As a second step, a finer evaluation to establish the optimum light dosimetry was performed. Eight further groups were employed to analyze the photodynamic effects at 15, 30, 45, 60, 75, 90, 105 and 120 s of irradiation (0.45, 0.9, 1.35, 1.8, 2.25, 2.7 and 3.6 J/cm2) and once again 0.9 J/cm2 (30 s of irradiation) provided the best survival rate (Figure 1). Figure 1 Dose–response 24 h after aPDT in G. mellonella infected by C. albicans Can14. Larvae were infected with 1×106 CFU/larva of C. albicans Can14. The best
survival rate was found when the fluence of 0.9 J/cm2 was applied. As a third step, a further comprehensive experimental procedure was designed to assess the effects of aPDT, mediated by the optimum dose (1 mM MB and red light at 0.9 J/cm2), on host curve survival when infected by the wild-type strain C. albicans Can14 and the fluconazole resistant isolate C. albicans PI3K Inhibitor Library cost Can37. We observed that MB-mediated aPDT, prolonged the larval survival when compared to non-PDT treated larvae, however a statistically significant difference between PDT and control groups was observed only for C. albicans Can14 (Figure 2). Figure 2 Killing of G. mellonella by C. albicans exposed
to antimicrobial PDT. In the aPDT group, the larvae received the PS injection 90 min after the infection with C. albicans. In order to allow a good dispersion of the PS into the insect body, we waited at least 30 additional min after the PS injection prior to the light irradiation. A control group received PS without light exposure. Larvae were
this website maintained at 37°C. a) C. albicans Can14 wild-type strain SC5314, b) C. albicans Can37 clinical isolate from oropharyngeal candidiasis and fluconazole resistant. Since it was observed that fluconazole resistant strain (Can37) showed reduced sensitivity to PDT, we evaluated the number of CFU within the hemolymph to determine if the fungal burden was reduced even if survival was not significantly increased. We compared the hemolymph burden of aPDT-treated larvae with non-treated larvae and a significant reduction in the CFU number was observed post-PDT RG7420 treatment (Figure 3). These results confirmed that aPDT was able to reduced fungal cell viability (0.2 Log) immediately upon light exposure, suggesting that singlet oxygen and other ROS were produced, leading to cell damage [21, 22]. Figure 3 Number of fungal cells in G. mellonella hemolymph immediately post exposed to antimicrobial PDT treatment. Larvae were infected with 1.41×106 CFU/larva of C. albicans Can37 and were maintained at 37°C. After 90 min post-infection, the PS was injected. We waited an additional 30 min prior to light irradiation. After light irradiation, the bacterial burden was measured immediately. Fungal burden was quantified from pools of three larvae hemolymph. aPDT exposed groups resulted in a significant fungal burden reduction when compared to the control group that was not exposed to light.