Antigenic variation in Renibacterium salmoninarum p57: functional and diagnostic implications.

Gregory D. Wiens¹ and Ole Bendik Dale²

¹National Center for Cool and Cold Water Aquaculture, Kearneysville, WV 25430

²National Veterinary Institute, Oslo, Norway

The 57-kDa protein (p57) is an important diagnostic antigen and is also implicated in the pathogenesis of salmonid bacterial kidney disease. Little is known about the nature and extent of antigenic variation in p57. Previously, we reported that p57 produced by R. salmoninarum strain 684 contains a mutation that disrupts monoclonal antibody 4C11 binding. Sequence analysis of the 5' and coding regions of the 684 msa1 and msa2 genes identified a single C-to-A nucleotide mutation in both genes as compared to the ATCC 33209 strain. This mutation creates an Ala(139)-to-Glu substitution in the amino-terminal region of p57. Purified p57 from strain 684 displayed enhanced binding for Chinook salmon leukocytes suggesting that the mutation may confer a selective advantage. Here, we examine antigenic variation in a panel of 24 R. salmoninarum isolates obtained from diverse geographic locations world-wide. Six p57-specific monoclonal antibodies (4C11, 4D3, 3H1, 4H8, 4D10 and 1A1) were used to probe dot and western blots to determine the relative expression, size and cellular association of p57. Full length p57 was produced by all 24 isolates and for each isolate, the protein was associated with the bacterial cell surface. The epitopes recognized by four Mabs, 4D3, 4H8, 3H1 and 1A1, were conserved among all strains tested. The 4D10 epitope was disrupted in one isolate from British Columbia, while the 4C11 epitope was lost in 5 of 8 strains isolated from Norway. The 5 Norwegian isolates were genetically similar sharing the following traits: one tandem repeat in the ETRA locus, a sequovar-four 16-23S rRNA intervening sequence, an alter msa1 5’ region, and they lack additional msa1-like genes. These results suggest that while p57 is not highly polymorphic, some antigenic variation exists that may be geographically restricted in distribution.