The HepHIV 2023 Madrid conference: A call to action for political leadership in reaching the sustainable development goals on earlier testing and linkage to care for HIV, viral hepatitis, and sexually transmitted infections.

INTRODUCTION AND OBJECTIVES: The HepHIV 2023 Conference, held in Madrid in November 2023, highlighted how Europe is not on track to meet the United Nations (UN) sustainable development goals and Joint UN Programme on HIV/AIDS (UNAIDS) targets. This article presents the outcomes of the conference, which focus on ways to improve testing and linkage to care for HIV, viral hepatitis, and other sexually transmitted infections. HIV-related stigma and discrimination, a major barrier to progress, was a key concept of the conference and on the agenda of the Spanish Presidency of the European Union. METHODS: The HepHIV 2023 organizing committee, alongside the Spanish Ministry of Health, oversaw the conference organization and prepared the scientific programme based on abstract rankings. Key outcomes are derived from conference presentations and discussions. RESULTS: Conference presentations covered the obstacles that HIV-related stigma and discrimination continue to pose to access to services, models for data collection to better monitor progress in the future, and examples of legislative action that can be taken at national levels. Diversification of testing approaches was also highlighted, to reach key populations, (e.g. migrant populations), to increase testing offered in healthcare settings (e.g. emergency departments), and to account for different stages of epidemics across the region. CONCLUSION: With a strong call for intensified action to address the impact of HIV-related stigma and discrimination on testing uptake, the conference concluded that strengthened collaboration is required between governments and implementers around testing and linkage to care. There is also an ongoing need to ensure sustainable political commitment and appropriate resource allocation to address gaps and inequalities in access for key populations and to focus on the implementation of integrated responses to HIV, viral hepatitis, and sexually transmitted infections.

Identifying priority groups for pre-exposure prophylaxis among sex workers in Flanders, Belgium: insights into routine HIV and sexually transmitted infection data in community-based clinics.

OBJECTIVES: The burden of HIV and other sexually transmitted infections (STIs) remains high in sex workers globally, calling for strengthening targeted prevention strategies, including HIV pre-exposure prophylaxis (PrEP). The study's objective was to assess HIV and STI burden among female, male and transgender sex workers in Flanders, Belgium, to guide targeting of PrEP strategies for sex workers. METHODS: We conducted a retrospective analysis of routine data collected between January 2016 and December 2019 by community-based organisations providing sexual healthcare services for sex workers in Flanders. HIV prevalence stratified by gender was assessed and associations with sociodemographic characteristics were explored using bivariable and multivariable logistic regression. Positivity rates of chlamydia, gonorrhoea and syphilis tests were used as proxy indicators for STI burden. RESULTS: The study included a total of 6028 sex workers, comprising 5617 (93.2%) female, 218 (3.6%) male and 193 (3.2%) transgender sex workers. The HIV prevalence was 0.3% among female, 8.9% among male and 12.3% among transgender sex workers. Engaging in escort sex work and originating from South America or Sub-Saharan Africa were associated with a higher likelihood of having acquired HIV. The positivity rate for gonorrhoea was higher among male sex workers (5.2% vs 2.2%) and syphilis was more frequently detected among male and transgender sex workers (3.0% and 6.1% vs 0.5%), all compared with female sex workers. CONCLUSIONS: HIV combination prevention, including improved access to PrEP, should be strengthened among sex workers in Flanders, with particular attention to male and transgender sex workers.

Self-sampling with oral rinse to detect oropharyngeal Neisseria gonorrhoeae among men who have sex with men: results from an exploratory study in Belgium (the SSONG Study).

