Log (Lactobacillus crispatus/ Gardnerella vaginalis): a new indicator of diagnosing bacterial vaginosis.

To explore a new marker which can detect bacterial vaginosis (BV) with high sensitivity and specificity quantitatively. According to the Nugent Score, vaginal secretions from study participants were divided into BV, healthy, and BV-intermediate groups. First, we compared the obvious differences and high abundance of bacteria in the three groups using 16S rRNA-sequencing, and screened out candidate markers. Then, quantitative detection of these candidate markers from the three groups was done using real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR), followed by evaluation of the sensitivity and specificity. Finally, we verified the new markers using clinical cases. Gardnerella vaginalis, Atopobium vaginae, Lactobacillus, Megasphaera were screened out by 16S rRNA-sequencing. RT-qPCR data were transformed and analyzed through ROC curves. PCR results for these bacteria were log-transformed using Lactobacillus crispatus as the numerator and other BV-related bacteria as the denominator. Four new indicators were found. Of these, log L. crispatus/G. vaginalis (L/G) <0 was the best indicator. The sensitivity, specificity, positive predictive value, and negative predictive value of our system were 93.5%, 97.2%, 96.6 and 94.6%, respectively. Combination of data for 16S rRNA-sequencing and RT-qPCR revealed four indicators for BV detection. Of these, log L/G < 0 was the best indicator. Creating a molecular-diagnostic system independent of the Nugent Score for BV could have an important impact on the clinical management of BV.Abbreviation: log L. crispatus/G. vaginalis (logL/G); Bacterial vaginosis (BV); vaginal secretions (VSs); polymerase chain reaction (PCR); rRNA-sequencing (rRNA-seq); real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR); operational taxonomic unit (OTU); non-metric multidimensional scaling (NMDS); receiver operating characteristic (ROC).

Quantification of Gardnerella vaginalis, Atopobium vaginae and Lactobacillus spp. in bacterial vaginosis.

INTRODUCTION: The aim of the study was to investigate prevalence of bacteria most frequently associated with bacterial vaginosis using Amsel's criteria as well as to quantify these bacteria by real-time PCR and to explore the difference in their quantity between healthy and bacterial vaginosis samples. METHODOLOGY: For classification of vaginal discharge samples Amsel's criteria have been used. To detect and quantify Gardnerella vaginalis Atopobium vaginae, Lactobacillus spp. and total vaginal microbiome, real-time PCR has been applied. RESULTS: According to results of our study Amsel's criteria matched well with real-time PCR diversification of healthy women and women with BV. Nevertheless, real-time PCR has been more sensitive in diagnosis of bacterial vaginosis. DNA quantification of bacteria demonstrated that mutual abundance of G.vaginalis and A. vaginae was good bacterial vaginosis marker . On the contrary, Lactobacillus spp. was present in high amount in both healthy and bacterial vaginosis samples, but ratio of investigated bacteria was different between them. In fact, G. vaginalis and A. vaginae comprised only 0.1% of total microbiome in healthy, whereas Lactobacillus spp. took 99.3% of it. Nonetheless, in bacterial vaginosis, G. vaginalis and A. vaginae made up 34.4% of total microbiome, while Lactobacillus spp. was 21.6%. CONCLUSIONS: According to the results of our study real-time PCR analysis was more sensitive in diagnosis of bacterial vaginosis than Amsel's method, as well as it represented fine tool in making a difference between microbial entities in healthy and bacterial vaginosis samples.

The vaginal microbiota composition of women undergoing assisted reproduction: a prospective cohort study.

OBJECTIVE: To evaluate the impact of vaginal microbiota on pregnancy outcomes in women undergoing assisted reproduction. DESIGN: A prospective cohort study. SETTING: A university-based assisted reproductive technology (ART) centre. POPULATION: 223 women undergoing ART treatment. METHODS: Prior to embryo transfer, vaginal samples were collected from the posterior fornix. Vaginal microbiota identification was carried out using next-generation sequencing and categorised according to the V3-V4 hypervariable region in the 16S rRNA gene region. MAIN OUTCOME MEASURES: ART clinical outcomes (implantation, clinical pregnancy rates and live birth rates). RESULTS: The live birth rate in women with community state type (CST)-I (39%) was higher than that in women with CST-III (21.5%) but the difference was not statistically significant (P = 0.052). The relative abundance of Lactobacillus was lower in women who failed to become pregnant (NP group) (67.71%) than in women who became pregnant (PR group) (79.72%). However, this difference was not statistically significant (P = 0.06). In the NP group, the relative abundance of Streptococcus (7.81%) and Gardnerella (9.40%) was higher than that in the PR group (relative abundance of Streptococcus and Gardnerella was 2.28% and 5.56%, respectively). The abundance of Streptococcus was found to be statistically significantly different between the two study groups (P = 0.014). Linear discriminant analysis (LDA) further validated that Streptococcus had the highest contribution (LDA score >4.0) to the difference between these two groups. CONCLUSIONS: Streptococcus has the highest contribution to the distinction between the PR and NP groups. TWEETABLE ABSTRACT: A relatively high abundance of Streptococcus in the vaginal microbiota may be associated with a lower ART success rate.

