Spontaneous resolution of Trichomonas vaginalis infection in men.

OBJECTIVES: We aimed to investigate the early natural history of Trichomonas vaginalis in men recently testing positive for this infection by a nucleic acid amplification test (NAAT). We hypothesised that 50% of men would spontaneously resolve their infection (in the absence of treatment) on repeat T. vaginalis NAAT. METHODS: Men ages ≥18 years at the Jefferson County Health Department Sexual Health Clinic testing positive for T. vaginalis by NAAT during standard-of-care (SOC) within the past 30 days and presenting to the clinic for treatment were approached. At enrolment, participants completed a questionnaire, provided urine for repeat T. vaginalis NAAT, and were treated with 2 g oral metronidazole. Those with a repeat positive enrolment NAAT were seen for a 4-week test-of-cure (TOC) visit. At TOC, men provided urine for repeat NAAT. We determined the proportion of men with spontaneous resolution of T. vaginalis and evaluated predictors of spontaneous resolution. In those with a repeat positive enrolment T. vaginalis NAAT, we evaluated the proportion with persistent infection at TOC as a secondary outcome. RESULTS: Between October 2021 and January 2023, 53 men with a recent positive SOC T. vaginalis NAAT were approached; 37 (69.8%) participated. The mean participant age was 32.9 years (SD 9.9); all identified as Black. The majority (97.3%) reported sex with women only; 35.1% reported sex with >1 partner in the last month. At enrolment, 26/37 (70.3%) had a repeat positive T. vaginalis NAAT in the absence of treatment after an average of 8.4 days (SD 5.9). Sexual partner gender, number of recent sexual partners, genital symptoms, unprotected sex with any partner and recent antibiotic use were not associated with spontaneous resolution. Of the 26 men attending a TOC visit, 17 (65.4%) returned and all except one (94.1%) were cured. CONCLUSION: Most men do not spontaneously clear T. vaginalis infection during early repeat testing.

Challenges in Male Partner Referral Among Trichomonas vaginalis -Infected Women.

ABSTRACT: This study assessed feasibility of male partner referral by Trichomonas vaginalis -infected women. Of 93 women approached, only 20 enrolled. Only 1 male partner contacted the study but was unable to be reached for scheduling. Other public health interventions are necessary to engaged T. vaginalis -infected women and their male partners in care.

In vitro Testing of Trichomonas vaginalis Drug Susceptibility: Evaluation of Minimal Lethal Concentration for Secnidazole that Correlates with Treatment Success.

ABSTRACT: We determined the in vitro minimum lethal concentration (MLC) of secnidazole (SEC) and assessed correlation with clinical susceptibility among T. vaginalis isolates obtained from 71 women, of whom 66 were successfully treated with this medication. An MLC ≤12.5 µg/ml correlated with clinical susceptibility in this study.

An Indirect Fluorescence Microscopy Method to Assess Vaginal Lactobacillus Concentrations.

Lactobacillus species are the main colonizers of the vaginal microbiota in healthy women. Their absolute quantification by culture-based methods is limited due to their fastidious growth. Flow cytometry can quantify the bacterial concentration of these bacteria but requires the acquisition of expensive equipment. More affordable non-culturable methods, such as fluorescence microscopy, are hampered by the small size of the bacteria. Herein, we developed an indirect fluorescence microscopy method to determine vaginal lactobacilli concentration by determining the correlation between surface area bacterial measurement and initial concentration of an easily cultivable bacterium (Escherichia coli) and applying it to lactobacilli fluorescence microscopy counts. In addition, vaginal lactobacilli were quantified by colony-forming units and flow cytometry in order to compare these results with the indirect method results. The colony-forming-unit values were lower than the results obtained from the other two techniques, while flow cytometry and fluorescence microscopy results agreed. Thus, our developed method was able to accurately quantify vaginal lactobacilli.

Optimized bacterial absolute quantification method by qPCR using an exogenous bacterial culture as a normalization strategy in triple-species BV-like biofilms.

