The Infection Process of Yersinia ruckeri: Reviewing the Pieces of the Jigsaw Puzzle.

Finding the keys to understanding the infectious process of Yersinia ruckeri was not a priority for many years due to the prompt development of an effective biotype 1 vaccine which was used mainly in Europe and USA. However, the gradual emergence of outbreaks in vaccinated fish, which have been reported since 2003, has awakened interest in the mechanism of virulence in this pathogen. Thus, during the last two decades, a large number of studies have considerably enriched our knowledge of many aspects of the pathogen and its interaction with the host. By means of both conventional and a variety of novel strategies, such as cell GFP tagging, bioluminescence imaging and optical projection tomography, it has been possible to determine three putative Y. ruckeri infection routes, the main point of entry for the bacterium being the gill lamellae. Moreover, a wide range of potential virulence factors have been highlighted by specific gene mutagenesis strategies or genome-wide transposon/plasmid insertion-based screening approaches, such us in vivo expression technology (IVET) and signature tagged mutagenesis (STM). Finally, recent proteomic and whole genomic analyses have allowed many of the genes and systems that are potentially implicated in the organism's pathogenicity and its adaptation to the host environmental conditions to be elucidated. Altogether, these studies contribute to a better understanding of the infectious process of Y. ruckeri in fish, which is crucial for the development of more effective strategies for preventing or treating enteric redmouth disease (ERM).

Temperature-Dependent Gene Expression in Yersinia ruckeri: Tracking Specific Genes by Bioluminescence During in Vivo Colonization.

Yersinia ruckeri is a bacterium causing fish infection processes at temperatures below the optimum for growth. A derivative Tn5 transposon was used to construct a library of Y. ruckeri mutants with transcriptional fusions between the interrupted genes and the promoterless luxCDABE and lacZY operons. In vitro analysis of ß-galactosidase activity allowed the identification of 168 clones having higher expression at 18°C than at 28°C. Among the interrupted genes a SAM-dependent methyltransferase, a diguanylated cyclase, three genes involved in legionaminic acid synthesis and three transcriptional regulators were defined. In order to determine, via bioluminescence emission, the in vivo expression of some of these genes, two of the selected mutants were studied. In one of them, the acrR gene coding a repressor involved in regulation of the AcrAB-TolC expulsion pump was interrupted. This mutant was found to be highly resistant to compounds such as chloramphenicol, tetracycline, and ciprofloxacin. Although acrR mutation was not related to virulence in Y. ruckeri, this mutant was useful to analyze acrR expression in fish tissues in vivo. The other gene studied was osmY which is activated under osmotic stress and is involved in virulence. In this case, complemented mutant was used for experiments with fish. In vivo analysis of bioluminescence emission by these two strains showed higher values for acrR in gut, liver and adipose tissue, whereas osmY showed higher luminescence in gut and, at the end of the infection process, in muscle tissue. Similar results were obtained in ex vivo assays using rainbow trout tissues. The results indicated that this kind of approach was useful for the identification of genes related to virulence in Y. ruckeri and also for the in vivo and in vitro studies of each of the selected genes.

Comparative genome analysis reveals important genetic differences among serotype O1 and serotype O2 strains of Y. ruckeri and provides insights into host adaptation and virulence.

Despite the existence of a commercial vaccine routinely used to protect salmonids against Yersinia ruckeri, outbreaks still occur, mainly caused by nonmotile and lipase-negative strains (serotype O1 biotype 2). Moreover, epizootics caused by other uncommon serotypes have also been reported. At the moment, one of the main concerns for the aquaculture industry is the expanding range of hosts of this pathogen and the emergence of new biotypes and serotypes causing mortality in fish farms and against which the vaccine cannot protect. The comparative analysis of the genome sequences of five Y. ruckeri strains (150, CSF007-82, ATCC29473, Big Creek 74, and SC09) isolated from different hosts and classified into different serotypes revealed important genetic differences between the genomes analyzed. Thus, a clear genetic differentiation was found between serotype O1 and O2 strains. The presence of 99 unique genes in Big Creek 74 and 261 in SC09 could explain the adaptation of these strains to salmon and catfish, respectively. Finally, the absence of 21 genes in ATCC29473 which are present in the other four virulent strains could underpin the attenuation described for this strain. The study reveals important genetic differences among the genomes analyzed. Further investigation of the genes highlighted in this study could provide insights into the understanding of the virulence and niche adaptive mechanisms of Y. ruckeri.

Genome Sequence of the Fish Pathogen Yersinia ruckeri Strain 150, Isolated from Diseased Rainbow Trout.

We present here the draft genome of a pathogenic Yersinia ruckeri strain, isolated from rainbow trout (Oncorhynchus mykiss) affected by enteric redmouth disease. The chromosome has 3,826,775 bp, a GC content of 46.88%, and is predicted to contain 3,538 coding sequences. The data will be useful for comparative pathogenicity studies.

Temperature-dependent expression of virulence genes in fish-pathogenic bacteria.

Virulence gene expression in pathogenic bacteria is modulated by environmental parameters. A key factor in this expression is temperature. Its effect on virulence gene expression in bacteria infecting warm-blooded hosts is well documented. Transcription of virulence genes in these bacteria is induced upon a shift from low environmental to a higher host temperature (37°C). Interestingly, host temperatures usually correspond to the optimum for growth of these pathogenic bacteria. On the contrary, in ectothermic hosts such as fish, molluscs, and amphibians, infection processes generally occur at a temperature lower than that for the optimal growth of the bacteria. Therefore, regulation of virulence gene expression in response to temperature shift has to be modulated in a different way to that which is found in bacteria infecting warm-blooded hosts. The current understanding of virulence gene expression and its regulation in response to temperature in fish-pathogenic bacteria is limited, but constant extension of our knowledge base is essential to enable a rational approach to the problem of the bacterial fish diseases affecting the aquaculture industry. This is an interesting issue and progress needs to be made in order to diminish the economic losses caused by these diseases. The intention of this review is, for the first time, to compile the scattered results existing in the field in order to lay the groundwork for future research. This article is an overview of those relevant virulence genes that are expressed at temperatures lower than that for optimal bacterial growth in different fish-pathogenic bacteria as well as the principal mechanisms that could be involved in their regulation.