specific microbial community structure that predicts sPTB, but differences in microbiota richness, diversity and Mollicutes prevalence were observed between groups. CONCLUSIONS: Although a causal relationship remains to be determined, our results confirm previous reports of an association between Mollicutes and sPTB and further suggest that a more diverse microbiome may be important in the pathogenesis of some cases.

Replication and refinement of a vaginal microbial signature of preterm birth in two racially distinct cohorts of US women.

Preterm birth (PTB) is the leading cause of neonatal morbidity and mortality. Previous studies have suggested that the maternal vaginal microbiota contributes to the pathophysiology of PTB, but conflicting results in recent years have raised doubts. We conducted a study of PTB compared with term birth in two cohorts of pregnant women: one predominantly Caucasian (n = 39) at low risk for PTB, the second predominantly African American and at high-risk (n = 96). We profiled the taxonomic composition of 2,179 vaginal swabs collected prospectively and weekly during gestation using 16S rRNA gene sequencing. Previously proposed associations between PTB and lower Lactobacillus and higher Gardnerella abundances replicated in the low-risk cohort, but not in the high-risk cohort. High-resolution bioinformatics enabled taxonomic assignment to the species and subspecies levels, revealing that Lactobacillus crispatus was associated with low risk of PTB in both cohorts, while Lactobacillus iners was not, and that a subspecies clade of Gardnerella vaginalis explained the genus association with PTB. Patterns of cooccurrence between L. crispatus and Gardnerella were highly exclusive, while Gardnerella and L. iners often coexisted at high frequencies. We argue that the vaginal microbiota is better represented by the quantitative frequencies of these key taxa than by classifying communities into five community state types. Our findings extend and corroborate the association between the vaginal microbiota and PTB, demonstrate the benefits of high-resolution statistical bioinformatics in clinical microbiome studies, and suggest that previous conflicting results may reflect the different risk profile of women of black race.

Gardnerella vaginalis bacteremia associated with severe acute encephalopathy in a young female patient.

Gardnerella vaginalis is a facultative anaerobic bacterium that inhabits the genitourinary tract of both healthy women and those with bacterial vaginosis. We report a case of G. vaginalis bacteremia associated with severe toxic encephalopathy in a young woman. Anaerobic blood cultures yielded pure growth of small gram-variable rods later identified as G. vaginalis by both rapid biochemical tests and 16S rRNA gene sequencing. The patient recovered after treatment with amoxicillin-clavulanate according to the in vitro susceptibility testing. The complete genome of G. vaginalis isolate from blood cultures was determined. In vitro G. vaginalis isolate produced elevated amounts of a pore-forming toxin vaginolysin compared to control G. vaginalis isolates. We hypothesize that this toxin, if produced in high amounts in blood, is able to disrupt the blood-brain barrier and exert a toxic activity on brain cells.

Gardnerella vaginalis diversity and ecology in relation to vaginal symptoms.

Gardnerella vaginalis was first described in 1953, and subsequently identified as the causative agent of a cluster of vaginal symptoms currently known as vaginosis. Research has so far failed to confirm whether and by which mechanism G. vaginalis initiates vaginosis, with, consequently, poor diagnostics and treatment outcomes. Recent molecular analyses of protein-coding genes demonstrate that the taxon G. vaginalis consists of at least four distinct species. This development may represent a critical turning point in clarifying ecological interactions and virulence factors contributing to symptoms and/or sequelae of vaginosis.

The influence of sexual activity on the vaginal microbiota and Gardnerella vaginalis clade diversity in young women.

