OmpK36-mediated carbapenem resistance in Klebsiella pneumoniae
File(s)
Author(s)
Wong, Joshua Liang Chao
Type
Thesis
Abstract
Klebsiella pneumoniae (KP) is a common nosocomial pathogen that is frequently responsible for pneumonia, urinary tract infections and bacteraemia in patients with long term health challenges. Such infections can progress to sepsis, often necessitating intensive care support. Antibiotics are the cornerstone of treatment, however, mounting resistance to a current last-line class, the carbapenems, threatens to render some infections untreatable. Robust carbapenem resistance is achieved by two divergent but synergistic mechanisms in high-risk, globally disseminated, resistant sequence types (ST), such as KP ST258. Firstly, plasmid-borne carbapenemase enzymes hydrolytically inactivate carbapenems in the bacterial periplasm. This is complemented by chromosomal mutations in two outer membrane (OM) porin genes, ompK35 and ompK36, that reduce drug diffusion into the periplasm. Whilst truncations in OmpK35 are well described widespread phenomena, loss of OmpK36 is rarely observed in clinical isolates. In this thesis I focus on identifying and characterising the precise molecular mechanisms by which mutations in the dominant KP porin, OmpK36, impair carbapenem diffusion. I start by focusing on OmpK36 pore-constriction and identify a di-amino acid Glycine-Aspartate insertion in the pore diameter-determining motif, loop 3. This structural modification reduces carbapenem diffusion and increases the minimum inhibitory concentration. When the fitness cost of OmpK36 pore-constriction is tested, in a novel murine ventilator-associated pneumonia model, I identify a fitness disadvantage only when tested in competition with wild-type strains. This contrasts with a recurrently emerging novel ompK36 mutation, a 25c>t, identified in a large KP ST258 genome collection. The 25c>t depletes OmpK36 from the OM by inducing an inhibitory mRNA secondary structure and leads to significant attenuation in vivo. Finally, bringing this together, when meropenem treatment is instituted in this translational model, both mutations are demonstrated to be advantageous to KP and result in treatment failure, whereas strains expressing wild-type OmpK36 are susceptible to therapy.
Version
Open Access
Date Issued
2022-05
Date Awarded
2023-01
Copyright Statement
Creative Commons Attribution NonCommercial Licence
License URL
Advisor
Frankel, Gad
Clements, Abigail
Brett, Stephen
Sponsor
Medical Research Council (Great Britain)
Grant Number
MR/R502376/1
Publisher Department
Department of Life Sciences
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)