RESUMEN
Mycoplasma genitalium is a sexually transmitted bacterium associated with nongonococcal urethritis (NGU) in men and cervicitis, endometritis, and pelvic inflammatory disease in women. Effective treatment is challenging due to the inherent, and increasingly acquired, antibiotic resistance in this pathogen. In our treatment trial conducted from 2007 to 2011 in Seattle, WA, we demonstrated poor efficacy of azithromycin (AZM) and doxycycline (DOX) against M. genitalium among men with NGU. In the present study, we cultured M. genitalium from 74 of 80 (92.5%) PCR-positive men at enrollment (V-1) and defined the MICs of AZM (N = 56 isolates) of DOX (N = 62 isolates). Susceptibility to AZM was bimodal; MICs were >8 µg/ml (44.6%) and <0.004 µg/ml (55.4%) for these isolates. The association of MIC with treatment efficacy was determined for men initially treated with either AZM (N = 30) or DOX (N = 24). Men treated with AZM were more likely to experience microbiologic treatment failure (P < 0.001) if infected with isolates that had AZM MICs of >8 µg/ml (18/18 men) than those with isolates that had AZM MICs of <0.004 µg/ml (1/12 men). Clinical treatment failure also was more likely to occur (P = 0.002) with AZM MICs of >8 µg/ml (12/18 men) than with AZM MICs of <0.004 µg/ml (1/12 men). In contrast, DOX MICs ranged from <0.125 to 2 µg/ml and were not correlated with microbiologic (P = 0.71) or clinical treatment (P = 0.41) failure, demonstrating no relationship between DOX MICs and treatment efficacy. Given the rapid spread of AZM resistance and the emergence of quinolone resistance, the current second-line therapy, monitoring MICs and evaluating other potential treatments for M. genitalium will be critical.
Asunto(s)
Infecciones por Mycoplasma , Mycoplasma genitalium , Uretritis , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Azitromicina , Doxiciclina , Farmacorresistencia Bacteriana , Femenino , Humanos , Masculino , Infecciones por Mycoplasma/tratamiento farmacológico , Resultado del Tratamiento , Uretritis/tratamiento farmacológicoRESUMEN
Mycoplasma genitalium is a sexually transmitted bacterial pathogen that infects men and women. Antigenic variation of MgpB and MgpC, the immunodominant adherence proteins of M. genitalium, is thought to contribute to immune evasion and chronic infection. We investigated the evolution of mgpB and mgpC sequences in men with non-gonococcal urethritis persistently infected with M. genitalium, including two men with anti-M. genitalium antibodies at enrollment and two that developed antibodies during follow-up. Each of the four patients was persistently infected with a different strain type and each patient produced antibodies targeting MgpB and MgpC. Amino acid sequence evolution in the variable regions of MgpB and MgpC occurred in all four patients with changes observed in single and multiple variable regions over time. Using the available crystal structure of MgpC of the G37 type strain we found that predicted conformational B cell epitopes localize predominantly to the variable region of MgpC, amino acids that changed during patient infection lie in these epitopes, and variant amino acids are in close proximity to the conserved sialic acid binding pocket. These findings support the hypothesis that sequence variation functions to avoid specific antibodies thereby contributing to persistence in the genital tract.
Asunto(s)
Adhesinas Bacterianas/genética , Infecciones por Mycoplasma/genética , Mycoplasma genitalium/genética , Uretritis/genética , Secuencia de Aminoácidos/genética , Animales , Anticuerpos Antiidiotipos/sangre , Anticuerpos Antiidiotipos/inmunología , Linfocitos B/inmunología , Linfocitos B/microbiología , Chlorocebus aethiops , Doxiciclina/farmacología , Evolución Molecular , Humanos , Infecciones por Mycoplasma/sangre , Infecciones por Mycoplasma/inmunología , Infecciones por Mycoplasma/microbiología , Mycoplasma genitalium/inmunología , Mycoplasma genitalium/patogenicidad , Reacción en Cadena de la Polimerasa , Uretritis/sangre , Uretritis/inmunología , Uretritis/microbiología , Células VeroRESUMEN
In a vaginal 16S ribosomal RNA gene quantitative PCR study of 17 pelvic inflammatory disease (PID) cases and 17 controls who tested positive for Chlamydia trachomatis, women who additionally tested positive for Atopobium vaginae, Sneathia spp., Megasphaera spp., Eggerthella-like bacterium or Prevotella amnii were more likely to develop PID.
