| 1. |
- Shipitsyna, Elena, et al.
(författare)
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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
- 2023
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Ingår i: Sexually Transmitted Infections. - : BMJ Publishing Group Ltd. - 1368-4973 .- 1472-3263. ; 99:3, s. 191-194
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Tidskriftsartikel (refereegranskat)abstract
- 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.
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| 2. |
- Hadfield, James, et al.
(författare)
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Comprehensive global genome dynamics of Chlamydia trachomatis show ancient diversification followed by contemporary mixing and recent lineage expansion
- 2017
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory Press. - 1088-9051 .- 1549-5469. ; 27:7, s. 1220-1229
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Tidskriftsartikel (refereegranskat)abstract
- 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.
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| 3. |
- Rumyantseva, Tatiana, et al.
(författare)
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Evaluation and subsequent optimizations of the quantitative AmpliSens Florocenosis/Bacterial vaginosis-FRT multiplex real-time PCR assay for diagnosis of bacterial vaginosis
- 2016
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Ingår i: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS). - Hoboken, USA : Wiley-Blackwell Publishing Inc.. - 0903-4641 .- 1600-0463. ; 124:12, s. 1099-1108
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Tidskriftsartikel (refereegranskat)abstract
- Traditional microscopy-based methods for diagnosis of bacterial vaginosis (BV) are underutilized in many settings, and molecular techniques may provide opportunities for rapid, objective, and accurate BV diagnosis. This study evaluated the quantitative AmpliSens Florocenosis/Bacterial vaginosis-FRT multiplex real-time PCR (Florocenosis-BV) assay. Vaginal samples from a previous study including unselected female subjects (n = 163) and using Amsel criteria and 454 pyrosequencing for BV diagnosis were examined with the Florocenosis-BV test and additionally tested for the presence and quantity of Gardnerella vaginalis clades 3 and 4. The Florocenosis-BV assay demonstrated 100% and 98% sensitivity compared with the Amsel criteria and 454 pyrosequencing, respectively, with 91% specificity. The modified Florocenosis-BV assay (detecting also G. vaginalis clades 3 and 4) resulted in 100% sensitivity vs the Amsel criteria and 454 pyrosequencing with specificity of 86% and 88%, respectively. Further optimizations of thresholds for the quantitative parameters used in the kit resulted in 99-100% accuracy vs Amsel criteria and 454 pyrosequencing for selected parameters. The Florocenosis-BV assay is an objective, accurate, sensitive, and specific method for BV diagnosis; however, the performance of the test can be further improved with some minor optimizations.
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| 4. |
- Seth-Smith, Helena M. B., et al.
(författare)
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Whole-genome sequences of Chlamydia trachomatis directly from clinical samples without culture
- 2013
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory Press (CSHL). - 1088-9051 .- 1549-5469. ; 23:5, s. 855-866
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Tidskriftsartikel (refereegranskat)abstract
- The use of whole-genome sequencing as a tool for the study of infectious bacteria is of growing clinical interest. Chlamydia trachomatis is responsible for sexually transmitted infections and the blinding disease trachoma, which affect hundreds of millions of people worldwide. Recombination is widespread within the genome of C. trachomatis, thus whole-genome sequencing is necessary to understand the evolution, diversity, and epidemiology of this pathogen. Culture of C trachomatis has, until now, been a prerequisite to obtain DNA for whole-genome sequencing; however, as C trachomatis is an obligate intracellular pathogen, this procedure is technically demanding and time consuming. Discarded clinical samples represent a large resource for sequencing the genomes of pathogens, yet clinical swabs frequently contain very low levels of C trachomatis DNA and large amounts of contaminating microbial and human DNA. To determine whether it is possible to obtain whole-genome sequences from bacteria without the need for culture, we have devised an approach that combines immunomagnetic separation (IMS) for targeted bacterial enrichment with multiple displacement amplification (MDA) for whole-genome amplification. Using IMS-MDA ill conjunction with high-throughput multiplexed Illumina sequencing, we have produced the first whole bacterial genome sequences direct from clinical samples. We also show that this method can be used to generate genome data from nonviable archived samples. This method will prove a useful tool in answering questions relating to the biology of many difficult-to-culture or fastidious bacteria of clinical concern.
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| 5. |
- Shalepo, Kira, et al.
(författare)
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Diagnosis of Chlamydia trachomatis in Russia--in-house PCR assays may be effective but overall optimization and quality assurance are urgently needed
- 2006
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Ingår i: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS). - : Wiley. - 0903-4641 .- 1600-0463. ; 114:7-8, s. 500-507
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Tidskriftsartikel (refereegranskat)abstract
- 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.
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| 6. |
- Shipitsyna, Elena, et al.
(författare)
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A profile of the FDA-approved and CE/IVD-marked Aptima Mycoplasma genitalium assay (Hologic) and key priorities in the management of M. genitalium infections
- 2020
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Ingår i: Expert Review of Molecular Diagnostics. - : Expert Reviews Ltd.. - 1473-7159 .- 1744-8352. ; 20:11, s. 1063-1074
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Tidskriftsartikel (refereegranskat)abstract
- INTRODUCTION: (MG) causes frequently asymptomatic STIs. MG prevalence figures are lacking and management is complicated by the lack of etiological diagnostics and high antimicrobial resistance in many countries. Appropriately validated, quality-assured, and FDA-approved MG diagnostic assays have been lacking.AREAS COVERED: The clinical and analytical performance characteristics of the Aptima® MG assay, the first FDA-approved MG nucleic acid amplification test (NAAT), are summarized. Key priorities in the management and control of MG infections are also discussed.EXPERT OPINION: Highly sensitive, specific, and quality-assured MG NAATs, e.g. the Aptima MG assay on the automated and flexible Panther® platform, are imperative to improve the management and control of MG infections internationally. This testing, combined with macrolide resistance testing (not yet available on the Panther platform), offers a rapid, high-throughput, and appropriate diagnosis of MG. Macrolide resistance-guided sequential treatment needs to be implemented for MG infections. Dual antimicrobial therapy, novel antimicrobials and, ideally, a vaccine may become essential.