Association of Vaginal Microbiota With Signs and Symptoms of the Genitourinary Syndrome of Menopause Across Reproductive Stages.

The genitourinary syndrome of menopause (GSM) describes signs and symptoms resulting from effects of estrogen deficiency on the female genitourinary tract, including the vagina, labia, urethra, and bladder. Signs/symptoms associated with GSM may occur during any reproductive stage from multiple etiologies but are most common during menopause due to low estrogen. Vaginal microbiota, particularly Lactobacillus spp., are beneficial to the female genital tract; however, their abundance declines during menopause. We aimed to longitudinally assess vaginal microbiota characterized by 16S rRNA gene amplicon sequencing and GSM-associated endpoints across reproductive stages. In a 2-year cohort study of 750 women aged 35-60 years at enrollment and 2 111 semiannual person-visits, low-Lactobacillus vaginal microbiota communities were observed at 21.2% (169/798), 22.9% (137/597), and 49.7% (356/716) of person-visits among pre-, peri-, and postmenopausal women, respectively (p < .001). Compared to communities that have high Gardnerella vaginalis relative abundance and diverse anaerobes, the following communities were associated with a lower covariate-adjusted odds of vaginal atrophy: L crispatus-dominated communities among postmenopausal women (odds ratio [OR] = 0.25; 95% confidence interval [CI]: 0.08, 0.81), L gasseri/L jensenii (OR = 0.21; 95% CI: 0.05, 0.94) and L iners (OR = 0.21; 95% CI: 0.05, 0.85) among perimenopausal women, and L iners-dominated communities (OR = 0.18; 95% CI: 0.04, 0.76) among premenopausal women. Postmenopausal women with L gasseri/L jensenii-dominated communities had the lowest odds of vaginal dryness (OR = 0.36; 95% CI: 0.12, 1.06) and low libido (OR = 0.28; 95% CI: 0.10, 0.74). Findings for urinary incontinence were inconsistent. Associations of vaginal microbiota with GSM signs/symptoms are most evident after menopause, suggesting an avenue for treatment and prevention.

Detection of co-infection of Gardnerella vaginalis and Atopobium vaginae using qualitative PCR: A better predictor of bacterial vaginosis.

The present study aimed to determine the utility of detection of co-infection of Gardnerella vaginalis and Atopobium vaginae using qualitative PCR for diagnosing bacterial vaginosis (BV). Vaginal samples (n = 385) categorized as positive (n = 108) or negative (n = 208) for bacterial vaginosis based on the Nugent scoring system, were analyzed for the presence of G. vaginalis and A. vaginae by conventional PCR. We compared the sensitivity, specificity, positive predictive value, negative predictive value and odds ratio for the detection of each bacterium alone with the combination of the two bacteria for diagnosing BV. The detection of co-infection of the two bacteria demonstrated a sensitivity of 96%, a specificity of 82.9%, a positive predictive value of 68.5%, a negative predictive value of 98.2% with an odds ratio of 116 (CI -32 - 409). In our study, we found a high sensitivity, specificity, negative predictive value and odds ratio for the detection of co-infection of A. vaginae and G. vaginalis for the diagnosis of BV.

Influence of Lactobacillus crispatus, Lactobacillus iners and Gardnerella vaginalis on bacterial vaginal composition in pregnant women.

PURPOSE: To investigate associations between bacterial species in the vagina in mid-trimester pregnant women from Brazil. METHODS: The vaginal microbiome in 613 subjects was identified by analysis of the V1-V3 region of bacterial 16S ribosomal RNA and the relative prevalence of individual bacteria were determined. RESULTS: The bacterial species present in the greatest number of women were Lactobacillus crispatus (306 women), L. iners (298 women) and Gardnerella vaginalis (179 women). When present in the vagina, L. crispatus was the most abundant bacterium more than 85% of the time. In contrast, L. iners and G. vaginalis were most abundant in 63% and 41% of women who were positive for these microorganisms, respectively (p < 0.0001 vs. L. crispatus). The proportion of L. crispatus was negatively associated with the proportions of L. iners, L. jensenii, L. gasseri, G. vaginalis, Megasphaera, Atopobium vaginae and Prevotella (p < 0.0001). In contrast, the proportion of G. vaginalis was positively associated with levels of Megasphaera, A. vaginae and Prevotella (p < 0.0001) while L. iners proportion was unrelated to the proportion of L. jensenii, G. vaginalis, Megasphaera, A. vaginae or Prevotella. CONCLUSION: The composition of the vaginal microbiota in mid-trimester pregnant women is influenced by the relative concentrations of L. crispatus, L. iners and G. vaginalis.

