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Phylogenetic Characterization and Prevalence of "Spirobacillus cienkowskii," a Red-Pigmented, Spiral-Shaped Bacterial Pathogen of Freshwater Daphnia Species

Jorge L. M. Rodrigues,^1, Meghan A. Duffy,^2, Alan J. Tessier,^2, Dieter Ebert,³ Laurence Mouton,³ and Thomas M. Schmidt^1,2*

Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, Michigan 48824,¹ W. K. Kellogg Biological Station, Michigan State University, Hickory Corners, Michigan 49060,² Zoologisches Institut der Universität Basel, Evolutionsbiologie, Vesalgasse 1, 4051 Basel, Switzerland³

Received 29 October 2007/ Accepted 2 January 2008

Microscopic examination of the hemolymph from diseased daphniids in 17 lakes in southwestern Michigan and five rock pools in southern Finland revealed the presence of tightly coiled bacteria that bore striking similarities to the drawings of a morphologically unique pathogen, "Spirobacillus cienkowskii," first described by Elya Metchnikoff more than 100 years ago. The uncultivated microbe was identified as a deeply branching member of the Deltaproteobacteria through phylogenetic analyses of two conserved genes: the 16S rRNA-encoding gene (rrs) and the β-subunit of topoisomerase (gyrB). Fluorescence in situ hybridization confirmed that the rRNA gene sequence originated from bacteria with the tightly coiled morphology. Microscopy and PCR amplification with pathogen-specific primers confirmed infections by this bacterium in four species of Daphnia: Daphnia dentifera, D. magna, D. pulicaria, and D. retrocurva. Extensive field surveys reveal that this bacterium is widespread geographically and able to infect many different cladoceran species. In a survey of populations of D. dentifera in lakes in Michigan, we found the bacterium in 17 of 18 populations studied. In these populations, 0 to 12% of the individuals were infected, with an average of 3% during mid-summer and early autumn. Infections were less common in rock pool populations of D. magna in southern Finland, where the pathogen was found in 5 of 137 populations. The broad geographic distribution, wide host range, and high virulence of S. cienkowskii suggest it plays an important role in the ecology and evolution of daphniids.

Published ahead of print on 11 January 2008.

Present address: Department of Biology, University of Texas at Arlington, Arlington, TX 76019.

Present address: School of Biology, Georgia Institute of Technology, 310 Ferst Drive, Atlanta, GA 30332.

Present address: Division of Environmental Biology, National Science Foundation, Arlington, VA 22230.