Evaluation of commercial, customized microdilution plates for Ureaplasma parvum, Ureaplasma urealyticum, and Mycoplasma hominis antimicrobial susceptibility testing and determination of antimicrobial resistance prevalence in France.

Antimicrobial susceptibility testing (AST) of human mycoplasmas using microdilution is time-consuming. In this study, we compared the performance of MICRONAUT-S plates (Biocentric-Bruker) designed for AST of Ureaplasma parvum, Ureaplasma urealyticum, and Mycoplasma hominis with the results using the Clinical & Laboratory Standards Institute (CLSI) reference method. Then, we investigated the prevalence and mechanisms of resistance to tetracyclines, fluoroquinolones, and macrolides in France in 2020 and 2021. The two methods were compared using 60 strains. For the resistance prevalence study, U. parvum-, U. urealyticum-, and M. hominis-positive clinical specimens were collected for 1 month each year in 22 French diagnostic laboratories. MICs were determined using the MICRONAUT-S plates. The tet(M) gene was screened using PCR, and fluoroquinolone resistance-associated mutations were screened using PCR and Sanger sequencing. Comparing the methods, 99.5% (679/680) MICs obtained using the MICRONAUT-S plates concurred with those obtained using the CLSI reference method. For 90 M. hominis isolates, the tetracycline, levofloxacin, and moxifloxacin resistance rates were 11.1%, 2.2%, and 2.2%, respectively, with no clindamycin resistance. For 248 U. parvum isolates, the levofloxacin and moxifloxacin resistance rates were 5.2% and 0.8%, respectively; they were 2.9% and 1.5% in 68 U. urealyticum isolates. Tetracycline resistance in U. urealyticum (11.8%) was significantly (P < 0.001) higher than in U. parvum (1.2%). No macrolide resistance was observed. Overall, the customized MICRONAUT-S plates are a reliable, convenient tool for AST of human mycoplasmas. Tetracycline and fluoroquinolone resistance remain limited in France. However, the prevalence of levofloxacin and moxifloxacin resistance has increased significantly in Ureaplasma spp. from 2010 to 2015 and requires monitoring. IMPORTANCE: Antimicrobial susceptibility testing of human urogenital mycoplasmas using the CLSI reference broth microdilution method is time-consuming and requires the laborious preparation of antimicrobial stock solutions. Here, we validated the use of reliable, convenient plates designed for antimicrobial susceptibility testing that allows the simultaneous determination of the MICs of eight antibiotics of interest. We then investigated the prevalence and mechanisms of resistance of each of these bacteria to tetracyclines, fluoroquinolones, and macrolides in France in 2020 and 2021. We showed that the prevalence of levofloxacin and moxifloxacin resistance has increased significantly in Ureaplasma spp. from 2010 to 2015 and requires ongoing monitoring.

Anorectal lymphogranuloma venereum among men who have sex with men: a 3-year nationwide survey, France, 2020 to 2022.

BackgroundIn France, lymphogranuloma venereum (LGV) testing switched from universal to selective testing in 2016.AimTo investigate changes in LGV-affected populations, we performed a nationwide survey based on temporarily reinstated universal LGV testing from 2020 to 2022.MethodsEach year, during three consecutive months, laboratories voluntarily sent anorectal Chlamydia trachomatis-positive samples from men and women to the National Reference Centre for bacterial sexually transmitted infections. We collected patients' demographic, clinical and biological data. Genovars L of C. trachomatis were detected using real-time PCR. In LGV-positive samples, the ompA gene was sequenced.ResultsIn 2020, LGV positivity was 12.7% (146/1,147), 15.2% (138/907) in 2021 and 13.3% (151/1,137) in 2022 (p > 0.05). It occurred predominantly in men who have sex with men (MSM), with rare cases among transgender women. The proportion of HIV-negative individuals was higher than that of those living with HIV. Asymptomatic rectal LGV increased from 36.1% (44/122) in 2020 to 52.4% (66/126) in 2022 (p = 0.03). Among users of pre-exposure prophylaxis (PrEP), LGV positivity was 13.8% (49/354) in 2020, 15.6% (38/244) in 2021 and 10.9% (36/331) in 2022, and up to 50% reported no anorectal symptoms. Diversity of the LGV ompA genotypes in the Paris region increased during the survey period. An unexpectedly high number of ompA genotype L1 variant was reported in 2022.ConclusionIn rectal samples from MSM in France, LGV positivity was stable, but the proportion of asymptomatic cases increased in 2022. This underscores the need of universal LGV testing and the importance of continuous surveillance.

Prevalence of Mycoplasma penetrans in Urogenital Samples From Men Screened for Bacterial Sexually Transmitted Infections.