OBJECTIVES: We aimed to assess whether a self-collected oral rinse was non-inferior to clinician-collected oropharyngeal swabs to detect Neisseria gonorrhoeae (Ng) using culture and nucleic acid amplification tests (NAAT) among men who have sex with men (MSM), and whether Ng may still be detected in oral rinses for a minimum of 5 days after collection. METHODS: MSM with a positive Ng result in an oropharyngeal or pooled sample (oropharynx, urethra and anorectum) were approached. Clinician-collected oropharyngeal swabs and oral rinses (15 mL sterile water) were taken. Ng culture and NAAT (Abbott 2000m RealTime System CT/NG assay and in-house PCR) were performed. Diagnostic accuracy was assessed using sensitivity and specificity, and agreement between both techniques using Cohen's kappa statistic. Aliquots of positive oral rinses were left at room temperature for a minimum of 5 days and reanalysed using NAAT. Lastly, participants filled in a questionnaire to explore perceptions of both methods. RESULTS: We included 100 participants between June 2022 and October 2023. 45 individuals (45 of 100) had a positive Ng result in either the oral rinses (42 of 45, 93%) or the swabs (36 of 45, 80%). Sensitivity was higher for oral rinses than swabs (sensitivity=0.93/0.80, specificity=1.0/1.0, respectively) and agreement between both techniques was good (kappa=0.75, p<0.001). Of the 42 positive oral rinses, 37 remained positive after a minimum of 5 days (88.1%). Using culture, 18 individuals had a positive Ng result in either the oral rinses (8 of 18, 44%) or the swabs (16 of 18, 88%). Most participants found the oral rinse easy or very easy to use and would be willing to use the oral rinse for home-based sampling. CONCLUSION: We detected more oropharyngeal Ng infections via NAAT using oral rinses than swab samples. However, swabs were better than oral rinses for culturing Ng. Oral rinses might allow for home-based self-sampling to detect oropharyngeal Ng.

Frequent chlamydia and gonorrhoea but very infrequent syphilis among men who have sex with men using HIV pre-exposure prophylaxis in West Africa.

OBJECTIVES: Although oral pre-exposure prophylaxis (PrEP) for HIV is being rolled out in West Africa, data on sexually transmitted infections (STIs) in PrEP users are scarce. We assessed the prevalence, incidence and determinants of bacterial STIs in men who have sex with men (MSM) taking PrEP in Burkina Faso, Côte d'Ivoire, Mali and Togo. METHODS: A prospective cohort study among MSM initiating PrEP as part of a comprehensive HIV prevention package was conducted between 2017 and 2021 in community-based clinics in the four study countries. Molecular screening for Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) was performed at months 0, 6 and 12. Serological testing for syphilis was performed every 3 months over the first year of follow-up. Determinants of CT and/or NG incidence were identified using Poisson generalised linear mixed models. RESULTS: A total of 598 participants with a median age of 24.7 years were included. Prevalence of CT and/or NG was 24.4% (95% CI 21.0 to 28.1), 22.4% (95% CI 18.4 to 26.8) and 29.0% (95% CI 24.2 to 34.1) at months 0, 6 and 12, respectively. The prevalence of syphilis ranged from 0.2% (95% CI 0.0 to 0.9) at month 0 to 0.8% (95% CI 0.2 to 2.4) at month 12. Ninety incident CT and/or NG infections occurred during a total follow-up time of 280.6 person-years (incidence rate 32.1 per 100 person-years, 95% CI 25.8 to 39.4). Three incident syphilis infections were detected during a total follow-up time of 459.7 person-years (incidence rate 0.7 per 100 person-years, 95% CI 0.1 to 1.9). CT and/or NG incidence was associated with condomless insertive anal sex (adjusted incidence rate ratio 1.96, 95% CI 1.04 to 3.71, p=0.038). CONCLUSIONS: CT and NG were frequent but syphilis was very infrequent in MSM using HIV PrEP in West Africa. HIV programme managers should integrate STI services into PrEP programmes.

Estimation of antimicrobial resistance of Mycoplasma genitalium, Belgium, 2022.

BackgroundAntimicrobial resistance (AMR) of Mycoplasma genitalium (MG) is a growing concern worldwide and surveillance is needed. In Belgium, samples are sent to the National Reference Centre of Sexually Transmitted Infections (NRC-STI) on a voluntary basis and representative or robust national AMR data are lacking.AimWe aimed to estimate the occurrence of resistant MG in Belgium.MethodsBetween July and November 2022, frozen remnants of MG-positive samples from 21 Belgian laboratories were analysed at the NRC-STI. Macrolide and fluoroquinolone resistance-associated mutations (RAMs) were assessed using Sanger sequencing of the 23SrRNA and parC gene. Differences in resistance patterns were correlated with surveillance methodology, socio-demographic and behavioural variables via Fisher's exact test and logistic regression analysis.ResultsOf the 244 MG-positive samples received, 232 could be sequenced for macrolide and fluoroquinolone RAMs. Over half of the sequenced samples (55.2%) were resistant to macrolides. All sequenced samples from men who have sex with men (MSM) (24/24) were macrolide-resistant. Fluoroquinolone RAMs were found in 25.9% of the samples and occurrence did not differ between socio-demographic and sexual behaviour characteristics.ConclusionAlthough limited in sample size, our data suggest no additional benefit of testing MG retrieved from MSM for macrolide resistance in Belgium, when making treatment decisions. The lower occurrence of macrolide resistance in other population groups, combined with emergence of fluoroquinolone RAMs support macrolide-resistance testing in these groups. Continued surveillance of resistance in MG in different population groups will be crucial to confirm our findings and to guide national testing and treatment strategies.