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| 7. |
- Shipitsyna, Elena, et al.
(författare)
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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
- 2020
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Ingår i: European Journal of Clinical Microbiology and Infectious Diseases. - : Springer. - 0934-9723 .- 1435-4373. ; 39:7, s. 1221-1230
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Tidskriftsartikel (refereegranskat)abstract
- 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.
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| 8. |
- Shipitsyna, Elena, et al.
(författare)
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Comparison of microscopy, culture and in-house PCR and NASBA assays for diagnosis of Neisseria gonorrhoeae in Russia
- 2008
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Ingår i: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS). - Oxford : Blackwell. - 0903-4641 .- 1600-0463. ; 116:2, s. 133-138
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Tidskriftsartikel (refereegranskat)abstract
- 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 (nΩ286) and urethral specimens from men (nΩ48) 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.
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| 9. |
- Shipitsyna, Elena, et al.
(författare)
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Composition of the Vaginal Microbiota in Women of Reproductive Age - Sensitive and Specific Molecular Diagnosis of Bacterial Vaginosis Is Possible?
- 2013
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 8:4, s. e60670-
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Tidskriftsartikel (refereegranskat)abstract
- Background and Objective: Bacterial vaginosis (BV) is the most common vaginal disorder, characterized by depletion of the normal lactobacillus-dominant microbiota and overgrowth of commensal anaerobic bacteria. This study aimed to investigate the composition of the vaginal microbiota in women of reproductive age (healthy women and women with BV), with the view of developing molecular criteria for BV diagnosis. Materials and Methods: Vaginal samples from 163 women (79 control, 73 BV and 11 intermediate (Lactobacillary grade II flora) cases) were analyzed using 454 pyrosequencing of the hypervariable regions V3-V4 of the 16S rRNA gene and 16 quantitative bacterial species/genus-specific real-time PCR assays. Sensitivities and specificities of potential BV markers were computed using the Amsel criteria as reference standard for BV. The use of quantitative thresholds for prediction of BV, determined for both relative abundance measured with 454 pyrosequencing and bacterial load measured with qPCR, was evaluated. Results: Relative to the healthy women, the BV patients had in their vaginal microbiota significantly higher prevalence, loads and relative abundances of the majority of BV associated bacteria. However, only Gardnerella vaginalis, Atopobium vaginae, Eggerthella, Prevotella, BVAB2 and Megasphaera type 1 detected at or above optimal thresholds were highly predictable for BV, with the best diagnostic accuracy shown for A. vaginae. The depletion of Lactobacillus species combined with the presence of either G. vaginalis or A. vaginae at diagnostic levels was a highly accurate BV predictor. Conclusions: Quantitative determination of the presence of G. vaginalis, A. vaginae, Eggerthella, Prevotella, BVAB2 and Megasphaera type 1 as well as the depletion of Lactobacillus was highly accurate for BV diagnosis. Measurements of abundance of normal and BV microbiota relative to total bacteria in vaginal fluid may provide more accurate BV diagnosis, and be used for test-of-cure, rather than qualitative detection or absolute counts of BV related microorganisms.
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| 10. |
- Shipitsyna, Elena, et al.
(författare)
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First evaluation of six nucleic acid amplification tests widely used in the diagnosis of Chlamydia trachomatis in Russia
- 2009
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Ingår i: Journal of the European Academy of Dermatology and Venereology. - : Wiley. - 0926-9959 .- 1468-3083. ; 23:3, s. 268-276
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Tidskriftsartikel (refereegranskat)abstract
- Abstract Background In Russia, nationally developed nucleic acid amplification tests (NAATs), which have never been validated to international commercially available NAATs, are mainly used in the diagnosis of Chlamydia trachomatis infection. Objective To evaluate the performance characteristics of six NAATs widely used to diagnose C. trachomatis infection in Russia. Materials and methods In total, 446 consecutive symptomatic patients (319 females and 127 males) were included. Five polymerase chain reaction (PCR) assays and one real-time nucleic acid sequence-based amplification (NASBA) assay were evaluated on cervical and vaginal samples from females and on urethral and first voided urine samples from males. As reference methods, the Cobas Amplicor PCR, as the main 'gold standard' method, and LightMix 480HT PCR were used. Results The overall prevalence of C. trachomatis infection was 12.6%. The Russian NAATs and the reference methods displayed a high level of concordance (97.9% to 99.2%). In comparison with the reference methods, the sensitivities, specificities, positive predictive values and negative predictive values of the Russian tests in different specimens ranged from 86.1% to 100%, 99.1% to 100%, 92.3% to 100% and 98.2% to 100%, respectively. Conclusions According to the reference methods, C. trachomatis NAATs developed and used in Russia have relatively good performance characteristics for both invasive and non-invasive samples. However, larger studies that include symptomatic and asymptomatic patients as well as genital and extra-genital samples, and in comparison with other internationally well-recognized, validated, and ideally Food and Drug Administration-approved C. trachomatis NAATs performed strictly according to the manufacturer's instructions, need to be conducted. Conflicts of interest None declared.
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