Sexually transmitted infections detected by multiplex real time PCR in asymptomatic women and association with cervical Intraepithelial neoplasia / Infecções sexualmente transmissíveis detectadas por PCR multiplex em tempo real em mulheres assintomáticas e associação com neoplasia intraepitelial cervical

Abstract Objective To determine the frequency of sexually transmitted infections (STIs) in asymptomatic women and the association of STIs with cervical intraepithelial neoplasia (CIN). Methods A cross-sectional studywas performed, enrollingwomen examined in a general gynecology clinic and in a colposcopy referral center fromOctober 2014 to October 2015. The colposcopy groupconsisted of 71women, and the general gynecologygroupconsisted of 55 women. Cervical samples were collected for cervical cytology and a multiplex realtime polymerase chain reaction (PCR) was developed to detect human papillomavirus (HPV) and the STIs caused by the following microorganisms: Chlamydia trachomatis, Mycoplasma hominis, Mycoplasma genitalium, Ureaplasma urealyticum, and Neisseria gonorrhoeae. A multivariate analysis was performed by logistic regression, considering the significance level of 0.05. Results The general frequency of STIs was: 46.8% (HPV); 27.8% (C. trachomatis); 28.6% (M. genitalium); 0.8% (M. hominis); 4.8% (U. urealyticum); and 4.8% (N. gonorrhoeae). The significant risk factors for CIN were: HPV infection (odds ratio [OR] = 2.53; p = 0.024); C. trachomatis (OR = 3.04; p = 0.009); M. genitalium (OR = 2.37; p = 0.04); and HPV and C. trachomatis coinfection (OR = 3.11; p = 0.023). After the multivariate analysis, a significant associationwas found betweenHPVand CIN(OR = 2.48; 95% confidence interval [95%CI]: 1.04-5.92; p = 0.04); and between C. trachomatis and CIN (OR = 2.69; 95%CI: 1.11-6.53; p = 0.028). Conclusion The frequency of STIs was high in asymptomatic patients. Infections by HPV and C. trachomatis were independently associated with the presence of CIN. The high frequency of STIs in asymptomatic women suggests the need for routine screening of these infections.

Resumo Objetivo Determinar a frequência de infecções sexualmente transmissíveis (ISTs) em mulheres assintomáticas e a associação destas infecções com a neoplasia intraepitelial cervical (NIC). Métodos Foi realizado um estudo transversal recrutando mulheres atendidas em uma clínica ginecológica geral e em um centro de referência para colposcopia, de outubro de 2014 a outubro de 2015. O grupo de colposcopia consistiu de 71 mulheres, e o grupo de ginecologia geral consistiu de 55 mulheres. Amostras cervicais foram coletadas para citologia cervical e uma reação em cadeia de polimerase (RCP) multiplex em tempo real para detecção do vírus do papiloma humano (HPV) e das ISTs provocadas pelos seguintes micro-organismos: Chlamydia trachomatis, Mycoplasma hominis, Mycoplasma genitalium, Ureaplasma urealyticum e Neisseria gonorrhoeae. Foi realizada uma análise multivariada por regressão logística, considerando-se o nível de significância de 0,05. Resultados A frequência geral de ISTs foi: 46,8% (HPV); 27,8% (C. trachomatis); 28,6% (M. genitalium); 0,8% (M. hominis); 4,8% (U. urealyticum); e 4,8% (N. gonorrhoeae). Os fatores de risco significantes para NIC foram: infecção pelo HPV (razão de probabilidades [RP] = 2,53; p = 0,024); C. trachomatis (RP = 3,04; p = 0,009); M. genitalium (RP = 2,37; p = 0,04); e coinfecção por HPV e C. trachomatis (RP = 3,11; p = 0,023). Após a análise multivariada, foi encontrada uma associação significante entre HPV e NIC (RP = 2.48; intervalo de confiança de 95% [IC95%]: 1,04-5,92; p = 0,04) e entre C. trachomatis e NIC (RP = 2,69; IC95%: 1,11-6,53; p = 0,028). Conclusões A frequência de ISTs foi alta em mulheres assintomáticas. Infecções por HPV e C. trachomatis foram independentemente associadas com a presença de NIC. A alta frequência de ISTs em mulheres assintomáticas sugere a necessidade de rastreamento rotineiro dessas infecções.

In-house quantitative real-time PCR for the diagnosis of hepatitis B virus and hepatitis C virus infections

Abstract The quantification of viral nucleic acids in serum by real-time PCR plays an important role in diagnosing hepatitis B virus and hepatitis C virus infection. In this study, we developed an assay using specific primers and probes to quantify hepatitis B virus DNA or hepatitis C virus RNA in serum from infected patients. For standardization and validation of the assay, an international panel of hepatitis B virus/hepatitis C virus and standard plasmids was used. A correlation coefficient of 0.983 and 0.963 for hepatitis B virus and hepatitis C virus, respectively, was obtained based on cycle threshold values and concentrations of DNA or RNA. The standard curve showed a linear relationship from 19 IU/mL to 1.9 × 109 IU/mL of serum, with a coefficient of determination (r2) of 0.99. In sera from patients infected with hepatitis B virus or hepatitis C virus viral loads (19 IU/mL and 1.9 × 109 IU/mL), we quantified viral loads with a detection limit of 1.9 × 102 IU/mL. The real-time quantitative PCR assay developed in this study provides an ideal system for routine diagnosis and confirmation of indeterminate serological results, especially in immunosuppressed patients.