Characterization of newly revealed sequences in the infectious myonecrosis virus genome in Litopenaeus vannamei

Abstract

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 59 terminus and 23 nt at the 39 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 59 RACE, an additional 4 nt were discovered; 39 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 59 end of the IMNV genome in IMNVinfected L. vannamei. An RNAi trigger targeting a 376 bp length of the 59 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.

Nucleic acid amplification‐based methods for diagnosis of shrimp viral diseases

“…It is a non‐enveloped, linear dsRNA containing virus with icosahedral symmetry and a diameter of 40 nm 190 . The RNA was reported to be in the size range of ~8 to 9 kb 191 . The genome contains two non‐overlapping ORFs, namely ORF‐1 encoding a capsid protein and a putative RNA‐binding protein, and a second, ORF‐2, which is responsible for encoding putative RNA‐dependent RNA polymerase, RdRp 191 .…”

“…190 The RNA was reported to be in the size range of 8 to 9 kb. 191 The genome contains two non-overlapping ORFs, namely ORF-1 encoding a capsid protein and a putative RNA-binding protein, and a second, ORF-2, which is responsible for encoding putative RNA-dependent RNA polymerase, RdRp. 191 The major host species for IMNV infection is P. vannamei, while other shrimp species including Farfantepenaeus subtilis, P. monodon and P. stylirostris are experimentally susceptible to IMNV.…”

“…191 The genome contains two non-overlapping ORFs, namely ORF-1 encoding a capsid protein and a putative RNA-binding protein, and a second, ORF-2, which is responsible for encoding putative RNA-dependent RNA polymerase, RdRp. 191 The major host species for IMNV infection is P. vannamei, while other shrimp species including Farfantepenaeus subtilis, P. monodon and P. stylirostris are experimentally susceptible to IMNV. 187,192 The infection is more prevalent in juvenile stages of some penaeid species farmed in marine and brackish water as well as in low salinity brackish water species, but the sub-adult and adult stages are also susceptible, and it causes mortality up to 40%-70%.…”