Antimicrobial photodynamic therapy effectiveness against susceptible and methicillin-resistant Staphylococcus aureus biofilms.

Staphylococcus aureus have a great ability to become rapidly resistant to conventional antimicrobial therapies. This study evaluated the efficacy of antimicrobial photodynamic therapy (aPDT) mediated by Curcumin (Cur) and light-emitting diode (LED) in the inactivation of biofilms of methicillin susceptible and resistant S. aureus (MSSA and MRSA, respectively). Biofilms were treated with Cur (20, 40 or 80 μM) and illuminated with LED source (455 ± 3 nm; 5.28 J/cm2) (aPDT groups), or treated either with Cur or LED only. Other samples were not exposed to Cur or LED (negative control). The biofilms viability after all experimental conditions were evaluated by counting the number of colonies (CFU/mL) and XTT assay. Additional samples were also evaluated by LIVE/DEAD® staining using confocal laser scanning microscopy (CLSM). Data were analyzed by ANOVAs followed by the Games-Howell post hoc test (α = 0.05). For both strains, all aPDT groups significantly reduced both CFU/mL and metabolic activity of biofilms compared to the negative control (p < 0.001). The results were enhanced when 80 μM of Cur was used. CLSM images showed that both bacteria biofilms submitted to aPDT had a large number of red-stained colonies, especially at aPDT80. In general, MRSA biofilms tended to be less susceptible to aPDT than MSSA biofilms. It can be concluded that aPDT mediated by Cur and LED was an efficient method to inactivate 48 -h biofilms of both S. aureus strains.

Teixeira CGS ，Sanitá PV ，Ribeiro APD ，Dias LM ，Jorge JH ，Pavarina AC ... -  《-》

The effect of antimicrobial photodynamic therapy on the expression of biofilm associated genes in Staphylococcus aureus strains isolated from wound infections in burn patients.

Burn patients are particularly susceptible to microbial infection. Staphylococcus aureus causes burn wound, impetigo and cellulitis. Although sub-lethal antimicrobial photodynamic therapy (aPDT) would not result in microorganism killing, it can considerably influence microbial virulence factor. Twelve methicillin-resistant S. aureus (MRSA) and methicillin-sensitive S. aureus (MSSA) isolated from burns patients. To determine the sub-lethal dose of aPDT, 12 clinical isolates of S. aureus photosensitized with 100 μg ml -1 toluidine blue O (TBO) and irradiated by light emitting diode (LED) with a wavelength of 630 ± 10 nm and energy densities of 52.0, 104.1, and 156.2 J/cm2, then bacterial viability was measured. The effects of sub-lethal aPDT on the expression levels of ica ABCD and ica R genes were assessed by quantitative Real-time PCR (qRT-PCR) method. Fifty and 100 μg ml-1 of TBO significantly reduced the mean cell survival in the MRSA (2.5 - 3 log10) and MSSA (2.75-3.1 log10) isolates. The average expression levels of icaA, ica B, ica C, and ica D in the MRSA and MSSA isolates were decreased by (12, 14, 11, and 9) and (13, 14.5, 12, and 9.5) fold change, respectively (P < 0.05). However, the expression of ica R gene was decreased by 6 and 8 folds change in MRSA and MSSA, respectively. The potential of TBO-mediated aPDT could reduce the expression of ica ABCD as important genes involved in biofilm formation and ica R gene as a repressor of the ica operon. Therefore, the use of aPDT agents as a complementary therapy in wound infections of burn patients is recommended.

Mahmoudi H ，Pourhajibagher M ，Alikhani MY ，Bahador A ... -  《-》

Dual wavelength irradiation antimicrobial photodynamic therapy using indocyanine green and metformin doped with nano-curcumin as an efficient adjunctive endodontic treatment modality.

