Complete Genome Sequence Of A Rodent Parvovirus,Polyomavirus And Gemycircularvirus In Hainan Island

Objective: Pathogenic and potentially pathogenic viruses carried by wild rodent in some part of Hainan province were genome analyzed,to provide data for preventing and controlling the prevalence of rodent-borne emerging infectious diseases.Methods: Based on the lab previous results of metagenomic,high-throughput sequencing analysis of wild rodent samples from part of Hainan Province,the reads of parvovirus(ParV),polyomavirus(PyV)and gemycircularvirus(GemyCV)were selected,respectively.The genome of the three virus were by PCR amplifticated,assemblied and analyzed,and the Nested PCR method was preliminarily established to detect to ParV and PyV.Results:(1)Genome analysis of rodent-borne parvovirus Based on the previous results of metagenomic,high-throughput sequencing and bioinformatics analysis of wild rodent pharyngeal swab samples collected in Lingao city in Hainan Province,1603 reads were found to have the homology with rodent ParV(80 %-94 %),indicating that it should be ParV(designated ParV/LG strain).The complete genome sequences of ParV/LG strain was obtained by PCR amplification and gene assembly.The genome of ParV/LG strain comprise of 4691 nucleotides(nt),encoding NS1 nonstructural protein(721aa)and VP1 viral protein(583aa).The G+C content of NS1 was 54.57 % and VP1 was 45.43 %.The Walker loop 448-GPATTGKS-455[GXXXXGK(T/S)],which is an ATP-or GTP-binding motif,was found in the NS1.Phylogenetic analysis based on NS1 and VP1 proteins was deduced indicated that ParV/LG strain was closely related to Kilham rat virus strain(KRV)(AF321230.1),their genome proteins were also related to each other(NS1,91 %;VP1,97 %),the genomic sequence of ParV/LG strain has been deposited in GenBank under the accession no.MK530648.1,named parvovirus Ro/Lingao/China.The Nested PCR based on the VP1 gene method was preliminarily established to detect to parvovirus of Lingao strain.(2)Genome analysis of rodent-borne polyomavirus Based on the previous results of metagenomic,high-throughput sequencing and bioinformatics analysis of pharyngeal swab samples in wild Rattus norvegicus of Haikou city in Hainan Province,114 reads were found to have the homology with rodents PyV(93 %-97 %),indicating that it should be PyV(designated PyV/HK strain).The complete genome sequences of PyV/HK strain was obtained by PCR amplification and gene assembly.The genome of PyV/HK strain comprise of 5318 nucleotides(nt),encoding early gene and late gene region,the early region encoding large T(LTAg),middle T(MTAg)antigens and small T(STAg),the late region encoding the major capsid proteins VP1,capsid proteins VP2 and VP3.Phylogenetic analysis based on NS1 and VP1 proteins was deduced indicated that PyV/HK strain was closely related to Rattus norvegicus polyomavirus 1 strain(RnorPyV1)(KR079545.1),LTAg,MTAg and VP1 proteins were also related to each other(99 %,99 %,99 %),the genomic sequence of PyV/HK strain has been deposited in GenBank under the accession no.MK372231.1,named polyomavirus sp RN/Haikou/China.The Nested PCR based on the VP1 gene method was preliminarily established to detect to polyomavirus of Haikou strain.(3)Genome analysis of rodent-borne gemycircularvivus Based on the previous results of metagenomic,high-throughput sequencing and bioinformatics analysis of pharyngeal swab samples in wild Rattus norvegicus of Haikou city in Hainan Province.298 reads were found to have the highest homology with GemyCV(81 %-97 %),indicating that it should be GemyCV(designated GemyCV/HK strain).The complete genome sequences of GemyCV/HK strain was obtained by PCR amplification and gene assembly.The genome of GemyCV/HK strain comprise of 2199 nucleotides(nt),encoding a capsid protein(Cap 322aa),a replication associated protein1(Rep1 194aa)and a replication associated protein 2(Rep2 112aa).The intergenic region(127-nt)contained a stem loop structure having an atypical nonamer [TAAAATTTA],GemyCV/HK strain had unique intron(140-nt).Phylogenetic analysis based on Cap and Rep proteins was deduced indicated that GemyCV/HK strain was closely related to gemycircularvirus SL1 strain(GemyCV-SL1)(KP133075.1),Cap and Rep proteins were also related to each other(97 %,99 %),the genomic sequence of GemyCV/HK strain has been deposited in GenBank under the accession no.MK293717.1,named gemycircularvirus RN/Haikou/China.Conclusion:(1)The whole genome sequences of parvovirus were obtained,phylogenetic analysis indicated that parvovirus of Lingao strain was closely related to Kilham rat virus(KRV)strain(AF321230.1)that causes disease in rats,it indicates that rodents carries ParV in Haikou,Hainan Province.The Nested PCR based on the VP1 gene method was preliminarily established to detect to parvovirus of Haikou strain.(2)The whole genome sequences of polyomavirus were obtained,phylogenetic analysis indicated that polyomavirus of Haikou strain was closely related to Rattus norvegicus polyomavirus 1(RnorPyV1)strain(KR079545.1),it indicates that rodents carries PyV in Haikou,Hainan Province.The Nested PCR based on the VP1 gene method was preliminarily established to detect to polyomavirus of Haikou strain.(3)The whole genome sequences of gemycircularvirus were obtained,phylogenetic analysis indicated that gemycircularvirus of Haikou strain was closely related to gemycircularvirus SL1(GemyCV-SL1)strain(KP133075.1),it indicates that rodents carries GemyCV in Haikou,Hainan Province.