Pseudomonas aeruginosa Citrate Synthase GltA Influences Antibiotic Tolerance and the Type III Secretion System through the Stringent Response

Rising antimicrobial resistance imposes a severe threat to human health. It is urgent to develop novel antimicrobial strategies by understanding bacterial regulation of virulence and antimicrobial resistance determinants. Carbohydrate metabolism plays essential roles in energy generation and providing carbon skeletons for amino acid syntheses. In addition, carbohydrate metabolism has been shown to influence bacterial susceptibility to antibiotics and virulence. In this study, we demonstrate that citrate synthase gltA mutation can increase the expression of the type III secretion system (T3SS) genes and antibiotic tolerance in Pseudomonas aeruginosa. The stringent response is activated in the gltA mutant, and deletion of the (p)ppGpp synthetase gene relA restores the antibiotic tolerance and expression of the T3SS genes to wild-type level. We further demonstrate that the intracellular level of cAMP is increased by the stringent response in the gltA mutant, which increases the expression of the T3SS master regulator gene exsA. Overall, our results reveal an essential role of GltA in metabolism, antibiotic tolerance, and virulence, as well as a novel regulatory mechanism of the stringent response-mediated regulation of the T3SS in P. aeruginosa.IMPORTANCE Rising antimicrobial resistance imposes a severe threat to human health. It is urgent to develop novel antimicrobial strategies by understanding bacterial regulation of virulence and antimicrobial resistance determinants. The stringent response plays an essential role in virulence and antibiotic tolerance. Pseudomonas aeruginosa is an opportunistic pathogen that causes acute and chronic infections in humans. The bacterium produces an arsenal of virulence factors and is highly resistant to a variety of antibiotics. In this study, we provide evidence that citrate synthase GltA plays a critical role in P. aeruginosa metabolism and influences the antibiotic tolerance and virulence. We further reveal a role of the stringent response in the regulation of the antibiotic tolerance and virulence. The significance of this work is in elucidation of novel regulatory pathways that control both antibiotic tolerance and virulence in P. aeruginosa.

Automated summary

Rising antimicrobial resistance poses a severe threat to human health, and understanding bacterial regulation of virulence and antimicrobial resistance determinants is urgently needed for developing new antimicrobial strategies. This study demonstrates that a mutation in citrate synthase gltA increases the expression of type III secretion system (T3SS) genes and antibiotic tolerance in Pseudomonas aeruginosa. The stringent response is activated in the gltA mutant, and restoring the antibiotic tolerance and T3SS gene expression to wild-type level requires deletion of the (p)ppGpp synthetase gene relA. Furthermore, increased intracellular cAMP level in the gltA mutant due to the stringent response enhances the expression of the T3SS master regulator gene exsA. This work reveals the important roles of GltA in metabolism, antibiotic tolerance, and virulence, as well as a novel regulatory mechanism of the stringent response-mediated regulation of the T3SS in P. aeruginosa.