Asunto(s)
Antibacterianos/uso terapéutico , Bacterias/efectos de los fármacos , Enfermedad Inflamatoria Pélvica/epidemiología , Reacción en Cadena de la Polimerasa/métodos , Vagina/microbiología , Vaginosis Bacteriana/epidemiología , Actinobacteria , Adulto , Bacterias/aislamiento & purificación , Estudios de Casos y Controles , ADN Bacteriano/genética , ADN Ribosómico/genética , Femenino , Humanos , Prevotella , ARN Ribosómico 16S/genética , Vaginosis Bacteriana/complicacionesRESUMEN
OBJECTIVES: We sought to determine whether the relationship between a history of vaginal douching and pelvic inflammatory disease (PID) is mediated by endometrial infection with one or more novel bacterial vaginosis (BV)-associated organisms among Atopobium vaginae, the BV-associated bacterium 1 (BVAB1), neathia (Leptotrichia) amnionii and Sneathia sanguinegens. METHODS: We first conducted log-binomial regression analyses to identify risk factors for endometrial infection in 535 adolescent and adult women with clinically suspected PID in the PID Evaluation and Clinical Health (PEACH) study. We then examined whether endometrial infection by the BV-associated organisms mediated the association between a history of vaginal douching and histologically confirmed PID using inverse probability weighted marginal structural models. RESULTS: Vaginal douching was significantly associated with endometrial infection with one or more of the targeted BV-associated organisms (relative risk (RR) 1.21, 95% CI: 1.08 to 1.35). The total effect estimate suggested that vaginal douching increased the risk of endometritis by 24% (RR 1.24, 95% CI: 1.03 to 1.49). The controlled direct effect of this association was attenuated with endometrial infection by one or more BV-associated organisms (adjusted RR (aRR) 1.00, 95% CI: 0.57 to 1.74) and endometrial infection by all four BV-associated organisms (aRR 0.95, 95% CI: 0.53 to 1.70) as intermediate variables. CONCLUSIONS: Endometrial infection with one or more of the novel BV-associated organisms partially mediated the relationship between vaginal douching and histologically confirmed endometritis in the PEACH study. Frequent vaginal douching may confer risk for endometritis through increasing the risk of endometrial infection by novel-BV-associated organisms. Other potential pathways should be explored.
Asunto(s)
Endometritis/epidemiología , Enfermedad Inflamatoria Pélvica/epidemiología , Ducha Vaginal/estadística & datos numéricos , Vaginosis Bacteriana/microbiología , Actinobacteria , Adolescente , Adulto , Endometritis/microbiología , Femenino , Fusobacterias , Infecciones por Bacterias Gramnegativas/epidemiología , Infecciones por Bacterias Gramnegativas/microbiología , Infecciones por Bacterias Grampositivas/epidemiología , Infecciones por Bacterias Grampositivas/microbiología , Humanos , Enfermedad Inflamatoria Pélvica/microbiología , Estados Unidos/epidemiología , Adulto JovenRESUMEN
OBJECTIVE: Many sexually transmitted infections increase risk of mother-to-child transmission (MTCT) of HIV, but the effect of Mycoplasma genitalium is not known. We hypothesized that M. genitalium infection would be common among HIV-infected pregnant women and could be associated with in-utero and intrapartum MTCT. DESIGN: Observational case-cohort study. METHODS: The current study used specimens from a Kenyan perinatal MTCT cohort (1999-2005) involving HIV-infected women and their infants, who received short-course zidovudine for prevention of MTCT. Vaginal swabs collected at 32 weeks gestation were tested for M. genitalium using a transcription-mediated amplification assay. Infant perinatal HIV infection was determined at birth and 4 weeks of age by DNA PCR. Using a case-cohort design, a random sample was generated with 3â:â1 controlâ:âcase ratio; prevalence and correlates of M. genitalium were assessed with chi-squared and t tests; predictors of infant outcomes were analyzed using logistic regression. RESULTS: Among 220 HIV-infected pregnant women evaluated, 47 women (21.4%) had M. genitalium. Antenatal M. genitalium infection was associated with higher HIV RNA in plasma (5.0 vs. 4.6âlog10 copies/ml in M. genitalium-positive vs. M. genitalium-negative women, Pâ=â0.02) at 32 weeks. Women with M. genitalium were less likely to report prior sexually transmitted infections and genital ulcers (both Pâ=â0.05). There was no association found between exposure to M. genitalium and perinatal MTCT (odds ratioâ=â0.72, 95% confidence interval 0.35, 1.51, Pâ=â0.39). CONCLUSION: Vaginal M. genitalium infection was frequently detected among Kenyan HIV-infected pregnant women and was associated with higher plasma HIV levels, but was not associated with perinatal transmission of HIV.
Asunto(s)
Infecciones por VIH/epidemiología , Infecciones por Mycoplasma/epidemiología , Mycoplasma genitalium/aislamiento & purificación , Complicaciones Infecciosas del Embarazo/epidemiología , Adulto , Estudios de Cohortes , Femenino , Humanos , Recién Nacido , Transmisión Vertical de Enfermedad Infecciosa/prevención & control , Kenia/epidemiología , Modelos Logísticos , Embarazo , Adulto Joven , Zidovudina/uso terapéuticoRESUMEN
From February 2015 to October 2017, among 20 men who have sex with men with Mycoplasma genitalium-associated nongonococcal urethritis, 15% had macrolide resistance and S83I ParC mutations. Azithromycin followed by moxifloxacin cleared Mycoplasma genitalium in 2 of 2 with and 11 of 13 without S83I mutations. Dual failures were cleared after doxycycline. S83I mutations were not associated with moxifloxacin failure.
