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Diseases of Aquatic Organisms

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DAO 75:37-50 (2007)  -  doi:10.3354/dao075037

Ultrastructure and molecular diagnosis of Spironucleus salmonis (Diplomonadida) from rainbow trout Oncorhynchus mykiss in Germany

M. Reza Saghari Fard1,2,*, Anders Jørgensen3, Erik Sterud3,4, Wilfrid Bleiss5, Sarah L. Poynton1,6

1Department of Inland Fisheries, Leibniz-Institute of Freshwater Ecology and Inland Fisheries, Müggelseedamm 310, 12587 Berlin, Germany
2Faculty of Agriculture and Horticulture, Humboldt University of Berlin, Invalidenstrasse 42, 10115 Berlin, Germany
3National Veterinary Institute, PO Box 8156 Dep, 0033 Oslo, Norway
4Standards Norway, PO Box 242, 1326 Lysaker, Norway
5Molecular Parasitology, Institute of Biology, Humboldt University of Berlin, Philippstrasse 13, 10115 Berlin, Germany
6Department of Molecular and Comparative Pathobiology, Johns Hopkins University School of Medicine, Broadway Research Building, 733 North Broadway, Room 807, Baltimore, Maryland 21205, USA

ABSTRACT: Diplomonad flagellates infect a wide range of fish hosts in aquaculture and in the wild in North America, Asia and Europe. Intestinal diplomonad infection in juvenile farmed trout can be associated with morbidity and mortality, and in Germany, diplomonads in trout are commonly reported, and yet are poorly characterised. We therefore undertook a comprehensive study of diplomonads from German rainbow trout Oncorhynchus mykiss, using scanning and transmission electron microscopy, and sequencing of the small subunit (ssu) rRNA gene. The diplomonad was identified as Spironucleus salmonis, formerly reported from Germany as Hexamita salmonis. Our new surface morphology studies showed that the cell surface was unadorned and a caudal projection was present. Transmission electron microscopy facilitated new observations of functional morphology, including vacuoles discharging from the body surface, and multi-lobed apices of the nuclei. We suggest the lobes form, via hydrostatic pressure on the nucleoplasm, in response to the beat of the anterior-medial flagella. The lobes serve to intertwine the nuclei, providing stability in the region of the cell exposed to internal mechanical stress. The ssu rRNA gene sequence clearly distinguished S. salmonis from S. barkhanus, S. salmonicida, and S. vortens from fish, and can be used for identification purposes. A 1405 bp sequence of the ssu rRNA gene from S. salmonis was obtained and included in a phylogenetic analysis of a selection of closely related diplomonads, showing that S. salmonis was recovered as sister taxon to S. vortens.

KEY WORDS: Diagnosis · Diplomonad · Flagellate · Oncorhynchus mykiss · Rainbow trout · Sequence · Spironucleus salmonis · ssu rDNA · Ultrastructure

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