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The main targets of photodynamic inactivation (PDI) are the external bacterial structures, cytoplasmic membrane and cell wall. In this work it was evaluated how the external bacterial structures influence the PDI efficiency. To reach this objective 8 bacteria with distinct external structures were selected; 4 Gram-negative bacteria (Escherichia coli, with typical Gram-negative external structures; Aeromonas salmonicida, Aeromonas hydrophila both with an S-layer and Rhodopirellula sp., with a peptidoglycan-less proteinaceous cell wall and with cytoplasm compartmentalization) and 4 Gram-positive bacteria (Staphylococcus aureus, with typical Gram-positive external structures; Truepera radiovictrix, Deinococcus geothermalis and Deinococcus radiodurans, all with thick cell walls that give them Gram-positive stains, but including a second complex multi-layered membrane and structurally analogous to that of Gram-negative bacteria). The studies were performed in the presence of 5,10,15,20-tetrakis(1-methylpyridinium-4-yl)porphyrin tetraiodide (Tetra-Py(+)-Me) at 5.0 μM with white light (40 W m(-2)). The susceptibility of each bacteria to PDI by Tetra-Py(+)-Me was dependent on bacteria external structures. Although all Gram-positive bacteria were inactivated to the detection limit (reduction of ∼8 log) after 60-180 min of irradiation, the inactivation followed distinct patterns. Among the Gram-negative bacteria, E. coli was the only species to be inactivated to the detection limit (∼8 log after 180 min). The efficiency of inactivation of the two species of Aeromonas was similar (reduction of ∼5-6 log after 270 min). Rhodopirellula was less susceptible (reduction of ∼4 log after 270 min). As previously observed, the Gram-positive bacteria are more easily inactivated than Gram-negative strains, and this is even true for T. radiovictrix, D. geothermalis and D. radiodurans, which have a complex multi-layered cell wall. The results support the theory that the outer cell structures are major bacterial targets for PDI. Moreover, the chemical composition of the external structures has a stronger effect on PDI efficiency than complexity and the number of layers of the external coating, and lipids seem to be an important target of PDI.

Aims: In this study, we propose (i) to study the photodynamic inactivation (PDI) efficiency of neutral and cationic porphyrin derivatives, (ii) to characterize the kinetics of the inactivation process using Bacillus cereus as a model endospore-producing bacterium and (iii) to conclude on the applicability of porphyrin derivatives in the inactivation of bacterial endospores., Methods and Results: The study of PDI of Bacillus cereus endospores, taken as model-endospores, using porphyrin derivatives differing in the number of positive charges and in the meso-substituent groups, showed that neutral, monocationic and dicationic porphyrins are quite ineffective, in contrast with the tri- and tetra-cationic molecules. The most effective porphyrin is a tricationic porphyrin with a meso-pentafluorophenyl group. With this photosensitizer (PS), at 0.5 micromol l(-1), a reduction of 3.5 log units occurs after only 4 min of irradiation. None of the porphyrin derivatives showed toxicity in the absence of light., Conclusions: Some porphyrin derivatives are efficient PSs for the inactivation of bacterial endospores and should be considered in further studies. Small modifications in the substituent groups, in addition to charge, significantly improve the effectiveness of the molecule as a PS for endospore inactivation., Significance and Impact of the Study: Tetrapyrrolic macrocycles should be regarded as worthy to explore for the PDI of spore-producing gram-positive bacteria. The development of molecules, more selective and effective, emerges as a new objective.