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The role of lipid a head group modification in Escherichia coli virulence and polymyxin resistance

Moran, Clare (2017) The role of lipid a head group modification in Escherichia coli virulence and polymyxin resistance. Honours thesis, Murdoch University.

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Abstract

The use of natural antimicrobials as antibiotics has divested considerable clinical benefit, with the consequence being that pathogenic species are well-prepared to counteract their effects in new, clinical settings.

When considering antimicrobial resistance, the ecological origins of an antibiotic class can direct the conditions under which resistance is promoted, or constrained. By inducing protective transcriptional regulation, bacteria quickly reconcile with otherwise bactericidal compounds in their local environment. The polymyxin antibiotics, Polymyxin B and Colistin, are highly efficacious against pathogenic Gram-negative bacteria, but are nevertheless overcome by such resistance mechanisms. This has been ascribed to the similarities between polymyxins and endogenous cationic antimicrobial peptides, which must be deflected by commensal bacteria and pathogens alike to survive within a variety of hosts.

To repel cationic antimicrobials, bacteria activate transcriptional pathways that reduce the intrinsic anionic charges of the cell envelope. Polymyxin resistant Klebsiella pneumoniae, and Escherichia coli possess both the arnBCADTEF-ugd and eptA operons, expression of which modifies lipid A with the cationic L-Ara4N and pEtN moieties respectively. Electrostatic repulsion of polymyxins by either moiety alone is sufficient for Pseudomonas aeruginosa and Acinetobacter baumannii to attain therapeutically dangerous levels of polymyxin resistance, raising questions as to why certain Enterobacteriaceae maintain both.

The aim of this study was to investigate non-pathogenic E. coli strain BW25113 and the isogenic single deletion strains BW25113ΔarnT, and BW25113ΔeptA. Each mutant strain lacks the final transferase necessary for modification with either L-Ara4N (ArnT), or pEtN (EptA). By constraining E. coli to a singular modification type, the relative influence each moiety exerts on both polymyxin resistance, and pathogenically relevant phenotypes can be uncovered.

Due to their role in eroding polymyxin efficacy, and enhancing pathogenesis, both ArnT and EptA are potential candidates for antivirulence drug design. Therefore, bioinformatics analyses of both enzymes were conducted between E. coli BW25113 and select Gram-negative pathogens.

Homologous enzymes demonstrated preservation of key sites implicated in catalysis, indicating that inhibitors designed to disrupt activity in E. coli may have a broad-spectrum action. Parallel phenotypic assessment of E. coli ArnT and EptA mutant strains under polymyxin exposure, acidification, and within the J774.A1 macrophage survival model suggests that E. coli can readily compensate for individual deficiencies in lipid A modification. This raises implications for the sole pursuit of either enzyme to conquer burgeoning polymyxin resistance, as the Enterobacteriaceae appear to be “ambidextrous” with regard to their preference for ArnT or EptA.

Publication Type: Thesis (Honours)
Murdoch Affiliation: School of Veterinary and Life Sciences
UNSD Goals: Goal 3: Good Health and Well-being
Supervisor: Coombs, Geoffrey
URI: http://researchrepository.murdoch.edu.au/id/eprint/39883
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