Assessment of vaginal health before and after treatment with vaginal suppository containing Lactiplantibacillus pentosus KCA1

Bacterial Vaginosis (BV) is a polymicrobial syndrome characterized by a shift in the vaginal flora from a predominant population of Lactobacilli to a replacement with anaerobes. Vaginal suppositories containing L. pentosus KCA1 were formulated using glycerol-gelatin base. A prospective and follow-up community-based study was done. Vaginal samples were collected from women of reproductive age (non-pregnant) between 18-45 years. A total of 360 women both symptomatic and asymptomatic provided High Vaginal Swabs (HVS) for BV screening using Nugent scoring method and 65 (18.1 %) participants were identified as having BV by microscopy with 45 (12.5 %) of them being consistent with BV (Nugent score of 7-10) and twenty (5.56 %) being intermediate with BV (Nugent score of 4-6). A baseline and intervention questionnaire were administered to the participants. Majority (81.5 %) of the participants affirmed that they have not heard about BV. Social burden reveals that 90.8 % affirmed that they feel frustrated, having BV symptoms recur after treatment and 83.1 % admitted that BV symptoms makes them feel embarrassed, ashamed and dirty. Majority (58.8 %) claimed that the state of their reproductive health is ‘very good’ four weeks after treatment, while 88.2 % of the participants rated their reproductive health after four weeks of treatment to be ‘somewhat better than now than a year ago’. In conclusion, it was revealed that BV has impact on the social status and quality of life of the participants except on their economic status. Knowledge on the awareness of BV among women of reproductive age is essential.

The Immune Landscape of Molecular Bacterial Vaginosis and HPV Natural History

Bacterial vaginosis (BV) is a highly prevalent condition that is associated with acquisition of sexually transmitted infections and adverse reproductive outcomes. It has been proposed that BV’s role as a pathogenic condition is mediated via bacteria-induced local inflammation. However, the complex interplay between vaginal microbes and host immune factors has yet to be clearly elucidated. We demonstrate that 16S rRNA amplicon sequencing and a novel pipeline can be used to generate a molecular Nugent BV score (molBV) corresponding to the Nugent score 0 - 10. This algorithm is independent of the region of 16S rRNA amplified, the sequencing platform and source population. We further identify two local immune cytokine patterns associated with this molecular Nugent score (q-values<0.001). The main immune response is represented by an elevated IL-1β/IP-10 ratio, whereas a second pattern consists of an increased TNF-α/MIP-1β ratio. To evaluate the biological significance of molBV-BV and the local immune response, we show that clearance of high-risk HPV (HR-HPV) infections (HZ=1.86, 95% CI: 1.19-2.9) was associated with immune profiles, but not molBV-BV when both were considered in the model. In contrast, the TNF-α/MIP-1β signature was associated with progression of incident infections to CIN2+ (OR = 2.81, 95% CI: 1.62-5.42), but not HR-HPV clearance. Thus, BV is a heterogeneous condition that activates different arms of the local immune response, which in turn are independent risk factors for HR-HPV clearance and progression.

Recent Semen Exposure Impacts the Cytokine Response and Bacterial Vaginosis in Women

