Performance of the first commercial dual resistance assay, AmpliSens Mycoplasma genitalium-ML/FQ-Resist-FL, for detection of potential macrolide and quinolone resistance-associated mutations and prevalence of M. genitalium resistance mutations in St. Petersburg, Russia.

OBJECTIVES: Antimicrobial resistance in Mycoplasma genitalium (MG) is a poorly surveyed and controlled global health concern. We evaluated the first commercial dual resistance assay, AmpliSens M. genitalium-ML/FQ-Resist-FL assay, for detection of potential macrolide and quinolone resistance-associated mutations (MRAMs and QRAMs, respectively) and estimated the prevalence of these mutations in MG in St. Petersburg, Russia. METHODS: Urogenital samples positive (n=145 from 2007 to 2020) and negative (n=56 from 2021) for MG in routine diagnostics were retrospectively analysed using the AmpliSens M. genitalium-ML/FQ-Resist-FL assay (Central Research Institute of Epidemiology, Moscow, Russia) and Sanger sequencing for validation. RESULTS: The AmpliSens M. genitalium-ML/FQ-Resist-FL assay detected potential MRAMs and QRAMs with sensitivities of 100% (CI95% 83.9 to 100) and 92.3% (CI95% 66.7 to 99.6) and specificities of 99.2% (CI95% 95.6 to 100) and 100% (CI95% 97.2 to 100), respectively, in clinical specimens with ≥1000 MG geq/mL. In total, MRAMs were detected in 13.8% (CI95% 9.1 to 20.3) of samples, with 23S rRNA A2058G being the most prevalent mutation (45.0% (CI95% 25.8 to 65.8)). QRAMs were found in 9.0% (CI95% 5.3 to 14.7) of samples, with S83I the most frequent mutation (53.8% (CI95% 29.1 to 76.8)). Dual resistance was observed in 5.5% (CI95% 2.8 to 10.5) of samples. Potential MRAM and dual resistance rates significantly increased over time: from 0% in 2007-2008 to 25% (p trend =0.0009) and 10% (p trend =0.0447), respectively, in 2018-2020. QRAM rate appeared to increase (from 0% to 13%), but significance was not reached (p trend =0.0605). CONCLUSIONS: The rapid increase in MG antimicrobial resistance in St. Petersburg, especially prominent for MRAMs, necessitates implementation of macrolide resistance-guided therapy in Russia. The first commercial dual resistance assay, AmpliSens M. genitalium-ML/FQ-Resist-FL assay, was sensitive and specific for detection of potential MRAMs and QRAMs and could be valuable in macrolide resistance-guided therapies and possibly for surveillance of QRAMs. International surveillance of antimicrobial resistance-associated mutations in MG, further research into clinical relevance of several parC mutations and novel treatments are essential.

Molecular Testing for the Diagnosis of Bacterial Vaginosis.

Previously established diagnostic approaches for the diagnosis of bacterial vaginosis (BV), such as the Amsel criteria or the Nugent scoring system, do not always correspond to modern trends in understanding the etiology and pathogenesis of polymicrobial conditions. Inter-examiner variability and interpretation of data complicate the wet mount microscopy method. Gram staining of smears does not always provide reliable information regarding bacterial taxa, biofilms, or vaginal dysbiosis. Therefore, the introduction of molecular techniques into clinical practice is extremely relevant. Molecular approaches allow not only the diagnosis of BV but also provide an assessment of microbial composition, which is especially important in the differential diagnosis of vaginal infections. The current review represents an expert opinion on BV diagnosis and is based on extensive experience in the field of vaginal infection diagnosis and treatment.

Vaginal Microbiota Molecular Profiling in Women with Bacterial Vaginosis: A Novel Diagnostic Tool.