Asunto(s)
Antibacterianos/uso terapéutico , Topoisomerasa de ADN IV/genética , Homosexualidad Masculina , Infecciones por Mycoplasma/tratamiento farmacológico , Mycoplasma genitalium/efectos de los fármacos , Uretritis/tratamiento farmacológico , Adulto , Antibacterianos/farmacología , Farmacorresistencia Bacteriana/efectos de los fármacos , Farmacorresistencia Bacteriana/genética , Humanos , Masculino , Mycoplasma genitalium/genética , Minorías Sexuales y de Género , Resultado del TratamientoRESUMEN
Mycoplasma genitalium is a sexually transmitted bacterium linked to adverse sexual and reproductive health outcomes in women and men. M. genitalium is difficult to culture, and in the absence of validated amplified molecular methods for diagnosis of infection, there is no reference standard available for use as a comparator for the validation of new M. genitalium diagnostic tests. We evaluated the analytical and clinical performance of three transcription-mediated amplification (TMA) tests for M. genitalium, each targeting unique rRNA sequences, for use as a composite comparator for clinical validation of the Aptima Mycoplasma genitalium (AMG) assay, an in vitro diagnostic (IVD) TMA test that targets 16 s rRNA of M. genitalium Analytical sensitivity, specificity, and strain inclusivity of all four TMA tests were determined using nine laboratory strains of M. genitalium and 56 nontarget bacteria, protozoa, and viruses. Analytical sensitivity of the tests for M. genitalium ranged from 0.017 to 0.040 genome equivalents/ml. None of the nontarget organisms evaluated cross-reacted with any test. A composite comparator reference standard consisting of the 3 alternate (Alt) TMA tests was used to evaluate the clinical performance of the AMG assay by testing residual vaginal swab, female urine, and male urine specimens obtained from 1,400 adult subjects from three U.S. clinical sites. Using this reference standard to establish infected specimen status, the sensitivity, specificity, and overall agreement of the AMG IVD assay were 100%, 99.9%, and 99.9%, respectively. These results demonstrate the utility of molecular composite reference standard methodology for the clinical validation of future IVD tests for this organism.
Asunto(s)
Infecciones por Mycoplasma/diagnóstico , Mycoplasma genitalium/genética , Técnicas de Amplificación de Ácido Nucleico/métodos , Técnicas de Amplificación de Ácido Nucleico/normas , Transcripción Genética , Adulto , Femenino , Humanos , Masculino , Infecciones por Mycoplasma/microbiología , Infecciones por Mycoplasma/orina , Pene/microbiología , ARN Ribosómico 16S/genética , Sensibilidad y Especificidad , Manejo de Especímenes , Vagina/microbiologíaRESUMEN
BACKGROUND: Although Mycoplasma genitalium (MG) is an acknowledged cause of nongonococcal urethritis (NGU), access to diagnostic testing is limited. Syndromic management is common, yet little is known about natural history. METHODS: Between August 2014 and April 2016, 13 heterosexual men aged ≥16 years with MG were identified within a cohort study of men with and without NGU attending an urban sexually transmitted diseases clinic. Men had 6-7 monthly visits. NGU was defined as ≥5 polymorphonuclear leukocytes per high-power field on urethral Gram stain plus either visible urethral discharge or urethral symptoms. Men with NGU received 1 g of azithromycin. Men with persistent NGU received moxifloxacin 400 mg for 14 days. First-void urine was retrospectively tested for MG using transcription-mediated amplification. Resistance-associated mutations were detected by polymerase chain reaction (PCR) and sequencing. Organism load was determined by quantitative PCR. RESULTS: Sixty-two percent of MG-positive men had macrolide resistance-mediating mutations (MRMM) at enrollment; 31% had parC mutations (all outside the quinolone resistance-determining region). MG persisted after azithromycin in 7 men, 6 of whom had MRMM. The median duration of persistence in the absence of curative therapy was 143 days (range, 21-228). Five men experienced symptom resolution after azithromycin, but MG persisted for another 89-186 days before moxifloxacin. Organism load was somewhat lower in MRMM than wild-type infections (P = .16). CONCLUSIONS: The high prevalence of macrolide resistance and long duration of infection after symptom resolution highlights the need for diagnostic MG testing of men with NGU to direct therapy.
