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Original Research ARTICLE Provisionally accepted The full-text will be published soon. Notify me

Front. Microbiol. | doi: 10.3389/fmicb.2019.01995

Microtiter Screening Reveals Oxygen-Dependent Antimicrobial Activity of Natural Products Against Mastitis-Causing Bacteria

 Scott A. Ferguson1, Ayana Menorca1, Essie M. Van Zuylen1, Chen-Yi Cheung1, Michelle McConnell1,  David Rennison2,  Margaret A. Brimble2, Kip Bodle3,  Scott McDougall4, Gregory M. Cook1* and  Adam Heikal5*
  • 1University of Otago, New Zealand
  • 2The University of Auckland, New Zealand
  • 3Other, New Zealand
  • 4Other, New Zealand
  • 5University of St Andrews, United Kingdom

In this study we investigated the influence of oxygen availability on a phenotypic microtiter screen to identify new, natural product inhibitors of growth for the bovine mastitis-causing microorganisms; Streptococcus uberis, Staphylococcus aureus, and Escherichia coli. Mastitis is a common disease in dairy cattle worldwide and is a major cause of reduced milk yield and antibiotic usage in dairy herds. Prevention of bovine mastitis commonly relies on the application of teat disinfectants that contain either iodine or chlorhexidine. These compounds are used extensively in human clinical settings and increased tolerance to chlorhexidine has been reported in both Gram-positive and Gram-negative microorganisms. As such new, non-human use alternatives are required for the agricultural industry. Our screening was conducted under normoxic (20% oxygen) and hypoxic (<1% oxygen) conditions to mimic the conditions on teat skin and within the mammary gland respectively, against two natural compound libraries. No compounds inhibited E. coli, under either oxygen condition. Against the Gram-positive microorganisms, 12 inhibitory compounds were identified under normoxic conditions, and 10 under hypoxic conditions. Data revealed a clear oxygen-dependency amongst compounds inhibiting growth, with only partial overlap between oxygen conditions. The oxygen-dependent inhibitory activity of a naturally occurring quinone, beta-lapachone, against
S. uberis was subsequently investigated and we demonstrated that this compound is only active under normoxic conditions with a minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of 32 µM and kills via a reactive oxygen species (ROS)-dependent mechanism as has been demonstrated in other microorganisms. These results demonstrate the importance of considering oxygen-availability in high-throughput inhibitor discovery.

Keywords: Mastitis, Streptococcus uberis, Staphycoccus aureus, antimicrobial, oxygen-dependent, natural product, Beta-lapachone

Received: 09 Apr 2019; Accepted: 14 Aug 2019.

Copyright: © 2019 Ferguson, Menorca, Van Zuylen, Cheung, McConnell, Rennison, Brimble, Bodle, McDougall, Cook and Heikal. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence:
Prof. Gregory M. Cook, University of Otago, Dunedin, 9016, Otago, New Zealand, greg.cook@otago.ac.nz
Dr. Adam Heikal, University of St Andrews, St Andrews, United Kingdom, heikaladamheikal@gmail.com