The adaptive resistance of Pseudomonas aeruginosa to
aminoglycosides is known to occur during chronic lung
infections in
cystic fibrosis patients in response to nonlethal concentrations of
aminoglycosides. Not only is it difficult to achieve high levels of
drug throughout the dehydrated mucus in the lung, but also steep
oxygen gradients exist across the mucus layer, further reducing the bactericidal activity of
aminoglycosides. In this study, microarray analysis was utilized to examine the gene responses of P. aeruginosa to lethal, inhibitory, and subinhibitory concentrations of
tobramycin under aerobic and anaerobic conditions. While prolonged exposure to subinhibitory concentrations of
tobramycin caused increased levels of expression predominantly of the efflux pump genes mexXY, the greatest increases in gene expression levels in response to lethal concentrations of
tobramycin involved a number of heat shock genes and the PA0779 gene (renamed here asrA), encoding an alternate
Lon protease. Microarray analysis of an asrA::luxCDABE transposon mutant revealed that the induction of heat shock genes in response to
tobramycin in this mutant was significantly decreased compared to that in the parent strain. The level of expression of asrA was induced from an
arabinose-inducible promoter to 35-fold greater than wild-type expression levels in the absence of
tobramycin, and this overexpression alone caused an increased expression of the heat shock genes, as determined by quantitative PCR (qPCR). This overexpression of asrA conferred short-term protection against lethal levels (4 μg/ml) of
tobramycin but did not affect the
tobramycin MIC. The RpoH heat shock
sigma factor was found to be involved in the regulation of asrA in response to both heat shock and
tobramycin at the posttranscriptional level. The results of this work suggest that the
tobramycin concentration has a significant impact on the gene expression of P. aeruginosa, with lethal concentrations resulting in immediate adaptations conferring short-term protection, such as the induction of the heat shock response, and with subinhibitory concentrations leading to more sustainable long-term protection mechanisms, such as increased efflux.