RESUMO
OBJECTIVE: To establish the real-time fluorescent PCR method for detecting enterovirus, enterovirus 71 and Coxsackievirus A 16 nucleic acid. METHODS: Primers and MGB probe were chosen for virus gene. The samples of 38 HFMD patients were analyzed by TaqMan-MGB PCR technique on a fluorescence real-time PCR instrument,and the results were compared with those by conventional RT-PCR. RESULTS: The real-time fluorescent PCR positive rates of EV, EV71 and Cox A16 were 73.7%, 60.5%, 13.2%; the conventional RT-PCR were 71.1%, 55.3%, 13.2%. There were no significant differences between the two methods. CONCLUSION: The real-time fluorescent PCR detecting method of EV, EV71 and Cox A16 nucleic acid have been established successfully.
Assuntos
Enterovirus Humano A/isolamento & purificação , Doença de Mão, Pé e Boca/diagnóstico , Reação em Cadeia da Polimerase/métodos , Criança , Pré-Escolar , Primers do DNA/química , Primers do DNA/genética , Enterovirus Humano A/genética , Feminino , Corantes Fluorescentes/química , Doença de Mão, Pé e Boca/patologia , Doença de Mão, Pé e Boca/virologia , Humanos , Lactente , MasculinoRESUMO
OBJECTIVE: To demonstrate molecular characterization of a newly isolated enterovirus. METHODS: Virus were isolated from patient with unknown-causing disease and tested by reverse transcription-polymerase chain reaction (RT-PCR) and 5'3'RACE (rapid amplification of cDNA ends, RACE), in an attempt to obtain the sequence of this newly isolated enterovirus. RESULTS: Sequence analysis showed that this newly isolated enterovirus shared 83%-94% nucleotide identity and 91%-100% amino acid identity with enterovirus 89. Phylogenetic analysis indicated that it was probably a new subtype of enterovirus 89. CONCLUSION: This newly isolated enterovirus in the stool specimen from patient has the same serotype with enterovirus 89, but it was probably a new subtype of enterovirus 89.