Quantitative Polymerase Chain Reaction (qPCR) is a widely used method in molecular biology to quantify target DNA sequences. Despite its accuracy, there are important experimental controls that should be considered to avoid biased results. One of them is gDNA loss during extraction, which is higher among samples with lower bacterial concentrations. Improvement in qPCR quantification procedures is mandatory to obtain reproducible and accurate results. Herein, we report an improved qPCR method for bacterial quantification of Gardnerella vaginalis, Prevotella bivia, and Fannyhessea vaginae, three key-bacterial vaginosis (BV)-associated bacteria (BVAB) thought to play important roles in the pathogenesis of this common vaginal infection. The formation of a mature biofilm on vaginal epithelial cells is an unique feature of BV and, despite over 60 years of research, the exact etiology of BV remains unknown. Here, we optimized a qPCR method that accurately quantified triple-species biofilms containing these key BVAB, after the addition of an exogenous bacterial control containing a fixed concentration of Escherichia coli, prior to gDNA extraction. This improved method minimized and normalized the inherent losses associated with bacterial centrifugation, which allows better sensitivity at lower bacterial concentrations.

The Role of Prevotella Species in Female Genital Tract Infections.

Female genital tract infections (FGTIs) include vaginal infections (e.g., bacterial vaginosis [BV]), endometritis, pelvic inflammatory disease [PID], and chorioamnionitis [amniotic fluid infection]. They commonly occur in women of reproductive age and are strongly associated with multiple adverse health outcomes including increased risk of HIV/sexually transmitted infection acquisition and transmission, infertility, and adverse birth outcomes such as preterm birth. These FGTIs are characterized by a disruption of the cervicovaginal microbiota which largely affects host immunity through the loss of protective, lactic acid-producing Lactobacillus spp. and the overgrowth of facultative and strict anaerobic bacteria. Prevotella species (spp.), anaerobic Gram-negative rods, are implicated in the pathogenesis of multiple bacterial FGTIs. Specifically, P. bivia, P. amnii, and P. timonensis have unique virulence factors in this setting, including resistance to antibiotics commonly used in treatment. Additionally, evidence suggests that the presence of Prevotella spp. in untreated BV cases can lead to infections of the upper female genital tract by ascension into the uterus. This narrative review aims to explore the most common Prevotella spp. in FGTIs, highlight their important role in the pathogenesis of FGTIs, and propose future research in this area.

Microbial interactions among Gardnerella, Prevotella and Fannyhessea prior to incident bacterial vaginosis: protocol for a prospective, observational study.

INTRODUCTION: The aetiology of bacterial vaginosis (BV), a biofilm-associated vaginal infection, remains unknown. Epidemiologic data suggest that it is sexually transmitted. BV is characterised by loss of lactic acid-producing lactobacilli and an increase in facultative and strict anaerobic bacteria. Gardnerella spp are present in 95%-100% of cases; Gardnerella vaginalis has been found to be more virulent than other BV-associated bacteria (BVAB) in vitro. However, G. vaginalis is found in women with normal vaginal microbiota and colonisation is not sufficient for BV development. We hypothesise that Gardnerella spp initiate BV biofilm formation, but incident BV (iBV) requires incorporation of other key BVAB (ie, Prevotella bivia, Fannyhessea vaginae) into the biofilm that alter the transcriptome of the polymicrobial consortium. This study will investigate the sequence of microbiologic events preceding iBV. METHODS AND ANALYSIS: This study will enrol 150 women aged 18-45 years with normal vaginal microbiota and no sexually transmitted infections at a sexual health research clinic in Birmingham, Alabama. Women will self-collect twice daily vaginal specimens up to 60 days. A combination of 16S rRNA gene sequencing, qPCR for Gardnerella spp, P. bivia and F. vaginae, and broad range 16S rRNA gene qPCR will be performed on twice daily vaginal specimens from women with iBV (Nugent score 7-10 on at least 2 consecutive days) and controls (with comparable age, race, contraceptive method and menstrual cycle days) maintaining normal vaginal microbiota to investigate changes in the vaginal microbiota over time for women with iBV. Participants will complete daily diaries on multiple factors including sexual activity. ETHICS AND DISSEMINATION: This protocol is approved by the University of Alabama at Birmingham Institutional Review Board (IRB-300004547) and written informed consent will be obtained from all participants. Findings will be presented at scientific conferences and published in peer-reviewed journals as well as disseminated to providers and patients in communities of interest.