Bacterial vaginosis (BV) is a most common microbiological syndrome. Multiplex next-generation sequencing (NGS) or molecular tests allow a complete and accurate vaginal microbiota profiling in order to determine the primary causative agent. Due to the high costs and limited availability of NGS, the multiplex real-time PCR draws more attention. The present study aimed to evaluate the microbial composition and dominant lactobacilli species in non-pregnant women with bacterial vaginosis using a multiplex RT-PCR test and determine its diagnostic significance. In total, 331 women complaining of vaginal discharge were included. BV was confirmed upon clinical examination and Nugent criteria. A real-time PCR test was carried out with a new Femoflor test, which identifies opportunistic bacteria, STD pathogens, and some viruses. According to the results, the rate of lactobacilli is significantly reduced in BV-affected patients when compared to healthy women. Moreover, the rate of L. crispatus significantly decreases, while the rate of L. iners remains high. Among obligate anaerobic bacteria, Gardnerella vaginalis was the most prevalent in women with BV. The Femoflor test demonstrated high sensitivity and specificity for diagnosing BV. Moreover, the test allows the identification of infection in women with intermediate vaginal microbiota, as well as STD pathogens, and viruses. Thus, the application of real-time PCR tests can be effectively used in vaginal microbiota evaluation in women with BV, intermediate vaginal microbiota, and healthy women. In addition, this test may be used as an alternative to the Amsel criteria and Nugent scoring method in diagnosing BV.

Local Immune Biomarker Expression Depending on the Uterine Microbiota in Patients with Idiopathic Infertility.

The endometrium has traditionally been considered sterile. Nowadays, active studies are performed on the female upper genital tract microbiota. Bacteria and/or viruses colonizing the endometrium are known to alter its functional properties, including receptivity and embryo implantation. Uterine cavity inflammation caused by microorganisms leads to disrupted cytokine expression, which, in turn, is mandatory for the successful implantation of the embryo. The present study assessed the vaginal and endometrial microbiota composition and its relation to the levels of cytokines produced by the endometrium in reproductive-aged women complaining of secondary infertility of unknown origin. The multiplex real-time PCR assay was applied for vaginal and endometrial microbiota analysis. The quantitative measurement of endometrial α-defensin (DEFa1), transforming growth factor (TGFß1), and basic fibroblast growth factor (bFGF2) was carried out using the ELISA (Cloud-Clone Corporation (Katy, TX, USA; manufactured in Wuhan, China). A reliable decline in endometrial TGFß1 and bFGF2 and an increase in DEFa1 were demonstrated in women with idiopathic infertility when compared to fertile patients. However, TGFß1, bFGF2, and DEFa1 expression correlated reliably only with the presence of Peptostreptococcus spp. and HPV in the uterine cavity. The obtained results highlight the importance of local immune biomarker determination in the assessment of certain bacteria and viruses' significance as causative agents of infertility.

Comprehensive global genome dynamics of Chlamydia trachomatis show ancient diversification followed by contemporary mixing and recent lineage expansion.

Chlamydia trachomatis is the world's most prevalent bacterial sexually transmitted infection and leading infectious cause of blindness, yet it is one of the least understood human pathogens, in part due to the difficulties of in vitro culturing and the lack of available tools for genetic manipulation. Genome sequencing has reinvigorated this field, shedding light on the contemporary history of this pathogen. Here, we analyze 563 full genomes, 455 of which are novel, to show that the history of the species comprises two phases, and conclude that the currently circulating lineages are the result of evolution in different genomic ecotypes. Temporal analysis indicates these lineages have recently expanded in the space of thousands of years, rather than the millions of years as previously thought, a finding that dramatically changes our understanding of this pathogen's history. Finally, at a time when almost every pathogen is becoming increasingly resistant to antimicrobials, we show that there is no evidence of circulating genomic resistance in C. trachomatis.

Significant shifts in the distribution of vaccine capsular polysaccharide types and rates of antimicrobial resistance of perinatal group B streptococci within the last decade in St. Petersburg, Russia.

