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Antimicrobial Agents and Chemotherapy, January 2008, p. 299-305, Vol. 52, No. 1
0066-4804/08/$08.00+0     doi:10.1128/AAC.00988-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Toluidine Blue-Mediated Photodynamic Effects on Staphylococcal Biofilms{triangledown}

Mrinalini Sharma,1,3* Livia Visai,1 Francesca Bragheri,2 Ilaria Cristiani,2 Pradeep Kumar Gupta,3 and Pietro Speziale1*

Department of Biochemistry,1 Department of Electronic Engineering, University of Pavia, Pavia 27100, Italy,2 Laser Biomedical Applications and Instrumentation Division, Raja Ramanna Centre for Advanced Technology, Indore 452013, India3

Received 30 July 2007/ Returned for modification 28 August 2007/ Accepted 23 October 2007

Staphylococci are important causes of nosocomial and medical-device-related infections. Their virulence is attributed to the elaboration of biofilms that protect the organisms from immune system clearance and to increased resistance to phagocytosis and antibiotics. Photodynamic treatment (PDT) has been proposed as an alternative approach for the inactivation of bacteria in biofilms. In this study, we have investigated the effect of the photodynamic action of toluidine blue O (TBO) on the viability and structure of biofilms of Staphylococcus epidermidis and of a methicillin-resistant Staphylococcus aureus strain. Significant inactivation of cells was observed when staphylococcal biofilms were exposed to TBO and laser simultaneously. The effect was found to be light dose dependent. Confocal laser scanning microscopic study suggested damage to bacterial cell membranes in photodynamically treated biofilms. In addition, scanning electron microscopy provided direct evidence for the disruption of biofilm structure and a decrease in cell numbers in photodynamically treated biofilms. Furthermore, the treatment of biofilms with tetrasodium EDTA followed by PDT enhanced the photodynamic efficacy of TBO in S. epidermidis, but not in S. aureus, biofilms. The results suggest that photodynamic treatment may be a useful approach for the inactivation of staphylococcal biofilms adhering to solid surfaces of medical implants.

* Corresponding author. Mailing address for Mrinalini Sharma: Laser Biomedical Applications and Instrumentation Division, Raja Ramanna Centre for Advanced Technology, Indore 452013, India. Phone: 91-0731-2488482. Fax: 91-0731-2488430. E-mail: msharma{at}cat.ernet.in. Mailing address for Pietro Speziale: Department of Biochemistry, Viale Taramelli 3/b, 27100 Pavia, Italy. Phone: 39 0382 987787. Fax: 39 0382 423108. E-mail: pspeziale{at}unipv.it

{triangledown} Published ahead of print on 29 October 2007.

Antimicrobial Agents and Chemotherapy, January 2008, p. 299-305, Vol. 52, No. 1
0066-4804/08/$08.00+0     doi:10.1128/AAC.00988-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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