Recruiting transgender men in the Southeastern United States for genital microbiome research: Lessons learned.

BACKGROUND: Transgender men (TGM) are underrepresented in genital microbiome research. Our prospective study in Birmingham, AL investigated genital microbiota changes over time in TGM initiating testosterone, including the development of incident bacterial vaginosis (iBV). Here, we present lessons learned from recruitment challenges encountered during the conduct of this study. METHODS: Inclusion criteria were assigned female sex at birth, TGM or non-binary identity, age ≥18 years, interested in injectable testosterone but willing to wait 7 days after enrollment before starting, and engaged with a testosterone-prescribing provider. Exclusion criteria were recent antibiotic use, HIV/STI infection, current vaginal infection, pregnancy, or past 6 months testosterone use. Recruitment initiatives included community advertisements via flyers, social media posts, and referrals from local gender health clinics. RESULTS: Between February 2022 and October 2023, 61 individuals contacted the study, 17 (27.9%) completed an in-person screening visit, and 10 (58.8%) of those screened were enrolled. The primary reasons for individuals failing study screening were having limited access to testosterone-prescribing providers, already being on testosterone, being unwilling to wait 7 days to initiate testosterone therapy, or desiring the use of topical testosterone. Engagement of non-White TGM was also minimal. CONCLUSION: Despite robust study inquiry by TGM, screening and enrollment challenges were faced including engagement by TGM not yet in care and specific study eligibility criteria. Excitement among TGM for research representation should be leveraged in future work by engaging transgender community stakeholders at the inception of study development, particularly regarding feasibility of study inclusion and exclusion criteria, as well as recruitment of TGM of color. These results also highlight the need for more clinical resources for prescribing gender-affirming hormone therapy, especially in the Southeastern US.

Genital tract infections, the vaginal microbiome and gestational age at birth among pregnant women in South Africa: a cohort study protocol.

INTRODUCTION: Preterm birth complications are the most common cause of death in children under 5 years. The presence of multiple microorganisms and genital tract inflammation could be the common mechanism driving early onset of labour. South Africa has high levels of preterm birth, genital tract infections and HIV infection among pregnant women. We plan to investigate associations between the presence of multiple lower genital tract microorganisms in pregnancy and gestational age at birth. METHODS AND ANALYSIS: This cohort study enrols around 600 pregnant women at one public healthcare facility in East London, South Africa. Eligible women are ≥18 years and at <27 weeks of gestation, confirmed by ultrasound. At enrolment and 30-34 weeks of pregnancy, participants receive on-site tests for Chlamydia trachomatis and Neisseria gonorrhoeae, with treatment if test results are positive. At these visits, additional vaginal specimens are taken for: PCR detection and quantification of Trichomonas vaginalis, Candida spp., Mycoplasma genitalium, M. hominis, Ureaplasma urealyticum and U. parvum; microscopy and Nugent scoring; and for 16S ribosomal RNA gene sequencing and quantification. Pregnancy outcomes are collected from a postnatal visit and birth registers. The primary outcome is gestational age at birth. Statistical analyses will explore associations between specific microorganisms and gestational age at birth. To explore the association with the quantity of microorganisms, we will construct an index of microorganism load and use mixed-effects regression models and classification and regression tree analysis to examine which combinations of microorganisms contribute to earlier gestational age at birth. ETHICS AND DISSEMINATION: This protocol has approvals from the University of Cape Town Research Ethics Committee and the Canton of Bern Ethics Committee. Results from this study will be uploaded to preprint servers, submitted to open access peer-reviewed journals and presented at regional and international conferences. TRIAL REGISTRATION NUMBER: NCT06131749; Pre-results.