RESUMEN
BACKGROUND: Some sexually transmitted infectious agents, such as Chlamydia trachomatis and Herpes simplex, cause local inflammation, and could contribute to Human Papillomavirus (HPV) and cervical lesion progression. Thus, the aim of this study was to determine any association between the presence of microorganisms of gynecological importance, sexual behavior, clinical and demographical variables to the development and progress of cervical lesions. METHODS: One hundred and thirty-two women between 14 and 78 years and living at Vitória da Conquista, Bahia, Brazil, were included (62 individuals with cervical lesions and 70 without lesions). They answered a questionnaire to provide data for a socioeconomic and sexual activity profile. Samples of cervical swabs were collected and analyzed by PCR to detect genital microorganisms and HPV. Quantitative PCR was used to detect and quantify Ureaplasma urealyticum and Ureaplasma parvum. Univariate and multiple logistic regression were performed to measure the association with the cervical lesions, and an odds ratio (OR) with 95% confidence intervals (95%CI) were calculated. The Mann-Whitney U test was also used to compare the microorganism load in the case and control groups. The significance level was 5% in all hypotheses tested. RESULTS: Cervical lesions were associated with: women in a stable sexual relationship (OR = 14.21, 95%CI = 3.67-55.018), positive PCR for HPV (OR = 16.81, 95%CI = 4.19-67.42), Trichomonas vaginalis (OR = 8.566, 95%CI = 2.04-35.94) and Gardnerella vaginalis (OR = 6.13, 95%CI = 1.53-24.61), adjusted by age and qPCR for U. parvum. U. parvum load showed a statistical difference between the case and control groups (p-value = 0.002). CONCLUSION: Variables such as stable relationship, HPV, T. vaginalis, G. vaginalis were associated with cervical lesions in epidemiological studies. U. parvum load was higher in woman with cervical lesions compared with women without lesions. Additional studies are needed to better understand the role of these factors in cervical lesion development.
Asunto(s)
Infecciones por Papillomavirus/diagnóstico , Enfermedades de Transmisión Sexual/diagnóstico , Enfermedades del Cuello del Útero/diagnóstico , Adolescente , Adulto , Anciano , Brasil , Cuello del Útero/microbiología , Cuello del Útero/virología , Coinfección/diagnóstico , Coinfección/microbiología , Coinfección/virología , ADN Bacteriano/aislamiento & purificación , ADN Bacteriano/metabolismo , ADN Viral/aislamiento & purificación , ADN Viral/metabolismo , Femenino , Gardnerella vaginalis/genética , Gardnerella vaginalis/aislamiento & purificación , Humanos , Modelos Logísticos , Persona de Mediana Edad , Oportunidad Relativa , Papillomaviridae/genética , Papillomaviridae/aislamiento & purificación , Infecciones por Papillomavirus/transmisión , Infecciones por Papillomavirus/virología , Reacción en Cadena en Tiempo Real de la Polimerasa , Enfermedades de Transmisión Sexual/microbiología , Enfermedades de Transmisión Sexual/transmisión , Enfermedades de Transmisión Sexual/virología , Encuestas y Cuestionarios , Trichomonas vaginalis/genética , Trichomonas vaginalis/aislamiento & purificación , Ureaplasma/genética , Ureaplasma/aislamiento & purificación , Ureaplasma urealyticum/genética , Ureaplasma urealyticum/aislamiento & purificación , Enfermedades del Cuello del Útero/microbiología , Enfermedades del Cuello del Útero/virología , Adulto JovenRESUMEN
BACKGROUND: The role of Mycoplasma hominis and M. genitalium in urogenital tract infections remains unknown. Furthermore these mollicutes present a complex relationship with the host immune response. The role of inflammatory cytokines in infections also makes them good candidates to investigate bacterial vaginosis and mycoplasma genital infections. Therefore, the aim of this study was to detect the above-mentioned mollicutes by quantitative Polymerase Chain Reaction (qPCR) methodologies in vaginal swabs and dosage of cytokines. METHODS: Vaginal swabs and peripheral blood were collected from 302 women, including healthy individuals. The molecular findings were correlated with some individual behavioral variables, clinical and demographic characteristics, presence of other important microorganisms in vaginal swabs, and levels of interleukin (IL)-1ß and IL-6. RESULTS: M. hominis and M. genitalium were detected in 31.8% and 28.1% of samples, respectively. The qPCR results were associated with clinical signs and symptoms of the infections studied. The frequency of Trichomonas vaginalis, Gardnerella vaginalis, Neisseria gonorrhoeae and Chlamydia trachomatis was 3.0%, 21.5%, 42.4%, and 1.7% respectively. Increased levels of IL-1ß were associated with the presence of M. hominis and signs and/or symptoms of the genital infection of women studied. CONCLUSION: IL-1ß production was associated with the detection of M. hominis by qPCR. The sexual behavior of women studied was associated with the detection of mycoplasma and other agents of genital infections.
