猪伪狂犬病病毒变异毒株HeNZK-2014的分离鉴定及gE基因序列分析

doi:10.16431/j.cnki.1671-7236.2017.07.026

Abstract

Abstract:

In order to learn the situation of pig pseudorabies virus (PRV) variant in this study, tissue samples such as lymph nodes which were collected from clinical pigs with suspected PRV infection were identified by PCR. PRV positive sample were inoculated on PK-15 cells after grinding and degerming, with further experiment including virus isolation, plague purification, PCR and IFA identification, TCID₅₀ confirmed by Reed-Muench method, inoculation test and observation of clinical symptoms in mice. The gE gene of purified PRV and brain tissue samples of dead mice were identified by sequencing analysis. The results showed that the virus grown on PK-15 cells could produce typical cytopathic effect (CPE) after 24 h; After three rounds of plaque purification,the isolate was PRV positive identified by PCR and IFA, and nominated as HeNZK-2014; The TCID₅₀ of the isolate was 10^-9.77/0.1 mL; The virus in 1×10⁸ TCID₅₀ inoculation was able to cause itching, tearing, death in infected mice, and PRV could be detected in tissues of dead mice; The molecular genetic variation analysis of gE gene by PCR amplification and clone sequencing indicated that the gE gene from brain tissue of infected mice shared 100.0% homology with HeNZK-2014, and located in a relatively independent branch with newly pandemic isolates in recent years after 2011, but was far from the classical strains before 2011, and both had two insertion of aspartic acid (D) at sites of 48 and 496 amino acids, which were considered to be the typical characteristics of PRV variants. This study successfully isolated a PRV variant, which laid a foundation for further research on vaccine development, prevention and control against PRV variants.

Key words: pig; pseudorabies virus (PRV); variant; isolation; identification; gE gene; genetic variation analysis

CLC Number:

S852.65

MA Zhen-yuan, LI Ning, YAN Ruo-qian, BAN Fu-guo, WANG Shu-juan, ZHAO Xue-li, XIE Cai-hua, WANG Dong-fang, WANG Hua-jun, CAO Wei-wei. Isolation and Identification of Variant HeNZK-2014 of Pseudorabies Virus and Sequence Analysis of gE Gene[J]. , 2017, 44(7): 2086-2095.

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Isolation and Identification of Variant HeNZK-2014 of Pseudorabies Virus and Sequence Analysis of gE Gene

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