| PCV2is the virus associated with postweaning multisystemic wasting syndrome(PMWS), porcine dermatitis and nephropathy syndrome (PDNS), porcine respiratorydisease complex (PRDC)，proliferative and necrotising pneumonia (PNP)，porcinerespiratory disease complex and so on. PCV2have a great impact on the swine industryaround the world,and caused serious damage. Currently, Wild boar have been developed,and were the utilization of resources in many parts of China. In order to understand theprevalence and biological characteristics of PCV2from domesticated wild boar in Anhuiprovince, PCV2Anhui strain from domesticated wild boar were isolated and identificated.Then it were sequenced and analyed the complete genome, comparing ORF1, ORF2sequence homology sex, and animal pathogenicity test,so as to clarify the characteristicsof PCV2Anhui strain from domesticated wild. These studies can provide the basis forthe study of porcine circovirus disease.1. The sample was inoculated into the PK15cells and passaged for eight generation.One PCV2strain was identificated by PCR, IFA and electron microscopy. The isolate ofPCV2was named as YZ0901. It is the first demonstration that PCV2infected thedomesticated wild boar in Anhui Province.2. The whole genomic sequences of YZ0901and the5PCV2Anhui isolates werecloning,sequencing and analysising.The results show, that full genomic sequences ofYZ0901was1767nt in length,and was belong the PCV2-1C. It has94.2-99.5%homologyin whole nucleotide sequence,96.7-100%in ORF1,89.6-89.3%in ORF2,with YZ0901and42strains of domestic and foreign. YZ0901has the existence of homologousrecombination.The ORF1nucleotide sequence of YZ0901have less variation.Thevariation was mainly concentrated in YZ0901ORF2.PCV2-1C gene cluster have changedin in the ORF2of the two main strains of the immune response zone. The genotype ofPCV2from domesticated wild boar were closely related to the genotypes of the localstrains.3. Based on the ORF2nucleotide sequence of porcine circovirus type2（PCV2）,twopairs of primers and a TaqMan probe were designed and synthesized. To the restructuringplasmid for standard product did standard curve, established TaqMan fluorescentQuantitative PCR for Porcine Circovirus Type2. TaqMan fluorescence quantitative PCRhas good repeatability, and sensitivity about6x101copy/uL. TaqMan fluorescencequantitative PCR had no cross reaction with classical swine fevervirus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV) and pseudorabies virus (PRV).Clinical applications showed this method was rapid and high sensitivity test. Thereal-timeTaqMan fluorescence quantitative PCR assay which is specific, sensitive andaccurate can be used for the diagnosis of PCV2infection. It can provide the technicalsupport for the research,such as PCV2tropism, cell culture characteristics and so on.4. In experimental animals, the antibodies, cytokines, and the organizations in the numberof virus particles have been detected,and the pathological changes have been observed.YZ0901has a lower level in the IFN-α, IFN-γ, IL-2, IL-10concentration, the antibodiesare more fluctuating. It increased more corpuscle splenic in splenic, have more theglomerular enlargement and the cell increased in kindy. It has the most virus particles inthe value (5.35×104copies/μL,5.995×104copies/μL) than the other groups. It showedwhen YZ0901strain infected mice, the cellular immune suppression have been produced atfirst,then the humoral immune suppression have been produced and have stronger tropismin the spleen, kidney than other groups.YZ0901and XC0901strains that were belong thePCV2b-1C have the poor immune response than PCV2b-1A(DY0801)、PCV2b-1B(BH0801). But the level of tropism in PCV2b-1C have stronger thanPCV2b-1A,and weaker than PCV2b-1B. |
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