Treponema pallidum Strains Among Women and Men Who Have Sex With Women in Amsterdam, the Netherlands and Antwerp, Belgium Between 2014 and 2020.

ABSTRACT: The Treponema pallidum strain distribution among men who have sex with women were similar to the strain distribution men who have sex with men. The most prevalent strains and percentage of strains belonging to the Nichols lineage are similar to previous studies in Amsterdam focusing on men who have sex with men.

Global epidemiology of antimicrobial resistance in commensal Neisseria species: A systematic review.

BACKGROUND: Commensal Neisseria species (spp). represent an important reservoir of antimicrobial resistance genes for pathogenic Neisseria spp. In this systematic review, we aimed to assess the antimicrobial susceptibility of commensal Neisseria spp. and how this has evolved over time. We also aimed to assess if commensal Neisseria spp. showed intrinsic resistance to four antimicrobials - penicillin, azithromycin, ceftriaxone and ciprofloxacin. METHODS: Pubmed and Google Scholar were searched following the PRISMA guidelines. Articles reporting MICs of commensal Neisseria spp. were included according to inclusion/exclusion criteria, and the quality of the articles was assessed using a pre-designed tool. Individual and summary measures of penicillin, azithromycin, ceftriaxone and ciprofloxacin MICs were collected. Additional data was sought to perform a comparison between the MICs of pathogenic and commensal Neisseria spp. RESULTS: A total of 15 studies met our criteria.We found no evidence of intrinsic AMR in commensal Neisseria spp. We did find evidence of an increasing trend in MICs of commensal Neisseria spp. over time for all antimicrobials assessed. These findings were similar in various countries. Eight additional studies were included to compare pathogenic and commensal Neisseria spp. CONCLUSION: The MICs of commensal Neisseria spp. appear to be increasing in multiple countries. Surveillance of MICs in commensals could be used as an early warning system for antimicrobial resistance emergence in pathogens. Our findings underline the need for antibiotic stewardship interventions, particularly in populations with high antimicrobial consumption.

Chlorhexidine Mouthwash Fails to Eradicate Oropharyngeal Gonorrhea in a Clinical Pilot Trial (MoNg).

ABSTRACT: This single-arm open-label pilot trial in Antwerp, Belgium, was ended early in accordance with the protocol because twice-daily gargling with chlorhexidine 0.2% for 6 days failed to eradicate Neisseria gonorrhoeae from the oropharynx of asymptomatic men who have sex with men (n = 3; efficacy of 0%; 95% confidence interval, 0%-56.1%).

An alarming high prevalence of resistance-associated mutations to macrolides and fluoroquinolones in Mycoplasma genitalium in Belgium: results from samples collected between 2015 and 2018.

OBJECTIVES: The number of reported cases of multiresistant Mycoplasma genitalium (MG) is increasing globally. The aim of this study was to estimate the prevalence of macrolide and possible fluoroquinolone resistance-associated mutations (RAMs) of MG in Belgium. METHODS: The study was performed retrospectively on two sets of MG-positive samples collected in Belgium between 2015 and 2018. The first set of samples originated from routine surveillance activities and the second set came from a cohort of men who have sex with men (MSM) using pre-exposure prophylaxis to prevent HIV transmission. Detection of RAMs to macrolides and fluoroquinolones was performed on all samples using DNA sequencing of the 23S ribosomal RNA gene, the gyrA gene and the parC gene. RESULTS: Seventy-one per cent of the MG samples contained a mutation conferring resistance to macrolides or fluoroquinolones (ParC position 83/87). RAMs were more frequently found among men compared with women for fluoroquinolones (23.9% vs 9.1%) and macrolides (78.4% vs 27.3%). Almost 90% of the MG infections among MSM possessed a RAM to macrolides (88.4%). In addition, 18.0% of the samples harboured both macrolides and fluoroquinolone RAMs; 3.0% in women and 24.2% in MSM. Being MSM was associated with macrolide RAMs (OR 15.3), fluoroquinolone RAMs (OR 3.8) and having a possible multiresistant MG infection (OR 7.2). CONCLUSION: The study shows an alarmingly high prevalence of MG with RAMs to macrolides and fluoroquinolones in Belgium. These results highlight the need to improve antimicrobial stewardship in Belgium in order to avoid the emergence of untreatable MG.