Asunto(s)
Antibacterianos/uso terapéutico , Infecciones Asintomáticas , Farmacorresistencia Bacteriana , Infecciones por Mycoplasma/tratamiento farmacológico , Uretritis/tratamiento farmacológico , Uretritis/microbiología , Adulto , Gonorrea , Heterosexualidad , Humanos , Macrólidos/uso terapéutico , Masculino , Mycoplasma genitalium , Estudios Retrospectivos , Adulto JovenRESUMEN
Antigenic variation of the immunodominant MgpB and MgpC proteins has been suggested to be a mechanism of immune evasion of the human pathogen Mycoplasma genitalium, a cause of several reproductive tract disease syndromes. Phase variation resulting in the loss of adherence has also been documented, but the molecular mechanisms underlying this process and its role in pathogenesis are still poorly understood. In this study, we isolated and characterized 40 spontaneous, nonadherent phase variants from in vitro-passaged M. genitalium cultures. In all cases, nonadherence was associated with the loss of MgpBC protein expression, attributable to sequence changes in the mgpBC expression site. Phase variants were grouped into seven classes on the basis of the nature of the mutation. Consistent with the established role of RecA in phase variation, 31 (79.5%) variants arose via recombination with MgPa repeat regions that contain mgpBC variable sequences. The remaining mutants arose via nonsense or frameshift mutations. As expected, revertants were obtained for phase variants that were predicted to be reversible but not for those that arose via an irreversible mechanism. Furthermore, phase variants were enriched in M. genitalium cultures exposed to antibodies reacting to the extracellular, conserved C terminus of MgpB but not in cultures exposed to antibodies reacting to an intracellular domain of MgpB or the cytoplasmic HU protein. Genetic characterization of the antibody-selected phase variants confirmed that they arose via reversible and irreversible recombination and point mutations within mgpBC These phase variants resisted antibody-mediated growth inhibition, suggesting that phase variation promotes immune evasion.
Asunto(s)
Anticuerpos Antibacterianos/inmunología , Infecciones por Mycoplasma/inmunología , Infecciones por Mycoplasma/microbiología , Mycoplasma genitalium/inmunología , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Secuencia de Bases , Eliminación de Gen , Expresión Génica , Marcación de Gen , Variación Genética , Humanos , Mutación , Mycoplasma genitalium/genética , Recombinación GenéticaRESUMEN
PROBLEM: Toll-like (TLR) receptor genetic variants have been implicated in bacterial vaginosis (BV). We determined whether TLR variants are associated with fastidious BV-associated microbes that are linked with infertility following pelvic inflammatory disease (PID). METHOD OF STUDY: Sneathia spp., Atopobium vaginae, BVAB1, and Ureaplasma urealyticum were measured in 250 women from the PID Evaluation and Clinical Health (PEACH) study. Relative risk (RR) and 95% confidence intervals (CI) were calculated adjusting for chlamydia and gonorrhea. Principal component analysis was used to adjust for population stratification. A false discovery rate q-value of 0.05 was significant. RESULTS: TLR2-1733C>A (P = .003) and TLR2-616A>G (P = .004) were associated with cervical A. vaginae. TLR2-1733C>A and TLR6-438C>T were associated with A. vaginae detection in the endometrium, but this was not significant after adjustment for multiple comparisons (FDR q-value = 0.06). CONCLUSION: Host gene variants in TLR2 signaling pathways were modestly associated with cervical A. vaginae in women with clinical PID.
Asunto(s)
Infecciones por Corynebacterium/genética , Corynebacterium/fisiología , Endometrio/inmunología , Receptor Toll-Like 2/genética , Vaginosis Bacteriana/genética , Adolescente , Adulto , Infecciones por Corynebacterium/inmunología , Estudios Transversales , Endometrio/microbiología , Femenino , Predisposición Genética a la Enfermedad , Genotipo , Humanos , Polimorfismo de Nucleótido Simple , Análisis de Componente Principal , Riesgo , Transducción de Señal , Receptores Toll-Like/genética , Vaginosis Bacteriana/inmunología , Adulto JovenRESUMEN
Mycoplasma genitalium is increasingly appreciated as a common cause of sexually transmitted disease syndromes, including urethritis in men and cervicitis, endometritis, pelvic inflammatory disease, and possibly preterm birth, tubal factor infertility, and ectopic pregnancy in women. Despite these disease associations, which parallel those of Chlamydia trachomatis and Neisseria gonorrhoeae, the mechanisms by which this pathogen elicits inflammation, causes cellular damage, and persists in its only natural host (humans) are unique and are not fully understood. The purpose of this review is to briefly provide a historical background on the discovery, microbiology, and recognition of M. genitalium as a pathogen, and then summarize the recent advances in our understanding of the molecular biology and pathogenesis of this unique urogenital organism. Collectively, the basic scientific discussions herein should provide a framework for understanding the clinical and epidemiological outcomes described in the accompanying articles in this supplemental issue.
