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

Abstract

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 β-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.

Transcriptome changes in response to temperature in the fish pathogen Photobacterium damselae subsp. damselae: Clues to understand the emergence of disease outbreaks at increased seawater temperatures

“…TolC and AcrAB form a tripartite multidrug and toxin secretion efflux pump. Expression of tolC and acrAB in the fish pathogen Yersinia ruckery is increased at 28°C with respect to 18°C with a concomitant increased resistance to antibiotics and toxic substances such as acriflavine [59]. The role of this system in the biology of Pdd remains unknown.…”

The repeat structure of two paralogous genes, Yersinia ruckeri invasin (yrInv) and a “Y. ruckeri invasin-like molecule”, (yrIlm) sheds light on the evolution of adhesive capacities of a fish pathogen

“…Many Y. ruckeri genes related to virulence were reported to be upregulated at 18°C, similarly as in other fish pathogens (Mendez et al, 2018). These include, among others, genes involved in Osmolarity is an important factor that can influence the expression of virulence genes.…”

“…A recent study showed a practical approach for identification of virulence . All these genes were previously implicated in virulence (Mendez et al, 2018).…”

“…Molecular approaches include the in vivo expression approach (IVET) and signature-tagged mutagenesis (STM) (Dahiya and Stevenson, 2010a;Mendez et al, 2018). The IVET is based on identification of genes that are expressed in vivo in the host during the infection process.…”

Temperature Restriction in Entomopathogenic Bacteria

“…Y. ruckeri is the causative agent of enteric red mouth disease in salmonid fish species (Kumar et al, 2015) and its optimum growth temperature is also 28 • C, while its optimum temperature for infectivity is 18 • C. Similarly, V. anguillarum has an optimum growth temperature of 25 • C whilst the optimum temperature for vibriosis disease is 15 • C (Lages et al, 2019). It appears that a lower temperature optimum for virulence is common in fish pathogens and virulence genes are found to be up-regulated at these lower temperatures (Guijarro et al, 2015;Mendez et al, 2018;Lages et al, 2019). The association of TRL operons with pathogens of poikilothermic hosts (e.g., fish) tempts speculation there is an as yet unknown advantage to this mechanism in such niches.…”