RESUMO
Rotaviruses continue being the most important pathogens responsible of diarrhea in young children worldwide. Seminested reverse transcription polymerase chain reaction (RT-PCR) is used to determine rotavirus genotype; however, this technique employs multistep procedures. The real-time RT-PCR is a fast and reliable tool that can be used as rotavirus genotyping tool, especially in rotavirus outbreaks. In this study, we tested a real-time RT-PCR to identify rotavirus genotype using a panel of 252 samples from patients with diarrheal disease caused by G9P[4] and G12P[8] genotypes, which were identified as emerging rotaviruses in 2 outbreaks in Chiapas, Mexico. Our results show that the real-time RT-PCR assay detected these rotaviruses, and it allowed us to identify mixed genotype infections, G/P combinations, and the viral abundance in some samples in which the seminested assay could not identify them. Therefore, the real-time RT-PCR is a molecular tool that can be great support during rotavirus outbreaks.
Assuntos
Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Infecções por Rotavirus/diagnóstico , Infecções por Rotavirus/virologia , Rotavirus/classificação , Rotavirus/genética , Fezes/virologia , Genes Virais , Genótipo , Humanos , Epidemiologia Molecular/métodos , RNA Viral , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reprodutibilidade dos Testes , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Sensibilidade e EspecificidadeRESUMO
Rotavirus produces diarrhea in children under 5 years old. Most of those conventional methods such as polyacrylamide gel electrophoresis (PAGE) and reverse transcription-polymerase chain reaction (RT-PCR) have been used for rotavirus detection. However, these techniques need a multi-step process to get the results. In comparison with conventional methods, the real-time RT-PCR is a highly sensitive method, which allows getting the results in only one day. In this study a real-time RT-PCR assay was tested using a panel of 440 samples from patients with acute gastroenteritis, and characterized by PAGE and RT-PCR. The results show that the real-time RT-PCR detected rotavirus from 73% of rotavirus-negative samples analyzed by PAGE and RT-PCR; thus, the percentage of rotavirus-positive samples increased to 81%. The results indicate that this real-time RT-PCR should be part of a routine analysis, and as a support of the diagnosis of rotavirus in Mexico.
