Adaptation of the neutral bacterial comet assay to assess antimicrobial-mediated DNA double-strand breaks in Escherichia coli

Dipesh Solanky, Shelley Haydel

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

This study aimed to determine the mechanism of action of a natural antibacterial clay mineral mixture, designated as CB, by investigating the induction of DNA double-strand breaks (DSBs) in Escherichia coli. To quantify DNA damage upon exposure to soluble antimicrobial compounds, we modified a bacterial neutral comet assay, which associates the general length of an electrophoresed chromosome, or comet, with the degree of DSB-associated DNA damage. To appropriately account for antimicrobial-mediated strand fragmentation, suitable control reactions consisting of exposures to water, ethanol, kanamycin, and bleomycin were developed and optimized for the assay. Bacterial exposure to the CB clay resulted in significantly longer comet lengths, compared to water and kanamycin exposures, suggesting that the induction of DNA DSBs contributes to the killing activity of this antibacterial clay mineral mixture. The comet assay protocol described herein provides a general technique for evaluating soluble antimicrobial-derived DNA damage and for comparing DNA fragmentation between experimental and control assays.

Original languageEnglish (US)
Pages (from-to)257-261
Number of pages5
JournalJournal of Microbiological Methods
Volume91
Issue number2
DOIs
StatePublished - Nov 2012

Keywords

  • Antibacterial mineral leachate
  • Antibiotic
  • DNA double-strand breaks
  • E. coli
  • Neutral comet assay
  • Single cell gel electrophoresis

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology
  • Microbiology (medical)

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