Role of Bacterial Lipocalin Proteins in Antimicrobial Resistance

Abstract

Burkholderia cenocepacia, an opportunistic Gram-negative bacterium that causes serious respiratory infections in patients with cystic fibrosis, produces the extracellular bacterial lipocalin protein BcnA upon exposure to
sublethal concentrations of bactericidal antibiotics. BcnA captures a range of antibiotics outside bacterial cells, providing a global extracellular mechanism of antimicrobial resistance. In this thesis, I report that watersoluble
and liposoluble forms of vitamin E inhibit antibiotic binding by BcnA.
In vitro, both vitamin E forms bind strongly to BcnA and contribute to reduce the MICs of antibiotics. Expression of BcnA was required for the adjuvant effect of vitamin E. In vivo, vitamin E and norfloxacin treatment significantly increased Galleria mellonella larva survival upon infection in a BcnAdependent
manner. Together, my findings suggest that vitamin E can be used to increase killing by bactericidal antibiotics through interference with
lipocalin binding.

The transcription of bcnA gene increases upon antibiotic stress, which stimulates the oxidative stress and membrane lipid peroxidation. BcnA lipocalin, and associated proteins BcoA cytochrome and BarA reductase are involved in membrane peroxidation stress response to antibiotics and other forms of oxidative stress. I report that bcnA, bcoA and barA deletion mutants
display enhanced membrane lipid peroxidation and fail to survive under conditions that stimulate peroxidative stress (e.g. Tellurite treatment, cold stress and salt stress). Peroxidation also affects the functionality of the bacterial outer membrane. Absence of BcnA compromises the permeability of the outer membrane, resulting in the release of extracellular DNA leading to a depletion aggregation phenotype and cell death. Together, my findings uncover a novel peroxidation quenching mechanism based on B. cenocepacia BcnA, BcoA and BarA, which protects the bacterial cell envelope against lipid peroxidation stimulated by antibiotic or metal stress, in the presence of oxygen.

Date of Award	Dec 2019
Original language	English
Awarding Institution	Queen's University Belfast
Supervisor	Miguel A. Valvano (Supervisor)