Prophages and Growth Dynamics Confound Experimental Results with Antibiotic-Tolerant Persister Cells

mBio. 2017;8(6):e01964-17 DOI 10.1128/mBio.01964-17


Journal Homepage

Journal Title: mBio

ISSN: 2150-7511 (Online)

Publisher: American Society for Microbiology

LCC Subject Category: Science: Microbiology

Country of publisher: United States

Language of fulltext: English

Full-text formats available: PDF, HTML



Alexander Harms
Cinzia Fino
Michael A. Sørensen
Szabolcs Semsey
Kenn Gerdes
Joerg Vogel


Blind peer review

Editorial Board

Instructions for authors

Time From Submission to Publication: 12 weeks


Abstract | Full Text

Bacterial persisters are phenotypic variants that survive antibiotic treatment in a dormant state and can be formed by multiple pathways. We recently proposed that the second messenger (p)ppGpp drives Escherichia coli persister formation through protease Lon and activation of toxin-antitoxin (TA) modules. This model found considerable support among researchers studying persisters but also generated controversy as part of recent debates in the field. In this study, we therefore used our previous work as a model to critically examine common experimental procedures to understand and overcome the inconsistencies often observed between results of different laboratories. Our results show that seemingly simple antibiotic killing assays are very sensitive to variations in culture conditions and bacterial growth phase. Additionally, we found that some assay conditions cause the killing of antibiotic-tolerant persisters via induction of cryptic prophages. Similarly, the inadvertent infection of mutant strains with bacteriophage ϕ80, a notorious laboratory contaminant, apparently caused several of the phenotypes that we reported in our previous studies. We therefore reconstructed all infected mutants and probed the validity of our model of persister formation in a refined assay setup that uses robust culture conditions and unravels the dynamics of persister cells through all bacterial growth stages. Our results confirm the importance of (p)ppGpp and Lon but no longer support a role of TA modules in E. coli persister formation under unstressed conditions. We anticipate that the results and approaches reported in our study will lay the ground for future work in the field.