Flavobacterium psychrophilum is a devastating bacterial pathogen of salmonids reared in freshwater worldwide. So far, serological diversity between isolates has been described but the underlying molecular factors remain unknown. By combining complete genome sequence analysis and the serotyping method proposed by Lorenzen and Olesen (1997) for a set of 34 strains, we identified key molecular determinants of the serotypes. This knowledge allowed us to develop a robust multiplex PCR-based serotyping scheme, which was applied to 244 bacterial isolates. The results revealed a striking association between PCR-serotype and fish host species and illustrate the use of this approach as a simple and cost-effective method for the determination of F. psychrophilum serogroups. PCR-based serotyping could be a useful tool in a range of applications such as disease surveillance, selection of salmonids for bacterial coldwater disease resistance and future vaccine formulation.

Flavobacterium psychrophilum is the causative agent of "bacterial cold water disease" and "rainbow trout fry syndrome" in salmonid farming worldwide. These diseases, especially rainbow trout fry syndrome, are among the main hazards for French aquaculture. In this study, a multilocus sequence typing approach (MLST) was used to evaluate the genetic diversity of this bacterium. Seven housekeeping genes in a set of 66 isolates were investigated. They were recently collected from rainbow trout during clinical episodes in French farms from the two main geographical areas of production. A total of 5808 bp of sequence were analyzed for each isolate and showed relatively low levels of gene (H=0.4313) and nucleotide (π×100=0.31%) diversities. MLST identified 15 sequence types (STs), of which 14 have never been described. eBURST analysis separated the 15 STs in one clonal complex of 8 genetically related STs (with ST2 as founder) and 7 singletons. Genetic diversity was largely due to recombination, as demonstrated by a pairwise homoplasy index (PHI=5.35×10(-9)) significantly different from zero (p<0.05). The evolution of standardized association index (I(A)(S)) (all isolates: 0.6088, p<0.05; single representative of STs: 0.4567, p<0.05; and clusters of STs: 0.084, p>0.05), showed an epidemic structure of the population. These results emphasized the expansion of a limited number of dominant genetic variants in French clinical F. psychrophilum isolates from a single host species, with no geographic relationships.

Yersinia ruckeri is the causative agent of yersiniosis, a disease reported in a number of fish species, especially rainbow trout. This study was undertaken to describe the phenotypes of Y. ruckeri on French rainbow trout farms. More than 100 isolates, collected during recent outbreaks on trout farms, were characterized by phenotypic tests, namely using biochemical tests of the API 20E system, serotyping, biotyping (tests for motility and lipase activity) and by describing the pattern of susceptibility to several antibiotics. The isolates showed a low phenotypic diversity with a prevalent serotype (O1) and API 20E profile 5 1(3)07 100. As in other European countries, Biotype 2 (BT2), which lacks both motility and secreted lipase activity, was found to be present in France. The emergence of 'French' BT2 was different than that observed for other European countries (Finland, Spain, Denmark and the UK). The antibiotic pattern was uniform for all isolates, regardless of the geographical area studied. The results indicate that no resistance has yet emerged, and the efficacy of the antibiotic generally used against yersiniosis in France, trimethoprim/sulfamethoxasol, is not compromised (minimum inhibitory concentrations [MIC] of between 0.016 and 0.128 µg ml-1). Enrofloxacin and doxycycline, not used as a first-line treatment in fish diseases, have reasonably good efficacies (with MICs ≤0.128 and 0.256, respectively).

Yersinia ruckeri is a pathogen that has an impact on aquaculture worldwide. The disease caused by this bacterial species, yersiniosis or redmouth disease, generates substantial economic losses due to the associated mortality and veterinary costs. For predicting outbreaks and improving control strategies, it is important to characterize the population structure of the bacteria. The phenotypic and genetic homogeneities described previously indicate a clonal population structure as observed in other fish bacteria. In this study, the pulsed-field gel electrophoresis (PFGE) and multi locus sequence typing (MLST) methods were used to describe a population of isolates from outbreaks on French fish farms. For the PFGE analysis, two enzymes (NotI and AscI) were used separately and together. Results from combining the enzymes showed the great homogeneity of the outbreak population with a similarity > 80.0% but a high variability within the cluster (cut-off value = 80.0%) with a total of 43 pulsotypes described and an index of diversity = 0.93. The dominant pulsotypes described with NotI (PtN4 and PtN7) have already been described in other European countries (Finland, Germany, Denmark, Spain and Italy). The MLST approach showed two dominant sequence types (ST31 and ST36), an epidemic structure of the French Y. ruckeri population and a preferentially clonal evolution for rainbow trout isolates. Our results point to multiple types of selection pressure on the Y. ruckeri population attributable to geographical origin, ecological niche specialization and movements of farmed fish.