DAO 33:33-41 (1998)  -  doi:10.3354/dao033033

Purification of Piscirickettsia salmonis and partial characterization of antigens

M. N. Barnes1,*, M. L. Landolt1, D. B. Powell2, J. R. Winton3,**

1School of Fisheries, University of Washington, PO Box 357980, Seattle, Washington 98195, USA
2Alpharma Inc., 1720 130th Avenue N.E., Bellevue, Washington 98005, USA
3Western Fisheries Research Center, 6505 N.E. 65th St., Seattle, Washington 98115, USA
*Present address: Alaska District Veterinary Command, 1060 Gaffney Road, Ft. Wainwright, Alaska 99703, USA
**Addressee for correspondence. E-mail:

Piscirickettsia salmonis is the etiological agent of salmonid rickettsial septicemia, an economically significant disease affecting the salmon aquaculture industry. As with other rickettsial pathogens, antigenic analysis of P. salmonis has been limited by the inherent difficulties of purifying an intracellular organism away from host cell material. In this report, we describe the use of diatrizoate meglumine and diatrizoate sodium (DMDS) density gradient centrifugation to purify P. salmonis grown in chinook salmon embryo (CHSE-214) cells. Plaque assay titers and total protein assays confirmed that viable P. salmonis was consistently concentrated in a visible band within the DMDS density gradient at a density of 1.15 to 1.16 g ml-1. Recovery of purified, viable organisms from DMDS density gradients varied from 0.6 to 3%. Preparations of uninfected CHSE-214 cells, CHSE-214 cells infected with P. salmonis, and gradient-purified P. salmonis were compared using sodium dodecyl sulfate polyacrylamide gel electrophoresis to assess the degree of purification and to identify P. salmonis-specific proteins. Although gradient-purified P. salmonis preparations were not completely free of host cell material, 8 bacterial proteins were identified. Polyclonal rabbit antiserum was used in an immunoblot of proteins from purified P. salmonis to identify 3 major and 5 minor antigens. The major antigens of 56, 30 and 20 kDa were potential candidates for experimental vaccines and development of novel diagnostic assays.

Piscirickettsia salmonis · Rickettsia · Fish disease · Salmonid · Antigen

Full text in pdf format