Clinical chorioamnionitis at term X: microbiology, clinical signs, placental pathology, and neonatal bacteremia - implications for clinical care.

OBJECTIVES: Clinical chorioamnionitis at term is considered the most common infection-related diagnosis in labor and delivery units worldwide. The syndrome affects 5-12% of all term pregnancies and is a leading cause of maternal morbidity and mortality as well as neonatal death and sepsis. The objectives of this study were to determine the (1) amniotic fluid microbiology using cultivation and molecular microbiologic techniques; (2) diagnostic accuracy of the clinical criteria used to identify patients with intra-amniotic infection; (3) relationship between acute inflammatory lesions of the placenta (maternal and fetal inflammatory responses) and amniotic fluid microbiology and inflammatory markers; and (4) frequency of neonatal bacteremia. METHODS: This retrospective cross-sectional study included 43 women with the diagnosis of clinical chorioamnionitis at term. The presence of microorganisms in the amniotic cavity was determined through the analysis of amniotic fluid samples by cultivation for aerobes, anaerobes, and genital mycoplasmas. A broad-range polymerase chain reaction coupled with electrospray ionization mass spectrometry was also used to detect bacteria, select viruses, and fungi. Intra-amniotic inflammation was defined as an elevated amniotic fluid interleukin-6 (IL-6) concentration ≥2.6 ng/mL. RESULTS: (1) Intra-amniotic infection (defined as the combination of microorganisms detected in amniotic fluid and an elevated IL-6 concentration) was present in 63% (27/43) of cases; (2) the most common microorganisms found in the amniotic fluid samples were Ureaplasma species, followed by Gardnerella vaginalis; (3) sterile intra-amniotic inflammation (elevated IL-6 in amniotic fluid but without detectable microorganisms) was present in 5% (2/43) of cases; (4) 26% of patients with the diagnosis of clinical chorioamnionitis had no evidence of intra-amniotic infection or intra-amniotic inflammation; (5) intra-amniotic infection was more common when the membranes were ruptured than when they were intact (78% [21/27] vs. 38% [6/16]; p=0.01); (6) the traditional criteria for the diagnosis of clinical chorioamnionitis had poor diagnostic performance in identifying proven intra-amniotic infection (overall accuracy, 40-58%); (7) neonatal bacteremia was diagnosed in 4.9% (2/41) of cases; and (8) a fetal inflammatory response defined as the presence of severe acute funisitis was observed in 33% (9/27) of cases. CONCLUSIONS: Clinical chorioamnionitis at term, a syndrome that can result from intra-amniotic infection, was diagnosed in approximately 63% of cases and sterile intra-amniotic inflammation in 5% of cases. However, a substantial number of patients had no evidence of intra-amniotic infection or intra-amniotic inflammation. Evidence of the fetal inflammatory response syndrome was frequently present, but microorganisms were detected in only 4.9% of cases based on cultures of aerobic and anaerobic bacteria in neonatal blood.

Rare case of osteomyelitis caused by Gardnerella vaginalis and Streptococcus parasanguinis in a postmenopausal woman.

Vertebral osteomyelitis is an infection of the vertebrae that can lead to spinal degeneration, most commonly caused by Staphylococcus aureus Here, we report an unusual case of pyogenic osteomyelitis caused by Gardnerella vaginalis and Streptococcus parasanguinis in a 61-year-old postmenopausal woman. The patient presented with a 2-week history of worsening lower back pain and fever and a recent episode of cystitis following re-engagement of sexual activity. Imaging revealed a deterioration of vertebrae discs and spinal canal stenosis at the L3-L4 levels with a formation of abscess in the right psoas muscle. Needle aspiration of the abscess identified G. vaginalis and S. parasanguinis and the patient was successfully treated with a 6-week course of ceftriaxone and metronidazole. This case describes an unusual coinfection of two pathogens that normally reside in the urogenital tract and oral cavity, respectively, and highlights the risk posed when these organisms breach the body's normal barriers.

Cervical Gardnerella vaginalis in women with preterm prelabor rupture of membranes.