Asunto(s)
Evasión Inmune , Infecciones por Mycoplasma/inmunología , Mycoplasma genitalium/genética , Mycoplasma genitalium/patogenicidad , Enfermedades Bacterianas de Transmisión Sexual/inmunología , Femenino , Genoma Bacteriano , Humanos , Inmunidad , Masculino , Mycoplasma genitalium/inmunología , Factores de Riesgo , Enfermedades Bacterianas de Transmisión Sexual/complicaciones , Uretritis/complicaciones , Uretritis/microbiología , Cervicitis Uterina/complicaciones , Cervicitis Uterina/microbiologíaRESUMEN
We assessed the association between recent bacterial vaginosis (BV) and incident Mycoplasma genitalium, a sexually transmitted bacterium associated with adverse female reproductive health outcomes. Female sex workers in Mombasa, Kenya, completed a monthly sexual behavior interview and clinical examination. During February 2005-February 2006, vaginal fluid specimens collected from women every other month were tested for M. genitalium by nucleic acid amplification testing. Vaginal microbiota were assessed monthly and categorized by Nugent score (0-3 = normal microbiota, 4-6 = intermediate microbiota disruption, and 7-10 = BV). A discrete failure time analysis for multiple events using logistic regression was employed to estimate the odds of incident M. genitalium infection at follow-up visits among women with BV (vs. normal microbiota) at the preceding visit. Among the 280 women, 54.3% were positive for human immunodeficiency virus. At baseline, 16.1% had prevalent M. genitalium infection and 40.4% had prevalent BV. There were 59 incident M. genitalium infections among 50 women, for an incidence rate of 34.6 cases per 100 person-years. Following adjustment for age, human immunodeficiency virus status, and time, prior BV was associated with a 3.5-fold increase in odds of incident M. genitalium (adjusted odds ratio = 3.49, 95% confidence interval: 1.86, 6.56). This strong association suggests that BV may enhance susceptibility to M. genitalium infection.
Asunto(s)
Mycoplasma genitalium/patogenicidad , Trabajadores Sexuales/estadística & datos numéricos , Vagina/microbiología , Vaginosis Bacteriana/microbiología , Adulto , Comorbilidad , Femenino , Infecciones por VIH/epidemiología , Humanos , Incidencia , Kenia/epidemiología , Mycoplasma genitalium/aislamiento & purificación , Prevalencia , Estudios Prospectivos , Salud Reproductiva , Factores de Riesgo , Conducta Sexual , Enfermedades de Transmisión Sexual/epidemiología , Vaginosis Bacteriana/complicaciones , Vaginosis Bacteriana/epidemiología , Vaginosis Bacteriana/transmisiónRESUMEN
Mycoplasma genitalium is an underappreciated cause of human reproductive tract disease, characterized by persistent, often asymptomatic, infection. Building on our previous experiments using a single female pig-tailed macaque as a model for M. genitalium infection (G. E. Wood, S. L. Iverson-Cabral, D. L. Patton, P. K. Cummings, Y. T. Cosgrove Sweeney, and P. A. Totten, Infect Immun 81:2938-2951, 2013, https://doi.org/10.1128/IAI.01322-12), we cervically inoculated eight additional animals, two of which were simultaneously inoculated in salpingeal tissue autotransplanted into abdominal pockets. Viable M. genitalium persisted in the lower genital tract for 8 weeks in three animals, 4 weeks in two, and 1 week in one; two primates resisted infection. In both animals inoculated in salpingeal pockets, viable M. genitalium was recovered for 2 weeks. Recovery of viable M. genitalium from lower genital tract specimens was improved by diluting the specimen in broth and by Vero cell coculture. Ascension to upper reproductive tract tissues was not detected, even among three persistently infected animals. M. genitalium-specific serum antibodies targeting the immunodominant MgpB and MgpC proteins appeared within 1 week in three animals inoculated both cervically and in salpingeal pockets and in one of three persistently infected animals inoculated only in the cervix. M. genitalium-specific IgG, but not IgA, was detected in cervical secretions of serum antibody-positive animals, predominantly against MgpB and MgpC, but was insufficient to clear M. genitalium lower tract infection. Our findings further support female pig-tailed macaques as a model of M. genitalium infection, persistence, and immune evasion.
