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Applied and Environmental Microbiology, April 2006, p. 2627-2636, Vol. 72, No. 4
0099-2240/06/$08.00+0     doi:10.1128/AEM.72.4.2627-2636.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Very Low Ethanol Concentrations Affect the Viability and Growth Recovery in Post-Stationary-Phase Staphylococcus aureus Populations

Indranil Chatterjee,1 Greg A. Somerville,2 Christine Heilmann,3 Hans-Georg Sahl,4 Hans H. Maurer,5 and Mathias Herrmann1*

Institute of Medical Microbiology and Hygiene, Institutes of Infectious Disease Medicine,1 Institute of Experimental and Clinical Pharmacology and Toxicology, University of Saarland, Homburg/Saar,5 Institute of Medical Microbiology, University of Münster, Münster,3 Institute for Medical Microbiology and Immunology, University of Bonn, Bonn, Germany,4 Department of Veterinary and Biomedical Sciences, University of Nebraska, Lincoln, Nebraska2

Received 11 November 2005/ Accepted 19 January 2006

Pharmaceuticals, culture media used for in vitro diagnostics and research, human body fluids, and environments can retain very low ethanol concentrations (VLEC) (≤0.1%, vol/vol). In contrast to the well-established effects of elevated ethanol concentrations on bacteria, little is known about the consequences of exposure to VLEC. We supplemented growth media for Staphylococcus aureus strain DSM20231 with VLEC (VLEC+ conditions) and determined ultramorphology, growth, and viability compared to those with unsupplemented media (VLEC– conditions) for prolonged culture times (up to 8 days). VLEC+-grown late-stationary-phase S. aureus displayed extensive alterations of cell integrity as shown by scanning electron microscopy. Surprisingly, while ethanol in the medium was completely metabolized during exponential phase, a profound delay of S. aureus post-stationary-phase recovery (>48 h) was observed. Concomitantly, under VLEC+ conditions, the concentration of acetate in the culture medium remained elevated while that of ammonia was reduced, contributing to an acidic culture medium and suggesting decreased amino acid catabolism. Interestingly, amino acid depletion was not uniformly affected: under VLEC+ conditions, glutamic acid, ornithine, and proline remained in the culture medium while the uptake of other amino acids was not affected. Supplementation with arginine, but not with other amino acids, was able to restore post-stationary-phase growth and viability. Taken together, these data demonstrate that VLEC have profound effects on the recovery of S. aureus even after ethanol depletion and delay the transition from primary to secondary metabolite catabolism. These data also suggest that the concentration of ethanol needed for bacteriostatic control of S. aureus is lower than that previously reported.

* Corresponding author. Mailing address: Kirrberger Str., Building 43, Homburg/Saar, 66421, Germany. Phone: 49 6841 162-3900. Fax: 49 6841-1623985. E-mail: mathias.herrmann{at}uniklinik-saarland.de.

Applied and Environmental Microbiology, April 2006, p. 2627-2636, Vol. 72, No. 4
0099-2240/06/$08.00+0     doi:10.1128/AEM.72.4.2627-2636.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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