Molecular detection and genetic analysis of betanodaviruses in bivalve mollusks

Betanodaviruses are small ssRNA viruses responsible of Viral Nervous Necrosis in marine fishes worldwide. Genetic analysis of a fragment of the coat protein gene sequences permit to divide Betanodavirus in four genotypes. Preview studies evidenced Betanodavirus infection in several wild asymptomatic fish species, also in the Mediterranean Sea (1) and in invertebrates from Korea, mostly crustaceans and in a mussel (2). Both fish and invertebrates could be a potential source of virus for farmed fish. In this study, we examined bivalve mollusks for the presence of Betanodavirus and genetically analyzed the detected viruses to find out whether these animals could be a source of genetically closely related Betanodaviruses transmissible horizontally to wild and farmed fish. A total of 46 samples were analyzed including species reared on the seabed (Tapes philippinarum; Crassostrea gigas) and in the water column (Mytilus galloprovincialis). Betanodavirus presence was investigated by RT-nested/PCR (3). Positive samples were sequenced to study the variable region of the viral RNA2 coding for the capside protein. No viruses were detected in mussels (M. galloprovincialis), whereas clams (T. philippinarum) and oysters (C. gigas), both species reared on the seabed, resulted highly positive (26.6%). Phylogenetic analysis showed the belonging of these viruses to the RGNNV genotype. Betanodaviruses from clams clustered with viruses detected in wild and farmed fish collected from the same region (Adriatic Sea). Oyster viruses clustered separately, however inside the same genotype; considering the foreign origin of oysters (France), they could act as vectors of the infection between different areas. This is the first study focusing on Betanodavirus in bivalve mollusks; it evidences important presence of Betanodaviruses in bivalve mollusks; in some cases viral strains are highly correlated to those found in fish. These results highlight the possible role of mollusks in the spread of Betanodavirus to wild and farmed fish.