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.

Multi-component prime-boost Chlamydia trachomatis vaccination regimes induce antibody and T cell responses and accelerate clearance of infection in a non-human primate model.

It is of international priority to develop a vaccine against sexually transmitted Chlamydia trachomatis infections to combat the continued global spread of the infection. The optimal immunization strategy still remains to be fully elucidated. The aim of this study was to evaluate immunization strategies in a nonhuman primate (NHP) model. Cynomolgus macaques (Macaqua fascicularis) were immunized following different multi-component prime-boost immunization-schedules and subsequently challenged with C. trachomatis SvD in the lower genital tract. The immunization antigens included the recombinant protein antigen CTH522 adjuvanted with CAF01 or aluminium hydroxide, MOMP DNA antigen and MOMP vector antigens (HuAd5 MOMP and MVA MOMP). All antigen constructs were highly immunogenic raising significant systemic C. trachomatis-specific IgG responses. In particularly the CTH522 protein vaccinated groups raised a fast and strong pecificsIgG in serum. The mapping of specific B cell epitopes within the MOMP showed that all vaccinated groups, recognized epitopes near or within the variable domains (VD) of MOMP, with a consistent VD4 response in all animals. Furthermore, serum from all vaccinated groups were able to in vitro neutralize both SvD, SvE and SvF. Antibody responses were reflected on the vaginal and ocular mucosa, which showed detectable levels of IgG. Vaccines also induced C. trachomatis-specific cell mediated responses, as shown by in vitro stimulation and intracellular cytokine staining of peripheral blood mononuclear cells (PBMCs). In general, the protein (CTH522) vaccinated groups established a multifunctional CD4 T cell response, whereas the DNA and Vector vaccinated groups also established a CD8 T cells response. Following vaginal challenge with C. trachomatis SvD, several of the vaccinated groups showed accelerated clearance of the infection, but especially the DNA group, boosted with CAF01 adjuvanted CTH522 to achieve a balanced CD4/CD8 T cell response combined with an IgG response, showed accelerated clearance of the infection.

Clinical Performance of Three Commercial Molecular Diagnostic Assays for the Detection of Fluoroquinolone Resistance-Associated Mutations in Mycoplasma genitalium.

The high prevalence of macrolide resistance in Mycoplasma genitalium results in an increased reliance on moxifloxacin, the second-line treatment; however, moxifloxacin resistance has also emerged. Because assays that can detect fluoroquinolone resistance-associated mutations will be useful for the management of macrolide-resistant M. genitalium infections, we evaluated the performance of three commercial assays (the Allplex MG & MoxiR Assay [Seegene], LightMix Modular parC kit [TIBMOLBIOL], and MGMO qPCR [NYtor) in comparison with parC gene Sanger sequencing used as the reference. Between January 2018 and December 2020, remnants of M. genitalium-positive clinical specimens received at the French National Reference Center for Bacterial Sexually Transmitted Infections were collected if a Sanger sequencing result was obtained for the parC gene. Overall, 368 M. genitalium-positive specimens were assessed. The clinical sensitivities for the detection of the ParC mutations that are likely of clinical significance were 91.8% (95% CI = 83.2 to 96.2), 98.6% (95% CI = 92.4 to 99.8), and 94.4% (95% CI = 86.6 to 97.8) for the Allplex MG & MoxiR, LightMix Modular parC, and MGMO qPCR kits, respectively, with no significant difference between the three kits. The clinical specificity of the Allplex MG & MoxiR and MGMO qPCR kits was 100% (95% CI = 97.7 to 100 and 98.7 to 100, respectively), which was significantly higher than the specificity of the LightMix Modular parC kit of 95.4% (95%CI = 92.3 to 97.3), for which the interpretation of melting curves may be misleading. These kits should be useful for the selection of antimicrobials in macrolide-resistant M. genitalium infections, although further developments may be necessary because parC mutations involved in fluoroquinolone resistance have not been precisely determined.

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.

Identification of 16S rRNA mutations in Mycoplasma genitalium potentially associated with tetracycline resistance in vivo but not selected in vitro in M. genitalium and Chlamydia trachomatis.

