Occurrence of an erythrocytic virus infection in cultured turbot Scophthalmus maximus

Abstract

Cytoplasmic inclusions, ranging in size from 0.3 to 1 μm, were found in circulating erythrocytes of diseased turbot Scophthalmus maximus (L.), reared on several farms in Galicia (NW Spam). Examination of the erythrocytes with electron microscopy showed virus-like particles within cytoplasmic membrane-bound vacuoles, at the cell membrane or extracellularly The particles, oval to spherical in shape, displayed a wide range of sizes, from 50 to 160 nm. Particles appeared to he formed at the cell membrane and were released by budding. In negatively stained preparations surface projections or spikes were recognizable on the external envelope. Virological studies carried out in affected turbot allowed the isolation of an enveloped RNA virus from the internal organs. Healthy turbot were infected intraperitoneally with this viral agent followed by observation of similar erythrocytic inclusions and virus-like particles 3 wk after infection. Erythrocytes obtained from healthy turbot were infected in vitro. Study of the sequence of morphologic events after virus infection revealed that 5 min after inoculation virus-like particles were observed within cytoplasmic vacuoles of erythrocytes. Virus-like particles were adsorbed to the cell surface after 30 min at 0̊C but not internalized. However, at 4 and at 150°C the particles were endocytosed into small vesicles After 2 to 3 d at 15°C, some erythrocytes showed irregular morphology and high electron-dense cytoplasmic areas as well as virus-like particles being released by budding at the cell membrane. The virus titrations showed that intracellular viral titres were almost constant along the experiment. However, extracellular viral titres increased through-out time reaching maximum values at between 36 and 60 h post-infection.