Asunto(s)
Infecciones por Mycoplasma/epidemiología , Mycoplasma genitalium/aislamiento & purificación , Mycoplasma hominis/aislamiento & purificación , Infecciones Urinarias/epidemiología , Vaginosis Bacteriana/epidemiología , Adolescente , Adulto , Anciano , Brasil/epidemiología , Infecciones por Chlamydia/epidemiología , Infecciones por Chlamydia/microbiología , Chlamydia trachomatis/aislamiento & purificación , Coinfección , Femenino , Enfermedades Urogenitales Femeninas/epidemiología , Enfermedades Urogenitales Femeninas/microbiología , Gardnerella vaginalis/aislamiento & purificación , Humanos , Persona de Mediana Edad , Infecciones por Mycoplasma/microbiología , Neisseria gonorrhoeae/aislamiento & purificación , Prevalencia , Enfermedades de Transmisión Sexual/epidemiología , Enfermedades de Transmisión Sexual/microbiología , Trichomonas vaginalis/aislamiento & purificación , Infecciones Urinarias/microbiología , Sistema Urogenital/microbiología , Vaginosis Bacteriana/microbiología , Adulto JovenRESUMEN
BACKGROUND: Bacterial pathogens have many strategies for infecting and persisting in host cells. Adhesion, invasion and intracellular life are important features in the biology of mollicutes. The intracellular location of Ureaplasma diversum may trigger disturbances in the host cell. This includes activation or inhibition of pro and anti-apoptotic factors, which facilitate the development of host damage. The aim of the present study was to associate U. diversum infection in HEp-2 cells and apoptosis induction. Cells were infected for 72hs with four U. diversum clinical isolates and an ATCC strain. The U. diversum invasion was analyzed by Confocal Laser Scanning Microscopy and gentamicin invasion assay. The apoptosis was evaluated using pro-apoptotic and anti-apoptotic gene expression, and FITC Annexin V/Dead Cell Apoptosis Kit. RESULTS: The number of internalized ureaplasma in HEp-2 cells increased significantly throughout the infection. The flow cytometry analysis with fluorochromes to detect membrane depolarization and gene expression for caspase 2, 3 and 9 increased in infected cells after 24 hours. However, after 72 hours a considerable decrease of apoptotic cells was observed. CONCLUSIONS: The data suggests that apoptosis may be initially induced by some isolates in association with HEp-2 cells, but over time, there was no evidence of apoptosis in the presence of ureaplasma and HEp-2 cells. The initial increase and then decrease in apoptosis could be related to bacterial pathogen-associated molecular pattern (PAMPS). Moreover, the isolates of U. diversum presented differences in the studied parameters for apoptosis. It was also observed that the amount of microorganisms was not proportional to the induction of apoptosis in HEp-2 cells.
