Loy Duan S, Liu Sijun, Mogler Mark A, Dustin Loy J, Blitvich Bradley J, Bartholomay Lyric C
1Department of Veterinary Microbiology and Preventive Medicine, Iowa State University, Ames, IA 50011, USA.
2Department of Entomology, Iowa State University, Ames, IA 50011, USA.
J Gen Virol. 2015 Jul;96(Pt 7):1821-9. doi: 10.1099/vir.0.000137. Epub 2015 Apr 2.
Infectious myonecrosis virus (IMNV) causes significant economic losses in farmed shrimp, where associated mortality in ponds can reach 70 %. To explore host/pathogen interactions, a next-generation sequencing approach using lymphoid organ tissue from IMNV-infected Litopenaeus vannamei shrimp was conducted. Preliminary sequence assembly of just the virus showed that there were at least an additional 639 bp at the 5' terminus and 23 nt at the 3' terminus as compared with the original description of the IMNV genome (7561 nt). Northern blot and reverse transcription-PCR analysis confirmed the presence of novel sequence at both ends of the genome. Using 5' RACE, an additional 4 nt were discovered; 3' RACE confirmed the presence of 22 bp rather than 23 bp of sequence. Based on these data, the IMNV genome is 8226 bp in length. dsRNA was used to trigger RNA interference (RNAi) and suppress expression of the newly revealed genome sections at the 5' end of the IMNV genome in IMNV-infected L. vannamei. An RNAi trigger targeting a 376 bp length of the 5' UTR did not improve survival of infected shrimp. In contrast, an RNAi trigger targeting a 381 bp sequence in ORF1 improved survival to 82.2 % as compared with 2.2 % survival in positive control animals. These studies revealed the importance of the new genome sections to produce high-titre infection, and associated disease and mortality, in infected shrimp.
传染性肌肉坏死病毒（IMNV）给养殖对虾造成了巨大的经济损失，池塘中的相关死亡率可达70%。为了探究宿主与病原体之间的相互作用，采用下一代测序方法，对感染IMNV的凡纳滨对虾的淋巴器官组织进行了研究。仅对病毒进行初步序列组装后发现，与IMNV基因组的原始描述（7561 nt）相比，其5'端至少还有639 bp，3'端还有23 nt。Northern印迹和逆转录PCR分析证实了基因组两端存在新序列。利用5' RACE发现了另外4 nt；3' RACE证实存在22 bp而非23 bp的序列。基于这些数据，IMNV基因组长度为8226 bp。双链RNA（dsRNA）被用于触发RNA干扰（RNAi），并抑制IMNV感染的凡纳滨对虾中IMNV基因组5'端新发现的基因组片段的表达。靶向5'非翻译区（UTR）376 bp长度的RNAi触发物并未提高受感染对虾的存活率。相比之下，靶向开放阅读框1（ORF1）中381 bp序列的RNAi触发物使存活率提高到了82.2%，而阳性对照动物的存活率为2.2%。这些研究揭示了新基因组片段对于在受感染对虾中产生高滴度感染以及相关疾病和死亡率的重要性。
