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Applied and Environmental Microbiology, March 2007, p. 1873-1881, Vol. 73, No. 6
0099-2240/07/$08.00+0     doi:10.1128/AEM.01801-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Piezotolerant Small-Colony Variants with Increased Thermotolerance, Antibiotic Susceptibility, and Low Invasiveness in a Clonal Staphylococcus aureus Population{triangledown}

Kimon A. G. Karatzas,1* Angelos Zervos,1 Chrysoula C. Tassou,2 Costas G. Mallidis,2 and Tom J. Humphrey1

Veterinary Pathology, Infection and Immunity, School of Clinical Veterinary Science, University of Bristol, Langford House, Langford, Bristol BS40 5DU, United Kingdom,1 National Agricultural Research Foundation, Institute of Technology of Agricultural Products, S. Venizelou 1, 141 23 Lycovrissi, Attiki, Greece2

Received 29 July 2006/ Accepted 13 January 2007

Following a pressure treatment of a clonal Staphylococcus aureus culture with 400 MPa for 30 min, piezotolerant variants were isolated. Among 21 randomly selected survivors, 9 were piezotolerant and all formed small colonies on several agar media. The majority of the isolates showed increased thermotolerance, impaired growth, and reduced antibiotic resistance compared to the wild type. However, several nonpiezotolerant isolates also demonstrated impaired growth and the small-colony phenotype. In agglutination tests for the detection of protein A and fibrinogen, the piezotolerant variants showed weaker agglutination reactions than the wild type and the other isolates. All variants also showed defective production of the typical S. aureus golden color, a characteristic which has previously been linked with virulence. They were also less able to invade intestinal epithelial cells than the wild type. These S. aureus variants showed phenotypic similarities to previously isolated Listeria monocytogenes piezotolerant mutants that contained mutations in ctsR. Because of these similarities, possible alterations in the ctsR hypermutable regions of the S. aureus variants were investigated through amplified fragment length polymorphism analysis. No mutations were identified, and subsequently we sequenced the ctsR and hrcA genes of three representative variants, finding no mutations. This work demonstrates that S. aureus probably possesses a strategy resulting in an abundance of multiple-stress-resistant variants within clonal populations. This strategy, however, seems to involve genes and regulatory mechanisms different from those previously reported for L. monocytogenes. We are in the process of identifying these mechanisms.

* Corresponding author. Mailing address: University of Bristol, School of Clinical Veterinary Science, Veterinary Pathology and Infection and Immunity, Langford House, Langford, Bristol BS40 5DU, United Kingdom. Phone: 44 117 9289245. Fax: 44 117 9289505. E-mail: k.a.karatzas{at}bris.ac.uk.

{triangledown} Published ahead of print on 26 January 2007.

Applied and Environmental Microbiology, March 2007, p. 1873-1881, Vol. 73, No. 6
0099-2240/07/$08.00+0     doi:10.1128/AEM.01801-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.





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