Cardiomyopathy Syndrome of Atlantic Salmon (Salmo salar L.) Is Caused by a Double-Stranded RNA Virus of the Totiviridae Family

Abstract

Cardiomyopathy syndrome (CMS) of farmed and wild Atlantic salmon (Salmo salar L.) is a disease of yet unknown etiology characterized by a necrotizing myocarditis involving the atrium and the spongious part of the heart ventricle. Here, we report the identification of a double-stranded RNA virus likely belonging to the family Totiviridae as the causative agent of the disease. The proposed name of the virus is piscine myocarditis virus (PMCV). On the basis of the RNA-dependent RNA polymerase (RdRp) sequence, PMCV grouped with Giardia lamblia virus and infectious myonecrosis virus of penaeid shrimp. The genome size of PMCV is 6,688 bp, with three open reading frames (ORFs). ORF1 likely encodes the major capsid protein, while ORF2 encodes the RdRp, possibly expressed as a fusion protein with the ORF1 product. ORF3 seems to be translated as a separate protein not described for any previous members of the family Totiviridae. Following experimental challenge with cell culture-grown virus, histopathological changes are observed in heart tissue by 6 weeks postchallenge (p.c.), with peak severity by 9 weeks p.c. Viral genome levels detected by real-time reverse transcription (RT)-PCR peak earlier at 6 to 7 weeks p.c. The virus genome is detected by in situ hybridization in degenerate cardiomyocytes from clinical cases of CMS. Virus genome levels in the hearts from clinical field cases correlate well with the severity of histopathological changes in heart tissue. The identification of the causative agent for CMS is important for improved disease surveillance and disease control and will serve as a basis for vaccine development against the disease.

Piscine myocarditis virus (PMCV) in wild Atlantic salmon Salmo salar

“…Sequencing revealed that both isolates from wild salmon were > 99% similar to the PMCV genome sequence reported by Haugland et al (2011) (GenBank accession numbers JQ745675-JQ745680). Using this genome as a reference, Isolate 1198 had a partial ORF3 se quence that was identical to the reference genome, but in both isolates a small number of unique single nucleotide differences were detected in the other 2 ORFs.…”

“…This was based on the presence of eosinophilic inclusions in nuclei of myocardial fibres in the vicinity of degenerated areas. Experimental transmission studies have supported this hypothesis (Bruno & Noguera 2009, Fritsvold et al 2009, and in 2010 a doublestranded RNA virus tentatively of the Totiviridae family was described as the causative agent of CMS (Løvoll et al 2010, Haugland et al 2011.…”

“…Samples that were PCR-positive were further investigated by sequencing. Partial sequences for all 3 open reading frames (ORFs) in the PMCV genome (Haugland et al 2011) were derived using direct sequencing of PCR products. The Qiagen OneStep RT-PCR kit was used with final primer concentrations 600 nM and the following PCR cycle: 30 min at 50°C (reverse transcription), 15 min at 95°C (inactivation of reverse transcriptase and activation of hotstart PCR DNA polymerase), 30 s at 94°C (template denaturation), 1 min at 55°C (primer annealing), 1 min at 72°C (primer elongation; 45 cycles total) and a final elongation step (3 min at 72°C).…”

Current Advances in Functional Genomics in Aquaculture

“…Similarly, viral haemorrhagic septicaemia (VHS) was irst reported in the early 1950s in salmonids while the causative agent was characterized later after 10 years in 1962 [24]. This trend was observed for several other diseases such as infectious hematopoietic necrosis virus (IHNV), nervous necrosis virus (NNV), heart and skeletal muscle inlammation (HSMI) and cardiac myopathy syndrome (CMS) in which identiication of the etiological agents took long after clinical signs were irst reported [25][26][27][28][29][30][31][32][33]. However, the upcoming of metagenomics has accelerated our discovery of novel pathogens in which the duration from observation of irst clinical signs to identiication of the etiological agent has been reduced.…”

Viral vaccines for farmed finfish

“…For example, infectious pancreatic necrosis (IPN) was removed as an OIE notifiable disease of aquatic animals in 2005 but IPN continues to cause losses to the aquaculture industry in Europe [15,86]. Examples of recently identified diseases include cadiomyopathy syndrome (CMS) [43], and hemorrhagic kidney syndrome (HKS) involving Toga-like virus in Atlantic salmon [11]. Another example of a recently identified virus that has been implicated in increased mortality in barbel fry (Barbus barbus) in Hungary includes a circovirus (single-stranded DNA containing virus) although a relation between the presence of the virus and clinical manifestations has yet to be established [65].…”