Elevated temperature treatment as a novel method for decreasing p57 on thecell surface of Renibacterium salmoninarum

Renibacterium salmoninarum is a Gram-positive diplo-bacillus and the causat ive agent of bacterial kidney disease, a prevalent disease of salmonid fish . Virulent isolates of R. salmoninarum have a hydrophobic cell surface and express the 57-58 kDa protein (p57). Here we have investigated parameters w hich effect cell hydrophobicity and p57 degradation. Incubation of R, salmo ninarum cells at 37 degrees C for >4 h decreased cell surface hydrophobicit y as measured by the salt aggregation assay, and decreased the amount of ce ll associated p57. Incubation of cells at lower temperatures (22, 17, 4 or -20 degrees C) for up to 16 h did not reduce hydrophobicity or the amount o f cell associated p57. Both the loss of cell surface hydrophobicity and the degradation of p57 were inhibited by pre-incubation with the serine protea se inhibitor phenylmethylsulfonyl fluoride (PMSF). Cell surface hydrophobic ity was specifically reconstituted by incubation with extracellular protein (ECP) concentrated from culture supernatant and was correlated with the re association of p57 onto the bacterial cell surface as determined by western blot and total protein stain analyses. The ability of p57 to reassociate s uggests that the bacterial cell surface is not irreversibly modified by the 37 degrees C treatment and that p57 contributes to the hydrophobic nature of R. salmoninarum. In summary, we describe parameters effecting the remova l of the p57 virulence factor and suggest the utility of this modification for generating a whole cell vaccine against bacterial kidney disease.