Molecular Characterization Analyses Of Porcine Sapoviruses And Their Infection In Porcine Kindey Cells

Sapovirus(SaV),belonging to genus Sapovirus within the family Caliciviridae,containing a positive sense and single stranded RNA genome.SaVs were transmitted through fecal-oral route and can cause acute gastroenteritis in humans and pigs of all ages,besides,recent epidemiological survey showed that SaVs detected in many kinds of mammals,including chimpanzee,pig,rat and sea lion,displayed close phylogenetic relationship with human SaVs and similar human SaVs were also detected from water and shellfish,all of which indicated that SaV infection was an important public health safety problem.Although SaV have been found for almost 50 years,since it is hard to isolated and cell-cultured SaVs,there are no available SaVs' vaccines or antiviral drugs.Therefore,on the one hand,SaVs genetic diversity analyses based on the complete genome sequences,including found of highly divergent SaVs and recombinant strains,and the SaVs classification,especially the genotyping of animal SaVs,both of which are important basis for SaV molecular epidemiology analyses.On the other hand,understanding of SaV interaction with host cell,including clarifying how SaVs entry into host cells and bile acids regulated the host cells' signal transduction during their early infection stage,both of which are vital for exploring SaVs' cell cultivation and preventive controlFor understanding the SaVs' genetic diversity in China,we analyzed diarrheic feces collected from piglets in Shanghai using viral metagenomics and found a SaV G? strain SH1703,this is the first time that G? was reported in China and SH1703 showed 63.6%and 64.1%homology with other two reported G? strains RV0042 and K7,respectively,based on complete genome sequences.Recombination is an important mechanism for SaV genetic evolution and divergence and plays an important role for viral pathogenecity and host tropism,recombinant SaV strains have been reported in many countries,SaV intra-recombinant G? was firstly found in China and was certified by RDP and Simplot programs that it happened at RdRp-VP1 junction site of JJ259 and CH430,pig infected with the SaV-positive samples performed diarrhea and small intestine inflammation.More and more SaV sequences were submitted in GenBank database and it is necessary for SaV classification,especially genotyping,in our study,we calculated the pairwise distances based on 137 SaV complete VP1 nucleotide sequences and finally divided 19 genogroups of SaV into 51 genotypesThe only one cell-culture SaV Cowden strain was introduced from Academician Linda of American Ohio state university,previous research indicated that SaV uptake didn't require bile acids,pretreatment with specific inhibition on LLC-PK1 cells showed that porcine SaV entry cells dependent on clathrin-and cholesterol-mediated endocytosis and requiring dynamin and actin rearrangement,and SaV uncoating depends on the low pH environment.Bile acids were essential during early stage of SaV replication,transcriptome analyses showed that adding bile acids strongly enhance the expression level of 47 signaling pathways including PI3K-Akt and 25 related genesIn conclusion,to the best of our knowledge,this is the first report that SaV G? and recombinant strains were detected in China and the complete genome of these two representative strains were submitted to GenBank database,19 genogroups of SaVs were updated and divided into 51 genotypes,which enriches the understanding of porcine SaVs in China,enhances the SaV full genome sequences in GenBank database and supplies the data and reference for SaV classification and molecular characterization analyses.We explored SaVs entry into cells and bile acids regulating SaV's early infection in host cells,which supplies the vital clues for finally clarifying the interaction mechanism between SaV and host.