Research On The PRRSV HuN4-F112 And Henan-A11-376 To Infect The MARC-145 Cells With Diferences

Porcine reproductive and respiratory syndrome(PRRS) is one of the devastating diseases affecting the swine industry worldwide. Since 2006, a highly pathogenic PRRS(HP-PRRS) which was characterized by high fever associated with high mobility and motality emerged in China and caused substaintial economic losses. Currently, the antenuated vaccine strains were obtained from the MARC-145 cells. In rescent years, PRRSV emerged many mutants which could not passage in the MARC-145 cells. Therefore, it is significant to research against these PRRSVs for scientific prevention and control of PRRS.The PRRSV HuN4-F112, high adaptive MARC-145 cells, while the PRRSV Henan-A11-376 could infect PAM cells but not MARC-145 cells. In the present study, six chimeric palasmids were constructed and five chimeric viruses, rHuN4-F112-TORF1 a, rHuN4-F112-TORF1 b, rHuN4-F112-TORF2-7, r376-TORF1 a and r376-TORF2-7, were successfully rescued by replaced gene regions(ORF1a, ORF1 b, and ORF2-7) each other based on the skeleton of the full length infectious cDNA clone of pHuN4-F112 and p376. The results showed that ORF1 b had a little effect on the infection of the two PRRSVs in MARC-145 cells, while ORF1 a and ORF2-7 play an important role to infect MARC-145 cells of the two PRRSVs.To determine the effect of non-structural proteins in ORF1 a gene region for the infection of the two PRRSVs in MARC-145 cells, six chimeric palasmids were constructed and three chimeric viruses, rHuN4-F112-TNSP1, rHuN4-F112-TNSP2 and rHuN4-F112-TNSP3-8, were successfully rescued by replaced gene regions(NSP1, NSP2, and NSP3-8) each other. The results indicated that any one of the three gene reigions(NSP1, NSP2, and NSP3-8) had no effect on infection of the two PRRSVs in MARC-145 cells.To determine the effect of the structural proteins region(ORF2-7) for infection of the two PRRSVs in MARC-145 cells, four chimeric palasmids were constructed and two chimeric viruses, rHuN4-F112-TORF5-7and r376-TORF2-4, were successfully rescued by replaced gene regions(ORF2-4 and ORF5-7) each other. The results showed that ORF5-7 gene region had no effect on the infection of the two PRRSVs in MARC-145 cells, while ORF2-4 played a key role for the infection of the two PRRSVs in MARC-145 cells.To determine the effect of minor structural proteins region(ORF2-4) for the two PRRSVs infection in MARC-145 cells, six chimeric palasmids were constructed and one chimeric viruse, rHuN4-F112-TORF4, was successfully rescued by replaced gene regions(coded ectodomain of GP2 a, ectodomain of GP3 and GP4 protein) each other. The results indicated that any one of the two regions(coded ectodomain of GP2 a and ectodomain of GP3) had an effect on the infection of rHu N4-F112 in MARC-145 cells but not r376. However, the region coded GP4 protein may have no effect on the infection of the two PRRSVs in MARC-145 cells.To determine the effect of any two regions in minor structural proteins region(ORF2-4) for the infection of the two PRRSVs in MARC-145 cells, six chimeric palasmids were constructed and four chimeric viruses, rHuN4-F112-TORF2a-ORF3、rHuN4-F112-TORF3-ORF4、r376- TORF2a-ORF3 and r376- TORF2a-ORF4 were successfully rescued by replaced gene regions(encoded GP2 a and ectodomain of GP3, ectodomain of GP2 a and the whole GP4 protein, and the whole GP3 and GP4 protein) each other. The results showed that the gene regions encoded GP2 a and ectodomain of GP3, and the whole GP3 and GP4 protein played an important role for infection of the two PRRSVs in MARC-145 cells.In the present study, the results indicated that ORF1 a and ORF2-4 played an important role for infecting MARC-145 cells of the two PRRSVs. Moreover, the gene regions, encoded both GP2 a and ectodomain of GP3 and the whole GP3 and GP4 protein, were the most important gene regions for infection of the two PRRSVs in MARC-145 cells. This study provided new method and idea for vaccine acclimation of those PRRSVs that could not infect MARC-145 cells.