OBJECTIVES: To examine the influence of sexual activity on the composition and consistency of the vaginal microbiota over time, and distribution of Gardnerella vaginalis clades in young women. METHODS: Fifty-two participants from a university cohort were selected. Vaginal swabs were self-collected every 3-months for up to 12 months with 184 specimens analysed. The vaginal microbiota was characterised using Roche 454 V3/4 region 16S rRNA sequencing, and G.vaginalis clade typing by qPCR. RESULTS: A Lactobacillus crispatus dominated vaginal microbiota was associated with Caucasian ethnicity (adjusted relative risk ratio[ARRR] = 7.28, 95%CI:1.37,38.57,p = 0.020). An L.iners (ARRR = 17.51, 95%CI:2.18,140.33,p = 0.007) or G.vaginalis (ARRR = 14.03, 95%CI:1.22,160.69, p = 0.034) dominated microbiota was associated with engaging in penile-vaginal sex. Microbiota dominated by L.crispatus, L.iners or other lactobacilli exhibited greater longitudinal consistency of the bacterial communities present compared to ones dominated by heterogeneous non-lactobacilli (p<0.030); sexual activity did not influence consistency. Women who developed BV were more likely to have clade GV4 compared to those reporting no sex/practiced non-coital activities (OR = 11.82, 95%CI:1.87,74.82,p = 0.009). Specimens were more likely to contain multiple G.vaginalis clades rather than a single clade if women engaged in penile-vaginal sex (RRR = 9.55, 95%CI:1.33,68.38,p = 0.025) or were diagnosed with BV (RRR = 31.5, 95%CI:1.69,586.87,p = 0.021). CONCLUSIONS: Sexual activity and ethnicity influenced the composition of the vaginal microbiota of these young, relatively sexually inexperienced women. Women had consistent vaginal microbiota over time if lactobacilli were the dominant spp. present. Penile-vaginal sex did not alter the consistency of microbial communities but increased G.vaginalis clade diversity in young women with and without BV, suggesting sexual transmission of commensal and potentially pathogenic clades.

Gardnerella vaginalis population dynamics in bacterial vaginosis.

Bacterial vaginosis (BV) is the leading cause of vaginal discharge and is associated with the facultative Gram-variable bacterium Gardnerella vaginalis, whose population structure consists of four clades. Our goal was to determine if these clades differ with regard to abundance during BV. We performed a short-term longitudinal study of BV. Patients were evaluated according to the Amsel criteria and Nugent scoring at initial diagnosis, immediately after treatment and at a 40- to 45-day follow-up visit. G. vaginalis clade abundance was determined by quantitative real-time polymerase chain reactions (qPCRs). Among all specimens, the abundance of clades 1 and 4 were higher than that of clades 2 and 3 (P < 0.001). In general, the abundance of each clade increased with the degree of vaginal dysbiosis, as determined by the Nugent score and was greater in women with Amsel 4 compared with those with Amsel 0. Only clade 1 abundance was greater when Amsel 0 or 1 specimens were compared with Amsel 2 or 3 specimens (P < 0.01). Following antimicrobial treatment, abundance of clades 1 (P < 0.001) and 4 (P < 0.05) decreased regardless of the clinical and microbiological outcome, whereas clade 2 only decreased in women who had a sustained treatment response for 40-45 days (P < 0.01). Recurrent BV was characterized by post-treatment increases of clade 1 and 2 (P < 0.01). Clades 1 and 4 predominate in vaginal specimens. Clade abundance differs with regard to the Nugent score, the Amsel criteria, and response to therapy and BV recurrence.

A molecular portrait of maternal sepsis from Byzantine Troy.

Pregnancy complications are poorly represented in the archeological record, despite their importance in contemporary and ancient societies. While excavating a Byzantine cemetery in Troy, we discovered calcified abscesses among a woman's remains. Scanning electron microscopy of the tissue revealed 'ghost cells', resulting from dystrophic calcification, which preserved ancient maternal, fetal and bacterial DNA of a severe infection, likely chorioamnionitis. Gardnerella vaginalis and Staphylococcus saprophyticus dominated the abscesses. Phylogenomic analyses of ancient, historical, and contemporary data showed that G. vaginalis Troy fell within contemporary genetic diversity, whereas S. saprophyticus Troy belongs to a lineage that does not appear to be commonly associated with human disease today. We speculate that the ecology of S. saprophyticus infection may have differed in the ancient world as a result of close contacts between humans and domesticated animals. These results highlight the complex and dynamic interactions with our microbial milieu that underlie severe maternal infections.

