The Crohn's disease-associated Escherichia coli strain LF82 relies on SOS and stringent responses to survive, multiply and tolerate antibiotics within macrophages.

Gaëlle Demarre,Silvia De Monte,Victoria Prudent,Guy Tran Van Nhieu,Hanna Schenk,Sylvie Rimsky,Olivier Espéli,Emilie Rousseau,Nicolas Barnich,Marie-Agnès Bringer

doi:10.1371/journal.ppat.1008123

Abstract

Adherent Invasive Escherichia coli (AIEC) strains recovered from Crohn's disease lesions survive and multiply within macrophages. A reference strain for this pathovar, AIEC LF82, forms microcolonies within phagolysosomes, an environment that prevents commensal E. coli multiplication. Little is known about the LF82 intracellular growth status, and signals leading to macrophage intra-vacuolar multiplication. We used single-cell analysis, genetic dissection and mathematical models to monitor the growth status and cell cycle regulation of intracellular LF82. We found that within macrophages, bacteria may replicate or undergo non-growing phenotypic switches. This switch results from stringent response firing immediately after uptake by macrophages or at later stages, following genotoxic damage and SOS induction during intracellular replication. Importantly, non-growers resist treatment with various antibiotics. Thus, intracellular challenges induce AIEC LF82 phenotypic heterogeneity and non-growing bacteria that could provide a reservoir for antibiotic-tolerant bacteria responsible for relapsing infections.

Highlights

Adherent Invasive Escherichia coli (AIEC) strains recovered from Crohn’s disease (CD) lesions are able to adhere to and invade cultured intestinal epithelial cells and to survive and multiply within macrophages [1,2]
We demonstrated that growth of AIEC LF82 inside the toxic mature phagolysosomes of macrophages requires sequential adaptations and phenotype
We observed that this method slightly underestimates dead bacteria inside macrophages because of a weak propidium iodide (PI) labeling (S2A Fig)

Summary

Introduction

Adherent Invasive Escherichia coli (AIEC) strains recovered from Crohn’s disease (CD) lesions are able to adhere to and invade cultured intestinal epithelial cells and to survive and multiply within macrophages [1,2]. Previous work performed with murine macrophage cell lines has revealed that the prototype AIEC strain LF82, multiplies in a vacuole presenting the characteristics of a mature phagolysosome [5,6]. In such an environment, AIEC should encounter acidic, oxidative, genotoxic and proteic stresses. Screening of genes involved in LF82 fitness within macrophage has revealed that HtrA, DsbA, or Fis proteins are required for optimum fitness, [7,8,9] These observations confirmed that LF82 encounter stresses in the phagolysosomes. The impact of these stresses on the survival and growth of LF82 inside phagolysomes has not yet been investigated

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Discussion

Conclusion