Recombinant VP1 protein of duck hepatitis virus 1 expressed in Pichia pastoris and its immunogenicity in ducks

Abstract

Summary. -The VP1 gene of duck hepatitis virus type 1 (DHV-1) strain VJ09 was amplified by reverse transcription PCR from the liver of a duckling with clinical symptoms of viral hepatitis. The resulting VP1 cDNA was 720 bp in length and encoded a 240-amino-acid protein. In VP1 gene-based phylogenetic analysis, the VJ09 strain grouped with DHV-1 genotype C. The VP1 gene was inserted into the expression vector pPICZαA and expressed in Pichia pastoris. The expressed VP1 protein was purified and identified by western blot analysis. To evaluate the recombinant VP1's immunogenic potential in ducklings, the antibodies raised in the immunized ducklings were titrated by ELISA, and lymphocyte proliferation and virus neutralization assays were performed. The results show that the recombinant VP1 protein induced a significant immune response in ducklings and this could be a candidate for the development of a subunit vaccine against DHV-1 genotype C.Keywords: duck hepatitis virus 1; genotype C; VP1 protein; Pichia pastoris; duck; immunogenicity * Corresponding authors. E-mail: wangchen2001@126.com, xiangchaoc@126.com; phone: +86-379-65507210. † Equal contribution. Abbreviations: AOX1 = alcohol oxidase 1; CAV-9 = coxsackievirus A9; DHV = duck hepatitis virus; DHV-1 = duck hepatitis virus type 1; FMDV = foot-and-mouth disease virus

Expression of duck hepatitis A virus type 1 VP3 protein mediated by avian adeno-associated virus and its immunogenicity in ducklings

“…The serological and challenge results observed in the ducklings vaccinated with rAAAV-VP3 in ovo indicated that the VP3 protein expressed in ducklings could induce sufficient immune protection for DHAV challenge. The VP1 protein is the major capsid protein and contains most of the motifs known to interact with cellular receptors and neutralizing antibodies (Oberste et al, 2003), and is the target gene for genetic engineering vaccine design (Wang et al, 2014;Fu et al, 2012). Our results show that the VP3 protein can serve as a candidate for the development of genetic engineering vaccine.…”

“…The VP1 protein has been sufficiently proved to be the major host-protective antigen containing T and B cell epitopes that induce protective neutralizing antibodies (Zhang et al, 2015;Wu et al, 2015;. Immunization of ducklings with the recombinant VP1 protein expressed via pichia pastoris (Wang et al, 2014), adeno virus (Yin et al, 2013), and suicidal DNA plasmid (Fu et al, 2012) has been shown to elicit strong protective immunity and provide enough protection against viral challenge (Wang et al, 2014;Fu et al, 2012;Yin et al, 2013). However, there are few studies about structural protein VP3 of DHAV.…”

“…Though the attenuated vaccine could induce strong immunity and provide enough protection for ducklings (Crighton and Woolcock, 1978), the probability of reversion to virulence still exists. To overcome these limitations, immunization of ducklings against DHAV with recombinant genetically engineered vaccines has already been reported; such as the suicidal DNA vaccine (Fu et al, 2012) and the recombinant Pichia pastoris subunit vaccine (Wang et al, 2014).…”