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Applied and Environmental Microbiology, November 2005, p. 6918-6925, Vol. 71, No. 11
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.11.6918-6925.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Photodynamic Inactivation of Bacillus Spores, Mediated by Phenothiazinium Dyes

Tatiana N. Demidova^1,2 and Michael R. Hamblin^1,3,4*

Wellman Center for Photomedicine, Massachusetts General Hospital, Boston, Massachusetts,¹ Graduate Program in Cell, Molecular and Developmental Biology, Sackler School of Graduate Biomedical Sciences, Tufts University School of Medicine, Boston, Massachusetts,² Department of Dermatology, Harvard Medical School, Boston, Massachusetts,³ Harvard-MIT Division of Health Sciences and Technology, Cambridge, Massachusetts⁴

Received 18 October 2004/ Accepted 28 June 2005

Spore formation is a sophisticated mechanism by which some bacteria survive conditions of stress and starvation by producing a multilayered protective capsule enclosing their condensed DNA. Spores are highly resistant to damage by heat, radiation, and commonly employed antibacterial agents. Previously, spores have also been shown to be resistant to photodynamic inactivation using dyes and light that easily destroy the corresponding vegetative bacteria. We have discovered that Bacillus spores are susceptible to photoinactivation by phenothiazinium dyes and low doses of red light. Dimethylmethylene blue, methylene blue, new methylene blue, and toluidine blue O are all effective, while alternative photosensitizers such as Rose Bengal, polylysine chlorin(e6) conjugate, a tricationic porphyrin, and a benzoporphyrin derivative, which easily kill vegetative cells, are ineffective. Spores of Bacillus cereus and B. thuringiensis are most susceptible, B. subtilis and B. atrophaeus are also killed, and B. megaterium is resistant. Photoinactivation is most effective when excess dye is washed from the spores, showing that the dye binds to the spores and that excess dye in solution can quench light delivery. The relatively mild conditions needed for spore killing could have applications for treating wounds contaminated by anthrax spores, for which conventional sporicides would have unacceptable tissue toxicity.

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* Corresponding author. Mailing address: Wellman Center for Photomedicine, Massachusetts General Hospital, BAR414, 40 Blossom Street, Boston, MA 02114. Phone: (617) 726-6182. Fax: (617) 726-8566. E-mail: hamblin{at}helix.mgh.harvard.edu.

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Applied and Environmental Microbiology, November 2005, p. 6918-6925, Vol. 71, No. 11
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.11.6918-6925.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

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