Comparison of Two Transmission Electron Microscopy Methods to Visualize Drug-Induced Alterations of Gram-Negative Bacterial Morphology
Hang Thi Nguyen,
Lisa A. O’Donovan,
Henrietta Venter,
Cecilia C. Russell,
Adam McCluskey,
Stephen W. Page,
Darren J. Trott,
Abiodun D. Ogunniyi
Affiliations
Hang Thi Nguyen
Australia Centre for Antimicrobial Resistance Ecology, School of Animal and Veterinary Sciences, Roseworthy Campus, The University of Adelaide, Roseworthy, SA 5371, Australia
Lisa A. O’Donovan
ARC Centre of Excellence in Plant Energy Biology, School of Agriculture Food & Wine, Waite Campus, The University of Adelaide, Urrbrae, SA 5064, Australia
Henrietta Venter
Health and Biomedical Innovation, Clinical and Health Sciences, University of South Australia, Adelaide, SA 5000, Australia
Cecilia C. Russell
Chemistry School of Environmental and Life Sciences, University of Newcastle, Callaghan, NSW 2308, Australia
Adam McCluskey
Chemistry School of Environmental and Life Sciences, University of Newcastle, Callaghan, NSW 2308, Australia
Stephen W. Page
Neoculi Pty Ltd., Burwood, VIC 3125, Australia
Darren J. Trott
Australia Centre for Antimicrobial Resistance Ecology, School of Animal and Veterinary Sciences, Roseworthy Campus, The University of Adelaide, Roseworthy, SA 5371, Australia
Abiodun D. Ogunniyi
Australia Centre for Antimicrobial Resistance Ecology, School of Animal and Veterinary Sciences, Roseworthy Campus, The University of Adelaide, Roseworthy, SA 5371, Australia
In this study, we optimized and compared different transmission electron microscopy (TEM) methods to visualize changes to Gram-negative bacterial morphology induced by treatment with a robenidine analogue (NCL195) and colistin combination. Aldehyde-fixed bacterial cells (untreated, treated with colistin or NCL195 + colistin) were prepared using conventional TEM methods and compared with ultrathin Tokuyasu cryo-sections. The results of this study indicate superiority of ultrathin cryo-sections in visualizing the membrane ultrastructure of Escherichia coli and Pseudomonas aeruginosa, with a clear delineation of the outer and inner membrane as well as the peptidoglycan layer. We suggest that the use of ultrathin cryo-sectioning can be used to better visualize and understand drug interaction mechanisms on the bacterial cell membrane.