Indocyanine green (ICG) doped with nano-curcumin (N-CUR) can increase photosensitivity in antimicrobial photodynamic therapy (aPDT). Since metformin (Met) provides a synergistic advantage with photosensitivity, it was conjugated with N-CUR@ICG. Aim of the study was to evaluate the photosensitizing effect of N-CUR@ICG-Met used as a new photosensitizer in dual wavelengths irradiation (diode laser and light-emitting diode, LED) aPDT in root canals infected with Enterococcus faecalis biofilm. Following synthesis and confirmation of N-CUR@ICG-Met by Scanning electrone microscope (SEM), Fourier transform infrared (FT-IR) spectroscopy, X-ray diffraction (XRD) and Zetasizer analysis, the mature microbial biofilm was formed. The quantitative and qualitative evaluations of E. faecalis biofilm were made using microbial viability and SEM analysis of the following groups of treatment modalities (n = 5): 1- N-CUR, 2- ICG, 3- Met, 4- N-CUR@ICG, 5- N-CUR@ICG-Met, 6- Diode laser, 7- LED, 8- aPDT/diode, 9- aPDT/LED, 10- aPDT/diode laser + LED, 11- aPDT/LED + diode laser, 12- 2.5 % sodium hypochlorite (NaOCl). In addition, E. faecalis not treated served as negative control. Data were statistically analyzed using One-way Analysis of Variance (ANOVA) and the post-Hoc Bonferroni tests with a level of significance set at P < 0.05. SEM, FT-IR, XRD, Zetasizer analysis confirmed successful doping of ICG-Met onto/into N-CUR. The treatment modality of N-CUR@ICG-Met mediated aPDT/diode laser, aPDT/LED, aPDT/diode laser + LED, and aPDT/LED + diode laser statistically decreased the cell viability of E. faecalis for 69.40 %, 75.52 %, 82.74 %, and 83.84 %, respectively compared with the negative control group (P < 0.05). The double wavelengths irradiation technique, which exposed the N-CUR@ICG-Met with diode laser irradiation and immediately after with LED (i.e. group No. 10), as well as, N-CUR@ICG-Met with LED irradiation and immediately after with diode laser (i.e. group No. 11) reduced the viable E. faecalis count in biofilm structures statistically more than the other treatment modalities (P < 0.05). N-CUR@ICG-Met as a new photosensitizer in dual wavelengths irradiation method may improve anti-biofilm activity of aPDT against E. faecalis.

Pourhajibagher M ，Plotino G ，Chiniforush N ，Bahador A ... -  《-》

Ultrasonic irradiation enhanced the efficacy of antimicrobial photodynamic therapy against methicillin-resistant Staphylococcus aureus biofilm.

Antimicrobial photodynamic therapy (aPDT) is a non-pharmacological antimicrobial regimen based on light, photosensitizer and oxygen. It has become a potential method to inactivate multidrug-resistant bacteria. However, limited by the delivery of photosensitizer (PS) in biofilm, eradicating biofilm-associated infections by aPDT remains challenging. This study aimed to explore the feasibility of combining ultrasonic irradiation with aPDT to enhance the efficacy of aPDT against methicillin-resistant staphylococcus aureus (MRSA) biofilm. A cationic benzylidene cyclopentanone photosensitizer with much higher selectivity to bacterial cells than mammalian cells were applied at the concentration of 10 μM. 532 nm laser (40 mW/cm2, 10 min) and 1 MHz ultrasound (500 mW/cm2, 10 min, simultaneously with aPDT) were employed against MRSA biofilms in vitro. In addition to combined with ultrasonic irradiation and aPDT, MRSA biofilms were treated with laser irradiation only, photosensitizer only, ultrasonic irradiation only, ultrasonic irradiation and photosensitizer, and aPDT respectively. The antibacterial efficacy was determined by XTT assay, and the penetration depth of PS in biofilm was observed using a photoluminescence spectrometer and a confocal laser scanning microscopy (CLSM). In addition, the viability of human dermal fibroblasts (WS-1 cells) after the same treatments mentioned above and the uptake of P3 by WS-1 cells after ultrasonic irradiation were detected by CCK-8 and CLSM in vitro. Results showed that the percent decrease in metabolic activity resulting from the US + aPDT group (75.76%) was higher than the sum of the aPDT group (44.14%) and the US group (9.88%), suggesting synergistic effects. Meanwhile, the diffusion of PS in the biofilm of MRSA was significantly increased by 1 MHz ultrasonic irradiation. Ultrasonic irradiation neither induced the PS uptake by WS-1 cells nor reduced the viability of WS-1 cells. These results suggested that 1 MHz ultrasonic irradiation significantly enhanced the efficacy of aPDT against MRSA biofilm by increasing the penetration depth of PS. In addition, the antibacterial efficacy of aPDT can be enhanced by ultrasonic irradiation, the US + aPDT treatment demonstrated encouraging in vivo antibacterial efficacy (1.73 log10 CFU/mL reduction). In conclusion, the combination of aPDT and 1 MHz ultrasound is a potential and promising strategy to eradicate biofilm-associated infections of MRSA.

Xu Y ，Liu S ，Zhao H ，Li Y ，Cui C ，Chou W ，Zhao Y ，Yang J ，Qiu H ，Zeng J ，Chen D ，Wu S ，Tan Y ，Wang Y ，Gu Y ... -  《-》

Curcumin-mediated anti-microbial photodynamic therapy against Candida dubliniensis biofilms.

Sanitá PV ，Pavarina AC ，Dovigo LN ，Ribeiro APD ，Andrade MC ，Mima EGO ... -  《-》