More Easily Cultivated Than Identified: Classical Isolation With Molecular Identification of Vaginal Bacteria.

BACKGROUND: Women with bacterial vaginosis (BV) have complex communities of anaerobic bacteria. There are no cultivated isolates of several bacteria identified using molecular methods and associated with BV. It is unclear whether this is due to the inability to adequately propagate these bacteria or to correctly identify them in culture. METHODS: Vaginal fluid from 15 women was plated on 6 different media using classical cultivation approaches. Individual isolates were identified by 16S ribosomal RNA (rRNA) gene sequencing and compared with validly described species. Bacterial community profiles in vaginal samples were determined using broad-range 16S rRNA gene polymerase chain reaction and pyrosequencing. RESULTS: We isolated and identified 101 distinct bacterial strains spanning 6 phyla including (1) novel strains with <98% 16S rRNA sequence identity to validly described species, (2) closely related species within a genus, (3) bacteria previously isolated from body sites other than the vagina, and (4) known bacteria formerly isolated from the vagina. Pyrosequencing showed that novel strains Peptoniphilaceae DNF01163 and Prevotellaceae DNF00733 were prevalent in women with BV. CONCLUSIONS: We isolated a diverse set of novel and clinically significant anaerobes from the human vagina using conventional approaches with systematic molecular identification. Several previously "uncultivated" bacteria are amenable to conventional cultivation.

Abnormal vaginal microbiota may be associated with poor reproductive outcomes: a prospective study in IVF patients.

STUDY QUESTION: What is the diagnostic performance of qPCR assays compared with Nugent scoring for abnormal vaginal microbiota and for predicting the success rate of IVF treatment? SUMMARY ANSWER: The vaginal microbiota of IVF patients can be characterized with qPCR tests which may be promising tools for diagnosing abnormal vaginal microbiota and for prediction of clinical pregnancy in IVF treatment. WHAT IS KNOWN ALREADY: Bacterial vaginosis (BV) is a common genital disorder with a prevalence of approximately 19% in the infertile population. BV is often sub-clinical with a change of the vaginal microbiota from being Lactobacillus spp. dominated to a more heterogeneous environment with anaerobic bacteria, such as Gardnerella vaginalis and Atopobium vaginae. Few studies have been conducted in infertile women, and some have suggested a negative impact on fecundity in the presence of BV. STUDY DESIGN, SIZE, DURATION: A cohort of 130 infertile patients, 90% Caucasians, attending two Danish fertility clinics for in vitro fertilization (IVF) treatment from April 2014-December 2014 were prospectively enrolled in the trial. PARTICIPANTS/MATERIALS, SETTING AND METHODS: Vaginal swabs from IVF patients were obtained from the posterior fornix. Gram stained slides were assessed according to Nugent's criteria. PCR primers were specific for four common Lactobacillus spp., G. vaginalis and A. vaginae. Threshold levels were established using ROC curve analysis. MAIN RESULTS AND THE ROLE OF CHANCE: The prevalence of BV defined by Nugent score was 21% (27/130), whereas the prevalence of an abnormal vaginal microbiota was 28% (36/130) defined by qPCR with high concentrations of Gardnerella vaginalis and/or Atopobium vaginae. The qPCR diagnostic approach had a sensitivity and specificity of respectively 93% and 93% for Nugent-defined BV. Furthermore, qPCR enabled the stratification of Nugent intermediate flora. Eighty-four patients completed IVF treatment. The overall clinical pregnancy rate was 35% (29/84). Interestingly, only 9% (2/22) with qPCR defined abnormal vaginal microbiota obtained a clinical pregnancy (P = 0.004). LIMITATIONS, REASONS FOR CAUTION: Although a total of 130 IVF patients were included in the study, a larger sample size is needed to draw firm conclusions regarding the possible adverse effect of an abnormal vaginal microbiota in relation to the clinical pregnancy rate and other reproductive outcomes. WIDER IMPLICATIONS OF THE FINDINGS: Abnormal vaginal microbiota may negatively affect the clinical pregnancy rate in IVF patients. If a negative correlation between abnormal vaginal microbiota and the clinical pregnancy rate is corroborated, patients could be screened and subsequently treated for abnormal vaginal microbiota prior to fertility treatment. STUDY FUNDING/COMPETING INTERESTS: This study was funded by The AP Møller Maersk Foundation for the advancement of Medical Science and Hospital of Central Jutland Research Fund, Denmark. No competing interests. TRIAL REGISTRATION NUMBER: The project was registered at clinicaltrials.gov (file number NCT02042352).