OBJECTIVE: To determine the association between microbial invasion of the amniotic cavity (MIAC) and/or intra-amniotic inflammation (IAI) and the cervical prevalence of Gardnerella vaginalis DNA in pregnancies with preterm prelabor rupture of membrane (PPROM). METHOD: In total, 405 women with singleton pregnancies complicated with PPROM were included. Cervical fluid and amniotic fluid samples were collected at the time of admission. Bacterial and G. vaginalis DNA were assessed in the cervical fluid samples using quantitative PCR technique. Concentrations of interleukin-6 and MIAC were evaluated in the amniotic fluid samples. Loads of G. vaginalis DNA ≥ 1% of the total cervical bacterial DNA were used to define the cervical prevalence of G. vaginalis as abundant. Based on the MIAC and IAI, women were categorized into four groups: with intra-amniotic infection (both MIAC and IAI), with sterile IAI (IAI without MIAC), with MIAC without IAI, and without either MIAC or IAI. RESULTS: The presence of the abundant cervical G. vaginalis was related to MIAC (with: 65% vs. without: 44%; p = 0.0004) but not IAI (with: 52% vs. without: 48%; p = 0.70). Women with MIAC without IAI had the highest load of the cervical G. vaginalis DNA (median 2.0 × 104 copies DNA/mL) and the highest presence of abundant cervical G. vaginalis (73%). CONCLUSIONS: In women with PPROM, the presence of cervical G. vaginalis was associated with MIAC, mainly without the concurrent presence of IAI.

A specific bacterial DNA signature in the vagina of Australian women in midpregnancy predicts high risk of spontaneous preterm birth (the Predict1000 study).

BACKGROUND: Intrauterine infection accounts for a quarter of the cases of spontaneous preterm birth; however, at present, it is not possible to efficiently identify pregnant women at risk to deliver preventative treatments. OBJECTIVE: This study aimed to establish a vaginal microbial DNA test for Australian women in midpregnancy that will identify those at increased risk of spontaneous preterm birth. STUDY DESIGN: A total of 1000 women with singleton pregnancies were recruited in Perth, Australia. Midvaginal swabs were collected between 12 and 23 weeks' gestation. DNA was extracted for the detection of 23 risk-related microbial DNA targets by quantitative polymerase chain reaction. Obstetrical history, pregnancy outcome, and demographics were recorded. RESULTS: After excluding 64 women owing to losses to follow-up and insufficient sample for microbial analyses, the final cohort consisted of 936 women of predominantly white race (74.3%). The overall preterm birth rate was 12.6% (118 births); the spontaneous preterm birth rate at <37 weeks' gestation was 6.2% (2.9% at ≤34 weeks' gestation), whereas the preterm premature rupture of the membranes rate was 4.2%. No single individual microbial target predicted increased spontaneous preterm birth risk. Conversely, women who subsequently delivered at term had higher amounts of Lactobacillus crispatus, Lactobacillus gasseri, or Lactobacillus jensenii DNA in their vaginal swabs (13.8% spontaneous preterm birth vs 31.2% term; P=.005). In the remaining women, a specific microbial DNA signature was identified that was strongly predictive of spontaneous preterm birth risk, consisting of DNA from Gardnerella vaginalis (clade 4), Lactobacillus iners, and Ureaplasma parvum (serovars 3 and 6). Risk prediction was improved if Fusobacterium nucleatum detection was included in the test algorithm. The final algorithm, which we called the Gardnerella Lactobacillus Ureaplasma (GLU) test, was able to detect women at risk of spontaneous preterm birth at <37 and ≤34 weeks' gestation, with sensitivities of 37.9% and 44.4%, respectively, and likelihood ratios (plus or minus) of 2.22 per 0.75 and 2.52 per 0.67, respectively. Preterm premature rupture of the membranes was more than twice as common in GLU-positive women. Adjusting for maternal demographics, ethnicity, and clinical history did not improve prediction. Only a history of spontaneous preterm birth was more effective at predicting spontaneous preterm birth than a GLU-positive result (odds ratio, 3.6). CONCLUSION: We have identified a vaginal bacterial DNA signature that identifies women with a singleton pregnancy who are at increased risk of spontaneous preterm birth and may benefit from targeted antimicrobial therapy.

Associação entre Vaginose Bacteriana e Anormalidades Citológicas nos Exames Citopatológicos Analisados em um Laboratório Escola de Goiânia-GO / Association Between Bacterial Vaginosis and Cytological Abnormalities in Cytopathological Exams Analyzed in a Laboratory School of Goiânia-GO / Asociación entre Vaginosis Bacteriana y Anormalidades Citológicas en Exámenes Citopatológicos Analizados en una Escuela de Laboratorio de Goiânia-GO

