Discovery of LarvalGnathostoma nipponicumin Frogs and Snakes from Jeju-do (Province), Republic of Korea

Gnathostoma species nematodes are clinically important and can cause food-borne zoonotic parasitism in humans.More than 12 species, including Gnathostoma spinigerum, have been reported as valid species [1].In the Republic of Korea, Kim [2] detected 2 G. spinigerum larvae in a snakehead, Channa argus argus, from Kimhae, Gyeongsangnam-do (Province) [2].The third stage larvae of G. hispidum were found in loaches and snakes from China [3,4] and in pit-viper snakes, Agkistrodon brevicaudus, from Korea [5].Larval G. nipponicum were also detected in imported Chinese loaches and also in tiger keelback snakes (Rhabdophis tigrinus tigrinus) from Hongcheon-gun, Korea [6,7].On the other hand, G. nipponicum adults were found in the stomach of the Jeju weasel, Mustela sibilica quelpartis, road-killed in Jeju-do [8].In the present study, we performed a survey on intermediate hosts of G. nipponicum in Jeju-do, the Republic of Korea.We collected 82 tadpoles, 23 black-spotted pond frogs (Rana nigromaculata), 6 red-tongue viper snakes (Agkistrodon ussuriensis), 7 tiger keelback snakes, and 2 cat snakes (Elaphe dione) from the vicinities of Jeju National University in August 2009 and 2010.They all were tranferred to the laboratory of Department of Veterinary Parasitology, Jeju National University College of Veterinary Medicine, Jeju, Korea, and examined by the pepsin-HCl digestion method.Some collected gnathostome larvae were fixed with 10% formalin, cleared in alcohol-glycerin solution, mounted in glycerin-jelly, and observed under a light microscope equipped with a micrometer.To observe the surface ultrastructure, some worms were washed several times with 0.2 M cacodylate buffer (pH 7.2) and fixed with 2.5% glutaraldehyde at 4˚C.After washing 3 times with the buffer, they were dehydrated through a graded alcohol series (50%, 70%, 80%, 90%, 95%, and absolute alcohol), dried with hexamethyldisilazane, coated (JFC-1100E ion sputtering device) with gold, and observed with a scanning electron microscope (SEM, Philips XL-30S, London, UK) at a 15-kV accelerating voltage.Total 5 gnathostome larvae were detected in 3 (50%) of 6 A.