Evaluation of circulating group B streptococcus (GBS) strains is important to assess the potential effects of GBS prevention strategies in a certain region. This study aimed at estimating the distributions of capsular polysaccharide (CPS) types and pilus profiles, and the rates of antimicrobial resistance among GBS strains isolated from colonized pregnant women and newborns in 2010-2011 and 2017-2018 in St. Petersburg, Russia. A total of 261 GBS isolates have been investigated. Antibiotic susceptibility testing was performed using the disc-diffusion method. CPS types and pilus profiles were determined by using PCR. Over the 9-year period, the resistance of GBS to both erythromycin and clindamycin has significantly increased, exceeding 30% in 2017-2018. The most prevalent CPS types were Ia, III, and V. Significant shifts were observed in the frequency of CPS types III (decreased) and V (increased), which resulted in a significant reduction (from 77 to 63%) in the potential coverage by a trivalent vaccine (including serotypes Ia, Ib, and III), whereas that of a pentavalent vaccine (including serotypes Ia, Ib, II, III, and V) remained largely unchanged (approximately 95%). The most common pilus profiles were PI-1/2a, PI-2a, and PI-1a/2b, and pilus genotype distribution has not changed with time. High and steadily growing resistance of perinatal GBS strains to clindamycin requires restricting its use to penicillin-allergic women at high risk for anaphylaxis and testing the GBS strains for their susceptibility to this antibiotic. A pentavalent CPS-based vaccine covers the vast majority of perinatal GBS strains in Russia.

Bacterial vaginosis-associated vaginal microbiota is an age-independent risk factor for Chlamydia trachomatis, Mycoplasma genitalium and Trichomonas vaginalis infections in low-risk women, St. Petersburg, Russia.

The large majority of studies investigating associations between bacterial vaginosis (BV) and sexually transmitted infections (STIs) have been conducted among predominantly young women with high risk for STIs. Since a risky sexual behavior is a significant risk factor for both STIs and BV, this creates a bias toward an increased association between BV and STIs. This study evaluated associations between BV-associated vaginal microbiota and STIs (Chlamydia trachomatis, Mycoplasma genitalium, Trichomonas vaginalis, and Neisseria gonorrhoeae) in a population of women with low risk for STIs and investigated STI outcomes depending on the dominating Lactobacillus species. Repository cervicovaginal samples collected from reproductive-age women from January 2014 to February 2019 were characterized for vaginal microbiota types and the STIs using multiplex real-time PCR assays. In total, 95 STI-positive and 91 STI-negative samples were included. A significant, age-independent association between BV-associated vaginal microbiota and the presence of C. trachomatis, M. genitalium, and T. vaginalis infections was identified (age-adjusted odds ratios 2.92 [95% confidence interval (CI) 1.24-7.03], 2.88 [95% CI 1.19-7.16], and 9.75 × 107 [95% CI 13.03-∞], respectively). Normal vaginal microbiota dominated by Lactobacillus crispatus, L. gasseri, or L. jensenii was a strong protective factor against C. trachomatis and/or M. genitalium infections, whereas L. iners-dominated microbiota was not significantly associated with C. trachomatis and/or M. genitalium positivity. The results of the present study confirm that STI prevention strategies should include interventions that also reduce the incidence of BV and promote a protective vaginal microbiota in both high- and low-risk women.

Secure application of graphene in medicine.

OBJECTIVE: The objective of our study was to explore the possible graphene impact on microorganism growth as well as on laboratory animal overall condition. Materials and technique: The experiments applied samples of graphene three concentrations and two 15 × 15 mm quartz glasses one of which carrying deposited graphene lattice. We have also used 5% blood agar, thioglycollate broth, bacterial suspensions of standard turbidity containing pure clinical isolate of microorganisms. Three white male 6-month-old laboratory rats were used to estimate the graphene impact on the overall animal condition. RESULTS: Graphene did not contain any microorganisms, does not destroy erythrocytes placed within the artificial nutritional medium, graphene lattice did not add any properties to the quartz glasses which could allow the Proteus spread all over its surface. It was also established that graphene did not show any native antibacterial impact. No significant reaction was noticed in animals after graphene administration to laboratory rats neither at the injection spot nor at the overall level. CONCLUSION: Our data confirm the applicability of graphene both in scientific and practical biomedical purposes.

Human papillomavirus prevalence and type-distribution in cervical glandular neoplasias: Results from a European multinational epidemiological study.

