Isolation And Identification Of Porcine Circovirus Type2in Partial Jiangsu Area Of China And Studies On Its Pathogenicity

PCV2has been one of the most studied pig pathogens since1997, now it represents the second most important pig viral pathogen after PRRSV and spreads worldwide. Since2000it was reported in China, the morbidity and mortality caused by PCV2have been high. In order to monitor the genetic variation of PCV2in pigs, clinical samples were collected in partial Jiangsu area from2012to2013. Using PCR, the nucleic acid fragments of PCV2, PRV, PPV, CSFV and PRRSV were detected from collected clinical samples. The PCV2positive sample tissues were homogenized and inoculated to the Dulac cells that were not contaminated by PCVs. We analyzed the genetic evolution of PCV2isolates and compared differences between PCV2b isolates when infect Dulac cells and commercial pigs, respectively. This study provides a data for epidemiological investigation and pathogenicity of PCV2in China.1. Epidemiological investigation of PCV2and other common viruses in partial Jiangsu area during2012and2013164suspicious samples from cases of swine collected from2012to2013in partial Jiangsu area of China were detected for the common viruses by PCR. The results showed that70cases were PCV2positive, with the positive rate42.7%;29cases were CSFV positive, with the positive rate17.7%;16cases were PRV positive, with the positive rate9.8%;12cases were PPV positive, with the positive rate7.3%;4cases were PRRSV positive, with the positive rate2.4%. The positive rate of PCV2/CSFV co-infection rate was5.4%(9of164); PCV2/PRV mixed infection rate was4.8%(8of164); PCV2/PPV mixed infection rate was2.4%.2. Genetic analysis of PCV2isolates in partial Jiangsu areaTo investigate the genetic variation of PCV2in partial Jiangsu area of China, thirty-eight PCV2strains isolated from the infected pigs in above area were indentified by sequencing. The results showed that these PCV2isolates could be classified into two genotypes, PCV2b (11of cluster 1B,26of cluster1C) and PCV2a (1strain), with genome length of1767bp (37strains) and1768bp (1strain), respectively. A comparison of the deduced amino acid residues encoded by ORF2indicated that PCV2genotypes have their specific mutation sites. At genotype level, PCV2cluster1C was the dominant at above area between2012and2013.3. Studies on pathogenicity of PCV2b isolatesStrain121108(1B) and strain130405(1C) were selected to compare the pathogenic differences of PCV2b1B and1C subtype strains of our laboratory. To compare their growth on passaged cells, the Dulac cell line was chose to detect the TCID50and copies of PCV2DNA at day1,2,3,4,5post-inoculation. To determine differences in infection kinetics of the two PCV2strains in experimental animals,8pigs were randomly assigned to one of three groups:negative controls (n=2); pigs infected with PCV2b1B (121108; n=3), pigs infected with PCV2b1C (130405; n=3). Blood samples were collected at day post-inoculation (dpi)7,14and21and tested for the presence of PCV2antibody and copies of PCV2DNA.The results showed that when infected Dulac cells, the TCID50and PCV2DNA copy numbers of strain121108in the cell supernatant were10times higher than those of strain130405(P<0.01). In the animal infection assay, strain121108initially replicated at higher levels and both strains reached peak replication levels at dpi7. Pigs infected with121108strain (PCV2b1B) had significantly higher amounts of viral DNA in serum at both dpi14and dpi21, compared to pigs infected with130405(PCV2b1C)(P<0.05). By21days post-challenge, gross and microscopic lesions of lymph nodes and lungs in pigs of121108strain challenge group were significantly more severe than those found in pigs of130405strain challenge group. There were multinucleated giant cells, large number of eosinophilia and lymphocyte depletion in lymph nodes in pigs of121108strain challenge group. There were no multinucleated giant cells and less lymphocyte depletion in pigs of130405strain challenge group. The immunohistochemistry results also show that more PCV2antigen was detected in lymph nodes, thymus and tonsils in pigs of121108strain challenge group. In conclusion, results of this study showed that, the infection of PCV2b1B cluster strain (121108) was stronger than that of PCV2b1C cluster strain (130405).