Pseudomonas aeruginosa Biofilm Antibiotic Resistance Gene ndvB Expression Requires the RpoS Stationary-Phase Sigma Factor

Abstract

Chronic, biofilm-based bacterial infections are exceptionally difficult to eradicate due to the high degree of antibiotic recalcitrance exhibited by cells in biofilm communities. In the opportunistic pathogen Pseudomonas aeruginosa, biofilm recalcitrance is multifactorial and arises in part from the preferential expression of resistance genes in biofilms compared to exponential-phase planktonic cells. One such mechanism involves ndvB, which we have previously shown to be expressed specifically in biofilms. In this study, we investigated the regulatory basis of this lifestyle-specific expression by developing an unstable green fluorescent protein (GFP) transcriptional reporter to observe the expression pattern of ndvB We found that in addition to its expression in biofilms, ndvB was upregulated in planktonic cells as they enter stationary phase. The transcription of ndvB in both growth phases was shown to be dependent on the stationary-phase sigma factor RpoS, and mutation of a putative RpoS binding site in the ndvB promoter abolished the activity of the promoter in stationary-phase cells. Overall, we have expanded our understanding of the temporal expression of ndvB in P. aeruginosa and have uncovered a regulatory basis for its growth phase-dependent expression.IMPORTANCE Bacterial biofilms are more resistant to antibiotics than free-living planktonic cells, and understanding the mechanistic basis of this resistance can inform treatments of biofilm-based infections. In addition to chemical and structural barriers that can inhibit antibiotic entry, the upregulation of specific genes in biofilms contributes to the resistance. We investigated this biofilm-specific gene induction by examining expression patterns of ndvB, a gene involved in biofilm resistance of the opportunistic pathogen Pseudomonas aeruginosa We characterized ndvB expression in planktonic and biofilm growth conditions with an unstable green fluorescent protein (GFP) reporter and found that the expression of ndvB in biofilms is dependent on the stationary-phase sigma factor RpoS. Overall, our results support the physiological similarity between biofilms and stationary-phase cells and suggest that the induction of some stationary-phase genes in biofilms may contribute to their increased antibiotic resistance.

Keywords: Pseudomonas aeruginosa; RpoS; antibiotic resistance; biofilms; ndvB; stationary phase.