Comparison of the Membrane-Filtration Fluorescent Antibody Test, the Enzyme-Linked Immunosorbent Assay, and the Polymerase Chain Reaction to Detect Renibacterium Salmoninarum in Salmonid Ovarian Fluid

Ovarian fluid samples from naturally infected chinook salmon ( Oncorhynchus tshawytscha ) were examined for the presence of Renibacterium salmoninarumby the membrane-filtration fluorescent antibody test (MF-FAT), an antigen capture enzyme-linked immunosorbent assay (ELISA), and a nested polymerase chain reaction (PCR). On the basis of the MF-FAT, 64% (66/103) samples contained detectable levels of R. salmon- inarum cells. Among the positive fish, the R. salmoninarum concentrations ranged from 25 cells/ml to 4.3 3 10 9 cells/ml. A soluble antigenic fraction of R. salmoninarum was detected in 39% of the fish (40/103) by the ELISA. The ELISA is considered one of the most sensitive detection methods for bacterial kidney disease in tissues, yet it did not detect R. salmoninarum antigen consistently at bacterial cell concentrations below about 1.3 3 10 4 cells/ml according to the MF-FAT counts. When total DNA was extracted and tested in a nested PCR designed to amplify a 320-base-pair region of the gene encoding a soluble 57-kD protein of R. salmoni- narum, 100% of the 100 samples tested were positive. The results provided strong evidence that R. salmoni- narum may be present in ovarian fluids thought to be free of the bacterium on the basis of standard diagnostic methods.