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Antimicrobial Agents and Chemotherapy, December 2009, p. 4979-4986, Vol. 53, No. 12
0066-4804/09/$08.00+0 doi:10.1128/AAC.00497-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Atomic Force Microscopy Investigation of the Morphology and Topography of Colistin-Heteroresistant Acinetobacter baumannii Strains as a Function of Growth Phase and in Response to Colistin Treatment
Rachel L. Soon,1
Roger L. Nation,1
Patrick G. Hartley,2
Ian Larson,3,
* and
Jian Li1,*
Facility for Anti-Infective Drug Development and Innovation,1 Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, 381 Royal Parade, Parkville, Victoria 3052, Australia,3 CSIRO Molecular and Health Technologies, Ian Wark Laboratory, Bayview Ave., Clayton, Victoria 3169 Australia2
Received 16 April 2009/ Returned for modification 5 July 2009/ Accepted 21 September 2009
The prevalence of infections caused by multidrug-resistant gram-negative Acinetobacter baumannii strains and the lack of novel antibiotics under development are posing a global dilemma, forcing a resurgence of the last-line antibiotic colistin. Our aim was to use atomic force microscopy (AFM) to investigate the morphology and topography of paired colistin-susceptible and -resistant cells from colistin-heteroresistant A. baumannii strains as a function of bacterial growth phase and colistin exposure. An optimal AFM bacterial sample preparation protocol was established and applied to examine three paired strains. Images revealed rod-shaped colistin-susceptible cells (1.65 ± 0.27 µm by 0.98 ± 0.07 µm) at mid-logarithmic phase, in contrast to spherical colistin-resistant cells (1.03 ± 0.09 µm); the latter were also more diverse in appearance and exhibited a rougher surface topography (7.05 ± 1.3 nm versus 11.4 ± 2.5 nm for susceptible versus resistant, respectively). Cellular elongation up to 
18 µm at stationary phase was more commonly observed in susceptible strains, although these "worm-like" cells were also observed occasionally in the resistant population. The effects of colistin exposure on the cell surface of colistin-susceptible and -resistant cells were found to be similar; topographical changes were minor in response to 0.5 µg/ml colistin; however, at 4 µg/ml colistin, a significant degree of surface disruption was detected. At 32 µg/ml colistin, cellular clumping and surface smoothening were evident. Our study has demonstrated for the first time substantial morphological and topographical differences between colistin-susceptible and -resistant cells from heteroresistant A. baumannii strains. These results contribute to an understanding of colistin action and resistance in regard to this problematic pathogen.
* Corresponding author. Mailing address: Facility for Anti-Infective Drug Development and Innovation, Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, 381 Royal Parade, Parkville, Victoria 3052, Australia. Phone for J. Li: 61 3 9903 9702. Fax: 61 3 9903 9629. E-mail: Jian.Li{at}pharm.monash.edu.au. Phone for I. Larson: 61 3 9903 9570. Fax: 61 3 9903 9629. E-mail: Ian.Larson{at}pharm.monash.edu.au
Published ahead of print on 28 September 2009.
I. Larson and J. Li are joint senior authors.
Antimicrobial Agents and Chemotherapy, December 2009, p. 4979-4986, Vol. 53, No. 12
0066-4804/09/$08.00+0 doi:10.1128/AAC.00497-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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