Interplay between Antibiotic Efficacy and Drug-Induced Lysis Underlies Enhanced Biofilm Formation at Subinhibitory Drug Concentrations.

Wen Yu,Kelsey M Hallinen,Kevin B Wood

doi:10.1128/aac.01603-17

Wen Yu, Kelsey M Hallinen + Show 1 more

Open Access

https://doi.org/10.1128/aac.01603-17

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Abstract

ABSTRACTSubinhibitory concentrations of antibiotics have been shown to enhance biofilm formation in multiple bacterial species. While antibiotic exposure has been associated with modulated expression of many biofilm-related genes, the mechanisms of drug-induced biofilm formation remain a focus of ongoing research efforts and may vary significantly across species. In this work, we investigate antibiotic-induced biofilm formation in Enterococcus faecalis, a leading cause of nosocomial infections. We show that biofilm formation is enhanced by subinhibitory concentrations of cell wall synthesis inhibitors but not by inhibitors of protein, DNA, folic acid, or RNA synthesis. Furthermore, enhanced biofilm is associated with increased cell lysis, increases in extracellular DNA (eDNA) levels, and increases in the density of living cells in the biofilm. In addition, we observe similar enhancement of biofilm formation when cells are treated with nonantibiotic surfactants that induce cell lysis. These findings suggest that antibiotic-induced biofilm formation is governed by a trade-off between drug toxicity and the beneficial effects of cell lysis. To understand this trade-off, we developed a simple mathematical model that predicts changes in antibiotic-induced biofilm formation due to external perturbations, and we verified these predictions experimentally. Specifically, we demonstrate that perturbations that reduce eDNA (DNase treatment) or decrease the number of living cells in the planktonic phase (a second antibiotic) decrease biofilm induction, while chemical inhibitors of cell lysis increase relative biofilm induction and shift the peak to higher antibiotic concentrations. Overall, our results offer experimental evidence linking cell wall synthesis inhibitors, cell lysis, increased eDNA levels, and biofilm formation in E. faecalis while also providing a predictive quantitative model that sheds light on the interplay between cell lysis and antibiotic efficacy in developing biofilms.

Highlights

Subinhibitory concentrations of antibiotics have been shown to enhance biofilm formation in multiple bacterial species
We find that subinhibitory concentrations of cell wall synthesis inhibitors, but not other drug classes, promote biofilm formation associated with increased cell lysis and increased extracellular DNA (eDNA) and extracellular RNA levels
Our work demonstrates that biofilm formation in E. faecalis is enhanced by subinhibitory concentrations of cell wall synthesis inhibitors but not by inhibitors of protein, DNA, folic acid, or RNA synthesis

Summary

Introduction

Subinhibitory concentrations of antibiotics have been shown to enhance biofilm formation in multiple bacterial species. A recent study in Staphylococcus aureus showed that ␤-lactams administered at subinhibitory concentrations promoted biofilm formation and induced eDNA release in an autolysindependent manner [27] Taken together, these results suggest that, for some drugs, biofilm induction hinges on a balance between the inhibitory effects of antibiotics, which reduce biofilm formation at sufficiently high concentrations, and the potential of antibiotic-induced cell lysis to promote biofilm formation, presumably through release of eDNA. These results suggest that, for some drugs, biofilm induction hinges on a balance between the inhibitory effects of antibiotics, which reduce biofilm formation at sufficiently high concentrations, and the potential of antibiotic-induced cell lysis to promote biofilm formation, presumably through release of eDNA We investigate this balance in E. faecalis biofilms exposed to multiple classes of antibiotics. Our results suggest that inhibitors of cell wall synthesis promote biofilm formation via increased cell lysis, and they offer a quantitative predictive framework for understanding the trade-offs between drug toxicity and lysis-induced biofilm induction

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