Detection And Sequence Analysis Of Buffalo Astrovirus In Parts Of Guangxi In 2019

Astrovirus(Ast V)is a single positive-strand RNA virus without envelope capsule and the diameter of Ast Vs about 28-30 nm.,Ast Vs can cause diarrhea in mammals and hepatitis in birds.In recent years,it has been found that some Mamastrovirus strains could cause extra-intestinal infection and even cause neurological symptoms.In order to understand the prevalence of Water Buffalo Astrovirus(BufAstV)in Guangxi.in this study,297 fecal samples were collected from 15 farms located in Nanning,Guiguang,Beihai,Hengxian and Lingshan of Guangxi Zhuang Autonomous Region in 2019.The samples deteceted of BufAstV was performed by nested polymerase chain reaction(Nest-PCR).The full-length ORF2 gene of BufAstV was amplified by rapid-amplification of c DNA ends(RACE).Base on the new-generation sequence technology(NGS),the whole genome of two BufAstV positive samples were sequenced.The genetic evolution analysis of the above results was carried out.The results showed that the BufAstV positive rate of the samples was 11%(33/297),and the positive rate of 1-150 d calves was 18.7%(27/144)was higher than cattle of other age.In addition,the multiple infection of different pathogens was common.33 BufAstV-positive samples were detected other viruses by PCR and the results showed thatmultiple infection rate was 45.4%(15/33).The multiple infection pathogens with BufAstV including Bovine Viral Diarrhea Virus(BVDV)(30.3%,10/33),Bovine Enterovirus(BEV)(24.2%,8/33)and Bovine Rotavirus(BRV)(21.2%,7/33)In this study,19 partial ORFlb genes of BufAstV were sequenced,and the nucleotide homology between different strains in this study was 53.9%-99.3%.Phylogenetic analysis found that the BufAstV ORF1 b genes in this study were divided into four clusters:Mamastrovirus 13,Mamastrovirus 27,Mamastrovirus 32 and Mamastrovirus33.Base on the 19 partial ORF1 b sequences as above,the ORF2 gene of BufAstV was amplified by RACE method.4 full-length ORF2 genes were obtained which named BufAstV/GX/NN/ORF2/S2(2271 bp),BufAstV/GX/NN/ORF2/12(2322 bp),BufAstV/GX/NN/ORF2/17(2217 bp)and BufAstV/GX/NN/ORF2/14(2289 bp),respectively,The nucleotide homology between these four ORF2 genes was 37.9%-53.8%.Phylogenetic analysis showed that BufAstV/GX/NN/S2,BufAstV/GX/NN/12 and BufAstV/GX/NN/17 were clustered in Mamastrovirus 33 genotypes,however,BufAstV/GX/NN/ORF2/14 was clustered in Mamastrovirus 13 which belong to Neurotropic Astrovirus.Two full-length genome sequences of BufAstV named BufAstV/GX/NN/14 and BufAstV/GX/NN/12 were obtained by NGS technology,The genome length of BufAstV/GX/NN/14 and BufAstV/GX/NN/12 were 6406 bp and 6230 bp,respectively,including three open reading frames(ORF).Phylogenetic analysis showed that BufAstV/GX/NN/14 belonged to Mamastrovirus 13 and BufAstV/GX/NN/12 belonged to Mamastrovirus 33.In this study,297 buffalo feces samples were detected BufAstV from five regions of Guangxi,and the BufAstV positive rate was 11%.19 partial ORF1 b and 4 complete ORF2 genes were sequenced.In addition,2 full-length genome sequences of BufAstV which belong to different genotypes of Mamastrovirus were obtained.This is the first time to find out the infection of Neurotropic Astrovirus from mammal in China.In addition,this study obtained the complete genome sequence of BufAstV from the feces samples of buffalo and provides important scientific basis for understanding its epidemic prevalent and molecular characteristics of BufAstV.