Asunto(s)
Infecciones por Mycoplasma/microbiología , Mycoplasma genitalium , Animales , Anticuerpos Antibacterianos/sangre , Anticuerpos Antibacterianos/inmunología , Carga Bacteriana , Biopsia , Línea Celular , Cuello del Útero/microbiología , Modelos Animales de Enfermedad , Femenino , Genitales Femeninos/microbiología , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Macaca nemestrina , Enfermedades de los Monos/microbiología , Infecciones por Mycoplasma/patología , Mycoplasma genitalium/inmunologíaRESUMEN
OBJECTIVES: As pelvic inflammatory disease (PID) aetiology is not completely understood, we examined the relationship between select novel bacteria, PID and long-term sequelae. METHODS: Fastidious bacterial vaginosis (BV)-associated bacteria (Sneathia (Leptotrichia) sanguinegens, Sneathia amnionii, Atopobium vaginae and BV-associated bacteria 1 (BVAB1)), as well as Ureaplasma urealyticum and Ureaplasma parvum were identified in cervical and endometrial specimens using organism-specific PCR assays among 545 women enrolled in the PID Evaluation and Clinical Health study. Risk ratios and 95% CIs were constructed to determine associations between bacteria, histologically confirmed endometritis, recurrent PID and infertility, adjusting for age, race, gonorrhoea and chlamydia. Infertility models were additionally adjusted for baseline infertility. RESULTS: Persistent detection of BV-associated bacteria was common (range 58% for A. vaginae to 82% for BVAB1) and elevated the risk for persistent endometritis (RRadj 8.5, 95% CI 1.6 to 44.6) 30â days post-cefoxitin/doxycycline treatment, independent of gonorrhoea and chlamydia. In models adjusted for gonorrhoea and chlamydia, endometrial BV-associated bacteria were associated with recurrent PID (RRadj 4.7, 95% CI 1.7 to 12.8), and women who tested positive in the cervix and/or endometrium were more likely to develop infertility (RRadj 3.4, 95% CI 1.1 to 10.4). Associations between ureaplasmas and PID sequelae were modest. CONCLUSIONS: To our knowledge, this is the first prospective study to demonstrate that S. sanguinegens, S. amnionii, BVAB1 and A. vaginae are associated with PID, failure of the Centers for Disease Control and Prevention-recommended treatment to eliminate short-term endometritis, recurrent PID and infertility. Optimal antibiotic regimens for PID may require coverage of novel BV-associated microbes.
Asunto(s)
Endometritis/microbiología , Infertilidad Femenina/microbiología , Enfermedad Inflamatoria Pélvica/microbiología , Vagina/microbiología , Vaginosis Bacteriana/microbiología , Adolescente , Adulto , Antibacterianos/uso terapéutico , Cefoxitina/uso terapéutico , Doxiciclina/uso terapéutico , Quimioterapia Combinada , Endometritis/tratamiento farmacológico , Endometritis/epidemiología , Femenino , Humanos , Infertilidad Femenina/prevención & control , Enfermedad Inflamatoria Pélvica/tratamiento farmacológico , Enfermedad Inflamatoria Pélvica/epidemiología , Estudios Prospectivos , Estados Unidos/epidemiología , Vaginosis Bacteriana/tratamiento farmacológico , Vaginosis Bacteriana/epidemiología , Adulto JovenRESUMEN
Mycoplasma genitalium is a sexually transmitted pathogen and is associated with reproductive tract disease that can be chronic in nature despite the induction of a strong antibody response. Persistent infection exacerbates the likelihood of transmission, increases the risk of ascension to the upper tract, and suggests that M. genitalium may possess immune evasion mechanism(s). Antibodies from infected patients predominantly target the MgpB adhesin, which is encoded by a gene that recombines with homologous donor sequences, thereby generating sequence variation within and among strains. We have previously characterized mgpB heterogeneity over the course of persistent infection and have correlated the induction of variant-specific antibodies with the loss of that particular variant from the infected host. In the current study, we examined the membrane topology, antibody accessibility, distribution of amino acid diversity, and the location of functional and antigenic epitopes within the MgpB adhesin. Our results indicate that MgpB contains a single transmembrane domain, that the majority of the protein is surface exposed and antibody accessible, and that the attachment domain is located within the extracellular C-terminus. Not unexpectedly, amino acid diversity was concentrated within and around the three previously defined variable regions (B, EF, and G) of MgpB; while nonsynonymous mutations were twice as frequent as synonymous mutations in regions B and G, region EF had equal numbers of nonsynonymous and synonymous mutations. Interestingly, antibodies produced during persistent infection reacted predominantly with the conserved C-terminus and variable region B. In contrast, infection-induced antibodies reacted poorly with the N-terminus, variable regions EF and G, and intervening conserved regions despite the presence of predicted B cell epitopes. Overall, this study provides an important foundation to define how different segments of the MgpB adhesin contribute to functionality, variability, and immunogenicity during persistent M. genitalium infection.
