Novel RNA viruses causing muscle lesions in red claw crayfish (Cherax quadricarinatus)

Abstract

In 2014, a new syndrome was recognized in redclaw crayfish (Cherax quadricarinatus) from farms in northern Queensland. Crayfish started to die with mortality reaching 40% in approximately three weeks after transportation and translocation stress. In trying to identify the cause of the disease, sequence dependent and independent PCRs failed to identify any aetiology. However, using next gen sequencing of RNA, two novel RNA viruses, namely chequa iflavirus and bunya‐like virus were found in the stressed, dying crayfish. Chequa iflavirus is a positive sense, single stranded RNA virus in the order Picornavirales, marginally in the genus Iflavirus. This is the first iflavirus identified from crustracea. The bunya‐like virus found is related to Whenzhou Shrimp Virus 2, a bi‐segmented, unaligned, negative sense, single strand RNA virus. Efficient and sensitive detection methods for these viruses based on RNA dependent RNA polymerase (RdRp) was developed for large scale screening of crayfish on farm. Due to a lack of virus‐free crayfish, house cricket, Acheta domesticus was trialled as a bioassay animal to prove River’s postulates for chequa iflavirus and bunya‐like virus. However, it was unsuccessful as the immune system of cricket might able to destroy and leave no viable virus or enough traces of RNA to be detected by reverse transcription polymerase chain reaction (RT‐PCR). To solve the viral infection problem with chequa iflavirus infection on farm, some possible viral reduction strategies for farmers were trialled. The polyphenol flavonoid quercetin, inhabiting water temperature at 32°C all had statistically significant effects reducing viral copy numbers (P<0.018; P<0.05 respectively). While RNA interference based on RdRp sequence appeared to drop viral copy number 80% at four weeks, it was not statistical significant (P˃0.05). For the crayfish farmers, quercetin is the most favoured treatment as it is abundant, cheap and readily incorporated into the diet and should ameliorate the original, transport‐stress mortality which started this study. Broodstock could be warmed to 32°C in the hatchery to reduce viral dose. Although RNAi treatment was most expensive to implement, it could be delivered with limited amounts of RNAi at the hatchery.