Morphology and distribution of blood fluke eggs and associated pathology in the gills of cultured Pacific bluefin tuna, Thunnus orientalis

Abstract

Infestations of blood flukes of the genus Cardicola have been observed in juvenile Pacific bluefin tuna (PBT) cultured in Japan. Infected fish harbor large numbers of parasite eggs in their gills. Although the link between blood fluke infection and juvenile mortality is not clear, accumulation of parasite eggs appears to be pathogenic to the fish. We investigated the origins, general morphology/distribution, and histopathology of these eggs in artificially produced 0yr old PBT. Dead and live fish were sampled on several occasions from two culture facilities in Wakayama prefecture, Japan. The number of eggs in each gill filament was enumerated under a microscope. In addition, we estimated the total number of eggs by dissolving the gills in a weak NaOH solution. We observed two morphologically distinct egg types in the gill filaments, smaller, oval shaped eggs in the gill lamellae and larger, crescent shaped eggs that occurred primarily in the filamentary arteries. Based on the ITS2 sequence, the ovoid and crescent shaped eggs were identified as C. orientalis and C. opisthorchis, respectively. Eggs of the former species were more abundant (maximum: 6400 per filament) than the latter (maximum: 1400), but the number was highly variable among filaments. The eggs of the latter species were relatively evenly distributed among the filaments. In a heavily infected individual, we estimated a total of >4.5 million eggs were present in the gills on one side of the fish. The number of eggs from the two species was positively correlated to each other and the dead fish tended to harbor more eggs than the live fish. Histological observation revealed host responses around the eggs, including encapsulation by fibroblasts and nodule formation, as seen in response to other aporocotylid eggs. In addition, we observed widespread fusion of gill lamellae and blockage of the filamentary arteries in some instances. Our results provide information that can be used for routine diagnosis of Cardicola blood flukes in cultured tuna and suggest they represent a risk to juvenile PBT.