Asunto(s)
Apoptosis/fisiología , Infecciones por Ureaplasma/fisiopatología , Ureaplasma/patogenicidad , Citoesqueleto de Actina/ultraestructura , Adhesión Bacteriana , Caspasa 2/metabolismo , Caspasa 3/metabolismo , Caspasa 9/metabolismo , Supervivencia Celular , Femenino , Citometría de Flujo , Expresión Génica , Gentamicinas/farmacología , Células HeLa/microbiología , Humanos , Microscopía Confocal , Moléculas de Patrón Molecular Asociado a Patógenos/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Estadísticas no Paramétricas , Factores de Tiempo , Factor de Necrosis Tumoral alfa/metabolismo , Ureaplasma/efectos de los fármacosRESUMEN
BACKGROUND: Bacterial pathogens have many strategies for infecting and persisting in host cells. Adhesion, invasion and intracellular life are important features in the biology of mollicutes. The intracellular location ofUreaplasma diversum may trigger disturbances in the host cell. This includes activation or inhibition of pro and anti-apoptotic factors, which facilitate the development of host damage. The aim of the present study was to associate U. diversum infection in HEp-2 cells and apoptosis induction. Cells were infected for 72hs with four U. diversum clinical isolates and an ATCC strain. The U. diversuminvasion was analyzed by Confocal Laser Scanning Microscopy and gentamicin invasion assay. The apoptosis was evaluated using pro-apoptotic and anti-apoptotic gene expression, and FITC Annexin V/Dead Cell Apoptosis Kit. RESULTS: The number of internalized ureaplasma in HEp-2 cells increased significantly throughout the infection. The flow cytometry analysis with fluorochromes to detect membrane depolarization and gene expression for caspase 2, 3 and 9 increased in infected cells after 24 hours. However, after 72 hours a considerable decrease of apoptotic cells was observed. CONCLUSIONS: The data suggests that apoptosis may be initially induced by some isolates in association with HEp-2 cells, but over time, there was no evidence of apoptosis in the presence of ureaplasma and HEp-2 cells. The initial increase and then decrease in apoptosis could be related to bacterial pathogen-associated molecular pattern (PAMPS). Moreover, the isolates of U. diversum presented differences in the studied parameters for apoptosis. It was also observed that the amount of microorganisms was not proportional to the induction of apoptosis in HEp-2 cells.
Asunto(s)
Humanos , Femenino , Ureaplasma/patogenicidad , Infecciones por Ureaplasma/fisiopatología , Apoptosis/fisiología , Factores de Tiempo , Ureaplasma/efectos de los fármacos , Adhesión Bacteriana , Citoesqueleto de Actina/ultraestructura , Gentamicinas/farmacología , Células HeLa/microbiología , Expresión Génica , Supervivencia Celular , Factor de Necrosis Tumoral alfa/metabolismo , Estadísticas no Paramétricas , Microscopía Confocal , Caspasa 3/metabolismo , Caspasa 2/metabolismo , Caspasa 9/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Citometría de Flujo , Moléculas de Patrón Molecular Asociado a Patógenos/metabolismoRESUMEN
Ureaplasma diversum in veterinary studies is an undesirable microbe, which may cause infection in bulls and may result in seminal vesiculitis, balanopostitis, and alterations in spermatozoids, whereas in cows, it may cause placentitis, fetal alveolitis, abortion, and birth of weak calves. U. diversum is released through organic secretions, especially semen, preputial and vaginal mucus, conjunctival secretion, and milk. The aim of the present study was to develop a TaqMan probe, highly sensitive and specific quantitative PCR (qPCR) assay for the detection and quantification of U. diversum from genital swabs of bovines. Primers and probes specific to U. diversum 16S rRNA gene were designed. The specificity, detection limit, intra- and inter-assay variability of qPCR to detect this ureaplasma was compared with the results of the conventional PCR assay (cPCR). Swabs of vaginal mucus from 169 cows were tested. The qPCR assay detected as few as 10 copies of U. diversum and was 100-fold more sensitive than the cPCR. No cross-reactivity with other Mollicutes or eubacteria was observed. U. diversum was detected in 79 swabs (46.42%) by qPCR, while using cPCR it was detected in 42 (25%) samples. The difference in cPCR and qPCR ureaplasma detection between healthy and sick animals was not statistically significant. But the U. diversum load in samples from animals with genital disorders was higher than in healthy animals. The qPCR assay developed herein is highly sensitive and specific for the detection and quantification of U. diversum in vaginal bovine samples.