Beyond the E. coli Paradigm: Why Pseudomonas Persistence Follows Its Own Rules

For decades, the study of bacterial persistence has relied heavily on Escherichia coli as the primary model organism. Microbiologists have largely operated under the assumption that the mechanisms governing antibiotic tolerance in E. coli would translate across other Gram-negative species. However, a recent comparative study by Princeton’s Gabrielle Leon and Mark Brynildsen challenges this paradigm, revealing that Pseudomonas aeruginosa exhibits significant divergence in how it survives fluoroquinolone treatment compared to the E. coli workhorse.

The researchers focused on DNA damage repair systems, which are essential because fluoroquinolones like ciprofloxacin kill bacteria by targeting DNA gyrase and topoisomerase IV, leading to double-stranded breaks. In E. coli, the story is consistent: homologous recombination and the induction of the SOS response are used for persister survival regardless of whether the cells are in a growing exponential-phase or a non-growing stationary-phase.

When the team applied the same conditions to P. aeruginosa, using MOPS minimal media with succinate, they found a surprising growth-phase dependency. While homologous recombination (driven by recA and recB) and the SOS response were indeed critical for P. aeruginosa persisters in stationary-phase, these systems were entirely dispensable during the exponential-phase. In growing cultures of P. aeruginosa, mutants lacking these repair systems survived ciprofloxacin treatment just as well as the wild-type. This suggests that exponential-phase P. aeruginosa persisters either utilize unknown repair mechanisms or avoid DNA damage altogether, perhaps through altered target activity or increased efflux.

The study delved into the role of non-homologous end-joining (NHEJ), a repair pathway present in P. aeruginosa but notably absent in E. coli. Despite the presence of Ku and LigD machinery, which are known to repair double-stranded breaks without a template, the researchers found that NHEJ does not contribute to fluoroquinolone persistence in P. aeruginosa in any growth phase. Interestingly, when they attempted to overexpress Ku and LigD using an IPTG-inducible promoter, it proved toxic to the cells. This toxicity suggests that the NHEJ machinery might interfere with normal repair processes, such as homologous recombination, by binding to spontaneous DNA breaks and creating lethal lesions.

These findings are not merely laboratory artifacts; the researchers confirmed that the importance of homologous recombination in stationary-phase holds true across different fluoroquinolones, such as levofloxacin.  It is also important in clinical isolates like MRSN 1612. This isolate, which belongs to a different clade than the common PAO1 lab strain, showed the same selective requirement for recA during stationary-phase persistence.

The clinical implications are significant. P. aeruginosa is a leading cause of hospital-acquired infections and chronic lung infections in cystic fibrosis patients, where persistence is thought to contribute to treatment failure and the subsequent evolution of resistance. While the conservation of repair mechanisms in stationary-phase suggests that targeting the SOS response or homologous recombination could be a viable strategy to eradicate persisters across different species, the divergence in exponential-phase serves as a warning.

Ultimately, this research highlights that we cannot view bacterial persistence as a monolithic phenomenon. The tools a cell uses to repair its genome are not chosen from a universal manual; rather, they depend on the species and its metabolic state. If we think of DNA repair as a toolkit for fixing a broken machine, E. coli keeps the same wrenches on hand whether the engine is idling or racing, while P. aeruginosa seems to swap its entire toolkit depending on the speed of the motor. This complexity underscores the need for species-specific studies to develop effective anti-persister therapies.

Leon G, Brynildsen MP. Conservation and divergence of ciprofloxacin persister survival mechanisms between Pseudomonas aeruginosa and Escherichia coli. PLoS Genet. 2025 Sep 2;21(9):e1011840. doi: 10.1371/journal.pgen.1011840. PMID: 40892829; PMCID: PMC12413089. https://journals.plos.org/plosgenetics/article?id=10.1371/journal.pgen.1011840