Optimization of the antimicrobial effect of blue light on methicillin-resistantStaphylococcus aureus(MRSA)in vitro

Abstract

Background and Objective In previous studies, we showed that irradiation with 405 nm or 470 nm light suppresses up to 92% methicillin-resistant Staphylococcus aureus (MRSA) growth in vitro and that the remaining bacteria re-colonize. In this study, the aim was to develop a protocol that yields 100% MRSA growth suppression. Materials and Methods We cultured 3 × 106 and 5 × 106 CFU/ml USA300 strain of MRSA and then irradiated each plate with varying fluences of 1–60 J/cm2 of 405 nm or 470 nm light, either once or twice at 6 hours intervals. Next, we plated 7 × 106 CFU/ml and irradiated it with 45, 50, 55, or 60 J/cm2 fluence, once, twice, or thrice at the same 6 hours intervals. In a third experiment, the same culture density was irradiated with 0, 165, 180, 220, or 240 J/cm2, either once, twice, or thrice. Results Irradiation with either wavelength significantly reduced the bacterial colonies regardless of bacterial density (P < 0.05). At 3 × 106 CFU/ml density, nearly 40% and 50% growth of MRSA were suppressed with as little as 3 J/cm2 of 405 nm and 470 nm wavelengths, respectively. Moreover, 100% of the colonies were suppressed with a single exposure to 55 or 60 J/cm2 of 470 nm light or double treatment with 50, 55, or 60 J/cm2 of 405 nm wavelength. At 5 × 106 CFU/ml density, irradiating twice with 50, 55, or 60 J/cm2 of either wavelength suppressed bacterial growth completely, lower fluences did not. The denser 7 × 106 CFU/ml culture required higher doses to achieve 100% suppression, either one shot with 220 J/cm2 of 470 nm light or two shots of the same dose using 405 nm. Conclusion The bactericidal effect of blue light can be optimized to yield 100% bacterial growth suppression, but with relatively high fluences for dense bacterial cultures, such as 7 × 106 CFU/ml.

The relative antimicrobial effect of blue 405 nm LED and blue 405 nm laser on methicillin-resistant Staphylococcus aureus in vitro

“…The strain was identified by standard procedures, including PCR. As detailed in our previous reports [27,29,30], bacteria cultures were diluted to a concentration of 5× 10 6 CFU/ml, in 0.9 % sterile saline, and then streaked onto individual wells in 24-well microplates containing tryptic soy agar before irradiation with 405-nm LED or laser.…”

“…Standardized digital images of MRSA colonies were taken 24 h after incubation, colonies were counted, irradiated groups were compared to controls, and percent survival calculated and plotted using Sigma Scan Pro 5 software (Systat Software, Inc., Point Richmond, CA, USA) as detailed in our previous reports [27,29,30]. To ensure consistency of colony magnification, the camera was positioned 10 cm perpendicularly above each plate.…”

“…Our laboratory and others have shown repeatedly that blue light in the range of 400-470 nm has antimicrobial effects on a diverse group of bacteria including Propionibacterium acnes, Pseudomonas aeruginosa, Porphyromonas gingivalis, Helicobacter pylori, and methicillin-resistant Staphylococcus aureus (MRSA) [16][17][18][19][20][21][22][23][24][25][26][27][28][29][30]. However, most of the studies in this area have been done with blue LEDs or photodynamic therapy involving the use of blue absorbing dyes, not laser diodes.…”

“…However, blue light is advantageous in that, whereas environmental surfaces are not readily amenable to antibiotic treatment, blue light irradiation can be applied easily to stem topical infections and eradicate the bacteria from contaminated equipment and environment. Although there is evidence that irradiation with 405-or 470-nm LED produces as much as 92 % MRSA growth inhibition in one application [19,21,27,29,30], there remains the need to achieve 100 % suppression at low fluences. Therefore, given the intense quest for an antimicrobial agent that could help stem the ongoing epidemic of MRSA, it would be immensely beneficial if a blue laser of either wavelength can be shown to have significantly more bacterial growth suppression than blue LED.…”

Antimicrobial blue light inactivation of pathogenic microbes: State of the art

“…In addition, at higher microbial inoculum oxygen might be consumed faster than it is re-supplied by diffusion from the surface of the initial suspension (Hessling et al, 2017; Vollmerhausen et al, 2017). Bumah et al investigated the effect of bacterial inoculum on the efficacy of aBL at 405 and 470 nm (Bumah et al, 2015a). At the inoculum of 3×10 6 CFU/mL, nearly 40% and 50% growth of MRSA were suppressed with as low as 3 J/cm 2 of 405 nm and 470 nm wavelengths.…”