Isolation And Identification Of PRRSV From Clinical Healthy Pigs

PRRS is one respiratory tract diseases to harm to pig farms, which characters are abortion, death of embryo, mummified fetuses, and respiratory problems in piglet. Pathogen of the disease was considered as porcine reproductive and respiratory syndrome virus (PRRSV). Since May 2006, PRRS has become one of the most economically important diseases in part of pig farms in Jiangxi, Hunan province. Kegong Tian reported that the reason of this deseas was highly pathogenic PRRSV which characters amino acid deletion in Nsp2 of 483 and 535-563. We found healthy pigs with mutant PRRSV in investigation of pig farms in 2007. In 2008 and 2009, there were healthy pigs with PRRSV in most pig farms. So some scholars thought that the Nsp2 deletion may not be the main reason of highly pathogenic PRRSV. We have a detection of mutant PRRSV in healthy pigs. These pigs are well in clinical, no one is died. So we sequenced the PRRSV which was isolated and analyzed if there were other mutation or not in PRRSV.This study included as following:1. Identification and separation of PRRSV: One hundred and fifty-seven samples were collected and tested with RT-PCR during the period from December 2008 to December 2009. The result is that one hundred and sixteen positive samples. Symptom of these field samples is that animal heat is 39.2℃-42.0℃. Majority cases appear rubefaction, lying and hypersomnia, most striking macroscopic lesions observed were lymph node hemorrhage or edema and diffuse interstitial pneumonia, intestine thin-walled, kidneys hemorrhage, spleen hemorrhage and pleuritis were observed. Six PRRSV strains were isolated from positive samples.2. According to the published genome sequence, designed primers, which amplified the gene of Nsp2, ORF4, ORF5, ORF6 and ORF7 of strain LY(One was separated from positive serum of healthy pig) and strain JX (another was separated form PRRS tipycal symptom deseased pig), the results shown that strain LY and JN were found 30 amino acid deletion in Nsp2 , the homology of nucleotide sequences and deduced amino acid sequences of gene among JXA1 was high, the homology of deduced amino acid sequences of ORF4, ORF5, ORF6, ORF7 gene between LY and JX was 92.5~100%, glycosylation site of ORF4, ORF5, ORF6 and ORF7 in strain LY fully consistent with strain JN. There are differences with JXA1 strain. Glycosylation site of ORF4 at 130 site is NFT in strain LY and JN which is NFS in strain JXA1. Glycosylation site of ORF5 at 34 site is NSS in strain LY and JN which is NNSS in strain JXA1.3. In order to study the immunity effect after immunized by classical swine fever vaccine in pigs with persistent infection of PRRS, the blood samples of finishing pigs, multiparous sow, replacement gilts and piglets in pig farm with PRRS were collected once a week. PRRSV were tested by RT-PCR, and the antibodies of PRRS and CSF were tested by ELISA after vaccination of CSFV vaccine. The results indicated that the detection rate of PRRSV was comparatively low, the level of PRRS antibody was high. The level of CSF antibody rose quickly after immunized by classical swine fever vaccine and the immunity effect was better. It did not product significant immunosuppression.