Cervical glandular neoplasias (CGN) present a challenge for cervical cancer prevention due to their complex histopathology and difficulties in detecting preinvasive stages with current screening practices. Reports of human papillomavirus (HPV) prevalence and type-distribution in CGN vary, providing uncertain evidence to support prophylactic vaccination and HPV screening. This study [108288/108290] assessed HPV prevalence and type-distribution in women diagnosed with cervical adenocarcinoma in situ (AIS, N = 49), adenosquamous carcinoma (ASC, N = 104), and various adenocarcinoma subtypes (ADC, N = 461) from 17 European countries, using centralised pathology review and sensitive HPV testing. The highest HPV-positivity rates were observed in AIS (93.9%), ASC (85.6%), and usual-type ADC (90.4%), with much lower rates in rarer ADC subtypes (clear-cell: 27.6%; serous: 30.4%; endometrioid: 12.9%; gastric-type: 0%). The most common HPV types were restricted to HPV16/18/45, accounting for 98.3% of all HPV-positive ADC. There were variations in HPV prevalence and ADC type-distribution by country. Age at diagnosis differed by ADC subtype, with usual-type diagnosed in younger women (median: 43 years) compared to rarer subtypes (medians between 57 and 66 years). Moreover, HPV-positive ADC cases were younger than HPV-negative ADC. The six years difference in median age for women with AIS compared to those with usual-type ADC suggests that cytological screening for AIS may be suboptimal. Since the great majority of CGN are HPV16/18/45-positive, the incorporation of prophylactic vaccination and HPV testing in cervical cancer screening are important prevention strategies. Our results suggest that special attention should be given to certain rarer ADC subtypes as most appear to be unrelated to HPV.

Differences in human papillomavirus type distribution in high-grade cervical intraepithelial neoplasia and invasive cervical cancer in Europe.

Knowledge of differences in human papillomavirus (HPV)-type prevalence between high-grade cervical intraepithelial neoplasia (HG-CIN) and invasive cervical cancer (ICC) is crucial for understanding the natural history of HPV-infected cervical lesions and the potential impact of HPV vaccination on cervical cancer prevention. More than 6,000 women diagnosed with HG-CIN or ICC from 17 European countries were enrolled in two parallel cross-sectional studies (108288/108290). Centralised histopathology review and standardised HPV-DNA typing were applied to formalin-fixed paraffin-embedded cervical specimens dated 2001-2008. The pooled prevalence of individual HPV types was estimated using meta-analytic methods. A total of 3,103 women were diagnosed with HG-CIN and a total of 3,162 with ICC (median ages: 34 and 49 years, respectively), of which 98.5 and 91.8% were HPV-positive, respectively. The most common HPV types in women with HG-CIN were HPV16/33/31 (59.9/10.5/9.0%) and in ICC were HPV16/18/45 (63.3/15.2/5.3%). In squamous cell carcinomas, HPV16/18/33 were most frequent (66.2/10.8/5.3%), and in adenocarcinomas, HPV16/18/45 (54.2/40.4/8.3%). The prevalence of HPV16/18/45 was 1.1/3.5/2.5 times higher in ICC than in HG-CIN. The difference in age at diagnosis between CIN3 and squamous cervical cancer for HPV18 (9 years) was significantly less compared to HPV31/33/'other' (23/20/17 years), and for HPV45 (1 year) than HPV16/31/33/'other' (15/23/20/17 years). In Europe, HPV16 predominates in both HG-CIN and ICC, whereas HPV18/45 are associated with a low median age of ICC. HPV18/45 are more frequent in ICC than HG-CIN and associated with a high median age of HG-CIN, with a narrow age interval between HG-CIN and ICC detection. These findings support the need for primary prevention of HPV16/18/45-related cervical lesions.

Antimicrobial resistance profiles and genes in uropathogenic Enterobacteriaceae in reproductive-age women in St. Petersburg, Russia: implication for treatment of pyelonephritis in pregnancy.

