Epidemiology Investigation And Pathogenicity Of Porcine Circovirus Type 2 In Guangxi

Porcine circovirus(PCV),a member of the Circoviridae family,is one of the smallest known animal DNA viruses.At present,PCV mainly includes three kinds of viruses,PCV1,PCV2 and PCV3.PCV2 can be divided into 6 gene subtypes including PCV2a,PCV2b,PCV2c,PCV2d,PCV2e and PCV2f.PCV2 is the main pathogen of porcine circovirus associated diseases(PCVAD).Pigs infected with PCV2 alone are usually mainly infected with subclinical infection,but it will cause immunosuppression of the body,which will aggravate the occurrence of clinical diseases in the case of co-infection with other viruses.Although PCV2 is a DNA virus,its mutation rate is very high.In recent years,the epidemic strain of PCV2 has been evolving from PCV2b to PCV2d.To understand the prevalence of PCV2 in pigs in guangxi region and genetic features,this study collected 495 clinical samples from 42 pig farms in guangxi during 2017to 2019.PCV2 was detected by PCR and the PCV2 total positive rate is 39.6%(196/495),and the positive rate was 35.56%(48/135)in 2017,40.00%(66/165)in2018 and 42.05%(82/195)in 2019,which show that PCV2 infection in guangxi region showed a trend of rising year by year.In order to understand the genetic characteristics of PCV2,partial clinical samples positive for PCV2 were amplified to ORF2 and genome-wide sequences,and 44 ORF2 sequences and 18genome-wide sequences were obtained.Homology analysis showed that the homology of the whole genome sequence of the strain obtained in this study was between 94.1%and 99.9%.The amino acid comparison of Cap protein showed that there were a total of 14 amino acid variation sites,most of which were on the antigen epitopes of Cap protein.Genetic and evolutionary analysis showed that the sequences of PCV2 strain obtained in this study belonged to the PCV2a,PCV2b and PCV2d subtypes,among which 4 belonged to PCV2a,15 to PCV2b and 25 to PCV2d,indicating that PCV2d strain was the main circulating strain in guangxi.In order to clarify the pathogenicity of the current epidemic strain,the current epidemic strain PCV2d in guangxi is isolated and identified,and the pathogenicity is studied by animal regression test.The results show that the isolated strain PCV2d-AG43 is obviously pathogenic to pigs,and the main clinical symptoms after the attack are rough back hair and decreased appetite.After 28 days of challenge,there is a significant weight loss,which is mainly manifested as lymph node hemorrhage and enlargement.After challenge,there is a marked viremia,which is highest at 14 days,followed by a decrease.PCV2 was detected in all tissues and organs(heart,liver,lungs,spleen,kidney,inguinal and mandibular lymph nodes)after the 28 days challenge,in which the combination of lung,mandibular lymph and inguinal lymph nodes was highly virulent,up to 10~6copies per gram.Histopathological observation of lung mainly show alveolar wall thickening and inflammatory cell infiltration.The inguinal lymph nodes show lymphocytosis.These results indicate that the PCV2d epidemic strain AG43 isolated from guangxi region in this study is highly pathogenic and may cause economic losses to the pig industry if it is not prevented and controlled in time.PCV2 has a high positive rate in the pig population.In addition to pig-to-pig transmission,whether there are other possible transmission routes needs further study.Mosquitoes play an important role in the spread of pathogens.In this study,94 mosquitoes from a pig farm in simao district,yunnan province were collected and the positive rate of PCV2 in mosquitoes is detected by PCR.The total positive rate of PCV2 in mosquitoes was 15.96%(15/94).In order to study the genetic characteristics of PCV2 in mosquitoes,ORF2 sequence was amplified and sequenced,and a total of 14 sequences were obtained.Homology analysis showed that the nucleotide homology of ORF2 gene in mosquitoes was 89.5-99.9%,and the homology of corresponding amino acids was 86.2%-99.6%.Amino acid comparison showed that there were 32 amino acid mutation sites of PCV2 Cap protein in mosquitoes.Genetic and evolutionary analysis showed that PCV2a,PCV2b and PCV2d subtypes of PCV2 existed in mosquitoes,among which PCV2d was the main genotype,which was consistent with the current epidemic trend of PCV2 in pigs.These results suggest that mosquitoes may act as mechanical vectors of PCV2 and play a role in the transmission of PCV2.Pig farms should improve the environment and enhance the elimination of mosquitoes.