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In recent years, with the rapid expansion of China's pet population, pet disease is a serious threat to the health of the group, causing huge losses to the industry. Canine parvovirus as a deadly infectious diseases, usually 50% 1 00% incidence rate, mortality rate 10% 5 0% is the dog populations hazardous largest one of the diseases. Disease pathogen of canine parvovirus (CPV) in the past three decades, CPV rapid evolution, DNA virus is relatively rare. Commercial vaccines currently in use protective decline, often appear immune failure Therefore, this study from 2010 to 2011, Beijing canine parvovirus molecular epidemiological investigation and analysis of genetic differences, future research CPV antigenic variation and the development of more effective vaccines has a very important significance. The established PCR method for the rapid detection of CPV, the lower detection limit of 38.4 pg / mL of virus DNA. 2010 to 2011 were collected randomly from different parts of the 327 clinical samples (faeces, urine, saliva, oral and nasal secretions, serum, etc.), the application of the established PCR detection method, filter out 51 CPV-positive samples, the prosecution the rate was 15.6%, colloidal gold immunochromatographic assay positive rate of 12.2% has a higher sensitivity of, visible PCR method CPV. VP2 as CPV nucleocapsid protein coding genes, it contains all of the structural proteins and antigenic sites, so choose the VP2 gene of canine parvovirus molecular epidemiology of target genes. Cloning and sequence analysis of the VP2 gene from 51 positive samples and found that the nucleotide sequence of the sample and the reference strains, the amino acid sequence homology of 98.2% to 100% and 97.7% to 100%. Sample between the nucleotide and amino acid homology was 99.0% to 100% and 98.8 to 100%, indicating that the the VP2 gene mutation in the Beijing area since 2010 less. The subtype analysis show that the CPV genotype still is CPV-2a, accounting for 90.2% (46/51), CPV-2b, the low percentage of 9.8% (5/51) and found no CPV-2c and CPV-2. Compared with the classic CPV-2a / b reference strains epidemic strain VP2 gene point mutations distributed mainly include 555Ile → Val 324Tyr → Ile, Tyr at 267Phe → 440Thr → Ala, 101Thr → Ile 87Leu → Met and 139Val → Ile, individual samples were in 187 (Pro → Ile, Gln), 188 (Ala → Gln), 308 (Val → Ile) bit replacement. The DNAStar Analysis addition to Ile 101, Ile 139, Ile 187, Gln 187, Gln 188, Tyr 267 mutation of little significance, Met 87, Ile 308, Ile 324, Ala 440, Val 555 are in the the potential VP2 protein antigen epitope region, hence the variance has a very important biological significance. Phylogenetic tree analysis results showed that 51 cases of samples in the CPV VP2 sequence branch, not with FPV in the same branch. CPV branch, a closer relationship with Taiwan, Beijing, Wuhan, Nanjing isolates, far isolates kinship with Brazil, France, Japan and Vietnam, the United States, Italy, Poland. Accordance with the amino acid homology analysis, the homology of 100% of the samples normalized to a group of 51 samples can be divided into six groups, each group representative of A1, B4, B6, B8, B25, E4 of 6 parts of a stool sample by synchronous culture infected the fetal cat kidney cells F81 results successfully isolated the five virus strains isolated virus morphology was observed PCR detection, IFA test and the TCID 50 detection, confirmed the separation of toxic strains for the CPV. The separation of five strains were named as CPV-BJ-A1 the CPV-BJ-B4 the CPV-BJ-B8, CPV-BJ-B25 and CPV-BJ-E4. VP2 full-length gene sequencing analysis showed that the five isolates were CPV-2a subtype, and the current widespread use of commercially available vaccine strains for the CPV-2 subtypes, suggesting that the pandemic strain vaccine strain differences may be caused by vaccine immunocompromised one of the reasons. In summary, the analysis of the reasons for the failure to investigate the clinical immunology and the development of targeted CPV vaccine for pets has laid a theoretical foundation for the past two years, Beijing, the pet canine parvovirus disease molecular epidemiology and genetic differences.
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