BACKGROUND: Urinary tract infections (UTIs) are common in women, and during pregnancy can cause significant morbidity. Growing and greatly varying antimicrobial resistance (AMR) of Enterobacteriaceae, responsible for most UTIs, necessitates regular local AMR surveillance. In obstetric population, where beta-lactams are the mainstay for treatment of severe UTIs, particular focus should be placed on beta-lactam resistance. This study aimed to evaluate AMR rates and frequency of extended spectrum beta-lactamase (ESBL) and carbapenemase genes in uropathogenic Enterobacteriaceae among reproductive-age women in St. Petersburg, Russia. MATERIALS/METHODS: Urine samples were collected from consecutive reproductive-age women, who attended the D.O. Ott Research Institute of Obstetrics, Gynecology and Reproductology from October 2017 to November 2019, and cultured according to routine procedures. Susceptibility to antibiotics and ESBL production was determined using the disc diffusion method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. All urine samples and Enterobacteriaceae isolates were tested for ESBL and carbapenemase genes using real-time PCR. RESULTS: Enterobacteriaceae were detected in 91 (56 pregnant and 35 non-pregnant) of 119 (76%) included women. The vast majority of Enterobacteriaceae strains were susceptible to nitrofurantoin, fosfomycin and meropenem (99-100%). The frequency of strains susceptible to penicillins and cephalosporins ranged from 59% to 82%; 78% of strains were susceptible to ciprofloxacin. ESBL production was phenotypically detected in 15 (16%) Enterobacteriaceae strains, with CTX-M genes revealed in all cases. In all corresponding urine samples, CTX-M genes were also detected. The remaining 104 urine samples were negative for CTX-M genes. In none of the isolates and urine samples, carbapenemase genes were present. CONCLUSIONS: The frequency of ESBL producing Enterobacteriaceae was relatively high (16%), with CTX-M genes detected in all cases in both urine and urine cultures. Rapid PCR detection of CTX-M genes directly in urine samples from women with pyelonephritis can be valuable for timely informing treatment choices.

Survey of diagnostic services for genital herpes in fourteen countries in Eastern Europe.

This paper reports survey-based data on the diagnosis and management of genital herpes simplex virus (HSV) infection in 14 countries of the Eastern European Network for Sexual and Reproductive Health (EE SRH). Only 43% of the countries could provide the number of genital HSV cases recorded at national level. Eighty-six percent of countries employed syndromic management in cases of genital ulcer disease. Most countries performed type-specific and/or non-type-specific enzyme immunoassays to detect HSV antibodies. Non-type-specific serology for diagnostic purposes should be actively discouraged. Direct detection methods for HSV, such as PCR, antigen detection and culture, are available in the region, but their usage was extremely low. Their use in Eastern European countries should be actively promoted. The availability of laboratory services must be improved, and countries in the region should implement consensus recommendations for the laboratory diagnosis of genital HSV infections in order to improve clinical practice.

Guidelines for the laboratory diagnosis of mycoplasma genitalium infections in East European countries.

The present guidelines aim to provide comprehensive information regarding laboratory diagnosis of Mycoplasma genitalium infections in East European countries. These guidelines are intended primarily for laboratory professionals testing specimens from patients at sexual health care clinics, but may also be useful for community-based screening programmes. Diagnosis of M. genitalium infection is performed exclusively using nucleic acid amplification tests (NAATs), owing to the poor and slow growth of the bacterium in culture. Because no internationally validated and approved commercial NAAT for M. genitalium detection is presently available, it is necessary that laboratories performing M. genitalium diagnostics not only carefully evaluate and validate their in-house PCRs before using them routinely, but also use comprehensive internal controls and take part in external quality assessment programmes. The guidelines were elaborated as a consensus document of the Eastern European Sexual and Reproductive Health (EE SRH) Network, and comprise one element of a series of guidelines aimed at optimizing, standardizing, and providing guidance on quality laboratory testing for reproductive tract infections.

Quantitation of all Four Gardnerella vaginalis Clades Detects Abnormal Vaginal Microbiota Characteristic of Bacterial Vaginosis More Accurately than Putative G. vaginalis Sialidase A Gene Count.