Introdução: A Gardnerella vaginalis facilita a infecção pelo papilomavírus humano (HPV). Objetivo: Verificar a associação entre anormalidades citológicas e presença de Gardnerella vaginalis nos esfregaços cervicovaginais encaminhados ao Laboratório Clínico da Pontifícia Universidade Católica de Goiás (LAC/PUC Goiás) estratificadas por faixa etária. Método: Estudo transversal realizado no LAC/PUC Goiás entre janeiro de 2013 a dezembro de 2015. Para análises estatísticas, a variável idade foi categorizada em ≤39 anos e >40 anos, utilizando o programa IBM SPSS Statistics (Version 2.0, 2011®) para o teste de qui-quadrado (X²), com intervalo de confiança de 95% e valor p<0,05. Resultados: Foram analisados 4.558 exames citopatológicos, a maioria com presença de Lactobacillus spp. (46,97%). A prevalência dos agentes patogênicos foi a Gardnerella vaginalis (79,6%), seguida de Candida spp. (16,8%), Trichomonas vaginalis (2,2%), Herpes simplex (0,4 %) e Chlamydia trachomatis (0,1%). As anormalidades citológicas foram observadas em 9,1%, sendo atypical squamous cells of undetermined significance (ASC-US) 2,57%, low grade squamous intraepithelial lesion (LSIL) 1,78%, atypical squamous cells of undetermined significance cannot exclude high grade squamous intraepithelial lesion (ASC-H) 3,52%, high grade squamous intraepithelial lesion (HSIL) 1,08%, atypical endocervical cells, favor neoplastic (AGC-NEO) 0,22% e carcinoma 0,02%. Houve uma associação significante entre anormalidades citológicas graves e mulheres ≥40 anos, OR 3,01 (IC 95% 2,0-4,58) (p<0,0001). Mulheres ≤40 anos mostraram significância à presença de Gardnerella vaginalis (p<0,0004). Conclusão: Uma elevada prevalência de Gardnerella vaginalis foi encontrada associada com as anormalidades citológicas, principalmente em mulheres sexualmente ativas.

Introduction:Gardnerella vaginalis facilitates human papillomavirus (HPV) infection. Objective: To verify the association between cytological abnormalities and the presence of Gardnerella vaginalis in cervicovaginal smears sent to the Clinical Laboratory of the Pontifical Catholic University of Goiás (LAC/PUC Goiás) stratified by age range. Method: Cross-sectional study carried out at LAC/PUC Goiás from January 2013 to December 2015. For statistical analysis, the variable age was categorized as ≤39 years and >40 years, using the IBM SPSS Statistics program (Version 2.0, 2011®) for the chi-square test (X²), with a 95% confidence interval and p<0.05. Results:4,558 cytopathological exams were analyzed, most of them with the presence of Lactobacillus spp (46.97%). The prevalence of pathogens was Gardnerella vaginalis (79.6%), followed by Candida spp. (16.8%), Trichomonas vaginalis (2.2%), Herpes simplex (0.4%) and Chlamydia trachomatis (0.1%). Cytological abnormalities were observed in 9.1%, being atypical squamous cells of undetermined significance (ASC-US) 2.57%, low grade squamous intraepithelial lesion (LSIL) 1.78%, atypical squamous cells of undetermined significance cannot exclude high intraepithelial lesion (ASC-H) 3.52%, high grade squamous intraepithelial lesion (HSIL) 1.08%, atypical endocervical cells, neoplastic favor (AGC-NEO) 0.22% and carcinoma 0.02%. There was a significant association between severe cytological abnormalities and women >40 years old OR 3.01 (95% CI 2.0-4.58) (p<0.0001). Women ≤40 years old showed the presence of Gardnerella vaginalis (p<0.0004). Conclusion:A high prevalence of Gardnerella vaginalis was found and its association with cytological abnormalities, especially in sexually active women.

Introducción:Gardnerella vaginalis facilita la infección por el virus del papiloma humano (VPH). Objetivo: Verificar la asociación entre anormalidades citológicas y la presencia de Gardnerella vaginalis en frotis cervicovaginales enviadas al Laboratorio Clínico de la Pontificia Universidad Católica de Goiás (LAC/PUC Goiás) estratificadas por grupo de edad. Método: Estudio transversal realizado en LAC/PUC Goiás desde enero de 2013 hasta diciembre de 2015. Para el análisis estadístico, la edad variable se clasificó como ≤39 años y >40 años, utilizando el programa IBM SPSS Statistics (Versión 2.0, 2011®) para la prueba de chi-cuadrado (X²), con un intervalo de confianza del 95% y p <0,05. Resultados: Se analizaron 4.558 exámenes citopatológicos. La prevalencia de Lactobacillusspp. con 46,97%. Los patógenos como Gardnerella vaginalis fueron 79,6%, Candidaspp. 16,8%, Trichomonas vaginalis 2,2%, Herpes simplex 0,4%, y Chlamydia trachomatis 0,1%. Se observaron anormalidades citológicas en 9,1%, con células escamosas atípicas de significado indeterminado (ASC-US) 2,57%, lesión intraepitelial escamosa de bajo grado (LSIL) 1,78%, células escamosas atípicas de significación indeterminada no pueden excluir lesión intraepitelial (ASC-H) 3,52%, lesión intraepitelial escamosa de alto grado (HSIL) 1,08%, células endocervicales atípicas, favor neoplásico (AGC-NEO) 0,22% y carcinoma 0,02%. Hubo una asociación significativa entre anormalidades citológicas severas y mujeres >40 años OR 3,01 (IC 95% 2,0-4,58) (p<0,0001). Las mujeres ≤40 años mostraron la presencia de Gardnerella vaginalis (p<0,0004). Conclusión: Se encontró una alta prevalencia de Gardnerella vaginalis y su asociación con anomalías citológicas, especialmente en mujeres sexualmente activas.