Identification of intrinsically metronidazole-resistant clades of Gardnerella vaginalis.

Gardnerella vaginalis is associated with bacterial vaginosis (BV), the most common cause of vaginal discharge. Metronidazole is a front-line therapy for BV, and treatment failure and recurrent disease are common problems. Whole-genome sequencing studies have revealed that G. vaginalis has a population structure that consists of 4 clades: clades 1 and 3 are associated with BV, whereas clades 2 and 4 are not. To determine if metronidazole susceptibility is associated with population structure, we analyzed 87 clinical isolates and found that metronidazole resistance (MIC ≥32 µg/mL) was highly associated with clade (P<0.0001), as 14/14 clade 3 isolates (100%) and 22/22 clade 4 isolates (100%) exhibited resistance, compared to only 16/37 clade 1 isolates (35%) and 1/14 clade 2 isolates (7.1%). The identification of intrinsically metronidazole-resistant G. vaginalis clades will facilitate future studies on the relationship between metronidazole resistance and BV treatment failure.

Development of a microarray-based tool to characterize vaginal bacterial fluctuations and application to a novel antibiotic treatment for bacterial vaginosis.

The healthy vaginal microbiota is generally dominated by lactobacilli that confer antimicrobial protection and play a crucial role in health. Bacterial vaginosis (BV) is the most prevalent lower genital tract infection in women in reproductive age and is characterized by a shift in the relative abundances of Lactobacillus spp. to a greater abundance of strictly anaerobic bacteria. In this study, we designed a new phylogenetic microarray-based tool (VaginArray) that includes 17 probe sets specific for the most representative bacterial groups of the human vaginal ecosystem. This tool was implemented using the ligase detection reaction-universal array (LDR-UA) approach. The entire probe set properly recognized the specific targets and showed an overall sensitivity of 6 to 12 ng per probe. The VaginArray was applied to assess the efficacy of rifaximin vaginal tablets for the treatment of BV, analyzing the vaginal bacterial communities of 22 BV-affected women treated with rifaximin vaginal tablets at a dosage of 25 mg/day for 5 days. Our results showed the ability of rifaximin to reduce the growth of various BV-related bacteria (Atopobium vaginae, Prevotella, Megasphaera, Mobiluncus, and Sneathia spp.), with the highest antibiotic susceptibility for A. vaginae and Sneathia spp. Moreover, we observed an increase of Lactobacillus crispatus levels in the subset of women who maintained remission after 1 month of therapy, opening new perspectives for the treatment of BV.

Recent progress in understanding the epidemiology of bacterial vaginosis.

PURPOSE OF REVIEW: Bacterial vaginosis epidemiology has been transformed by new theoretical insights and methodologies, such as molecular sequencing. We summarize the progress made in these domains. RECENT FINDINGS: The vaginal microbiome can be classified in five to eight clusters. Bacterial vaginosis-type clusters typically constitute one of these clusters, but in higher risk women, it can constitute up to three clusters. The vaginal microbiomes may be fairly stable or be subject to rapid changes in their constitutive makeup. Bacterial vaginosis does not appear to be a single entity. Certain bacterial communities are associated with particular symptoms of bacterial vaginosis that are paired with unique adverse outcomes. Biofilm-producing Gardnerella vaginalis are likely to play an important role in initiating the structured polymicrobial biofilm that is a hallmark of bacterial vaginosis. SUMMARY: Longitudinal studies currently underway should help elucidate how to best define bacterial vaginosis and its subtypes. Risk factors and outcomes associated with particular bacterial vaginosis subtypes should also be further clarified through these studies.