BACKGROUND: Bacterial vaginosis (BV) is a vaginal disorder characterized by a depletion of the normal lactobacillus-dominant microbiota and overgrowth of mainly anaerobic bacteria. OBJECTIVES: The study aimed to evaluate the distribution and abundance of the Gardnerella vaginalis clades and sialidase A gene in vaginal samples from Russian women, and investigate if the G. vaginalis sialidase A gene count detects an abnormal vaginal microbiota characteristic of BV more accurately than G. vaginalis load. METHODS: Vaginal samples from 299 non-pregnant patients of gynecological clinics were examined using Nugent scores and G. vaginalis clade and sialidase A gene quantitative real-time polymerase chain reactions (PCRs). Discriminatory power for BV microbiota was evaluated with receiver operating characteristic (ROC) analysis. RESULTS: The vaginal microbiota was characterized by Nugent scores as normal, intermediate, and BV microbiota in 162, 58, and 79 women, respectively. G. vaginalis clades 1, 2, 3, 4, and the sialidase A gene were detected in 56% (51-62%), 40% (34-45%), 20% (16-25%), 94% (91-96%), and 70% (64-75%) of vaginal samples, respectively. The frequency and abundance of clades 1, 2, 4, and the sialidase A gene as well as clade multiplicity were significantly associated with abnormal microbiota. The sialidase A gene was present in all multi-clade samples, in all single-clade samples comprising clades 1, 2, and 3, and in four of 84 (5% [2-12%]) samples comprising clade 4 only. Total G. vaginalis load showed significantly higher discriminatory power for abnormal microbiota than sialidase A gene count (areas under ROC curves 0.933 vs. 0.881; p = 0.0306). CONCLUSIONS: Quantifying all four G. vaginalis clades discriminates between BV microbiota and normal microbiota more accurately than measuring G. vaginalis sialidase A gene. Clade 4 is strongly associated with BV microbiota, despite most clade 4 strains lacking the sialidase A gene.

Risk of newly detected infections and cervical abnormalities in adult women seropositive or seronegative for naturally acquired HPV-16/18 antibodies.

BACKGROUND: Infections with human papillomavirus (HPV) types 16 and 18 account for ~70% of invasive cervical cancers but the degree of protection from naturally acquired anti-HPV antibodies is uncertain. We examined the risk of HPV infections as defined by HPV DNA detection and cervical abnormalities among women >25 years in the Human Papilloma VIrus Vaccine Immunogenicity ANd Efficacy trial's (VIVIANE, NCT00294047) control arm. METHODS: Serum anti-HPV-16/18 antibodies were determined at baseline and every 12 months in baseline DNA-negative women (N = 2687 for HPV-16 and 2705 for HPV-18) by enzyme-linked immunosorbent assay (ELISA) from blood samples. HPV infections were identified by polymerase chain reaction (PCR) every 6-months, and cervical abnormalities were confirmed by cytology every 12 months. Data were collected over a 7-year period. The association between the risk of type-specific infection and cervical abnormalities and serostatus was assessed using Cox proportional hazard models. RESULTS: Risk of newly detected HPV-16-associated 6-month persistent infections (PI) (hazard ratio [HR] = 0.56 [95%CI:0.32; 0.99]) and atypical squamous cells of undetermined significance (ASC-US+) (HR = 0.28 [0.12; 0.67]) were significantly lower in baseline seropositive vs baseline seronegative women. HPV-16-associated incident infections (HR = 0.81 [0.56; 1.16]) and 12-month PI (HR = 0.53 [0.24; 1.16]) showed the same trend. A similar trend of lower risk was observed in HPV-18-seropositive vs -seronegative women (HR = 0.95 [0.59; 1.51] for IIs, HR = 0.43 [0.16; 1.13] for 6-month PIs, HR = 0.31 [0.07; 1.36] for 12-month PIs, and HR = 0.61 [0.23; 1.61] for ASC-US+). CONCLUSIONS: Naturally acquired anti-HPV-16 antibodies were associated with a decreased risk of subsequent infection and cervical abnormalities in women >25 years. This possible protection was lower than that previously reported in 15- to 25-year-old women.

