DEVELOPMENT OF A POLYMERASE-CHAIN-REACTION ASSAY FOR IDENTIFICATION AND DETECTION OF THE FISH PATHOGEN FLAVOBACTERIUM-PSYCHROPHILUM

A PCR-based method was developed to identify and detect Flavobacterium psychrophilum, the causative agent of ''cold-water disease'' and ''ra inbow trout fry syndrome'' in salmonid fish. Two oligonucleotide prime rs were designed by comparing the 16S rRNA sequence of all taxa in the genus Flavobacterium and of representative species in most related ge nera within rRNA superfamily V. Purified chromosomal DNAs from all the se bacterial species, from 25 F. psychrophilum isolates and from sever al other fish-pathogenic bacteria were used to assess the specificity of the reaction. Amplification products were generated only with F. ps ychrophilum DNA. The detection level, equivalent to approximately 10 t o 100 bacterial cells, was increased 10-fold by hybridization with a r adioactive probe. Preliminary experiments demonstrated that this proce dure can also be applied to samples of infected tissue. This PCR assay is therefore a rapid, specific, and sensitive alternative to conventi onal plate culture methods for the identification and detection of F. psychrophilum.