Examination of cationic antimicrobial tolerance in Escherichia coli to identify phenotypic and genotypic adaptations
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Date
2019
Authors
Cartwright, Nicola Helen
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Abstract
Cationic antimicrobial (CA) agents describe a variety of positively charged antimicrobials that are widely used in many clinical, agricultural and industrial facilities to disinfect and prevent microbial growth. Increased tolerance to CAs by Gram-negative bacteria is a growing problem because CA tolerant bacteria frequently confer therapeutic antimicrobial cross-resistance. Previous studies have shown that CA tolerant bacteria frequently exhibit alterations in lipid modification pathways, up-regulation of efflux pumps and porins as a mechanism of tolerance but these changes have yet to be consistently identified in experiments containing the same species or strain exposed to different CAs. In this thesis, E. coli K12 was adapted to increasing concentrations of CAs, specifically, benzalkonium chloride, cetrimide bromide, chlorhexidine hydrochloride and colistin sulphate that belong to different antimicrobial classes to determine phenotypic and genotypic changes over 20-40 sub-cultures. It was revealed that CAs belonging to similar classes had similar growth phenotypes, antimicrobial cross-resistance and genotypic alterations. Experiments exploring the stability of CA-tolerant phenotypes when CA selection is removed over a 10-day period among revealed a dependence on previous CA exposure. Genotypic analysis involved identification of repeatedly identified single nucleotide variants (SNVs) in lipopolysaccharide biosynthesis pathways, antimicrobial transcriptional regulators, transposable elements, and to a lesser extent in efflux pump genes. This study suggests that CA adaptation may be dependent upon how each CA specifically disrupts the cell membrane, since each CA disrupts the membrane at potentially different outer membrane targets. It also reveals new insights and genetic markers associated with CA tolerance.
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Keywords
Cationic, Antimicrobial resistance, Escherichia coli