Isolation, Identification Of Japanese Encephalitis Virus In Children In Chongqing And Its Molecular Characteristics

Objective: To isolate and identify the Japanese encephalitis virus (JEV)from patients’ specimens in Chongqing in2011, and analyze the molecularcharacteristic of the isolates.Methods: A total of145specimens were collected from patients withepidemic encephalitis B in Children’s Hospital of Chongqing MedicalUniversity. The JEV strain was isolated and identified by the nested PCR,real-time PCR, inoculating BHK-21and2～3days suckling mice., thensequenced by TaKaRa company. Computer analyze of phylogenetic, nucleicacid data and deduced amino acid sequence were accomplished using theClustal X(1.8) and Mega5programs.Results: Only one strain was isolated from patient’s cerebrospinal fluidspecimen, named CQ11-66. The isolated led to cytopathogenic effect (CPE)on BHK-21cell and caused the2～3day old suckling mice to death.Comparison of ORF genome and E gene sequence of CQ11-66strain withother21JEV strains and30JEV isolates in GenBank, respectively, showed that ORF genome nucleotide sequence divergence ranges from1.68%to18.46%and deduced amino acid divergence ranges from0.32%to4.89%,while the E gene nucleotide divergence ranged from1.69%to18.74%andcorresponding amino acid divergence ranged from1.01%and5,13%.Comparison of the CQ11-66ORF genome with the vaccine strainSA14-14-2, a total of201nucleotide differences and42amino aciddifferences were found. Comparison of the CQ11-66strain E gene withvaccine strain SA14-14-2also performed. Overall36nucleotide changeswere detected, which resulted in15amino acid changes. Phylogeneticanalysis through the ORF genome and the E genome nucleotide sequenceshowed that the CQ11-66strain belonged to genotype Ⅲ。Conclusion: Only one strain, CQ11-66, was isolated from patient’scerebrospinal fluid specimen,. There are some divergences betweenCQ11-66strain and other JEV isolates, Phylogenetic analysis revealedCQ11-66strain belongs to genotype Ⅲ.