BackgroundThe presence of semen in the vagina from unprotected sex may influence the immune and microbial environment of the female genital tract. Inflammatory cytokine concentrations and BV-associated bacteria in female genital secretions may influence HIV risk, although the effect of recent sexual intercourse on incident BV and the cytokine milieu of cervicovaginal secretions has rarely been measured in previous studies. Here, we investigated the extent to which partner semen impacts the cytokine response and incident BV.MethodsAt baseline, we assessed the recency of semen exposure in menstrual cup supernatants by quantifying prostate specific antigen (PSA) levels using ELISA in 248 HIV-uninfected women at high risk for HIV infection. Luminex was used to measure 48 cytokines in menstrual cup supernatants and vaginal swabs to diagnose BV by Nugent score. Point-of-care screening for Chlamydia trachomatis and Neisseria gonorrhoeae was conducted using GeneXpert while OSOM was used for Trichomonas vaginalis detection. Multivariable models, adjusted for age, sexually transmitted infections, BV, current contraception use and condom use, were used to assess the impact of semen exposure on biomarkers of inflammation and BV.ResultsPresence of PSA, indicating recent semen exposure within 48 hours prior to sampling, was observed in menstrual cup supernatants of 17% (43/248) of women. Of these women, 70% (30/43) had self-reported condom use at their last sex act and 84% (36/43) had BV (Nugent score &gt;7). PSA presence was significantly associated with prevalent BV (Relative Risk (RR), 2.609; 95% Confidence Interval (CI), 1.104 - 6.165; p = 0.029). Furthermore, women with detectable PSA had high median concentrations of macrophage inflammatory protein- beta (MIP-1α, p=0.047) and low median concentration of the stem cell growth factor beta (SCGF-β, p=0.038) compared to those without PSA.ConclusionA degree of discordance between self-reports of consistent condom use and PSA positivity was observed. There was also evidence of a relationship between recent semen exposure, BV prevalence and altered cytokine concentrations. These findings suggest that PSA, as a semen biomarker, should be taken into consideration when investigating biological markers in the female genital tract and self-reported condom use in studies on reproductive and sexual health.

New Insights into Vaginal Environment During Pregnancy

During pregnancy, the vaginal ecosystem undergoes marked changes, including a significant enrichment with Lactobacillus spp. and profound alterations in metabolic profiles. A deep comprehension of the vaginal environment may shed light on the physiology of pregnancy and may provide novel biomarkers to identify subjects at risk of complications (e.g., miscarriage, preterm birth). In this study, we characterized the vaginal ecosystem in Caucasian women with a normal pregnancy (n = 64) at three different gestational ages (i.e., first, second and third trimester) and in subjects (n = 10) suffering a spontaneous first trimester miscarriage. We assessed the vaginal bacterial composition (Nugent score), the vaginal metabolic profiles (1H-NMR spectroscopy) and the vaginal levels of two cytokines (IL-6 and IL-8). Throughout pregnancy, the vaginal microbiota became less diverse, being mainly dominated by lactobacilli. This shift was clearly associated with marked changes in the vaginal metabolome: over the weeks, a progressive reduction in the levels of dysbiosis-associated metabolites (e.g., biogenic amines, alcohols, propionate, acetate) was observed. At the same time, several metabolites, typically found in healthy vaginal conditions, reached the highest concentrations at the end of pregnancy (e.g., lactate, glycine, phenylalanine, leucine, isoleucine). Lower levels of glucose were an additional fingerprint of a normal vaginal environment. The vaginal levels of IL-6 and IL-8 were significantly associated with the number of vaginal leukocytes, as well as with the presence of vaginal symptoms, but not with a condition of dysbiosis. Moreover, IL-8 concentration seemed to be a good predictor of the presence of vaginal Candida spp. Cytokine concentrations were negatively correlated to lactate, serine, and glycine concentrations, whereas the levels of 4-hydroxyphenyllactate, glucose, O-acetylcholine, and choline were positively correlated with Candida vaginal loads. Finally, we found that most cases of spontaneous abortion were associated with an abnormal vaginal microbiome, with higher levels of selected metabolites in the vaginal environment (e.g., inosine, fumarate, xanthine, benzoate, ascorbate). No association with higher pro-inflammatory cytokines was found. In conclusion, our analysis provides new insights into the pathophysiology of pregnancy and highlights potential biomarkers to enable the diagnosis of early pregnancy loss.

Predictive Value of Sialidase in the Diagnosis of Sexually Transmitted Infections: a Cross-section Study Based on Vaginal Microecology Evaluation System

