The efficiency of 5,10,15,20-tetrakis(1-methylpyridinium-4-yl)porphyrin tetra-iodide (Tetra-Py(+) -Me) in the photodynamic inactivation of single-species biofilms of Staphylococcus aureus, Pseudomonas aeruginosa and Candida albicans and mixed biofilms of S. aureus and C. albicans was evaluated. The effect on the extracellular matrix of P. aeruginosa was also assessed. Irradiation with white light up to an energy dose of 64.8 J cm(-2) in the presence of 20 μM of Tetra-Py(+) -Me Tetra-Py(+) -Me caused significant inactivation in all single-species biofilms (3 - 6 log reductions), although the susceptibility was attenuated in relation to planktonic cells. In mixed biofilms, the inactivation of S. aureus was as efficient as in single-species biofilms but the susceptibility of C. albicans decreased. In P. aeruginosa biofilms, a reduction of 81% in the polysaccharides content of the matrix was observed after treatment with a 20 μM PS concentration and a total light dose of 64.8 J cm(-2) . The results show that the Tetra-Py(+) -Me causes significant inactivation of the microorganisms, either in biofilms or in the planktonic form, and demonstrate that polysaccharides of the biofilm matrix may be a primary target of photodynamic damage. This article is protected by copyright. All rights reserved.