| In summer 2006, a unknown severe disease designated'high fever'occurred in several pig farms and subsequently overwhelmed almost half of China, including Jiangsu, Hebei, and Jiangxi provinces. The typical clinical symptoms showed in diseased pigs were rubefaction, blood spots on mouth, back and the inner thigh, high fever, disorder in the respiratory tract and nervous system. And this disease caused substantial economic losses in domestic pig industry in 2006 with more than 2,000,000 infected cases in a very short time.Based on the gene sequences published in GenBank, five pairs of primers were designed here to identify the pathogen cuased'high fever'. Using PCR/RT-PCR analysis, 22 strains of PRRSV were obtained. The representative PRRSV isolated from the lymph tissues of sick piglets with apparent respiratory and nervous system syndrome, in Shuyang county (named as SHY-1 strain), was characterized both biologically and phylogenicly. The sequence analysis revealed that SHY-1 was an American-origin strain, and artificial infection showed that this isolate can cause typical clinical syndromes in piglets.According to the published genome sequence in GenBank, specific primers were designed to amply the ORF5 gene and hypervariable region in Nsp2. With these primers reverse transcription PCR was then performed with all PRRSV isolated in Jiangsu province during 2006-2007, followed by the sequencing of amplified fragments. The results revealed that the ORF5 gene of all 22 isolates exhibited the same length (603 bp), and except strain SIY-1, the other 21 viruses showed 98.5%-100% identity with each other, 87.7%-97% with other domestic isolates and 85.6%-90.5% with overseas isolates. However, the strain SIY-1 showed identity of 88.1%-89.1% with the other 21 viruses and 99.5% and 88.6%-98.8% with American reference strain VR-2332 and domestic isolates, repectively.22 PRRSV isolates only exhibited an identity of 54.1-56.6% with European strain. Nineteen isolates, used for amplification of their Nsp2 gene, exhibited 3 nucleotides deletion at 2778-2780 nt (coding for phenylalanine) and 87 nucleotides deletion in 2947-3033 nt (encoding 27 amino acids). Among them, 2 strains isolated in Yangzhou both showed a deletion at the position 2747-2836 nt (coding for 30 amino acids) compared with other 17 isolates. There was an 96.8-99.4% identity among these 19 isolates, however, 77.3%-78.6% identity with VR-2332 strain and 96.8%-99.4% with strains isolated from Hebei and Zhengzhou in 2007 were detected.Besides, 13 pairs of primers were designed and synthesized to obtain the full-length genomic sequences of strains SHY-1 and JSDT-1, according to the genomic sequence of China strain published in GenBank. SHY-1 was identified by sequencing as the American-type PRRSV, with 15323 nts in genome full-length which codes 8 open reading frames (ORF). Compared with the reference strain VR-2332, the isolate showed a deletion of 3 nucleotides (coding for phenylalanine) in 2778-2780 nt and 87 nucleotides (encoding 27 amino acids) in 2947-3033 nt, similar to the strain JSDT-1 isolated from Zhenjiang, another American-origin strain with 98.8% homology with SHY-1 in the genomic sequence. Homology of these two strains with VR-2332 were 89.3% and 88.7%. Respectively, according to the phylogenetic analysis, the homology of the two strains with isolates from Henan were 89.0%-99.4%, and 89.0%-92.0% from Korean, American isolates. The homology of the two strains and European isolates were 59.8% to 60.4%.
|
PDF Full Text Request