Poster Session - Abstract # 21
Genetic Adaptation to Tobramycin Alters the Antibiotic Susceptibility of Quorum Sensing (LasR)-Null Mutants in Pseudomonas aeruginosa
Brielle M. McKee, Rhea Abisado, Kade Townsend, Kate Woods, Alexandra Holder, Josephine R. Chandler
Department of Molecular Biosciences, University of Kansas, 1200 Sunnyside Ave., Lawrence, KS 66045
The LasR-I quorum-sensing system contributes to Pseudomonas aeruginosa virulence and has been shown to increase resistance to the antibiotic tobramycin. Paradoxically, lasR-null mutations are commonly isolated from chronic infections of patients treated over sustained periods with tobramycin, suggesting there may be some mechanism allowing the lasR mutants to persist under antibiotic selection. We explore the hypothesis that adaptive mutations conferring tobramycin resistance change the role of lasR in tobramycin resistance. We identified that a single-nucleotide mutation in the translation elongation factor gene fusA1 G61A (FusA1A21T) confers a specific advantage to lasR mutants under tobramycin selection, a phenomenon known as sign epistasis. Under tobramycin selection a lasR-null mutation decreases fitness in the ancestral PA14 parent but this same mutation increases fitness in a fusA1 G61A mutant background. This mutation resides in a motif of fusA1 known as the Walker-A P-loop, which binds phosphoryl groups and is essential for the GTPase function of FusA1. This process is also important for activation of the stringent response, which is employed by bacteria during nutrient starvation to slow down translation and activate stress response pathways. We demonstrate that in fusA1 G61A mutants, important stringent response-controlled genes are not activated, and lead to increased oxidative stress in lasR mutants. These results suggest that the fusA1 G61A mutation inhibits activation of stringent response, which may provide a potential explanation for how this adapted mutation alters lasR regulation of tobramycin. Overall, this highlights the importance of adaptation on the evolutionary trajectory of quorum sensing and may explain how quorum sensing-null mutants persist in antibiotic-treated patients. Future experiments are focused on proving the effects of the fusA1 G61A mutation on stringent response and the mechanism by which this effects the regulation of lasR.