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Abstract
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Flavobacterium psychrophilum is the causative agent of bacterial coldwater disease (BCWD) in North America and rainbow trout fry syndrome in Europe. Seventy-five isolates of F. psychrophilum were collected over a 16-year period from farmed salmonids with tail rot, necrotic myositis, and cephalic/scleral osteochondrosis in Ontario, Canada. The isolates were characterized morphologically, biochemically, serologically, and genotypically. Although the collection of isolates was homogeneous by morphological and serological criteria, two distinct biovars were identified by API-ZYM testing. Previously two genetic lineages had been described: lineage I including type strain F. psychrophilum ATCC 49418 plus 13 strains (avirulent) and lineage II including F. psychrophilum CSF 259-93 plus 19 strains (virulent). Using PCR-restriction fragment length polymorphism assay, the 194 bp ribosomal RNA fragment of F. psychrophilum ATCC 49418 were digested completely by MnlI (MaeIII produced two fait bands) but the virulent strain digested only by MaeIII restriction enzymes. In the current study, a 194-bp PCR product was generated from all Ontario isolates and was digested individually with MaeIII or with MnlI. Analysis of the PCR-RFLP results showed that there are four restriction patterns in the isolates with the correlation between biovar I and digestion with Mae III (lineage II) and biovar II and digestion with Mnl I(lineage I). MnlI digestion produced two fragments of 105 and 62 bp in 16 of 33 isolates belonging to biovar II, but digested only 2 of the PCR products from biovar I isolates. Digestion with MaeIII gave rise to fragments of 128 and 64 bp in 40 of 45 isolates belonging to biovar I whereas only 1 isolate belonging to biovar II had a PCR product that was digested with MaeIII. One isolate of biovar I cut by both, two isolates of biovar I and 16 isolates of biovar II cut by neither of restriction enzymes. The sequence analysis of 194 bp ribosomal RNA fragment led to the concl
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