| Porcine reproductive and respiratory syndrome(PRRS)is a highly infectious respiratory disease in pigs caused by porcine reproductive and respiratory syndrome virus(PRRSV), it is recognized as a serious swine disease and is characterized with either reproductive failure in pregnant sows and respiratory tract distress. Initially outbreak and described in the USA in 1987, erupts popularly in our country most districts in 2006, brings the enormous economic loss for our country's pig-breeding industry, in order to clarify the PRRSV popular poisonous origin and the variation reason, explores the related prevention and control measure. This research isolated and identified the strains of PRRSV and samples were collected from two farms in Anhui province with suspected clinical signs of porcine reproductive and respiratory syndrome in 2008, clone and sequential analysis for its Nsp2,ORF7 gene; vaccination healthy pigs with isolate strains, carries on the histopathologic diagnosis and analyzes infection and the distributed situation of the PRRSV in the piglets in vivo with immunohistochemical method, for the Anhui Province PRRSV strains origin,the molecular epidemiology and the variation situation and so on provides the scientific basis, to further study the virus in the animal in vivo continually infection condition and expounded that PRRSV sends the pathogenesis to lay the rationale.Samples were collected from two farms in Anhui province with suspected clinical signs of porcine reproductive and respiratory syndrome in 2008, and observation specific cytopathogenic effect after 4-5 passages in Marc-145 cells. The isolated strains were amplified from infected cell culture by porcine reproductive and respiratory syndrome virus (Nsp2 1594～1680) RT-PCR and reacted specifically by indirect immunofluorescence assay and showed the virion is enveloped with the diameter of about 50-80nm by electronic microscopy. Our data supported that two porcine reproductive and respiratory syndrome viruses were isolated, which designated as HS08 and ZJ08 respectively.The Nsp2 and ORF7 genes of the isolates were amplified using RT-PCR which was followed with cloning and sequencing. The two isolates were similar to each other (nucleotide homology>99.0%), and they contained a discontinous 30-amino acid deletion in the Nsp2 gene. The two isolates had 80.7% and 80.2% nucleotide homology with strains CH-la, respectively, in the Nsp2 gene. They had 98.4% and 99.5% nucleotide homology with the highly pathogenic strain HuN isolated in 2007 in the ORF7 gene, and they had mutations of K46→R46, H109→Q109 and V117→A117 in the ORF7 gene. Phylogenetic analysis also revealed that the two isolates belonged to the cluster of highly pathogenic strains emerging in China in recent years.Six 60-day-old piglets with antigen and antibody were negative of PRRSV stochastical divided into 3 groups, each group of each two heads. Through the nasal vaccination cell cultures medium of isolate strains ZJ08 and HS08 and 2 mL/head dosage respectively in A and B groups, C group as the control groups. The body temperature in experimental group of piglets are rise to 41℃after vaccinate the first day, and continue the high fever, is highest when may reach 42.3℃, it can detect PRRSV antibodies in the 6th day, and antibody levels achieves high in the 10th day. Discovery to various internal organs of the experimental group pigs lesions lightly than the clinical observation through observation of the clinical symptoms, the pathological change, the pathological section. the lung mainly assumes interstitial pneumonia, lung congestion, interalveolar septum thickening, considerable inflammatory cell infiltrate; Hepatic congestion, hepatic cell degeneration, inflammatory cell infiltrate; kidney,spleen,lymph node hemorrhage; Identification lung, liver, kidney, spleen, lymph node, tonsil were have varying degree positive reaction with immunohistochemical method, of which the strongest masculine in the lung. |
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