Comparison of microscopy, culture and in-house PCR and NASBA assays for diagnosis of Neisseria gonorrhoeae in Russia.

This study aimed to assess the laboratory diagnosis of Neisseria gonorrhoeae in St. Petersburg, Russia. In total, 334 consecutive symptomatic patients were enrolled. Cervical and urethral specimens from women (n=286) and urethral specimens from men (n=48) were analyzed by microscopy, culture and two in-house NAATs, i.e. polymerase chain reaction (PCR) and nucleic acid sequence-based amplification (NASBA), developed in Russia. All N. gonorrhoeae-positive samples were confirmed using porA pseudogene and 16S rRNA gene sequencing. All methods displayed 100% specificity, i.e. positive predictive values of 100%. Compared to the PCR (most sensitive method in the present study), in women the sensitivity of both microscopy and culture was 31.8%, and that of NASBA was 90.9%. In men, microscopy, culture and NASBA displayed a sensitivity of 75%, 50% and 100%, respectively. The negative predictive values of microscopy, culture, and NASBA were 97.3%, 97.3%, and 99.6% in women, and 97.8%, 95.7%, and 100% in men, respectively. According to the PCR, the prevalences of N. gonorrhoeae were 4.5% (women) and 8.3% (men). In conclusion, both the investigated Russian NAATs displayed a high sensitivity and specificity. However, in general the diagnosis of gonorrhoea in Russia is suboptimal and crucially requires validation, improvements and quality assurance.

Prevalence of macrolide and fluoroquinolone resistance-mediating mutations in Mycoplasma genitalium in five cities in Russia and Estonia.

BACKGROUND AND OBJECTIVE: Resistance in the sexually transmitted bacterium Mycoplasma genitalium to all recommended therapeutic antimicrobials have rapidly emerged. However, to date, internationally reported resistance surveillance data for M. genitalium strains circulating in Eastern Europe are entirely lacking. The aim of this study was to estimate the prevalence of macrolide and fluoroquinolone resistance-associated mutations in M. genitalium in four cities in Russia and one in Estonia, 2013-2016. MATERIALS AND METHODS: Consecutive urogenital samples found positive for M. genitalium during diagnostic testing were retrospectively analyzed for resistance-associated mutations in the 23S rRNA and parC genes using pyrosequencing and conventional Sanger sequencing, respectively. RESULTS: In total, 867 M. genitalium positive samples from 2013-2016 were analyzed. Macrolide resistance-associated mutations were detected in 4.6% of the samples from Russia (0.7-6.8% in different cities) and in 10% of the samples from Estonia. The mutations A2059G and A2058G were highly predominating in both Russia and Estonia, accounting together for 90.9% of the cases positive for nucleotide substitutions in the 23S rRNA gene. The rates of possible fluoroquinolone resistance-associated mutations were 6.2% in Russia (2.5-7.6% in different cities) and 5% in Estonia. The mutations S83I and S83N were the most frequent ones in Russia (24.4% each), whereas D87N highly predominated in Estonia (83.3% of all fluoroquinolone resistance-associated mutations). Approximately 1% of the samples in both countries harbored both macrolide and possible fluoroquinolone resistance-associated mutations, with A2058G and S83I being the most frequent combination (37.5%). CONCLUSIONS: The prevalence of macrolide and fluoroquinolone resistance-associated mutations in M. genitalium was 4.6% and 6.2%, respectively, in Russia, and 10% and 5%, respectively, in Estonia. Despite the relatively low rates of macrolide and fluoroquinolone resistance in these countries, antimicrobial resistance surveillance and testing for resistance-associated mutations in M. genitalium positive cases would be valuable.

Diagnosis of Chlamydia trachomatis in Russia--in-house PCR assays may be effective but overall optimization and quality assurance are urgently needed.

