| Porcine reproductive and respiratory syndrome (PRRS), known as an acute and highly contacting infectious disease, that the height variation of which was the main control factor of the failure on the prevention and control. On PRRSV the two main protein (Nsp2/ORF5) were target genes for molecular epidemiological studies and variance genetic analysis, therefore important and meaningful to the overall research on PRRS mutation. This study had determined pathological samples from pigs in Guangxi during 2012 to 2015 by gene sequencing and analysis on ORF5 and Nsp2. And we will carry out the PRRSV antibody detection and analysis from different pigs. The purpose of this study was to comprehend the epidemic tendency and mutation in Guangxi in recent years popular trend and variation, that could provide certain theoretical data and theoretical basis and control and lay the foundation for the prevention and control of the disease in the future.In this study, Suspected sample infected with PRRSV were detected by RT-PCR, and 91 PRRS positive samples were detected (18.13% of total).18 fragments of Nsp2 and 48 ORF5 were obtained by gene sequencing. For Nsp2, the comparison results of nucleotide homologies and amino acid sequence showed identities of nucleotide and deduced amino acid were 51.9～99.1% and 30.4-98.7% with reference sequence available in GenBank, respectively. While compared to the previous PRRS sequences of collection from home and abroad, higher nucleotide homology between Guangxi isolates and JXA1 was obtained.The results showed that the 48 sequences of ORF5 gene displayed 61.8-99.8% nucleotide identity and 52.7～99.0% amino acid homologies with reference sequence available in GenBank. The Phylogeny based on genetic variation (mainlyORF5 then Nsp2) indicated that PRRSV strains collected from China could be classified into 3 subgroups of American genotype, most of them were subgroup IV represented by JXA1 strain, followed by subgroup III represented by HB-1(sh)/2002 strain.The amino acid analysis of Nsp2 showed that 15 of the 18 strains contained a 29+1 amino acid deletion which was considered as hallmarker of HP-PRRSV. In ORF5-coding GP5 protein, most of these virus strains were found mutated, abundant mutations that occurred within primary neutralizing epitope (PNE) on B cells, T cells and potential glycosylation site. Since 2012, the mutation rate of amino acid sites of 48 stains isolated in the study increased significantly compared to 89 strains on GenBank (NCBI) of GX isolates in the last decade. We found there were 4 amino acid sites mutate frequently, including R151K(18/48, the mutation rate was 37.5%), E170Q(14/48, the mutation rate was 29.17%), V185A(48/48, the mutation rate was 100%); Q196R/L(33/48, the mutation rate was 68.75%). And in the 89 reference strains, the mutation of the amino acid were R151K(2/89, the mutation rate was 2.24%); E170Q (9/89, the mutation rate was 10.11%); V185A(63/89, the mutation rate was 70.78%); Q196R/L(7/89, the mutation rate was 7.86%). Speculated that these would be a mutation trend about pandemic strain of Guangxi. And these mutations can lead to internal structure change of protein, the lack of a sugar base site may cause neutralizing antibody generating speed and quantity. Evolutionary tree analysis results, the closest genetic relationship was the three strains of plant GXBS120815, GXYL130617, GXYL130624 and the NCDA30 strain. The direction of mutation of amino acid site from 151,170,189,191,192, was consistent with that of NADC30 strain. Speculated that the like NADC-30 strain was prevalenting in Guangxi and may be derived from the same strain of NADC30 strain. And the direction of site mutation could be used as a reference index which Whether the reference strain and the NCDA30 strain were derived from the same strain.To investigate the immunity under immune pressure on farm animals and the virus evolution, serum antibody against IBRV (2228)and the viral pathogen were detected from medium-to large-scale or small-scale farm and slaughterhouse collected blood and tissue samples of spring and autumn during 2014 to 2015, by the RT-PCR. Classified according to the scale of swine breeding and source, The results of this study indicated that positive samples of PRRSV antibodies was 1698, positive rate was 76.21%, the ratio of S/P> 2.5 samples was 8.66%. In the breeding pig farm and more than 500 large-scale pig farm, the highest of positive rate was 92.5%, and the minimum ratio of S/P> 2.5 samples was 2.03%. The lowest of antibody qualified rate came from Scatter-fee, and positive rate was 51.31～68.42%, and the immune effect of Vaccine is the worst, Highest proportion of S/P> 2.5 samples was 16.28%. The results showed that the best and stablest immunologic effect was found in breeding pig farm and medium- to large-scale farm, rather than smaller scale, which was bad and unstable, the immune effec from Scatter-fee was worst and unstable. There would be a risk of wild virus infection.Pigs Classified according to age, sex, purposes, swine, the results of this study indicated that positive samples of PRRSV antibodies was 1648, positive rate was 73.97%, the percentage of S/P> 2.5 samples was 8.59%. The best immune effect was boar pigs, the positive rate was 83.33～94.44%, the value of S/P samples was 1.38～1.44, the highest proportion of S/P>2.5 samples was 2.32%. Positive of weaned pig antibody was 66.07～70.05%, and the lowest proportion S/P> 2.5 samples was 13.53%. Data analysis showed that the breeding herd was stable state, the antibody levels between different groups was uneven. The pigs were stable with a risk of wild strain virus infection.Analysis the PRRSV vaccine immune antibodies from different vaccine manufacturers, immune effect was significant difference, The positive rate of antibody from the bidding company were 71.81-82.32% and 73.15-88.55% during 2014 to 2015, and total positive rate was 79.27%. The firms antibody positive rate of 79.22%-89.91%,83.11-91.60%, and total positive rate was 83.17%. Antibody positive rate of weak poison vaccine was 85.4% and 87.23% from 2014 to 2015, antibody positive rate of inactivated seedlings was 73.33% and 76.68%. Immune effect was significant difference between different vaccines, PRRS weak poison vaccine immunization effect was superior to inactivated vaccines.Inspection of the 55 samples, The results of this study indicated that 11 ORF5 gene sequences was got by the RT-PCR techniques, According to identity and phylogenetic tree analysis,11 ORF5 sequences were amplified and sequenced for further analysis, with JXA1 strains the highest homology. Mainly concentrated in the Ⅳ subsets of HP-PRRSV with Represented by JXA1 strains, and a few distributed in the Ⅲ subgroup with the representative by HB 1 (sh) /2002 strains, which consistent with the PRRS trend in the whole Guangxi area. |
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