China Animal Husbandry and Veterinary Medicine ›› 2021, Vol. 48 ›› Issue (5): 1794-1803.doi: 10.16431/j.cnki.1671-7236.2021.05.031

• Basic Veterinary Medicine • Previous Articles     Next Articles

Isolation and Identification of Porcine Pseudorabies Virus and Genetic Variation Analysis of Partial Virulence Gene

XU Ning2, GE Guiyang2, LI Dongli2, LIU Yi2, GONG Qinglong2, MA Baoyi2, SHENG Chenyan2, SHI Kun1, LI Jianming1, LENG Xue1, DU Rui3   

  1. 1. College of Chinese Medicine Materials, Jilin Agricultural University, Changchun 130118, China;
    2. College of Animal Science and Technology, Jilin Agricultural University, Changchun 130118, China;
    3. Laboratory of Production and Product Application of Sika Deer of Jilin Province, Changchun 130118, China
  • Received:2020-11-29 Online:2021-05-20 Published:2021-05-20

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Abstract: To investigate the genetic variation of Porcine pseudorabies virus (PRV) in Jilin province,in this study,the virus isolation and identification were performed by cell passage,PCR and sequencing in clinical samples were collected from pigs with suspected porcine pseudorabies.The viruses titer was determined by TCID50,and the pathogenicity of the isolated strains to rabbits was determined by the rabbit infection test.Futhermore,PCR amplification and sequencing of TK,gB,gC,gD and gE genes were performed to analyze their genetic variation.The results showed that 3 samples of clinical samples had cytopathy after passage by PK15 cells.PCR identification and sequencing analysis showed that 3 strains of PRV were isolated and identified and named as JL03,JL12 and JL15 strain,respectively.The titers of the 3 strains were 106.5,106.5 and 107.5 TCID50/mL on PK15 cells.Six rabbits were inoculated with 3 isolated strains (103.5 TCID50 each rabbit),all showed symptoms such as elevated body temperature,itching at the injection site and paralysis of limbs,and all died 3 to 4 days after virus inoculation.The results of the deduced amino acid sequence analysis of the TK,gB,gC,gD and gE genes of the 3 isolated viruses showed that the TK gene had no amino acid variation compared with the reference strains.Amino acid deletions,insertions or mutations occurred in some positions of gB,gC,gD and gE genes.Among them,an aspartic acid insertion in the 48th and 496th positions of the gE gene were consistent with the variation characteristics of epidemic strains reported in China.Nucleotide genetic phylogenetic tree analysis showed that the TK,gB,gC,gD and gE genes of the 3 isolates were highly homologous with those of the reference strains JS-2012 and ZJ01 isolated after 2012 in China,and belong to the same branch.They were far from foreign strains NIA3 and Becker, and located in different branches,and the genetic relationship with the early domestic virus strains SC and Ea was between them.In short,we successfully isolated and identified three PRV variants which had strong pathogenicity to rabbits.These results provided new data for the epidemiological study of PRV in Jilin province and lay the foundation for relevant research.

Key words: Porcine pseudorabies virus; isolation and identification; phylogenetic analysis

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