ASF Serological Survey Of Some Large Scale Pig Farms And Virus Genetic Evolution Analysis In Hunan Province

African swine fever is an acute,febrile,and highly contagious disease characterized by systemic hemorrhage,respiratory disorders,and neurological symptoms caused by African swine fever virus(ASFV)in domestic and wild boars.ASF is an epidemic disease that requires explicit reporting in the Land Animal Health Code of the World Organization for Animal Health,and China lists it as a class Ⅰ animal disease.Since ASF was first discovered and reported in Kenya in East Africa in 1921,its spread has gradually expanded to the world.In August 2018,the first domestic ASF outbreak was reported in Shenbei District,Shenyang City,Liaoning Province,China.In a short period,ASF infection cases were also reported in other provinces and regions.ASF has caused huge economic losses to the pig industry in China and has aroused widespread concern in the industry.In order to understand the situation of ASF antibody in the serum of swine herds in Hunan province and the variation of ASFV strains in Hunan Province,in this study,we collected 600 serum samples from 10 large-scale farms and 10 ASFV positive tissue samples in Hunan Province.The sera were tested for ASFV antibody level by ELISA.The ten samples were all confirmed as ASFV positive by fluorescence quantitative PCR,then sent to Shanghai Bojie Medical Technology Co.,Ltd.Medical Laboratory.Partial genes including A179 L,A238L,A224 L,B646L,B602 L,B438L,EP402 R,CP204L,CP530 R,E183L,KP177 R,MGF110-1L,MGF360-1L,MGF360-16 R,NP419L,O174 L and TRS regions were sequenced by Sanger sequencing method.Blast analysis was performed to explore the nucleotide and amino acid homology between the obtained sequences and the domestic and foreign ASFV strains uploaded in NCBI.The results showed that,the positive rate of ASFV antibody of the 600 serum samples was 44.33%,and the farm positive rate in 10 large-scale farms was 50%.ASFV antibody levels varied greatly in deferent regions,some regions have a high rate,while some areas showed negative ASFV antibody level.In positive farms,ASFV antibody in breeding boars,sows and gilts were all at a relatively high level.In addition,the average S/P was between 1.5 and 1.7,the dispersion was between 35% and 50%,which indicted that all these three pig herds were susceptible to ASFV.Genetic evolution tree based on the data from the 10 ASFV positive samp-les showed that the sequencing strains were in the same branch with the dom-estic gene type II strains.According to homological analysis of nucleotide and amino acid sequences,we found that designated HN-HH8 strain had an insert at position 669 in B438 L gene,resulting in a change(LCLAHNLPASVEKGI AG to FVFSAQSTCVGRKRDCR)in the amino acid sequence at position 217,the translated amino acid sequence was terminated early.In addition,the NP419 gene of the designated HN-YZ13 strain had a base an insert at position 654,resulting in its amino acid sequence changed at position 205 to 234(YLHRK PLQWIAGQANAKTDSSELHFYVF to LFTPQTSAMDRGAGECEDRFLRAALL RV).The designated HN-YZ1 strain had no insertion of a repeat sequence(GG AATATATA)in the TRS region,showing intra-provincial difference.In summary,ASFV infection exists in some large-scale farms in Hunan Province,the positive rate of ASFV antibody is different in different regions and farms.ASFV strains epidemic in Hunan Province have high homology in nucleotide and amino acid sequences with other reported domestic strains,but there are some variations.The present study provides data of serological and pathogenic characteristics of ASFV in Hunan Province,which will benefit to guide the establishment of an effective prevention and control system and the vaccine development.