Background: The purpose of this study was to determine whether any indexes of vaginal microecology evaluation system (VMES) could predict sexually transmitted infections (STIs).Methods: A total of 1032 women, who presented to Gynecology Outpatient Clinic of the First Affiliated Hospital of Xi'an Jiaotong University between November 2016 and November 2020, were included in the study. Incident STIs was defined as any case of Chlamydia trachomatis (CT), Mycoplasma genitalium (MG) and Neisseria gonorrhoeae (NG), and were confirmed using the RNA-based simultaneous amplification and testing (SAT) assay. VEMS is structured in two parts, one is the morphological characterizations encompassing bacterial density, flora diversity, predominant flora, pathogenic microorganisms and indicators of inflammation, the other is the functional indexes, including vaginal pH, cleanliness, hydrogen peroxide (H2O2), sialidase, β-glucuronidase, leukocyte esterase, and acetylglucosaminidase. Bacterial vaginosis (BV) was diagnosed by Gram stain (Nugent score). Associations were mainly assessed using logistic regression (LR).Results: SAT assay detected STIs in 64 (6.2%) of the 1032 samples tested and 136 (13.2%) women had a clinical BV diagnosis using Nugent score. Multivariate logistic regression analysis revealed that women with sialidase-positive were more likely to test positive for STIs (aOR=3.081, 95% CI=1.586-5.984, P=0.001). Of 896 women without clinical BV, significant association was also found for sialidase and STIs (aOR =4.133, 95% CI= 1.140-14.978, P=0.031).Conclusions: Sialidase may be a useful indicator to help clinicians identify these women who are at risk for STIs, especially in the absence of BV population.

Multiplex PCR, Clinical Amsel’s Criteria and Microbiological Nugent Score for Diagnosis of Bacterial Vaginosis and Detection of its Virulence

Background: bacterial vaginosis is universally the commonest vaginal infection in reproductively active females. It causes major consequences as preterm labor, predisposing to sexually-transmitted infections and HIV infections. Although it is a public health concern, no one knows exactly its pathogenesis when some say that it is just disturbance in vaginal floral balance predisposing to clinical symptoms and signs, where the predominant Lactobacilli in vagina become replaced by other facultative and anaerobic bacteria. Objective: To evaluate different diagnostic tests, Amsel’s criteria and PCR and their ability to diagnose bacterial vaginosis in comparison to the gold standard test, Nugent score in terms of sensitivity, specificity, NPV and PPV. Also test for prevalence of BV among included females. Methodology: screening was done for all females in presence or absence of Amsel’s criteria. Wet mount along with Gram stained films were examined in Microbiology lab and Nugent score was calculated for every patient. Cervico-vaginal aspirate samples were collected for detection of G. vaginalis, Lactobacilli and Sialidase enzyme by multiplex PCR. Results: Using Nugent score patients were categorized into bacterial vaginosis (BV) group (32%), non-BV group (51%) and 17 % were in intermediate group. Sensitivity, specificity, PPV and NPV of whole Amsel’s criteria (100, 80.4, 76.2 and 100 % respectively) were better than using any criterion alone. Using multiplex PCR, detection of G. vaginalis (100%) and sialidase gene (93.7%) were higher in BV group and Lactobacilli gene (100%) higher in non-BV group with statistically high significant difference. Multiplex PCR detection of G. vaginalis has sensitivity (100%), specificity (92.2%), PPV (88.9%) and NPV (100%) for diagnosis of BV in relation to the gold standard test, Nugent score. Conclusion: using Amsel’s criteria as a whole is better than using individual criteria for diagnosing BV with highest sensitivity, specificity, NPV and PPV. Combined PCR detection of G. vaginalis and sialidase gene can predict occurrence of virulent BV infection. BV is associated with significant loss of protective Lactobacilli.

Vaginal Dysbiosis and Partial Bacterial Vaginosis: The Interpretation of the “Grey Zones” of Clinical Practice

Bacterial vaginosis (BV) affects one-third of reproductive age women, increasing the risk of acquiring sexually transmitted infections (STIs) and posing a risk for reproductive health. The current diagnosis with Gram stain (Nugent Score) identifies a transitional stage named partial BV or intermediate microbiota, raising the problem of how to clinically handle it. We retrospectively analyzed cervicovaginal swabs from 985 immunocompetent non-pregnant symptomaticspp. women (vaginal discharge, burning, itching) by Nugent score and qPCR for BV, aerobic or fungal vaginitis, and STIs (Mycoplasmas spp., Chlamydia t., Trichomonas v., and Neisseria g.). Nugent scores 0–3 and 7–10 were confirmed in 99.3% and 89.7% cases, respectively, by qPCR. Among Nugent scores 4–6 (partial BV), qPCR identified 46.1% of BV cases, with 37.3% of cases negative for BV, and only 16.7% of partial BV. Gram staining and qPCR were discordant (p value = 0.0001) mainly in the partial BV. Among the qPCR BV cases, the presence of aerobic vaginitis and STIs was identified, with a significant association (p < 0.0001) between the STIs and partial BV/overt BV. qPCR is more informative and accurate, and its use as an alternative or in combination with Gram staining could help clinicians in having an overview of the complex vaginal microbiota and in the interpretation of partial BV that can correspond to vaginitis and/or STIs.

