Isolation, Identification Of PRRSV And The Influence Of PRRSV Infection On The Expression Of Pr Pc In Marc-145 Cells

Porcine reproductive and respiratory syndrome(PRRS) can causes a serious breeding sows and piglets topregnancy of respiratory symptoms, and a high mortality rate of infectious disease characterized by iscomposed of porcine reproductive and respiratory syndrome virus(PRRSV). Which has been almostepidemiced all over the countries and regions after firstly report it in the United States since 1987, andbecomed a danger infectious disease of pigs and swine industry. PRRSV have a high variability like otherRNA viruses with posterior capsule, gene sequence of PRRSV has different degree of moduation compaireddifferent the strains, espcialli, type I and type II of PRRSV. In this sudy, we isolated the lungs and blood ofswines which have high fever and respiratory syndrome from a pig form of Qinghai province. Then isolatingthe virus using Marc-145 cells and identifying RT-PCR, immunofluorescence and electron microscopic thevirus. the results indicateded that isolated viruses is a Amerian PRRSV strain,named QH08. We designed 7pairs specific primers to amplify full-length genome of the QH08, and spliced the 7 fragments, results showedthat full-lengh of QH08 genome is 15320 bp. The genomic nucleotide homology was 98% compare to JXA1,the represent strain of highly pathogenic PRRSV(HP–PRRSV), and QH08 have two discontinuous deletion of90 nucleotides in NSP2 area, so QH08 PRRSV strain belong to America HP-PRRSV type.For study of adaptability, proliferation virulence of the PRRSV QH08 strain on Marc-145, this studycompaired the different characteristic of QH08 strain, GW-HR strain, CH-1R strains of Heli and NEVBAcompany, GW-HR strain in Marc-145 cells by comparing the TCID50 and the time of CPE turns up. Theseresults indicated that:(1) the proliferation, adaptability and virulence of different PRRSV increased from 1 to10 generation, and virulence of QH08 strain is the strongest than other 3 strains.(2) overrall, the time of CPEof four PRRSV becomed short from 1 to 10 generations proved that virulence and replication of PRRSVstrains increased with generations in Marc- 145 cells.At the same time, we studied the influence of PRRSV infection on the expression of cellular prion proteinin Marc-145 cells. Cellular prion protein(PrPC) represents a potential regulator of cellular immunity and hasbeen confirmed to be an invasive receptor exploited by bacteria such as Brucella abortus to facilitate theinfection in host cells. In this study, Marc-145 cells were incubated by PRRSV QH08 strain, we investagatedthe expression profile of PrPC from the protein level and mRNA level by using flow cytometry and QPCRassay. The result is that PrPC is expressed on the surfaces of Marc-145 cells and its expression profiles greatlychanged after PRRSV incubation with these cells. PrPC expression reached the peak at 12 hour after PRRSVinoculation and then decreased sharply to recover to the original status while CD163, a PRRSV receptor inMarc-145 cells, achieved the climax at 48 hour after virus infection and then slowly lowered its expressionlevel. The study was first time to relate PrPC and PRRSV together, and researched their relationship; There is afirst report that PrPC may be play a role in virus infection the host cells, which provided a new sight for PrPCphysiology in virus infection and a new theoretical basis for prevention and control of PRRSV.