Polymerase chain reaction detection of Renibacterium salmoninarum in fish: Validation of a modified protocol

A PCR protocol (McIntosh et al., 1996. Appl. Environ. Microbiol. 62, 3929–2932), designed to detect the P57 gene of Renibacterium salmoninarum, causative agent of Bacterial Kidney Disease, was modified slightly by redesign of the forward primer and recommended conditions to prevent possible false positives observed when tested against pure cultures of Yersinia ruckeri. The modified PCR, in combination with an improved DNAzol™-based DNA extraction technique, was very specific and sensitive (detecting between 5 and 72 R. salmoninarum cfu per mg head kidney tissue). It could detect infected fish using only 50 mg head kidney material, making the technique suitable for sampling small fish. It was also shown that samples could be pooled from up to five rainbow trout prior to PCR without noticeably affecting the sensitivity of the assay, providing at least 50 mg head kidney tissue was included from each of the fish that were pooled.