Prevalence and factors associated with Trichomonas vaginalis infection in indigenous Brazilian women.

There is a scarcity of studies on the prevalence of Trichomonas vaginalis (TV) in indigenous populations of Brazil. We conducted a cross-sectional study between January and December 2018, on indigenous women living nearby an urban center of the Midwest region of Brazil and determined the prevalence of TV. Factors associated with TV infection and a comparison of molecular and direct microscopy diagnoses were determined. 241 indigenous women aged above 18 years participated in the study. Cervical and vaginal brush samples were collected to diagnose TV through polymerase chain reaction (PCR). Direct microscopy for detection of TV, and cellular changes was performed. A sociodemographic and behavioral questionnaire was applied at the beginning of the study. All the data were analyzed using Statistical Package for the Social Sciences. The result obtained showed that 27.8% [95% CI: 22.2-33.9] were positive for TV on PCR, while 7.41% [95% CI: 4.1-11] showed positive on direct microscopy. Direct microcopy also found 21 (8.71%) and 8 (3.31%) women infected with Gardnerella vaginalis and Candida albicans, respectively. In addition, 10 women presented atypical squamous cells of unknown significance and 14 lesions suggestive of HPV. Single women, under the age of 30 and who do not use condoms, were found to have a greater chance of getting TV infection. The high prevalence TV found in this population is comparable to highly vulnerable populations, as prisoners, sex workers and women in regions with low socioeconomic levels, moreover, seems to be an underdiagnosis of this infection. Therefore, a routine test program, as well as a review of the diagnostic method used, is encouraged for proper management.

Glycan cross-feeding supports mutualism between Fusobacterium and the vaginal microbiota.

Women with bacterial vaginosis (BV), an imbalance of the vaginal microbiome, are more likely to be colonized by potential pathogens such as Fusobacterium nucleatum, a bacterium linked with intrauterine infection and preterm birth. However, the conditions and mechanisms supporting pathogen colonization during vaginal dysbiosis remain obscure. We demonstrate that sialidase activity, a diagnostic feature of BV, promoted F. nucleatum foraging and growth on mammalian sialoglycans, a nutrient resource that was otherwise inaccessible because of the lack of endogenous F. nucleatum sialidase. In mice with sialidase-producing vaginal microbiotas, mutant F. nucleatum unable to consume sialic acids was impaired in vaginal colonization. These experiments in mice also led to the discovery that F. nucleatum may also "give back" to the community by reinforcing sialidase activity, a biochemical feature of human dysbiosis. Using human vaginal bacterial communities, we show that F. nucleatum supported robust outgrowth of Gardnerella vaginalis, a major sialidase producer and one of the most abundant organisms in BV. These results illustrate that mutually beneficial relationships between vaginal bacteria support pathogen colonization and may help maintain features of dysbiosis. These findings challenge the simplistic dogma that the mere absence of "healthy" lactobacilli is the sole mechanism that creates a permissive environment for pathogens during vaginal dysbiosis. Given the ubiquity of F. nucleatum in the human mouth, these studies also suggest a possible mechanism underlying links between vaginal dysbiosis and oral sex.

Comparison between a novel molecular tool and conventional methods for diagnostic classification of bacterial vaginosis: is integration of the two approaches necessary for a better evaluation?

The etiological cause of bacterial vaginosis (BV) is the change of the vaginal ecosystem characterized by a decrease of lactobacilli and an increase of other germs, such as Gardnerella vaginalis and Atopobium vaginae. Molecular tools have revolutionized the diagnosis of these conditions. The aim of this paper was to compare results obtained from 158 vaginal swabs collected from women aged between 18 and 59 years old and subjected to microscopic evaluation (Nugent Score), culture and to the multiparametric molecular assay Vaginitis and Vaginosis Multiplex-Tandem (MT) PCR (AU27117) - Nuclear Laser Medicine. In 50 samples we also used matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for bacterial microbiome identification. Our results showed a moderate concordance between traditional and molecular methods for diagnosis of candidiasis and a lower concordance for BV and normal flora. MALDI TOF MS allowed us to discriminate more than 10 species of lactobacilli with a greater abundance of Lactobacillus gasseri, Lactobacillus paracasei spp. paracasei, Lactobacillus pentosus and Lactobacillus crispatus in BV and altered flora. This work underlined how the integration of different assays and metagenomics studies can greatly expand our current understanding of vaginal microbial diversity, providing more reliable diagnostic criteria for BV and its intermediate condition diagnosis.

