Photodynamic treatment by the cationic TMPyP photosensitizer was undertaken on the multiple antibiotic-resistant bacteria Acinetobacter baumannii and Escherichia coli. Total eradication of the bacterial cultures was determined immediately after initiation of illumination when these bacteria were treated with 5, 10, 15, 20-tetra (4-N methylpyridyl)porphine (TMPyP) at a concentration of 29.4 micromol/L and illuminated by blue, green, or red light. Total eradication of both bacteria was obtained also after treatment of bacterial cultures with 3.7 micromol/L TMPyP and illumination with blue light (400-450 nm). On the other hand, an 8- or 16- to 20-fold higher light intensity, respectively, was required for total eradication upon illumination with green (480-550 nm) or red light (600-700 nm). A 407-nm blue light only 7 and 9 joules/cm2, respectively, was needed for total eradication of both bacteria even at a concentration of 3.7 micromol/L TMPyP. X-ray-linked microanalysis demonstrated loss of potassium and a flood of sodium and chloride into the cells, indicating serious damage to the cytoplasmic membrane. Transmission electron microscopy (TEM) revealed structural changes and damage to the membrane of treated E. coli. In A. baumannii-treated cells, mesosomes and black dots that resemble aggregation of polyphosphate polymers could be seen. DNA breakage appeared only after a long period of illumination, when the bacterial cell was no longer viable. It can be concluded that cytoplasmic membrane damage and not DNA breakage is the major cause for bacterial death upon photosensitization.

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Propionibacterium acnes is a Gram-positive, microaerophilic bacterium that causes skin wounds. It is known to naturally produce high amounts of intracellular porphyrins. The results of the present study confirm that the investigated strain of P. acnes is capable of producing endogenic porphyrins with no need for any trigger molecules. Extracts from growing cultures have demonstrated emission peaks around 612 nm when excited at 405 nm, which are characteristic for porphyrins. Endogenic porphyrins were determined and quantified after their extraction from the bacterial cells by fluorescence intensity and by elution retention time on high-performance liquid chromatography (HPLC). The porphyrins produced by P. acnes are mostly coproporphyrin, as shown by the HPLC elution patterns. Addition of delta-aminolevulinic acid (ALA) enhanced intracellular porphyrin synthesis and higher amounts of coproporphyrin have been found. Eradication of P. acnes by its endogenic porphyrins was examined after illumination with intense blue light at 407-420 nm. The viability of 24 h cultures grown anaerobically in liquid medium was reduced by less than two orders of magnitude when illuminated once with a light dose of 75 J cm(-2). Better photodynamic effects were obtained when cultures were illuminated twice or three times consecutively with a light dose of 75 J cm(-2) and an interval of 24 h between illuminations. The viability of the culture under these conditions decreased by four orders of magnitude after two illuminations and by five orders of magnitude after three illuminations. When ALA-triggered cultures were illuminated with intense blue light at a light dose of 75 J cm(-2) the viability of the treated cultures decreased by seven orders of magnitude. This decrease in viability can occur even after a single exposure of illumination for the indicated light intensity. X-ray microanalysis and transmission electron microscopy revealed structural damages to membranes in the illuminated P. acnes. Illumination of the endogenous coproporphyrin with blue light (407-420 nm) apparently plays a major role in P. acnes photoinactivation. A treatment protocol with a series of several illuminations or illumination after application of ALA may be suitable for curing acne. Treatment by both pathways may overcome the resistance of P. acnes to antibiotic treatment.

Photodynamic treatment of the gram-negative bacteria Escherichia coli B and Acinetobacter baumannii and the gram-positive bacterium Staphylococcus aureus was performed using two newly devised and synthesized antioxidant carrier photosensitizers (antioxidant carrier sensitizers-2 [ACS-2] and antioxidant carrier sensitizers-3 [ACS-3]), which are butyl hydroxy toluene and propyl gallate substituted haematoporphyrins, respectively. It was found that ACS-2 is less reactive than other photosensitizers previously used for the same purpose, whereas ACS-3 is very effective against the multidrug-resistant bacterium A. baumannii, causing its complete eradication at a low fluence (approximately 7.5 J/cm2) of blue light (407-420 nm) and a low concentration (10 microM). At a higher fluence (approximately 37.5 J/cm2) complete eradication of E. coli B can be obtained under the same conditions. Furthermore, X-ray microanalysis and ultrastructural changes indicate that ACS-3, especially in the case of photodynamic treatment of A. baumannii, interferes with membrane functions and causes the inactivation of the bacterium. ACS-3 may be suggested as a specific photosensitization agent for photoinactivation of gram-negative bacteria.

Abstract— Deinococcus radiodurans is an extremely resistant bacteria to both the mutagenic and lethal effects of ionizing radiation. In the present study, we examined photoinactivation of D. radiodurans by two photosensitizers, the hydrophilic cationic 5, 10, 15, 20‐tetra(4‐N‐methylpyridyl)porphine (TMPyP) and deuteroporphyrin (Dp), which is more hydrophobic. The effects of illumination with various lights at different wavelengths were also examined. Efficient photoinactivation was obtained when D. radiodurans cells were irradiated by blue light (400–450 nm) and treated with TMPyP at a concentration of 0.73 μM. Under these conditions the viability of the culture was found to be decreased by almost seven orders of magnitude after 2 min of treatment at an intensity of only 2 J/cm2. Treatment of the bacterial cells by TMPyP or Dp resulted in an efflux of potassium and magnesium from the cells as well as phosphate loss as a function of light dose. These results can be regarded as a consequence of membrane damage, because transmission electron microscopy revealed cell membrane damage shortly after exposure of the treated bacteria to light. In addition, asymmetric septation and filamentous chromosomal DNA were seen. It seems that the cytoplasmic membrane damage in this bacterial species plays a more important role than in other gram‐positive or even gram‐negative bacteria, rendering this radiation‐resistant bacterium susceptible to photosensitization.