Asunto(s)
Adhesinas Bacterianas , Reacciones Antígeno-Anticuerpo , Sitios de Unión de Anticuerpos , Membrana Celular/metabolismo , Epítopos/inmunología , Mycoplasma genitalium , Adhesinas Bacterianas/química , Adhesinas Bacterianas/inmunología , Adhesinas Bacterianas/metabolismo , Secuencia de Aminoácidos/fisiología , Aminoácidos/genética , Aminoácidos/inmunología , Animales , Reacciones Antígeno-Anticuerpo/inmunología , Mapeo Epitopo , Epítopos/química , Epítopos/metabolismo , Femenino , Macaca nemestrina , Datos de Secuencia Molecular , Enfermedades de los Monos/inmunología , Infecciones por Mycoplasma/inmunología , Mycoplasma genitalium/inmunología , Estructura Terciaria de Proteína , Análisis de Secuencia de ADN , Homología de SecuenciaRESUMEN
OBJECTIVE: Ureaplasma urealyticum biovar 2 (UU-2), but not Ureaplasma parvum (UP), has been associated with non-gonococcal urethritis (NGU), but little is known about species-specific responses to standard therapies. We examined species-specific treatment outcomes and followed men with treatment failure for 9â weeks. METHODS: From May 2007 to July 2011, men aged ≥16 attending a sexually transmitted disease (STD) clinic in Seattle, Washington, with NGU (urethral discharge or urethral symptoms plus ≥5 polymorphonuclear leucocytes /high-powered field) enrolled in a double-blind, randomised trial. Participants received active azithromycin (1â g) + placebo doxycycline or active doxycycline (100â mg twice a day ×7â days) + placebo azithromycin. Ureaplasma were detected in culture followed by species-specific PCR. Outcomes were assessed at 3, 6 and 9â weeks. At 3â weeks, men with persistent Ureaplasma detection received 'reverse therapy' (e.g., active doxycycline if they first received active azithromycin). At 6â weeks, persistently positive men received moxifloxacin (400â mg×7â days). RESULTS: Of 490 men, 107 (22%) and 60 (12%) were infected with UU-2 and UP, respectively, and returned at 3â weeks. Persistent detection was similar for UU-2-infected men initially treated with azithromycin or doxycycline (25% vs. 31%; p=0.53), but differed somewhat for men with UP (45% vs. 24%; p=0.11). At 6â weeks, 57% of UU-2-infected and 63% of UP-infected men who received both drugs had persistent detection. Failure after moxifloxacin occurred in 30% and 36%, respectively. Persistent detection of UU-2 or UP was not associated with signs/symptoms of NGU. CONCLUSIONS: Persistent detection after treatment with doxycycline, azithromycin and moxifloxacin was common for UU and UP, but not associated with persistent urethritis. TRIAL REGISTRATION NUMBER: NCT00358462.
Asunto(s)
Antibacterianos/administración & dosificación , Azitromicina/administración & dosificación , Doxiciclina/administración & dosificación , Ureaplasma urealyticum/efectos de los fármacos , Uretritis/tratamiento farmacológico , Estudios de Seguimiento , Humanos , Masculino , Resultado del Tratamiento , Uretritis/microbiología , Washingtón/epidemiologíaRESUMEN
The human pathogen Mycoplasma genitalium employs homologous recombination to generate antigenic diversity in the immunodominant MgpB and MgpC proteins. Only recently, some of the molecular factors involved in this process have been characterized, but nothing is known about its regulation. Here, we show that M. genitalium expresses N-terminally truncated RecA isoforms via alternative translation initiation, but only the full-length protein is essential for gene variation. We also demonstrate that overexpression of MG428 positively regulates the expression of recombination genes, including recA, ruvA, ruvB and ORF2, a gene of unknown function co-transcribed with ruvAB. The co-ordinated induction of these genes correlated with an increase of mgpBC gene variation. In contrast, cells lacking MG428 were unable to generate variants despite expressing normal levels of RecA. Similarly, deletion analyses of the recA upstream region defined sequences required for gene variation without abolishing RecA expression. The requirement of these sequences is consistent with the presence of promoter elements associated with MG428-dependent recA induction. Sequences upstream of recA also influence the relative abundance of RecA isoforms, possibly through translational regulation. Overall, these results suggest that MG428 is a positive regulator of recombination and that precise control of recA expression is required to initiate mgpBC variation.
Asunto(s)
Proteínas Bacterianas/genética , Regulación Bacteriana de la Expresión Génica , Mycoplasma genitalium/genética , Recombinación Genética , Factores de Transcripción/metabolismo , Secuencia de Aminoácidos , Eliminación de Gen , Perfilación de la Expresión Génica , Datos de Secuencia Molecular , Recombinasas/biosíntesis , Alineación de Secuencia , Factores de Transcripción/genéticaRESUMEN
Mycoplasma genitalium is an emerging sexually transmitted pathogen associated with reproductive tract disease in men and women, and it can persist for months to years despite the development of a robust antibody response. Mechanisms that may contribute to persistence in vivo include phase and antigenic variation of the MgpB and MgpC adhesins. These processes occur by segmental recombination between discrete variable regions within mgpB and mgpC and multiple archived donor sequences termed MgPa repeats (MgPars). The molecular factors governing mgpB and mgpC variation are poorly understood and obscured by the paucity of recombination genes conserved in the M. genitalium genome. Recently, we demonstrated the requirement for RecA using a quantitative PCR (qPCR) assay developed to measure recombination between the mgpB and mgpC genes and MgPars. Here, we expand these studies by examining the roles of M. genitalium ruvA and ruvB homologs. Deletion of ruvA and ruvB impaired the ability to generate mgpB and mgpC phase and sequence variants, and these deficiencies could be complemented with wild-type copies, including the ruvA gene from Mycoplasma pneumoniae. In contrast, ruvA and ruvB deletions did not affect the sensitivity to UV irradiation, reinforcing our previous findings that the recombinational repair pathway plays a minor role in M. genitalium. Reverse transcription-PCR (RT-PCR) and primer extension analyses also revealed a complex transcriptional organization of the RuvAB system of M. genitalium, which is cotranscribed with two novel open reading frames (ORFs) (termed ORF1 and ORF2 herein) conserved only in M. pneumoniae. These findings suggest that these novel ORFs may play a role in recombination in these two closely related bacteria.