OBJECTIVES: Tetracyclines are widely used for the treatment of bacterial sexually transmitted infections (STIs) and recently have been used successfully for post-exposure prophylaxis of STIs in MSM. We investigated the in vitro and in vivo development of tetracycline resistance in Chlamydia trachomatis and Mycoplasma genitalium and evaluated 16S rRNA mutations associated with acquired resistance in other bacteria. METHODS: In vitro selection of resistant mutants of reference strains of C. trachomatis and M. genitalium was undertaken by serial passage in medium containing subinhibitory concentrations of tetracycline or doxycycline, respectively. The 16S rRNA gene of the two microorganisms was amplified and sequenced at different passages, as were those of 43 C. trachomatis- and 106 M. genitalium-positive specimens collected in France from 2013 to 2019. RESULTS: No tetracycline- or doxycycline-resistant strains of C. trachomatis and M. genitalium, respectively, were obtained after 30 serial passages. The tetracycline and doxycycline MICs were unchanged and analysis of the 16S rRNA gene, the molecular target of tetracyclines, of C. trachomatis and M. genitalium revealed no mutation. No mutation in the 16S rRNA gene was detected in C. trachomatis-positive specimens. However, six M. genitalium-positive specimens harboured a mutation potentially associated with tetracycline resistance without known prior tetracycline treatment for patients. CONCLUSIONS: Tetracyclines did not select in vitro-resistant mutants of C. trachomatis or M. genitalium. However, 16S rRNA mutations either responsible for or associated with tetracycline resistance in other bacteria, including mycoplasma species, were identified in several M. genitalium-positive specimens.

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.

Did L Strains Responsible for Lymphogranuloma Venereum Proctitis Spread Among People With Genital Chlamydia trachomatis Infection in France in 2013?

We retrospectively analyzed 1802 nonrectal Chlamydia trachomatis-positive specimens to determine if the L strains responsible for rectal Lymphogranuloma venereum in men who have sex with men could spread to the heterosexual population. No evidence for Lymphogranuloma venereum transmission among heterosexuals in France was observed in 2013. L2b strains seem to be restricted to the men who have sex with men population.

Surface lipoproteome of Mycoplasma hominis PG21 and differential expression after contact with human dendritic cells.

AIM: To assess the lipoproteins that are involved in the interaction between Mycoplasma hominis and human dendritic cells. MATERIALS & METHODS: The surface lipoproteome of M. hominis PG21 was characterized by using Triton X-114 extraction and LC-MS/MS identification. The transcriptional changes in lipoprotein genes upon contact with human dendritic cells were determined by using reverse transcription quantitative PCR after identification of reference genes suitable for normalization. RESULTS: A large-scale overexpression of lipoprotein genes was observed with 21 upregulated transcripts. Seven genes of unknown function were M. hominis species specific and six genes were putatively associated with increased nutrient capture from the host cell and adhesion. CONCLUSION: M. hominis regulates lipoprotein gene expression and may use species-specific mechanisms during the host colonization process.

Direct detection of macrolide resistance in Mycoplasma genitalium isolates from clinical specimens from France by use of real-time PCR and melting curve analysis.

Mycoplasma genitalium is a sexually transmitted organism commonly treated with azithromycin. However, macrolide resistance has been reported and is associated with point mutations in the 23S rRNA gene. To evaluate the prevalence of macrolide resistance in M. genitalium isolates from clinical specimens from France, we first used a previously reported high-resolution melting assay. Because susceptible and resistant M. genitalium isolates were hardly discriminated in M. genitalium-positive clinical specimens, we developed a new molecular assay for the rapid detection of macrolide resistance. An assay using real-time PCR based on fluorescence resonance energy transfer (FRET) coupled with melting curve analysis was designed. The assay was first validated on characterized macrolide-resistant M. genitalium isolates and then applied to 202 urogenital M. genitalium-positive specimens collected from 178 patients from France in 2011 and 2012. Resistant genotypes were confirmed by 23S rRNA gene sequencing. Among the 202 M. genitalium-positive specimens, 155 were amplified, demonstrating a sensitivity of 76.7%. A substitution in the 23S rRNA gene was found in 14.2% of the patient samples. Nine and six patients had M. genitalium isolates with a substitution at positions 2059 and 2058, respectively. In four cases, a mixed population of wild-type and mutated M. genitalium isolates was observed. The prevalence of M. genitalium macrolide resistance has been stable in France since its detection in 2006. Our FRET PCR assay is able to discriminate between wild-type and resistant genotypes directly from clinical specimens. This assay will allow clinicians to shorten the time to the initiation of effective disease treatment.