Changes in the vaginal microbiota following antibiotic treatment for Mycoplasma genitalium, Chlamydia trachomatis and bacterial vaginosis.

The human vagina harbor a rich microbiota. The optimal state is dominated by lactobacilli that help to maintain health and prevent various diseases. However, the microbiota may rapidly change to a polymicrobial state that has been linked to a number of diseases. In the present study, the temporal changes of the vaginal microbiota in patients treated for sexually transmitted diseases or bacterial vaginosis (BV) and in untreated controls were studied for 26 days. The patients included 52 women treated with azithromycin, tetracyclines or moxifloxacin for present or suspected infection with Chlamydia trachomatis or Mycoplasma genitalium. Women with concurrent BV were also treated with metronidazole. The controls were 10 healthy women of matching age. The microbiota was analyzed by 16S rRNA gene deep sequencing, specific qPCRs and microscopy. There was generally good correlation between Nugent score and community state type (CST) and qPCR confirmed the sequencing results. By sequencing, more than 600 different taxa were found, but only 33 constituted more than 1 ‰ of the sequences. In both patients and controls the microbiota could be divided into three different community state types, CST-I, CST-III and CST-IV. Without metronidazole, the microbiota remained relatively stable regarding CST although changes were seen during menstrual periods. Administration of metronidazole changed the microbiota from CST-IV to CST-III in approximately 50% of the treated patients. In contrast, the CST was generally unaffected by azithromycin or tetracyclines. In 30% of the BV patients, Gardnerella vaginalis was not eradicated by metronidazole. The majority of women colonized with Ureaplasma parvum remained positive after azithromycin while U. urealyticum was eradicated.

Production and characterization of a monoclonal antibody against the sialidase of Gardnerella vaginalis using a synthetic peptide in a MAP8 format.

Bacterial vaginosis is one of the most frequent vaginal infections. Its main etiological agent is Gardnerella vaginalis, which produces several virulence factors involved in vaginal infection and colonization, in particular, sialidase (SLD), a potential clinical biomarker that participates in immune response modulation and mucus degradation. The main objective of this work was the production and evaluation of a monoclonal antibody against G. vaginalis sialidase and its validation in immunoassays. For immunization of mice, a synthetic multiantigenic peptide was used, and hybridomas were generated. After fusion, hybridomas were evaluated for antibody production and cloned by limited dilution. One clone producing IgG1 was selected and characterized by indirect ELISA, dot blot, and Western blot, and we also tested clinical isolates and HeLa cells infected with G. vaginalis. The results showed that the anti-SLD antibody recognized a single protein of ~90 kDa that correlated with the estimated molecular weight of SLD. In addition, anti-SLD antibody recognized SLD from complete bacteria and from culture supernatants of infected Hela cells. In conclusion, our results showed that the anti-SLD antibody recognized SLD from different sources and could be considered a new tool for the diagnosis of bacterial vaginosis. KEY POINTS: • Anti-sialidase mAb was generated using a synthetic peptide • The mAb recognizes synthetic peptide and intact protein from multiple sources • The antibody was characterized by several immunological methods.

Bladder bacterial diversity differs in continent and incontinent women: a cross-sectional study.

