The development of next generation sequencing has greatly facilitated the exploration of viruses that infect marine hosts. For example, there are an increasing number of virus genomes belonging to the Totiviridae family described in crustacean hosts. In this study, two novel distinct dsRNA virus genomes were discovered in the Atlantic blue crab C. sapidus in the United States. The complete genome sequences of both viruses were determined, which are 6444 nt and 7421 nt in length, containing two overlapping open reading frames (ORFs) that encode capsid and RNA dependent RNA polymerase proteins with similarities to toti -like viruses. The viruses were tentatively named Callinectes sapidus toti -like virus 1 (CsTLV1) and Callinectes sapidus toti -like virus 2 (CsTLV2). Both genomes have typical elements required for -1 ribosomal frameshifting, which may induce the expression of an encoded ORF1-ORF2 (gag-pol) fusion protein. Phylogenetic analyses of CsTLV1 and CsTLV2 RdRp amino acid sequences suggested that they are members of new genera in the family Totiviridae .
The presence of CsTLV 1/2 genomes were detected in muscle, gill and hepatopancreases of blue crabs by RT-qPCR. The PCR assay was used investigate the geographic range of these viruses, which may be restricted to the northeast US. The presence of 40 nm totivirus -like viral particles in all these three tissues were validated by transmission electron microscope (TEM). The two virus genomes co-occur most, but not all, of the time. To our knowledge, this is the first report demonstrating the presence of two novel toti -like viruses in the Portunid crab - C. sapidus. Transmission and pathogenicity studies are currently underway.