CENTEX SHRIMP, Faculty of Science, Mahidol University, Rama VI Road, Bangkok, 10400, Thailand.
National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Thailand Science Park, Klong Luang, Patumthani 12120, Thailand.
J Gen Virol. 2013 Nov;94(Pt 11):2458-2468. doi: 10.1099/vir.0.054379-0. Epub 2013 Aug 15.
Yellow head virus (YHV) particles contain a nucleocapsid protein (p20) and two envelope glycoproteins (gp116 and gp64). The glycans attached to the two glycoproteins are N-linked and are complex and high mannose types, respectively. Here, we show that treatment with the N-linked glycosylation inhibitor tunicamycin in YHV-infected black tiger shrimp (Penaeus monodon) resulted in less severe yellow head disease and reduced mortality when compared with untreated control shrimp. Quantitative real-time reverse transcription PCR analysis also revealed lower YHV copy numbers in the haemolymph of treated than control shrimp. This was concurrent with less intense immuno-reactions in tissues of treated versus untreated shrimp using mAbs against all three YHV structural proteins. In addition, transmission electron microscopy of lymphoid organ tissue of the treated and untreated shrimp [eight collected at 36 h and eight at 48 h post-infection (p.i.)] revealed only unenveloped nucleocapsids in all but one of the treated shrimp (collected at 48 h p.i.). By contrast, all the untreated shrimp showed a mixture of many unenveloped and enveloped virions. These results were supported by purification of YHV from the cell-free haemolymph of treated and untreated shrimp followed by YHV structural protein analysis by SDS-PAGE. It revealed three expected structural protein bands (116, 64 and 20 kDa) from the untreated shrimp but no structural protein bands from the tunicamycin-treated shrimp (confirmed by Western blot analysis). Overall, the results indicated that blocking glycosylation with tunicamycin inhibited the formation of mature YHV virions and their subsequent release into shrimp haemolymph, reducing the severity of disease.
黄头病毒（YHV）颗粒包含核衣壳蛋白（p20）和两种包膜糖蛋白（gp116 和 gp64）。这两种糖蛋白上附着的糖均为 N-连接的，分别为复杂型和高甘露糖型。在这里，我们表明，与未处理的对照虾相比，在用 N-连接糖基化抑制剂衣霉素处理感染黄头病病毒的斑节对虾（Penaeus monodon）后，黄头病的严重程度降低，死亡率降低。定量实时逆转录 PCR 分析还显示，在用衣霉素处理的虾的血淋巴中 YHV 拷贝数低于对照虾。这与在用针对所有三种 YHV 结构蛋白的 mAb 处理的虾与未处理的虾的组织中的免疫反应强度较低相一致。此外，在用衣霉素处理和未处理的虾的淋巴器官组织的透射电子显微镜观察[在感染后 36 小时和 48 小时收集的 8 个虾和 8 个虾]中，除一个处理虾（在感染后 48 小时收集）外，仅在所有虾中发现未包膜的核衣壳。相比之下，所有未处理的虾均显示出许多未包膜和包膜的病毒粒子的混合物。这些结果得到了从用衣霉素处理和未处理的虾的无细胞血淋巴中纯化的 YHV 以及通过 SDS-PAGE 对 YHV 结构蛋白进行分析的支持。结果显示，未处理的虾中有三个预期的结构蛋白条带（116、64 和 20 kDa），但用衣霉素处理的虾中没有结构蛋白条带（通过 Western blot 分析证实）。总的来说，结果表明，用衣霉素阻断糖基化抑制了成熟 YHV 病毒粒子的形成及其随后向虾血淋巴中的释放，从而降低了疾病的严重程度。
