Isolation And Identification Of A Mutant Pseudorabies Virus Strain And Analysis Of Its Antigenic Differences

Pseudorabies(PR)is an acute infectious disease caused by pseudorabies virus(Pseudorabies Virus,PRV)with clinical feature of porcine nervous system disorders,respiratory diseases,reproductive disorders and fever.Porcine pseudorabies mainly affects sows and suckling piglets.Which give rise to reproductive disorders,suckling piglets with neurological symptoms,vomiting and diarrhea.At the end of 2011,a new epidemic of porcine pseudorabies which cause death byneurological symptoms,was emerged in Tianjin within the growth and fattening pigs that have been immunized with classic vaccines.So far,the new outbreak of the new porine pseudorabies has become a threat to the pig industry in China again.Knowing the new epidemic situation of porcine pseudorabies,understanding the genetic evolution and gene mutation status of the new porcine pseudorabies virus strain is helpful to guide the prevention and purification of porcine pseudorabies.In 2018,a suspected porcine pseudorabies outbreaked in a pig farm in Fuzhou,Fujian.The diseased pig organs were taken for nucleic acid detection and confirmed the wild-type PRV infection.In order to analyze this virus virulence,this study used a mouse regression test and susceptible cells inoculation and indirect immunofluorescence.As a result,we identified an epidemic PRV,named PRV-FJFZ strain,and this strain was further subjected to virulence determination and g E,TK,g B and g C gene amplification following sequencing analysis.In the end,our conclusion as follows.(1)The homology analysis of the nucleotide sequence and deduced amino acid sequence of PRV-FJFZ strain shows that it belongs to a newly popular gene type II PRV strains,which have at least 3 amino acid insertion,deletion or substitutionin the g E,TK,g B and g C genes.(2)With g E,TK,g B and g C gene sequence analysis,the PRV-FJFZ strain and other PRV strains found after 2011 in various domestic regions belong to the same independent branch of genotype II,which more closely related to isolates such as JS-2012 and ZJ01.(3)The virulence test of PRV-FJFZ strain showed that its TCID₅₀=10^-7.2/0.1m L,LD₅₀=10^-3.56/0.1m L,which means PRV-FJFZ strain is highly infectious to susceptible cells.In order to further analyzing the virulence of the PRV-FJFZ strain,we tested the sera cross immune protectionof wild and classic vaccinesbased on the former research of PRV-GD0304 strain which representing the new PRV genotype,and its TK g E and g I gene had been deleted,in our laboratory.(4)According to the analysis of the serotype identification criteria,the correlation coefficient R>70%of the cross-reactivitydegree between PRV-GD1406and PRV-Bartha K61 indicates that the new wild strain and the classic strain are in the same serotype.(5)The traditional strain PRV-Bartha K61 strain vaccine is still effective and has a medium affinity to the mutant epidemic strains.As a result,when using the PRV-Bartha vaccine,we suggesting a high antigen amount,and adjust the immunization program to ensure that the pig herd has a high Antibody levels.(6)PRV-GD0304 can inducing high level of antibody after vaccination,which has a high cross-neutralizing capacity to Bartha K61 strain,suggesting PRV-GD1406strain can expected to a good candidate vaccine for preventing mutant pseudorabies and expand the scope of application.(7)We used different sera(porcine anti-PRV-GD0304,porcine anti-PRV-Bartha K61)to neutralize the isolate PRV-FJFZ,and the results showed that the neutralization effect of porcine anti-PRV-GD0304 serum to the PRV-FJFZ strain is higher than that porcine anti-PRV-Bartha K61 in neutralizing the PRV-FJFZ strain,indicating that the PRV-GD0304 strain has a high affinity to PRV-FJFZ strain and their have a closer antigenicity.