Genetic Rescuing And Efficacy Evaluation Of Chimeric Classical Swine Fever Virus C Strain Containing E2 Hypervariable Antigenic Region 1 Representing The Prevalent Isolates

Classical swine fever(CSF),one of the most important infectious diseases of domestic pigs,is caused by classical swine fever virus(CSFV)and listed as a notifiable infectious disease by World Organization for Animal Health.The primary method to control classical swine fever in China is to vaccinate pigs with lapinized vaccine C strain.It has been reported that vaccine C strain belonging to genotype 1 has limited protection against field viruses of genotype 2 prevailent in recent years.Therefore,in order to offer better protection against new epidemic strains,this research used reverse genetics manipulation to construct a C-strain-based chimeric recombinant virus carrying a hypervariable antigenic region of the E2 gene from a field CSFV strain(fAR1).The main objectives were:1)to obtain a C-strain based chimeric virus containing fAR1(RecC-fAR1);2)to evaluate efficacy of the recombinant strain against the 2.1 d epidemic strain via immunization and challenge experiment;and 3)to compare serum neutralizing titers of pigs inoculated with RecC-fAR1 and the vaccine C strain.1.Identification of C-strain-based Chimeric Recombinant Virus Carrying E2 Hypervariable Region of a Field StrainThe hypervariable antigenic region 1(AR1)on QZ14 E2 was inserted into pA-RecC,an infectious clone of C strain constructed earlier in our lab and the recombinant virus RecC-fAR1 carrying QZ14 AR1 was successfully rescued.RT-PCR and IFA essays proved that the QZ14 AR1 region was successfully integrated into the viral genome,and that the recombinant virus could stably express the E2 protein carrying QZ14 AR1 in infected cells.This recombinant virus provided experimental materials for subsequent immunization and challenge experiments and serum neutralization tests.2.Efficacy of the Chimeric Strain RecC-fAR1 Against a Genotype 2.1d Field StrainIn the immunization and challenge experiment,how many?pigs were divided into three groups(?per group)and intramuscularily injected with RecC-fAR1,C strain,and DMEM respectively.On day 28 post immunization,pigs of each group was challenged with the field CSFV strain QZ14.,The pigs inoculated with RecC-fAR1 were in good condition without apparent symptoms and pathological lesions.Both RecC-fAR1 and C strains induced high levels of antibodies against CSFV E2,and RecC-fAR1 could produce higher levels to E2 of the field strain QZ14 than the C-strain.The pigs immunized with either RecC-fAR1 or C strain could resist challenge infection by the QZ14 strain.The experiment proves that the recombinant strain RecC-fAR1 can induce higher antibody responses to E2 of a field strain,and offers effective protection against field virus infection.3.Comparison of Neutralizing Antibodies of Recombinant Strain RecC-fAR1 and C Strain.Earlier studies in this lab showed that mutations in the AR1 region of the field strains will affect the neutralizing efficacy of the C strain immune sera.This study compared the differences of ELISA titers and neutralizing efficacy between RecC-fAR1 and C strains.The results showed that the ELISA and neutralizing titers of RecC-fAR1 immunized sera against the field strain QZ14 were significantly higher than those of C-strain immunized ones,while the ELISA and neutralizing titer against Shimen strain were not significantly different between the two serum sources.In summary,this study provides a candidate vaccine strain RecC-fAR1 that has the potential to provide better protection than its parental C-strain against CSFV of genotype 2 strains in the field.This study’s practical results are of reference value and practical significance for the control of classical swine fever virus of genotype 2.