Development of polymerase chain reaction assays for detection, identification, and differentiation of Piscirickettsia salm onis

A nested polymerase chain reaction (PCR) was developed to detect genomic DNA of Piscirickettsia salmonis, the causative agent of an epizootic disease in salmonids. The nested PCR assay, which used general bacterial 16s rDNA primers in the flrst amplification reaction, and P salmonisspecif~c primers in a second reaction, allowed detection of less than 1 P salmonis tissue culture infect~ous dose 50 (TCID,,). Using the P salmonis-specific primers in a single PCR reaction allowed the detection of 60 TCIDSo, The specificity of the PCR was assessed with a panel of 4 salrnonid and 15 bactenal genornic DNA preparations. Amplification products were produced only with l? salmonis DNA. Restriction fragment length polymorphism (RFLP) analysis of the complete 16s gene PCR products demonstrated that 1 isolate, EM-90, was unique. Two additional primers were developed and used in PCR assays that differentiated EM-90 from the 4 other P salmonis isolates tested.