Application of Ligilactobacillus salivarius CECT5713 to Achieve Term Pregnancies in Women with Repetitive Abortion or Infertility of Unknown Origin by Microbiological and Immunological Modulation of the Vaginal Ecosystem

In this study, the cervicovaginal environment of women with reproductive failure (repetitive abortion, infertility of unknown origin) was assessed and compared to that of healthy fertile women. Subsequently, the ability of Ligilactobacillus salivarius CECT5713 to increase pregnancy rates in women with reproductive failure was evaluated. Vaginal pH and Nugent score were higher in women with reproductive failure than in fertile women. The opposite was observed regarding the immune factors TGF-β 1, TFG-β 2, and VEFG. Lactobacilli were detected at a higher frequency and concentration in fertile women than in women with repetitive abortion or infertility. The metataxonomic study revealed that vaginal samples from fertile women were characterized by the high abundance of Lactobacillus sequences, while DNA from this genus was practically absent in one third of samples from women with reproductive failure. Daily oral administration of L. salivarius CECT5713 (~9 log10 CFU/day) to women with reproductive failure for a maximum of 6 months resulted in an overall successful pregnancy rate of 56%. The probiotic intervention modified key microbiological, biochemical, and immunological parameters in women who got pregnant. In conclusion, L. salivarius CECT5713 has proved to be a good candidate to improve reproductive success in women with reproductive failure.

Influence of intramuscular depot-medroxyprogesterone acetate initiation on vaginal microbiota in the postpartum period

Background The vaginal microbiome plays a key role in women’s reproductive health. Use of exogenous hormones, such as intramuscular depot-medroxyprogesterone acetate (DMPA-IM), may alter the composition of vaginal bacterial community. Methods Vaginal swabs were collected from postpartum Kenyan women initiating DMPA-IM or non-hormonal contraception (non-HC). Bacterial vaginosis was assessed by Nugent score (Nugent-BV) and bacterial community composition was evaluated using broad-range 16S rRNA gene PCR with high-throughput sequencing. Changes in Nugent score, alpha diversity (Shannon diversity index), and total bacterial load between contraceptive groups from enrollment to three-months post-initiation were estimated using multivariable linear mixed effects regression. Results Among 54 HIV-negative women, 33 choosing DMPA-IM and 21 choosing non-HC, Nugent-BV was more common among DMPA-IM users at enrollment. At follow-up, Nugent score had decreased significantly among DMPA-IM users (Δ=-1.89 (95%CI:-3.53, -0.25; p=0.02) while alpha diversity remained stable (Δ=0.03, 95%CI:-0.24, 0.30; p=0.83). Conversely, Nugent score remained relatively stable among non-HC users (Δ=-0.73, 95%CI:-2.18, 0.73; p=0.33) while alpha diversity decreased (Δ=-0.34, 95%CI:-0.67, -0.001; p=0.05). Total bacterial load decreased slightly in DMPA-IM users and increased slightly among non-HC users, resulting in a significant difference in change between the contraceptive groups (difference=-0.64 log10 gene copies/swab, 95%CI:-1.19, -0.08; p=0.02). While significant changes in Nugent score and alpha diversity were observed within contraceptive groups, changes between groups were not significantly different. Conclusions Postpartum vaginal bacterial diversity did not change in DMPA-IM users despite a reduction in Nugent-BV, but decreased significantly among women using non-HC. Choice of contraception may influence Lactobacillus recovery in postpartum women.