Poster Presentation BacPath 13: Molecular Analysis of Bacterial Pathogens Conference 2015

Genetic determinants of β-lactam tolerance via reversible cell morphotype transitions in Pseudomonas aeruginosa (#127)

Leigh G. Monahan 1 , Ibrahim Awadh 1 , Lynne Turnbull 1 , Ian G. Charles 1 , Cynthia B. Whitchurch 1
  1. The ithree Institute, University of Technology Sydney, Ultimo, NSW, Australia

Pseudomonas aeruginosa, a major human pathogen, is able to tolerate β-lactam antibiotics at high concentrations. Despite inhibiting the growth of the organism, these cell wall-targeting drugs exhibit little bactericidal activity and this has important implications for the treatment of P. aeruginosa infections. We recently showed that P. aeruginosa tolerates exposure to β-lactams by undergoing a rapid, population-wide transition from the normal bacillary form to a cell wall deficient (CWD) spherical morphotype. These β-lactam-induced spherical cells remain viable in the presence of the drug, and rapidly convert back to normal bacillary cells when it is removed or degraded. They can then resume normal growth and proliferation.

Importantly, we found that CWD cells of P. aeruginosa are highly susceptible to killing by other compounds, in particular antimicrobial peptides that target the exposed cytoplasmic membrane in the CWD form but show little to no activity against bacillary cells. Drug combinations that induce CWD cell formation en masse and then specifically target the CWD morphotype could serve as an effective new strategy for treating P. aeruginosa infections.

To investigate the molecular mechanisms underlying β-lactam tolerance in P. aeruginosa, we have screened a comprehensive nonredundant mutant library for mutants that are unable to survive or recover from β-lactam (meropenem) treatment through reversible transition to the CWD morphotype. Extensive characterisation of defective mutants revealed a total of 74 genuine meropenem tolerance genes in P. aeruginosa, each of which have a specific role either in CWD cell formation, survival or reversion after antibiotic treatment. Meropenem tolerance genes cluster into several distinct functional categories, including cell wall degradation, membrane transport and biotin synthesis. Not only does this work provide the first molecular insights into β-lactam tolerance in P. aeruginosa, it could serve as a starting point for drug development efforts to selectively target the CWD morphotype.