Mycoplasma penetrans prevalence was assessed in urogenital samples from men screened for Chlamydia trachomatis and Neisseria gonorrhoeae. Prevalence was 3.5% among men who have sex with men and 5.3% among human immunodeficiency virus (HIV)-positive patients, significantly higher than in HIV-negative individuals (0.4%, P = .0016). No association was found between M. penetrans and urogenital symptoms.

Prevalence of macrolide and fluoroquinolone resistance-associated mutations in Mycoplasma genitalium in metropolitan and overseas France.

OBJECTIVE: Limited macrolide and fluoroquinolone resistance data are available in France for Mycoplasma genitalium. We performed a multicentre cross-sectional study to investigate the prevalence of macrolide and fluoroquinolone resistance-associated mutations in M. genitalium-positive patients in metropolitan France between 2018 and 2020 and in overseas France in 2018 and 2019. METHODS: Each year, a 1-month prospective collection of M. genitalium-positive specimens was proposed to metropolitan French microbiology diagnostic laboratories, and a similar 3-month collection was proposed to overseas French laboratories. Resistance-associated mutations were detected using commercial kits and sequencing. RESULTS: A total of 1630 M. genitalium-positive specimens were analysed. In metropolitan France, the prevalence of macrolide resistance-associated mutations ranged between 34.7% (95% CI 29.4% to 40.4%) and 42.9% (95% CI 37.1% to 49.0%) between 2018 and 2020 and was significantly higher in men (95% CI 52.4% to 60.2%) than in women (95% CI 15.9% to 22.2%) (p<0.001). These prevalences were significantly higher than those of 6.1% (95% CI 3.7% to 10.3%) and 14.7% (95% CI 10.9% to 19.6%) observed in overseas France in 2018 and 2019 (p<0.001), where no difference between genders was noted. The prevalence of fluoroquinolone resistance-associated mutations was also significantly higher in metropolitan France (14.9% (95% CI 11.2% to 19.5%) to 16.1% (95% CI 12.1% to 21.2%)) than in overseas France (1.3% (95% CI 0.4% to 3.7%) and 2.6% (95% CI 1.3% to 5.3%) in 2018 and 2019, respectively) (p<0.001), with no difference between men and women regardless of the location. CONCLUSION: This study reports the high prevalence of macrolide and fluoroquinolone resistance-associated mutations in M. genitalium in metropolitan France and highlights the contrast with low prevalence in overseas France. In metropolitan France, macrolide resistance-associated mutation prevalence was three times higher in men than in women, which was likely to be driven by the proportion of men who have sex with men. This suggests that gender and sexual practice should also be taken into account for the management of M. genitalium infections.

Spread of clonal genovar E Chlamydia trachomatis among men who have sex with men.

In a previous study, we developed a Multi-Locus VNTRs Analysis (MLVA) typing system, called MLVA-5, for the discrimination of Chlamydia trachomatis genovar E strain. The results suggested the clonal spread of a MLVA-5 type 21 strain among men who have sex with men (MSM). We applied the MLVA-5 typing method on 157 French anorectal genovar E specimens and 19 Swedish specimens collected between 2010 and 2015. A total of 29 MLVA-5 types was obtained, with three predominant types among French samples: 78 specimens belonged to MLVA-5 type 21, two other types, 11 and 13, included 9 and 14 specimens, respectively. In 15 cases, one unique MLVA-5 type was observed for a single patient, 7 of which were new types not previously described. The distribution of MLVA-5 types according to sexual orientation showed that the 7 anorectal specimens from heterosexual patients belonged to 6 genotypes, and the 12 anorectal specimens from bisexual patients comprised eight types. The 95 anorectal specimens from MSM were distributed into 22 types, but 55 (57.9%) of them belonged to MLVA-5 type 21. Among the Swedish specimens from MSM, eight were from MLVA-type 21 (4 urines and 4 anorectal specimens). The results support the hypothesis of the spread of clonal genovar E strain among MSM.

Evaluation of four commercial real-time PCR assays for the detection of lymphogranuloma venereum in Chlamydia trachomatis-positive anorectal samples.

OBJECTIVES: Lymphogranuloma venereum (LGV) is a sexually transmitted infection (STI) caused by Chlamydia trachomatis (CT) genovars L. The identification of LGV is of therapeutic interest because treatment requires 3 weeks of doxycycline compared with 1 week for infection with a non-L strain. The aim of this study was to evaluate the performance of four commercial real-time PCR kits in comparison with the reference methods used for LGV diagnosis by the French National Reference Centre (NRC) for bacterial STIs. METHODS: A total of 215 French CT-positive anorectal specimens collected consecutively in 2017 were used (66 LGV and 149 non-LGV). Among these, 92 were collected from symptomatic men who have sex with men (MSM) and 123 from asymptomatic MSM using pre-exposure prophylaxis. Four commercial assays were evaluated; a single-plex assay RealCycler CHSL kit (Progenie Molecular), tested on all the specimens, and three multiplex kits, the RealCycler Universal ULCGEN (Progenie Molecular), the Allplex Genital Ulcer Assay (Seegene) and the VIASURE Haemophilus ducreyi + CT LGV Real Time PCR Detection kit (CerTest Biotec), tested on the 92 samples from symptomatic MSM. Clinical performance was determined in comparison to the in-house real time PCR targeting the pmpH and the ompA gene sequencing. RESULTS: Overall agreement ranged between 91.3% and 100% (95% CI 83.7-100%) with very good Kappa index values (>0.8). The clinical sensitivities and specificities varied between 91% and 100% (95% CI 80.8-100%), and 97% and 100% (95% CI 87.1-100%), respectively, with some kits performing better than others. DISCUSSION: The four assays showed very good performance for the detection of LGV on anorectal specimens.