The spread of Mycoplasma pneumoniae is polyclonal in both an endemic setting in France and in an epidemic setting in Israel.

Mycoplasma pneumoniae infections occur both endemically and epidemically, and macrolide resistance has been spreading for 10 years worldwide. A substantial increased incidence of M. pneumoniae infections has been reported in several countries since 2010. Whether this increased incidence is attributed to different or to the same M. pneumoniae genotype is unknown. We have developed a multilocus variable-number tandem-repeat (VNTR) analysis (MLVA) for the molecular typing of M. pneumoniae isolates. In this study, the MLVA typing method was modified and validated to be applicable directly to respiratory tract specimens without culture. This method was applied to 34 M. pneumoniae-positive specimens received at the Bordeaux Hospital, France, between 2007 and 2010 in an endemic setting, and to 63 M. pneumoniae-positive specimens collected during an epidemic surge of M. pneumoniae infections in 2010 in Jerusalem, Israel. The M. pneumoniae endemic spread was shown to be polyclonal in France, with 15 MLVA types identified. Strikingly, the Israeli epidemic surge was also a multi-clonal phenomenon, with 18 circulating MLVA types. The macrolide resistance-associated substitution, A2058G, was found in 22% of the Israeli patients. Macrolide-resistant M. pneumoniae belonged to four MLVA types, the MLVA type Z being the most frequent one. An association between the MLVA type Z and macrolide resistance might exist since macrolide resistance was present or generated during the course of illness in all patients infected with this MLVA type. In conclusion, the discriminatory power of the MLVA showed that the spread of M. pneumoniae strains in France in an endemic setting was polyclonal as well as the surge of M. pneumoniae infections in Israel in 2010.

Phenotypic and genotypic characterization of Salmonella enterica recovered from poultry meat in Tunisia and identification of new genetic traits.

Thirty-seven Salmonella enterica isolates obtained from poultry meat in Tunisia were included in this study for characterization of antibiotic resistance mechanisms. High percentages of resistance were detected to ampicillin, sulfonamides, tetracycline, nalidixic acid, and streptomycin (32.4%-89.2%), and lower percentages to amoxicillin-clavulanic acid, kanamycin, amikacin, trimethoprim-sulfamethoxazol, and chloramphenicol (2.7%-18.9%). All strains showed susceptibility to ceftazidime, cefotaxime, gentamicin, and ciprofloxacin. Class 1 integrons were detected in 30% of Salmonella isolates, and four different gene cassette arrangements were detected, including genes implicated in resistance to aminoglycosides (aadA1 and aadA2) and trimethoprim (dfrA1). Four different Pc variants (PcW, PcH1, PcH1(TTN-10), PcW(TGN-10)) with inactive P2 have been found among these isolates. Integron-positive isolates were ascribed to eight different serotypes. A Salmonella Schwarzengrund isolate harbored a new class 1 integron containing the qacH-dfrA1b-aadA1b-catB2 gene cassette arrangement, with the very unusual PcH1(TTN-10) promoter, which has been registered in GenBank (accession no. HQ874651). Different plasmid replicon types were demonstrated among integron-positive isolates: IncI1 (8 isolates), IncN (8), IncP (2), IncFIB (2), and IncFII (2). Ten different pulsed-field gel electrophoresis profiles were detected among the 11 integron-positive isolates and 8 different sequence types were identified by multilocus sequence typing, one of them (registered as ST867) was new, detected in 3 Salmonella Zanzibar isolates. A high diversity of clones is observed among poultry Salmonella isolates and a high proportion of them show a multiresistant phenotype with very diverse mobile genetic structures that could be implicated in bacterial dissemination in different environments.