Molecular and histological identification of Marteilioides infection in Suminoe Oyster Crassostrea ariakensis, Manila Clam Ruditapes philippinarum and Pacific Oyster Crassostrea gigas on the south coast of Korea Limpanont Yanin a,b,1 , Hyun-Sil Kang a,1 , Hyun-Ki Hong a , Hee-Do Jeung a , Bong-Kyu Kim a , Thanh Cuong Le a,d , Young-Ok Kim c , Kwang-Sik Choi a,⇑ a School of Marine Biomedical Science (BK21 PLUS), Jeju National University, 102 Jejudaehakno, Jeju 690-756, Republic of Korea b Department of Social and Environmental Medicine, Faculty of Tropical Medicine, Mahidol University, 420/6 Rajvithi Rd., Rajthevi, 10400 Bangkok, Thailand c South Sea Research Institute, Korea Institute of Ocean Science and Technology (KIOST), Geoje 656-830, Republic of Korea d Faculty of Aquaculture, Nha Trang University, 02 Nguyen Dinh Chieu, Nha Trang, Khanh Hoa, Vietnam article info Article history: Received 1 January 2013 Accepted 28 August 2013 Available online 7 September 2013 Keywords: Marteilioides chungmuensis Crassostrea gigas Ruditapes philippinarum Crassostrea ariakensis SSU rDNA Nested PCR abstract The oyster ovarian parasite Marteilioides chungmuensis has been reported from Korea and Japan, damag- ing the oyster industries. Recently, Marteilioides-like organisms have been identified in other commer- cially important marine bivalves. In this study, we surveyed Marteilioides infection in the Manila clam Ruditapes philippinarum, Suminoe oyster Crassostrea ariakensis, and Pacific oyster Crassostrea gigas, using histology and Marteilioides-specific small subunit (SSU) rDNA PCR. The SSU rDNA sequence of M. chungm- uensis (1716 bp) isolated from C. gigas in Tongyoung bay was 99.9% similar to that of M. chungmuensis reported in Japan. Inclusions of multi-nucleated bodies in the oocytes, typical of Marteilioides infection, were identified for the first time in Suminoe oysters. The SSU rDNA sequence of a Marteilioides-like organ- ism isolated from Suminoe oysters was 99.9% similar to that of M. chungmuensis. Marteilioides sp. was also observed from 7 Manila clams of 1840 individuals examined, and the DNA sequences of which were 98.2% similar to the known sequence of M. chungmuensis. Unlike Marteilioides infection of Pacific oysters, no remarkable pathological symptoms, such as large multiple lumps on the mantle, were observed in infected Suminoe oysters or Manila clams. Distribution of the infected Manila clams, Suminoe oysters and Pacific oysters was limited to small bays on the south coast, suggesting that the southern coast is the enzootic area of Marteilioides infection. Ó 2013 Elsevier Inc. All rights reserved. 1. Introduction Since the introduction of the modern suspended long-line and raft culture techniques in 1960, Pacific oyster (Crassostrea gigas) farming has been promoted on the southern coast of Korea. In 2009, oyster landing in Korea reached over 240,000 tones, more than 90% of which originated from the south coast, where the sus- pended long-line culture has been practiced widely in closed or semi-closed bays for the past decade (Choi, 2008). The ovarian par- asite Marteilioides chungmuensis has been identified as a major threat to the Korean oyster-farming industry; however, few studies have investigated its impact (Park and Chun, 1989; Ngo et al., 2003; Park et al., 2003). Recently, this ovarian parasite was identi- fied in other marine bivalves, such as the Iwakagi oyster Crassos- trea nippona (Itoh et al., 2004b) and Manila clam Ruditapes philippinarum (Lee et al., 2001; Itoh et al., 2005), suggesting that it is not limited to the Pacific oyster; other marine bivalves can be a host of M. chungmuensis. M. chungmuensis infection was first identified in Pacific oysters reared in a suspended long-line culture system in Tongyoung, off the south coast of Korea (Chun, 1972). The infected oysters exhib- ited multiple spherical or oval yellowish eccentric nodules on the mantle. Due to their abnormal appearance, the infected oysters were discarded during processing, resulting in economic loss to the oyster farmers. The pathogen isolated from nodules of the abnormal oysters was originally identified as an amoeba (Chun, 1972). The ultrastructure of the amoeba-like pathogen was exam- ined using transmitted electron microscopy (TEM), which revealed characteristics of paramyxean protozoa (Comps et al., 1986). Accordingly, Comps et al. (1986) established a new genus, Martei- lioides, in the Phylum Paramyxea, and the pathogen was recorded as a new species, M. chungmuensis (see the review by Berthe 0022-2011/$ - see front matter Ó 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.jip.2013.08.008 ⇑ Corresponding author. Fax: +82 64 756 3493. E-mail address: skchoi@jejunu.ac.kr (K.-S. Choi). 1 These authors contributed equally to this work. Journal of Invertebrate Pathology 114 (2013) 277–284 Contents lists available at ScienceDirect Journal of Invertebrate Pathology journal homepage: www.elsevier.com/locate/jip