Prevalence of lymphogranuloma venereum among anorectal Chlamydia trachomatis-positive MSM using pre-exposure prophylaxis for HIV.

OBJECTIVES: We evaluated the prevalence of lymphogranuloma venereum (LGV) in anorectal Chlamydia trachomatis-positive French men who have sex with men (MSM) using pre-exposure prophylaxis (PrEP) for HIV. Here, we describe the clinical, biological and behavioural characteristics of these patients. METHODS: Laboratories throughout French metropolitan areas performing routine testing for C. trachomatis sent positive anorectal specimens to the National Reference Centre for bacterial STIs for LGV real-time PCR targeting the pmpH gene. Identification of the C. trachomatis genovar was performed by ompA gene sequencing. For each patient, clinical, biological and sexual behaviour data were collected after obtaining written informed consent. RESULTS: In 2017, 486 anorectal C. trachomatis-positive specimens from MSM PrEP users were analysed. A strain of genovar L was detected in 91 cases (18.7%). Patients with LGV were significantly more symptomatic, had more sexual partners and more concurrent syphilis compared with their non-LGV counterparts. OmpA gene sequencing, successful in two-thirds of anorectal C. trachomatis-positive specimens, showed that the LGV cases were mainly of variant L2b (n=33), followed by genovar L2 (n=27) and genetic L2b ompA variants (n=16). In 11 cases, the results indicated the occurrence of genetic exchange between L and non-L genovars. CONCLUSIONS: LGV was diagnosed in 18.7% of anorectal C. trachomatis-positive specimens from French MSM using PrEP. LGV testing should be carried out for MSM diagnosed with chlamydia and with a large number of sexual partners, high-risk practices and anorectal symptoms. These patients should be presumptively treated as having LGV. This is the first surveillance study of LGV among MSM PrEP users and monitoring should continue.

Extra-rectal lymphogranuloma venereum in France: a clinical and molecular study.

OBJECTIVES: To describe a series of extrarectal lymphogranuloma venereum (LGV) cases diagnosed in France. METHODS: Consecutive LGV cases confirmed at the French Reference Centre for chlamydiae with an extrarectal sample from January 2010 to December 2015 were included. The first part of the study consisted of a retrospective case note review and analysis. In a second part, the complete ompA gene sequence of our samples was determined. RESULTS: There were 56 cases overall: 50 cases of genital LGV and six cases of pharyngeal LGV. Subjects were all men, median age 39 years, 27/53 were HIV-positive, 47/51 reported having sex with other men, 43/49 reported multiple sexual partners (a mean 25 in the last 6 months). Median time from symptom onset to diagnosis was 21 days. Subjects most commonly presented with inguinal adenopathy alone (19 of 50 genital cases) and adenopathy with genital ulcer (17 of 50). Three pharyngeal cases were symptomatic. Fever was reported in 11 cases. Inguinal abscess was reported in 22 of 42 cases presenting with lymphadenopathy. Co-infections were frequent: eight cases of syphilis, four non-LGV Chlamydia trachomatis infections, one case of gonorrhoea. Cure was always achieved with doxycycline therapy but prolonged treatment was necessary in eight cases with inguinal abscess. Genotyping according to ompA sequencing showed the co-circulation of genovars L2 (16 of 42 strains successfully typed) and L2b (24 of 42). There was no association between HIV status and disease severity or genovar distribution. CONCLUSION: In the span of 6 years, 56 extrarectal LGV cases were confirmed through genotyping in France. Extrarectal LGV seemed to share a common epidemiological background with rectal disease in terms of affected population and genovar distribution. HIV prevalence was lower than expected.

Changing Pattern of Chlamydia trachomatis Strains in Lymphogranuloma Venereum Outbreak, France, 2010-2015.

We describe a change in the molecular epidemiology of Chlamydia trachomatis strains involved in an outbreak of rectal lymphogranuloma venereum in France during January 2010-April 2015. Until 2012, the C. trachomatis L2b strain predominated; however, starting in 2013, most cases involved the L2 strain. We also identified 4 genetic L2b ompA variants.