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Diseases of Aquatic Organisms

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DAO 55:247-252 (2003)  -  doi:10.3354/dao055247

Detection of the oyster parasite Bonamia ostreae by fluorescent in situ hybridization

Ryan B. Carnegie1,3,*, Bruce J. Barber1, Daniel L. Distel1,2

1School of Marine Sciences, and
2Department of Biochemistry, Microbiology, and Molecular Biology, 5735 Hitchner Hall, University of Maine, Orono, Maine 04469-5735, USA
3Present address: Aquaculture Genetics and Breeding Technology Center, Virginia Institute of Marine Science, Gloucester Point, Virginia 23062, USA

ABSTRACT: Bonamia ostreae is an economically significant protistan parasite of the flat oyster Ostrea edulis in Europe and North America. Management of this parasite depends partly upon its reliable identification in wild and aquacultured oyster populations, but B. ostreae is small and difficult to detect by traditional microscopic methods. We designed a fluorescent in situ hybridization (FISH) assay to sensitively detect B. ostreae in standard histopathological sections of B. ostreae-infected oysters using fluorescently labeled DNA oligonucleotide probes. Hybridization using a cocktail of 3 presumptively B. ostreae-specific, fluorescein iso(thio)cyanate (FITC)-labeled oligonucleotides produced an unambiguous staining pattern of small green rings inside infected oyster hemocytes that was easily distinguished from host tissue background. This pattern is diagnostic for B. ostreae. A negative control cocktail of oligonucleotides containing 2 mismatches relative to target sequences, on the other hand, failed to hybridize at all. B. ostreae-specific probes did not cross-react with a related protist, Haplosporidium nelsoni.

KEY WORDS: Bonamia ostreae · Ostrea edulis · Fluorescent in situ hybridization

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