The Population-Level Effect of Screening for Mycoplasma genitalium on Antimicrobial Resistance: A Quasi-Experimental Study.

BACKGROUND: No studies have evaluated the utility and risks of screening for Mycoplasma genitalium in men who have sex with men taking preexposure prophylaxis (PrEP). We made use of a quasi-experimental design to evaluate the effect of screening for M. genitalium in a demonstration PrEP cohort with 3-monthly follow-up. METHODS: We compared the proportion of PrEP participants with M. genitalium clearance, the duration of persistence, proportion with incident symptoms, the incidence of fluoroquinolone and macrolide resistance, and the proportion of noncleared infections with resistance-associated mutations between 2 groups: those in whom the first episode of M. genitalium was treated and those in whom it was not treated. RESULTS: M. genitalium was detected in 70 of 179 individuals. The first episode of infection was treated in 46 individuals. Treatment was not significantly associated with the incidence of symptomatic infections or the acquisition of genotypic resistance. Treatment was associated with a higher probability of clearance of infection but at the expense of increasing the proportion of remaining infections that were resistant. In the nontreated group, the infections that did not clear were less likely to be fluoroquinolone resistant (1/6 [16.7%]) than those that did clear (4/4 [100%]; P = 0.048). In contrast, in the treated group, there was no significant difference in the proportion of fluoroquinolone resistance between the infections that persisted and cleared. CONCLUSIONS: If screening and treatment increase the ratio of resistant to susceptible M. genitalium in a population, then this could play a role in the spread of antimicrobial resistance.

Recurrent Sexually Transmitted Infections Among a Cohort of Men Who Have Sex With Men Using Preexposure Prophylaxis in Belgium Are Highly Associated With Sexualized Drug Use.

BACKGROUND: Men who have sex with men (MSM) experiencing recurrent sexually transmitted infections (STIs) may play a crucial role in the STI epidemic. However, there is limited understanding of what kind of behavior leads to recurrent STIs. METHODS: A total of 179 MSM using preexposure prophylaxis were followed up for 18 months and were screened quarterly for chlamydia, gonorrhea, and syphilis from 2015 to 2018 in Belgium. Participants were stratified into 3 different groups (no STI, one STI episode, recurrent STI episodes during the study). Sociodemographic and sexual behavioral characteristics were compared between the 3 groups, and significant associations with recurrent STI were explored using multivariate logistic regression models. RESULTS: A total of 62.0% (n = 111/179) of participants experienced at least one STI during the study, and more than 1 in 3 became reinfected with an STI at another visit (n = 66/179 [36.9%]). Participants experiencing recurrent STIs reported the highest frequency of sexualized drug use (86.4%) compared with participants experiencing one (60.0%) or no STI (47.1%). Therefore, sexualized drug use was highly associated with recurrent STIs (adjusted odds ratio [aOR]. 4.35). Other factors associated with recurrent STIs were being younger than 40 years (aOR, 3.29), had a high number (>4) of nonsteady partners with whom receptive (aOR, 1.17) or insertive (aOR, 1.12) condomless anal intercourse occurred in the last 3 months. CONCLUSIONS: Sexualized drug use was the greatest risk factor for having recurrent STIs. Tailoring prevention and care, including specialized services tackling problematic drug use in a sexual context, may help to curb the STI epidemic among MSM.

The Impact of PrEP on the Sex Lives of MSM at High Risk for HIV Infection: Results of a Belgian Cohort.

There is a need for an in-depth understanding of the impact of PrEP on users' sexual health and behaviour, beyond the focus on 'risk'. This mixed-method study was part of a Belgian PrEP demonstration project following 200 men who have sex with men (MSM) for at least 18 months. Taking a grounded-theory approach, 22 participants were interviewed and their transcripts analysed. The preliminary analysis guided the analysis of the questionnaire data. Overall, PrEP improved sexual health. Participants felt better protected against HIV, which enabled them to change their sexual behaviour. The reduction in condom use was moderated by interviewees' attitudes towards the risk for other STIs. Other changes included having more anal sex and experimentation with new sexual behaviours. While PrEP empowers MSM in taking care of their sexual health, comprehensive sexual health counselling is crucial to provide care for users who feel less in control over their sexual health.

