AUTHOR=Mendez Jessica , Cascales Desirée , Garcia-Torrico Ana I. , Guijarro Jose A. 

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

JOURNAL=Frontiers in Microbiology

VOLUME=9

YEAR=2018

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2018.01098

DOI=10.3389/fmicb.2018.01098

ISSN=1664-302X

ABSTRACT=<p><italic>Yersinia ruckeri</italic> is a bacterium causing fish infection processes at temperatures below the optimum for growth. A derivative <italic>Tn5</italic> transposon was used to construct a library of <italic>Y. ruckeri</italic> mutants with transcriptional fusions between the interrupted genes and the promoterless <italic>luxCDABE</italic> and <italic>lacZY</italic> operons. <italic>In vitro</italic> 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 <italic>in vivo</italic> expression of some of these genes, two of the selected mutants were studied. In one of them, the <italic>acrR</italic> 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 <italic>acrR</italic> mutation was not related to virulence in <italic>Y. ruckeri,</italic> this mutant was useful to analyze <italic>acrR</italic> expression in fish tissues <italic>in vivo</italic>. The other gene studied was <italic>osmY</italic> which is activated under osmotic stress and is involved in virulence. In this case, complemented mutant was used for experiments with fish. <italic>In vivo</italic> analysis of bioluminescence emission by these two strains showed higher values for <italic>acrR</italic> in gut, liver and adipose tissue, whereas <italic>osmY</italic> showed higher luminescence in gut and, at the end of the infection process, in muscle tissue. Similar results were obtained in <italic>ex vivo</italic> assays using rainbow trout tissues. The results indicated that this kind of approach was useful for the identification of genes related to virulence in <italic>Y. ruckeri</italic> and also for the <italic>in vivo</italic> and <italic>in vitro</italic> studies of each of the selected genes.</p>