DAO 67:149-154 (2005)  -  doi:10.3354/dao067149

Purification and antigenic characteristics of a rickettsia-like organism from the oyster Crassostrea ariakensis

Xinzhong Wu1,2,*,**, Jingfeng Sun2,**, Weizhu Zhang2, Bohai Wen3

1College of Animal Sciences, Zhejiang University, 268 Kaixuan Road, Hangzhou 310029, China
2The South China Sea Institute of Oceanology, The Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China
3Institute of Microbiology and Epidemiology, Beijing 100071, China
** The first two authors made equal contributions to this paper

ABSTRACT: A rickettsia-like organism (RLO) has been suggested to be the etiological agent responsible for heavy losses of the oyster Crassostrea ariakensis Gould in China. Because of the lack of molluscan cell lines for in vitro culture of intracellular prokaryotes, antigenic analysis of RLOs has been limited by the inherent difficulties of their purification. In this report, we describe the use of differential speed centrifugation and renografin density gradient centrifugation to purify the RLO directly from infected oyster tissues. The purity and integrity of purified prokaryotes were validated by transmission electron microscopy. Thirteen major constituent proteins, with molecular weights ranging between 17 and 99 kDa, were electrophoretically identified by silver staining, and 8 major proteins were identified with Coomassie blue R staining. Specific mouse polyclonal antiserum was prepared for serological characterization of the RLO and was used in an immunoblot assay, and 3 major antigen groups were identified. The present results advance our knowledge of RLO protein antigens, and several proteins have been identified that could potentially be useful for diagnostic assays or for production of experimental immunostimulants.

KEY WORDS: Rickettsia-like organism · Crassostrea ariakensis · Antigenic characteristics · Purification

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