BACKGROUND: Since the discovery of the bladder microbiome (urobiome), interest has grown in learning whether urobiome characteristics have a role in clinical phenotyping and provide opportunities for novel therapeutic approaches for women with common forms of urinary incontinence. OBJECTIVE: This study aimed to test the hypothesis that the bladder urobiome differs among women in the control cohort and women affected by urinary incontinence by assessing associations between urinary incontinence status and the cultured urobiome. STUDY DESIGN: With institutional review board oversight, urine specimens from 309 adult women were collected through transurethral catheterization. These women were categorized into 3 cohorts (continent control, stress urinary incontinence [SUI], and urgency urinary incontinence [UUI]) based on their responses to the validated Pelvic Floor Distress Inventory (PFDI) questionnaire. Among 309 women, 150 were in the continent control cohort, 50 were in the SUI cohort, and 109 were in the UUI cohort. Symptom severity was assessed by subscale scoring with the Urinary Distress Inventory (UDI), subscale of the Pelvic Floor Distress Inventory. Microbes were assessed by expanded quantitative urine culture protocol, which detects the most common bladder microbes (bacteria and yeast). Microbes were identified to the species level by matrix-assisted laser desorption and ionization time-of-flight mass spectrometry. Alpha diversity indices were calculated for culture-positive samples and compared across the 3 cohorts. The correlations of UDI scores, alpha diversity indices, and species abundance were estimated. RESULTS: Participants had a mean age of 53 years (range 22-90); most were whites (65%). Women with urinary incontinence were slightly older (control, 47; SUI, 54; UUI, 61). By design, UDI symptom scores differed (control, 8.43 [10.1]; SUI, 97.95 [55.36]; UUI, 93.71 [49.12]; P<.001). Among 309 participants, 216 (70%) had expanded quantitative urine culture-detected bacteria; furthermore, the urinary incontinence cohorts had a higher detection frequency than the control cohort (control, 57%; SUI, 86%; UUI, 81%; P<.001). In addition, the most frequently detected species among the cohorts were as follows: continent control, Lactobacillus iners (12.7%), Streptococcus anginosus (12.7%), L crispatus (10.7%), and L gasseri (10%); SUI, S anginosus (26%), L iners (18%), Staphylococcus epidermidis (18%), and L jensenii (16%); and UUI, S anginosus (30.3%), L gasseri (22%), Aerococcus urinae (18.3%), and Gardnerella vaginalis (17.4%). However, only Actinotignum schaalii (formerly Actinobaculum schaalii), A urinae, A sanguinicola, and Corynebacterium lipophile group were found at significantly higher mean abundances in 1 of the urinary incontinence cohorts when compared with the control cohort (Wilcoxon rank sum test; P<.02), and no individual genus differed significantly between the 2 urinary incontinence cohorts. Both urinary incontinence cohorts had increased alpha diversity similar to continent control cohort with indices of species richness, but not evenness, strongly associated with urinary incontinence. CONCLUSION: In adult women, the composition of the culturable bladder urobiome is associated with urinary incontinence, regardless of common incontinence subtype. Detection of more unique living microbes was associated with worsening incontinence symptom severity. Culturable species richness was significantly greater in the urinary incontinence cohorts than in the continent control cohort.

Molecular epidemiology and socio-demographic risk factors of sexually transmitted infections among women in Lebanon.

BACKGROUND: Sexually transmitted infections (STIs) cause a major public health problem that affect both men and women in developing and developed countries. The aim of the study was to estimate the prevalence of 11 STIs among women who voluntarily participated in the study, while seeking gynecological checkup. The existence of an association between the presence of pathogens and symptoms and various sociodemographic risk factors was assessed. METHODS: A total of 505 vaginal and cervical specimens were collected from women above 18 years of age, with or without symptoms related to gynecological infections. Nucleic acid was extracted and samples were tested by real-time PCR for the following pathogens: Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium, Ureaplasma urealyticum, Urealplasma parvum, Trichomonas vaginalis, Mycoplasma hominis, Mycoplasma girerdii, Gardnerella vaginalis, Candida albicans and Human Papillomavirus (HPV). Positive HPV samples underwent genotyping using a microarray system. RESULTS: Of the 505 samples, 312 (62%) were screened positive for at least one pathogen. Of these, 36% were positive for Gardnerella vaginalis, 35% for Ureaplasma parvum, 8% for Candida albicans, 6.7% for HPV, 4.6% for Ureaplasma urealyticum, 3.6% for Mycoplasma hominis, 2% for Trichomonas vaginalis, 0.8% for Chlamydia trachomatis, 0.4% for Mycoplasma girerdii, 0.2% for Mycoplasma genitalium and 0.2% for Neisseria gonorrhoeae. Lack of symptoms was reported in 187 women (37%), among whom 61% were infected. Thirty-four samples were HPV positive, with 17 high risk HPV genotypes (HR-HPV); the highest rates being recorded for types 16 (38%), 18 (21%) and 51 (18%). Out of the 34 HPV positives, 29 participants had HR-HPV. Association with various risk factors were reported. CONCLUSIONS: This is the first study that presents data about the presence of STIs among women in Lebanon and the MENA region by simultaneous detection of 11 pathogens. In the absence of systematic STI surveillance in Lebanon, concurrent screening for HPV and PAP smear is warranted.

The vaginal microbiota associates with the regression of untreated cervical intraepithelial neoplasia 2 lesions.

Emerging evidence suggests associations between the vaginal microbiota (VMB) composition, human papillomavirus (HPV) infection, and cervical intraepithelial neoplasia (CIN); however, causal inference remains uncertain. Here, we use bacterial DNA sequencing from serially collected vaginal samples from a cohort of 87 adolescent and young women aged 16-26 years with histologically confirmed, untreated CIN2 lesions to determine whether VMB composition affects rates of regression over 24 months. We show that women with a Lactobacillus-dominant microbiome at baseline are more likely to have regressive disease at 12 months. Lactobacillus spp. depletion and presence of specific anaerobic taxa including Megasphaera, Prevotella timonensis and Gardnerella vaginalis are associated with CIN2 persistence and slower regression. These findings suggest that VMB composition may be a future useful biomarker in predicting disease outcome and tailoring surveillance, whilst it may offer rational targets for the development of new prevention and treatment strategies.