Asunto(s)
Adhesinas Bacterianas/genética , Variación Antigénica , ADN Helicasas/genética , ADN Helicasas/metabolismo , Mycoplasma genitalium/enzimología , Mycoplasma genitalium/genética , Recombinación Genética , Eliminación de Gen , Regulación Bacteriana de la Expresión Génica , Prueba de Complementación Genética , Neumonía por Mycoplasma/genética , Transcripción GenéticaRESUMEN
OBJECTIVE: Doxycycline, one of two recommended therapies for non-gonococcal urethritis (NGU), consists of a 7-day course of therapy (100 mg BID). Since suboptimal adherence may contribute to poor treatment outcomes, we examined the association between self-reported imperfect adherence to doxycycline and clinical and microbiologic failure among men with NGU. METHODS: Men aged ≥16 years with NGU attending a Seattle, WA, sexually transmitted diseases clinic were enrolled in a double-blind, parallel-group superiority trial from January 2007 to July 2011. Men were randomised to active doxycycline/placebo azithromycin or placebo doxycycline/active azithromycin. Imperfect adherence was defined as missing ≥1 dose in 7 days. Urine was tested for Chlamydia trachomatis (CT), Mycoplasma genitalium (MG), and Ureaplasma urealyticum-biovar 2 (UU-2) using nucleic acid amplification tests. Clinical failure (symptoms and ≥5 PMNs/HPF or discharge) and microbiologic failure (positive tests for CT, MG, and/or UU-2) were determined after 3 weeks. RESULTS: 184 men with NGU were randomised to active doxycycline and provided data on adherence. Baseline prevalence of CT, MG and UU-2 was 26%, 13% and 27%, respectively. 28% of men reported imperfect adherence, and this was associated with microbiologic failure among men with CT (aRR=9.33; 95% CI 1.00 to 89.2) and UU-2 (aRR=3.08; 95% CI 1.31 to 7.26) but not MG. Imperfect adherence was not significantly associated with clinical failure overall or for any specific pathogens, but it was more common among imperfectly adherent men with CT (aRR=2.63; 0.93-7.41, p=0.07). CONCLUSIONS: Adherence may be important for microbiologic cure of select pathogens. Factors other than adherence should be considered for CT-negative men with persistent NGU.
Asunto(s)
Antibacterianos/administración & dosificación , Azitromicina/administración & dosificación , Doxiciclina/administración & dosificación , Cumplimiento de la Medicación , Uretritis/tratamiento farmacológico , Orina/microbiología , Adolescente , Adulto , Antibacterianos/efectos adversos , Azitromicina/efectos adversos , Infecciones por Chlamydia/tratamiento farmacológico , Infecciones por Chlamydia/orina , Estudios de Cohortes , Doxiciclina/efectos adversos , Humanos , Masculino , Persona de Mediana Edad , Infecciones por Mycoplasma/tratamiento farmacológico , Infecciones por Mycoplasma/orina , Técnicas de Amplificación de Ácido Nucleico , Reacción en Cadena de la Polimerasa , Estudios Prospectivos , Análisis de Regresión , Insuficiencia del Tratamiento , Infecciones por Ureaplasma/tratamiento farmacológico , Infecciones por Ureaplasma/orinaRESUMEN
Mycoplasma genitalium is a sexually transmitted pathogen associated with several acute and chronic reproductive tract disease syndromes in men and women. To evaluate the suitability of a pig-tailed macaque model of M. genitalium infection, we inoculated a pilot animal with M. genitalium strain G37 in the uterine cervix and in salpingeal pockets generated by transplanting autologous Fallopian tube tissue subcutaneously. Viable organisms were recovered throughout the 8-week experiment in cervicovaginal specimens and up to 2 weeks postinfection in salpingeal pockets. Humoral and cervicovaginal antibodies reacting to MgpB were induced postinoculation and persisted throughout the infection. The immunodominance of the MgpB adhesin and the accumulation of mgpB sequence diversity previously observed in persistent human infections prompted us to evaluate sequence variation in this animal model. We found that after 8 weeks of infection, sequences within mgpB variable region B were replaced by novel sequences generated by reciprocal recombination with an archived variant sequence located elsewhere on the chromosome. In contrast, mgpB region B of the same inoculum propagated for 8 weeks in vitro remained unchanged. Notably, serum IgG reacted strongly with a recombinant protein spanning MgpB region B of the inoculum, while reactivity to a recombinant protein representing the week 8 variant was reduced, suggesting that antibodies were involved in the clearance of bacteria expressing the original infecting sequence. Together these results suggest that the pig-tailed macaque is a suitable model to study M. genitalium pathogenesis, antibody-mediated selection of antigenic variants in vivo, and immune escape.