Division of Avian Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, the Chinese Academy of Agricultural Sciences, Harbin, PR China.
PLoS One. 2013 Jul 26;8(7):e70982. doi: 10.1371/journal.pone.0070982. Print 2013.
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.
传染性法氏囊病病毒（IBDV）是对家禽业具有全球重要意义的病原体。尽管衣壳蛋白 VP2 的 PDE 和 PFG 结构域对毒力和适应性有重要贡献，但 IBDV 的致病机制的详细分子基础仍未完全了解。由于 VP2 的 253 位和 284 位氨基酸残基并非毒力的唯一决定因素，因此我们假设 VP2 的 PDE 和 PFG 结构域中可能存在其他与毒力和适应性相关的残基。为了验证这一点，我们使用反向遗传学系统，通过 VP2 的 PDE 和 PFG 结构域的序列比较，选择了 5 个氨基酸变化，并分别引入到强毒株（rGx-F9VP2）中。然后，我们分别将选择的 249 位和 256 位氨基酸残基的反向突变引入到弱毒株（rGt）中。生成了 7 种修饰病毒，并在体外（CEF 细胞）和体内（SPF 鸡）进行了评估。对于 249 位氨基酸残基，Q249R 可以提高 rGx-F9VP2 的体外复制能力并降低其体内复制能力，而 R249Q 可以降低 rGt 的体外复制能力并提高其体内复制能力；同时，Q249R 降低了 rGx-F9VP2 的毒力，而 R249Q 提高了 rGt 的毒力，这表明 249 位氨基酸残基显著影响了 IBDV 的复制和毒力。对于 256 位氨基酸残基，I256V 可以提高 rGx-F9VP2 的体外复制能力并降低其体内复制能力，而 V256I 可以降低 rGt 的体外复制能力但不改变其体内复制能力；尽管 V256I 没有增加 rGt 的毒力，但 I256V 明显降低了强毒 IBDV 的毒力。本研究首次证明，VP2 的 249 位和 256 位氨基酸残基在不同程度上参与了 IBDV 的复制效率和毒力；这不仅有助于进一步了解致病机制，而且有助于设计针对 IBDV 的新型定制疫苗。
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