Abstract
Infectious bursal disease virus (IBDV) is a pathogen of worldwide significance to the poultry industry. Although the PDE and PFG domains of the capsid protein VP2 contribute significantly to virulence and fitness, the detailed molecular basis for the pathogenicity of IBDV is still not fully understood. Because residues 253 and 284 of VP2 are not the sole determinants of virulence, we hypothesized that other residues involved in virulence and fitness might exist in the PDE and PFG domains of VP2. To test this, five amino acid changes selected by sequence comparison of the PDE and PFG domains of VP2 were introduced individually using a reverse genetics system into the virulent strain (rGx-F9VP2). Then reverse mutations of the selected residues 249 and 256 were introduced individually into the attenuated strain (rGt). Seven modified viruses were generated and evaluated in vitro (CEF cells) and in vivo (SPF chicken). For residue 249, Q249R could elevate in vitro and reduce in vivo the replication of rGx-F9VP2 while R249Q could reduce in vitro and elevate in vivo the replication of rGt; meanwhile Q249R reduced the virulence of rGx-F9VP2 while R249Q increased the virulence of rGt, which indicated that residue 249 significantly contributed to the replication and virulence of IBDV. For residue 256, I256V could elevate in vitro and reduce in vivo the replication of rGx-F9VP2 while V256I could reduce in vitro but didn’t change in vivo the replication of rGt; although V256I didn’t increase the virulence of rGt, I256V obviously reduced the virulence of virulent IBDV. The present results demonstrate for the first time, to different extent, residues 249 and 256 of VP2 are involved in the replication efficiency and virulence of IBDV; this is not only beneficial to further understanding of pathogenic mechanism but also to the design of newly tailored vaccines against IBDV.
Highlights
Infectious bursal disease (IBD) is a highly contagious immunosuppressive disease in chickens that has caused significant losses to the commercial poultry industry worldwide [1,2]
Rescue of Modified infectious bursal disease virus (IBDV) To test the function of residues 242, 249, 256, 270, and 279 in the PDE and PFG domains of VP2, a set of mutated infectious clones with the backbone of the virulent strain were
Through identification with immunofluorescence assays (IFAs), electron microscopy assay, and sequencing, the modified IBDV were rescued in cells co-transfected with plasmids containing the rGx-F9VP2 backbone and pCGxBHRT; the rescued viruses were named rGxHT-242, rGxHT-249, rGxHT-256, rGxHT-270, and rGxHT-279, which contained mutations in the PDE domain (I242V, Q249R, and I256V) and the PFG domain (A270T and D279N) of VP2 in the backbone of the virulent strain rGx-F9VP2, respectively
Summary
Infectious bursal disease (IBD) is a highly contagious immunosuppressive disease in chickens that has caused significant losses to the commercial poultry industry worldwide [1,2]. IBDV is a member of the Birnaviridae family. It has a nonenveloped capsid structure containing a double-stranded RNA genome composed of two segments, A and B. The polyprotein is co-translationally self-cleaved to form the viral proteins VP2, VP3, and VP4 [9]. VP4 is a viral protease responsible for the self-processing of the IBDV polyprotein [10,11,12]. VP2 is the major structural protein and the only component of the icosahedral capsid [15,16]; it is responsible mostly for virulence, cell tropism [17,18,19,20,21,22,23], and antigenic variation [24]. The detailed molecular basis for the pathogenicity of vvIBDV is still not fully understood
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