Prevalence estimates of genital Chlamydia trachomatis infection in Belgium: results from two cross-sectional studies.

BACKGROUND: Chlamydia trachomatis (chlamydia) is the most diagnosed sexually transmitted infection in Belgium. Screening programs focus on young women, due to the implications of chronic asymptomatic infections for reproductive health. Thereby, the frequency of infections in men and older adults is underestimated. This study aimed to estimate the point-prevalence of chlamydia in the broader Belgian population, to inform evidence-based prevention and control strategies. METHODS: We conducted two cross-sectional prevalence studies of chlamydia infection in the population of Belgium aged 16-59 years, 2018-2020. In the CT1 study 12,000 representative individuals were randomly selected from the national register and invited by letter to collect a urine sample at home. The CT2 study used urine samples collected through the Belgian Health Examination Survey. Molecular detection of chlamydia DNA was performed using Xpert® or Abbott Real-Time CT/NG assays. Weighted estimated prevalence and 95% confidence interval (CI) was calculated per gender and age groups of 16/18-29, 30-44 and 45-59 years, relative to the general Belgian population. Data collected on sociodemographic variables and sexual behavior were used to identify potential risk factors for chlamydia infection through calculation of the odds ratio (OR). RESULTS: The population-wide weighted estimated prevalence was 1.54% (95% CI 0.78-3) in CT1 and 1.76% (95% CI 0.63-4) in CT2. We observed no statistically significant difference between men and women or age groups. Civil relationship status (OR = 14.1 (95% CI 1.78-112), p < 0.01), sexual intercourse with a casual partner (OR = 6.31 (95% CI 1.66-24.1), p < 0.01) and > 3 sexual partners in the last 12 months (OR = 4.53 (95% CI 1.10-18.6), p = 0.02) were associated with higher relative risk for chlamydia infection. CONCLUSION: Nationwide prevalence studies are relevant to assess the distribution of chlamydia and inform public health actions. The overall low prevalence and heterogeneous distribution of chlamydia in the general Belgian population needs to be considered for future strategies and potential harm of testing and treating asymptomatic individuals need to be taken into account. Effective case management should include appropriate treatment of symptomatic patients and partner notification, and prevention strategies should encourage behaviors such as condom use.

To Pool or Not to Pool Samples for Sexually Transmitted Infections Detection in Men Who Have Sex With Men? An Evaluation of a New Pooling Method Using the GeneXpert Instrument in West Africa.

BACKGROUND: Men who have sex with men (MSM) using preexposure prophylaxis (PrEP) are at risk for sexually transmitted infections (STIs). Therefore, PrEP services should include regular screening for Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) at urethra, anorectum, and pharynx. However, financial and logistic challenges arise in low-resource settings. We assessed a new STI sample pooling method using the GeneXpert instrument among MSM initiating PrEP in West Africa. METHODS: Urine, anorectal, and pharyngeal samples were pooled per individual for analysis. In case of an invalid result only (strategy 1) or a positive result of the pool (strategy 2), samples were analyzed individually to identify the infection's biological location. The results of 2 different pooling strategies were compared against the individual results obtained by a criterion standard. RESULTS: We found a prevalence of 14.5% for chlamydia and 11.5% for gonorrhea, with a predominance of infections being extragenital (77.6%). The majority of infections were asymptomatic (88.2%). The pooling strategy 1, had a sensitivity, specificity and agreement for CT of 95.4%, 98.7%, and 0.93, respectively; and 92.3%, 99.2%, and 0.93 for pooling strategy 2. For NG, these figures were 88.9%, 97.7%, and 0.85 for strategy 1, and 88.9%, 96.7%, and 0.81 for strategy 2. CONCLUSIONS: West African MSM have a high prevalence of extragenital and asymptomatic STIs. The GeneXpert method provides an opportunity to move from syndromic toward etiological STI diagnosis in low-income countries, as the platform is available in African countries for tuberculosis testing. Pooling will reduce costs of triple site testing.

Lymphogranuloma venereum is on the rise in Belgium among HIV negative men who have sex with men: surveillance data from 2011 until the end of June 2017.

