H-044. A Pseudomonas aeruginosa oxyR Mutant Requires Catalase (KatB) for Protection against H2O2 in Planktonic Culture While It Requires Alkyl Hydroperoxide Reductase (AhpCF) in Biofilms

W. Panmanee, D. J. Hassett;
Univ. of Cincinnati Coll. of Med., Cincinnati, OH.

Pseudomonas aeruginosa is an opportunistic pathogen and a leading cause of morbidity and mortality in cystic fibrosis (CF) patients. This is, in part, due to the formation of highly recalcitrant, antibiotic-resistant biofilms in their airways. A P. aeruginosa mutant lacking the H2O2-responsive transactivator, OxyR, is extremely sensitive to H2O2. OxyR regulates transcription of genes encoding the protective enzymes AhpB , KatB-AnkB, and AhpCF against H2O2 and organic peroxides, respectively. Predictably, the oxyR mutant in planktonic cells is very susceptible to H2O2 and more resistant in biofilm culture. In this study, we demonstrate that KatB plays a major role in protection of an oxyR mutant in planktonic culture when treated with H2O2, while AhpCF is more in protecting oxyR mutant biofilms. AhpCF afforded such organisms greater protection during biofilm culture. Interestingly, the periplasmic AhpB provided no such protection in either plantonic or biofilm cells. The total catalase activity of planktonic bacteria was lower in oxyR bacteria harboring AhpB but higher and no different in cells expressing AhpCF or KatB when compared to oxyR mutant bacteria. However, catalase in biofilm-grown cells was no greater in oxyR mutant organisms containing AhpB or AhpCF but slightly lower in KatB-containing strains. We also found that transcription of ahpB, ahpCF and katB genes respond to 10 mM H2O2 in plantonic stationary phase cell, resembling the metabolic state of biofilms. However, the response in biofilms was dose-dependent. We discovered that ahpB transcription was activated at low concentration (~5 mM) while the ahpCF and katB response to H2O2 was at higher concentration (~25 mM). Therefore, unlike previously thought, we conclude that catalase may not play an important role in the protection of oxyR biofilms from exposure to H2O2, but the cytoplasmic enzyme, AhpCF, does.

137/H. Gene Expression II: Regulatory Networks

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