In the present study, the performance of the cell culture method, two non-Russian direct immunofluorescence (DIF) assays, and three different in-house polymerase chain reaction (PCR) tests used in St. Petersburg, Russia, for detection of Chlamydia trachomatis in urogenital specimens was evaluated. A total of 650 patients were examined and it was most disquieting that previous C. trachomatis positivity with Russian DIF assays could - 7 days later - be confirmed only in 26% of the women and 30% of the men. Overall, the highest diagnostic sensitivity was obtained using PCR analysis. However, the sensitivity varied significantly: from 79% to 100% between the different PCR assays, sex of the patients, and type of samples. The highest sensitivity was obtained for female vaginal and male urine samples (100%). The specificity of the PCR assays varied from 97% to 100%. The sensitivity of cell culture and both the examined DIF assays was low, i.e. it varied from 46% to 56% and 55% to 75%, respectively. Meanwhile, cell culture was 100% specific and the DIFs showed a specificity varying from 99% to 100%. In conclusion, in a Russian perspective, adequate in-house PCR methods may be used quite effectively for detection of C. trachomatis in invasive as well as non-invasive clinical material. Simultaneous analysis of two different specimens from women resulted in a significantly increased detection rate of C. trachomatis. Nevertheless, in Russia the need for optimization and quality assurance of diagnostic methods for C. trachomatis, especially Russian DIF assays, has to be emphasized.

Efficacy, safety, and immunogenicity of the human papillomavirus 16/18 AS04-adjuvanted vaccine in women older than 25 years: 7-year follow-up of the phase 3, double-blind, randomised controlled VIVIANE study.

BACKGROUND: Although the risk of human papillomavirus (HPV) infection is greatest in young women, women older than 25 years remain at risk. We present data from the VIVIANE study of the HPV 16/18 AS04-adjuvanted vaccine in adult women after 7 years of follow-up. METHODS: In this phase 3, double-blind, randomised controlled trial, healthy women older than 25 years were enrolled (age stratified: 26-35 years, 36-45 years, and ≥46 years). Up to 15% in each age stratum had a history of HPV infection or disease. Women were randomly assigned (1:1) to receive HPV 16/18 vaccine or aluminium hydroxide control, with an internet-based system. The primary endpoint was vaccine efficacy against 6-month persistent infection or cervical intraepithelial neoplasia grade 1 or greater (CIN1+) associated with HPV 16/18. We did analyses in the according-to-protocol cohort for efficacy and total vaccinated cohort. Data for the combined primary endpoint in the according-to-protocol cohort for efficacy were considered significant when the lower limit of the 96·2% CI around the point estimate was greater than 30%. For all other endpoints and cohorts, data were considered significant when the lower limit of the 96·2% CI was greater than 0%. This study is registered with ClinicalTrials.gov, number NCT00294047. FINDINGS: The first participant was enrolled on Feb 16, 2006, and the last study visit took place on Jan 29, 2014. 4407 women were in the according-to-protocol cohort for efficacy (n=2209 vaccine, n=2198 control) and 5747 women in the total vaccinated cohort (n=2877 vaccine, n=2870 control). At month 84, in women seronegative for the corresponding HPV type in the according-to-protocol cohort for efficacy, vaccine efficacy against 6-month persistent infection or CIN1+ associated with HPV 16/18 was significant in all age groups combined (90·5%, 96·2% CI 78·6-96·5). Vaccine efficacy against HPV 16/18-related cytological abnormalities (atypical squamous cells of undetermined significance and low-grade squamous intraepithelial lesion) and CIN1+ was also significant. We also noted significant cross-protective efficacy against 6-month persistent infection with HPV 31 (65·8%, 96·2% CI 24·9-85·8) and HPV 45 (70·7%, 96·2% CI 34·2-88·4). In the total vaccinated cohort, vaccine efficacy against CIN1+ irrespective of HPV was significant (22·9%, 96·2% CI 4·8-37·7). Serious adverse events related to vaccination occurred in five (0·2%) of 2877 women in the vaccine group and eight (0·3%) of 2870 women in the control group. INTERPRETATION: In women older than 25 years, the HPV 16/18 vaccine continues to protect against infections, cytological abnormalities, and lesions associated with HPV 16/18 and CIN1+ irrespective of HPV type, and infection with non-vaccine types HPV 31 and HPV 45 over 7 years of follow-up. FUNDING: GlaxoSmithKline Biologicals SA.