Isolation And Identification Of Porcine Circovirus Type2and Challenge Of Commercial Pigs With PCV2

Porcine circovirus type2(PCV2) is the primary causative agent of associated diseases, they mainly include postweaing multisystemic wasting syndrome (PMWS), porcine dermatitis and nephropathy syndrome (PDNS), porcine respiratory disease complex (PRDC) and congenital tremor syndrome. PCV2is widespread both at home and abroad. The infection of PCV2may reduce the immunological ability of pigs, leading to the secondary infection of other pathogens. Diseases associated with PCV2could cause great economic loss in worldwide pig industry and handicape the progress of pig industry. PCY2infection was first reported in2000by Hongwu Lang in China, then PCV2infection was reported in Zhejiang, Beijing, Sichuan, Heilongjiang and so on.PMWS can threaten the health of pigs seriously which is caused by PCV2. The most common clinical signs are dyspnoea, unthrifty wasting, diarrhea, pallor, enlarged lymphnodes, and jaundice. PCV2has been one of the most important pathogens for swine recently, and being widely spreaded in the swine herds.The purpose of this study was to isolate and identify PCV2stains in eastern China, and provide effective information for prevention and treatment of related diseases. On the other hand, in this study,5-week-old commercial pigs were inoculated intranasally with PCV2, and immunized keyhole limpet hemocyanin (KLH) emulsifid in incomplete Freunds adjuvant to investigate the effect of immunostimulation on the induction of PMWS and reveale the pathogenic mechanism of PCV2.1Isolation and identification of porcine circovirus type2strainsThe lumphnode and lung tissues of pigs from slaughter house in Yangzhou were detected by PCR test. At the same time, porcine circovirus type2was isolated and identified from PCV2genomic-positive samples, consequently,5PCV2isolates were obtained, and their whole genomes were sequenced.Sequencing results showed that the PCV2isolates were in two the genotypes,4were PCV2group1, and the other was PCV2group2. Phylogenetic analysis revealed that the isolated strains are closed to each other, nucleotide homogenecity of complete genome ranged from95.0%-99.4%. Strain50belonged to PCV2group1B subtype, strain5847, stain L4and strain39belonged to PCV2group1C subtype, the three strains were more closed to the PCV2strains isolated from China, strain SD2belonged to PCV2group2E subtype, and it shared96.8-99%homology with other2E strains. Strain SD2was also near the PCV2group2D, refer to the phylogenetic tree. The results demonstrated that, PCV2group land PCV2group2are prevalent in China. These achievements will help to understand the epidemiology and control of PCV2infection, and lay the foundation of genetics and evolution of PCV2strains.2Challenge of commercial pigs with PCV2In this study, twelve5-week-old, PCV2sero-negative commercial piglets were assigned to four groups each with three pigs, group1was negative control, group2was KLH inoculation control, group3pigs were challenged with PCV2, group4pigs were challenged with PCV2and KLH. Group3and4were inoculated intranasally with PCV2. At the same time, the piglets were immunized with2mg KLH emulsified in incomplete Freunds adjuvant in group2and4, and also were given an intraperitoneal injection of10.0ml of thioglycollate broth (glycan) to stimulate peritoneal macrophage exudation into the peritoneal cavity. The study was terminated35days after inoculation.The results indicated that no obviously differences in clinical signs. PCV2-inoculated piglets seroconverted to PCV2within7days after inoculation. By quantitative polymerase chain reaction (Q-PCR), viral DNA load in samples was assessed. It demonstrated that lungs and lymphoid tissues of pigs in group3and4contained abundant PCV2antigen. Meanwhile the pathological lesions presented in pigs of group3were milder than those in group4. Granulomatous inflammatory reaction and lymphoid depletion that are typical lesions in pigs with PMWS were observed in the lymph nodes of piglets in group4. In the piglets of group4, more extensive replication of PCV2was documented by immunocytochemistry than that in group3. Thus, immune activation is an important contributor of the pathogenesis of PMWS in swine.