BACKGROUND: The number of cases of Lymphogranuloma venereum (LGV) is increasing in Europe. The described epidemic is mostly confined to HIV positive men who have sex with men (MSM). However, dissemination of LGV from HIV positive to HIV negative MSM could take place due to the implementation of pre-exposure prophylaxis (PrEP) and subsequent possible decrease in condom use. We describe here the LGV epidemiology in Belgium before the PrEP-era, starting from 2011 up to the end of the first half of 2017. METHODS: A descriptive analysis of the socio-demographic and clinical characteristics of all LGV cases was performed. Fisher's exact test was used to compare symptomatic to asymptomatic patients. Logistic regression models were used to check for trends over time for: number of LGV cases, HIV status and symptoms. RESULTS: The number of LGV cases rose by a factor four, from 21 in 2011 to 88 in 2016, and regression models showed a positive trend estimate of 14% increase per half year (p < 0.001). LGV decreased among HIV positive cases (odds ratio (OR): 0.79, p < 0.001) and increased among HIV negative cases (OR: 1.27, p < 0.001). In addition, a rise in the number of asymptomatic LGV cases (6.7%) was observed (OR:1.39, p = 0.047). Asymptomatic cases were also less likely to be HIV (p = 0.046) or Hepatitis C positive (p = 0.027). CONCLUSIONS: The rise of LGV in HIV negative MSM has now been documented. If we aim to halt the epidemic in HIV negative MSM, future public health strategies should include LGV testing of all Chlamydia trachomatis positive samples from MSM.

Evaluation of an enzymatic Chlamydia trachomatis point-of-care rapid assay in Rwanda: the BioChekSwab Rapid Test.

OBJECTIVES: We evaluated the performance of an enzymatic point-of-care rapid test for Chlamydia trachomatis (CT) (the BioChekSwab CT Rapid Test, EnZtek Diagnostics, Rio Vista, California, USA), which detects CT's Peptidase 123CBV enzyme and provides a result 15 min after specimen collection. METHODS: Two endocervical swabs, including one BioChekSwab, per person were obtained from 137 women who participated in a reproductive health study in Rwanda. The BioChekSwab was processed according to the manufacturer's instructions. A substrate was squirted over the swab by the study physician immediately after collection, and another reagent was released over the swab tip at arrival in the laboratory. The test was considered positive if a blue colour developed within 2 min. The other regular flocked endocervical swab was processed at the Institute of Tropical Medicine (ITM), Belgium, using a testing algorithm: Abbott RealTime CT/Neisseria gonorrhoeae (NG) assay with the confirmation of positive results by an in-house real-time PCR assay. RESULTS: Of the 137 women, nine were CT positive by the testing algorithm. All nine positive results were missed by the BioChekSwab assay and four false-positive results were obtained. The sensitivity was therefore 0% (95% CI 0% to 33.6%) and the specificity was 96.9% (95% CI 92.2% to 99.1%). CONCLUSIONS: The BioChekSwab Rapid Test, although ISO13485 certified and Conformitée Européenne (CE) labelled, lacked any sensitivity in our setting.

A fruitful alliance: the synergy between Atopobium vaginae and Gardnerella vaginalis in bacterial vaginosis-associated biofilm.

OBJECTIVES: Bacterial vaginosis (BV) is characterised by a change in the microbial composition of the vagina. The BV-associated organisms outnumber the health-associated Lactobacillus species and form a polymicrobial biofilm on the vaginal epithelium, possibly explaining the difficulties with antibiotic treatment. A better understanding of vaginal biofilm with emphasis on Atopobium vaginae and Gardnerella vaginalis may contribute to a better diagnosis and treatment of BV. METHODS: To this purpose, we evaluated the association between the presence of both bacteria by fluorescence in situ hybridisation (FISH) and BV by Nugent scoring in 463 vaginal slides of 120 participants participating in a clinical trial in Rwanda. RESULTS: A bacterial biofilm was detected in half of the samples using a universal bacterial probe. The biofilm contained A. vaginae in 54.1% and G. vaginalis in 82.0% of the samples. A. vaginae was accompanied by G. vaginalis in 99.5% of samples. The odds of having a Nugent score above 4 were increased for samples with dispersed G. vaginalis and/or A. vaginae present (OR 4.5; CI 2 to 10.3). The probability of having a high Nugent score was even higher when a combination of adherent G. vaginalis and dispersed A. vaginae was visualised (OR 75.6; CI 13.3 to 429.5) and highest when both bacteria were part of the biofilm (OR 119; CI 39.9 to 360.8). CONCLUSIONS: Our study, although not comprehensive at studying the polymicrobial biofilm in BV, provided a strong indication towards the importance of A. vaginae and the symbiosis of A. vaginae and G. vaginalis in this biofilm. TRIAL REGISTRATION NUMBER: NCT01796613.