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Control of wound sepsis is an important challenge in traumatology. However, increase in the drug-resistant bacteria makes this challenge considerably difficult in recent years. In this study, we attempted to control burn wound sepsis in rats by photodynamic treatment, which has been reported to be effective against some drug-resistant bacteria. A 20% TBSA (total body surface area) full-thickness burn was made in rat dorsal skin, and five days after injury, a suspension of P. aeruginosa was applied to the wound surface. At 30 min after infection, a methylene blue (MB) solution was applied to the wound surface; 5 min afterwards, the wound was illuminated with a 665-nm light emitting diode (LED) array for 10 min. This treatment (application of MB and illumination) was repeated 3 times successively. The averaged light intensity on the wound surface was 3.3 mW/cm 2 , the corresponding total light dose being 5.9 J/cm 2 . One week after injury, the numbers of bacteria in the blood and liver were counted by colony forming assay. In the liver, the number of bacteria of the treated group was significantly lower than that of the sham control group without photodynamic treatment. In the blood, no bacteria were detected in the treated group, while a certain amount of bacteria was detected in the control group. These results demonstrate the efficacy of MB-mediated PDT with a red LED array to control burn wound sepsis.

Control of infection in wounds is critically important to avoid transition to sepsis; however, recent rise of drug-resistant bacteria makes it difficult. Thus, antimicrobial photodynamic therapy (APDT) has recently received considerable attention. In this study, we examined methylene blue (MB)-mediated photodynamic inactivation of Psuedomonas aeruginosa in rat burned skin. Two days after infection, the wound surface was contacted with a MB solution at different concentrations, and thereafter the wound was irradiated with cw 665-nm light at a constant power density of 250 mW/cm 2 for different time durations. We obtained a two orders of magnitude decrease in the number of bacteria by PDT with a 2-h contact of 0.5-mM MB solution and a illumination of 480 J/cm 2 , demonstrating the efficacy of PDT against infection with Ps. aeruginosa in burns.

To obtain efficient antibacterial photodynamic effect in traumatic injuries such as burns, depth-resolved dosimetry of photosensitizer is required. In this study, we performed dual-wavelength photoacoustic (PA) measurement for rat burned skins injected with a photosensitizer. As a photosensitizer, methylene blue (MB) or porfimer sodium was injected into the subcutaneous tissue in rats with deep dermal burn. The wound was irradiated with red (665 nm or 630 nm) pulsed light to excite photosensitizers and green (532 nm) pulsed light to excite blood in the tissue; the latter signal was used to eliminate blood-associated component involved in the former signal. Acoustic attenuation was also compensated from the photosensitizer-associated PA signals. These signal processing was effective to obtain high-contrast image of a photosensitizer in the tissue. Behaviors of MB and porfimer sodium in the tissue were compared.

We investigated methylene blue-mediated photodynamic effect on Pseudomonas aeruginosa using 665-nm nanosecond pulsed light in vitro. The optimum excitation conditions were examined; a reduction of survival fraction up to 10-4 was obtained.

Pulsed photodynamic effect on Ps.aeruginosa has been investigated in vitro with the use of methylene blue (MB) as a photosensitizer. Bacterial suspensions were illuminated with 665-nm, nanosecond light pulses under the various drug and light dose conditions. Bacterial survival fraction decreased with increasing the drug and light doses, while no significant bacterial cell death was observed when the bacteria were washed after incubation with the drug. These results suggest a two-step photo-bactericidal mechanism; the photodynamic effect first induces membrane damage through which MB molecules were translocated into the bacterial cytoplasm, and then caused bacterial cell death.