Isolation,Identification and Phylogoenetic Analysis of the Genotype G6P[5]Bovine Rotavirus in Heilongjiang Province
Bovine rotavirus(BRV)is one of the main pathogens causing acute viral gastroenteritis in calves.In order to investigate the genotype and genetic evolution diversity of BRV in cattle in Heilongjiang province,the BRV positive stool samples were isolated from rhesus monkey embryonic kidney cells(MA-104),and the isolated virus strains were identified by reverse transcription-polymerase chain reaction(RT-PCR)detection,indirect immunofluorescence identification and electron microscope observation;further,the whole genome of the isolated virus was obtained by Illumina second-generation sequencing technology,and its sequence was analyzed and VP4 and VP7 evolutionary trees were constructed.The results showed that the cytopathic effect(CPE)of MA-104 cells was observed in the 5th generation,and CPE was stable in the 8th generation.The isolated virus strain was identified as BRV positive by RT-PCR and indirect immunofluorescence.The electron microscope observation showed that the typical wheel-shaped non-envelope virus particles were observed in the cell culture,consistent with the morphological characteristics of BRV,indicating that a BRV strain was successfully isolated and named BRV HLJ-J7;Genome sequencing analysis showed that its genotype was G6-P[5]-I2-R2-C2-M2-A11-N2-T6-E2-H3.The results of genetic evolution analysis showed that the VP4 gene had the closest genetic relationship with the Chinese isolate Y3,with 98.2%homology;VP7 gene has the closest genetic relationship with the Chinese strain LN19-4,with 98.8%homology.A G6P[5]type BRV was successfully isolated which laid a foundation for the study of genetic evolution and molecular epidemiology of BRV in Heilongjiang province.
bovine rotavirusseparation and identificationG6P[5]typegenetic evolution
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