Isolation And Identification Of Porcine Reproductive And Respiratory Syndrome Virus And Sequencing And Analysis Of Genomewide

Porcine reproductive and respiratory syndrome (PRRS), caused by porcine reproductive and respiratory syndrome virus (PRRSV), is a serious infectious disease of swine. It is characterized by severe reproductive failure in sows, and respiratory distress in piglets and growing pigs. Porcine reproductive and respiratory syndrome virus (PRRSV), the causative agent of PRRS, is a small-enveloped virus containing a single positive-stranded RNA genome. And the virus is genetically, antigenically, and pathogenically heterogenic. Substantial sequence divergence exists between the North American and European genotypes of the virus, showing only about 70 % nucleotide sequence identity. The typify strain are respectively VR-2332 and LV. PRRSV has highly aberration rate as same categories virus, exsting multilevel difference of gene between strain, especiallyEuropean strain. It indicates that there have many disparities by analyzing the virulence, gene sequence and antigenicity between isolated strains.Two doubted PRRSV strains were isolated and identified form visceral organs of piglets from the farms in Beijing and Guangdong. The cytopathogenic effect (CPE) observed on MARC-145 consists of the appearance of small rounded clumps of cellsraised above the remainder of deah from the monolayer gradually. After the isolate was inoculated on MARC-145 for several generations, the result of its TCID₅₀ test were 10^-5.48/0.1mL and 10^-5.71/0.1mL. The diameter of virus particle was about 60nm. The isolated virus was spherical and have envelope. Virus neutralization test showed that the isolate was neutralized by the serum to American genotype of PRRSV, not by the sera agaist PRV, HCV, PPV, CSFV respectively. According to the above results, the isolated virus was identificated as PRRSV and named PRRSV BJ strain and GD strain.Nine gene fragments of BJ strain and GD strain of procine reproductive and respiratory syndrome virus were amplified by RT-PCR and cloned into the PGEM-T-easy plasmid vector respectively and sequenced. Complete nucleotide sequence of genome of PRRSV BJ and GD were obtained by spicing sequence of each fragment in order (GenBank: EU825723, EU825724). The sequences of them were compared with that of other PRRSV strains in GenBank and draw the phylogenetic tree based on deduced amino acids of the PRRSV by bio-software. We try to analyses the genetic relationships and rule of inheritance between the different strains.It's useful for the epidemiological study and research of pathogenic mechanism of PRRSV.