Isolation And Identification Of Pseudorabies Virus In Fox,Racoon Dog And Mink And Expression Of GC Gene

Pseudorabies(PR)characterized by fever,unrelieved itchiness,respiratory system and reproductive system illness is a serious animal infectious disease causing great economic losses to the breeding industry.Since late 2011,a PR-like disease has occurred among fox,raccoon dog and mink populations on many farms in Liaoning,Hebei and Shandong provinces in China.To investigate the causative agent and its biological characteristics,several PRV strains like Fox1,Rac1 and Mink1 were identified.The herpesvirus like cytopathogenic effect was observed after inoculating the isolates onto MDCK cells.The mature virions with a diameter of 100 ~180 nm were observed by transmission electron microscopy.Typical sympotoms of PR was reproduced in rabbits and mice.The biological characteristics of Fox1,Rac1 and Mink1 were studied,and they were sensitive to chloroform as well as trypsin,and remained alive from pH 5 to pH 9 and showed tolerance to hot.Phylogenetic analyses based on gC and gE genes revealed nucleotide identity with recently reported Chinese isolates in pigs.3 insertions at position 48 and 497 of gE and 63~69 of gC,as a symbol of PRV variants were found in Rac1.It indicated that these PRV strains relating to fox,raccoon dog and mink PR were not vaccine strains like Bartha or early Chinese strains like Ea.However,the new variants have never been detected in foxes,raccoon dogs or minks before.gC gene of Rac1 was amplified and cloned into pET-32 a and pGEX-6p-1,respectivly.After induced by IPTG,the fusion proteins were highly expressed in BL21(DE3).The results indicated that the fusion protein with His-tag was about 70 kDa and about 76 kDa with GST-tag.It provided specific antigens for new diagnose and treatment methods.