Evaluation of methylene blue based photodynamic inactivation (PDI) against intracellular B-CoV and SARS-CoV2 viruses under different light sources in vitro as a basis for new local treatment strategies in the early phase of a Covid19 infection

Abstract

The local antiviral photodynamic inactivation (PDI) may prove to be a helpful tool reducing the viral load in the nose and throat area in the early phase of a Covid19 infection. Both the infectivity and the prognosis of SARS-CoV-2 infections in the early phase can depend on the viral load in this area. The aim of our study was to find a simplified PDI therapy option against corona viruses in this region with low dose methylene blue (MB) as photosensitizer and use of LED light instead of laser. As a substitute for SARS-CoV2 viruses we started with BCoV infected U373 cells first. We used an 810nm diode laser with 300mW/cm2 and 100J/cm² light dose as well as a 590 nm LED and a broadband LED with irradiation intensity of 10,000 lx each (irradiation time 2.5 and 10 min) and concentrations of the sensitizer of 0.001% and 0.0001%. The 0.001% MB sensitizer experiments showed similar results with all exposures. The logarithmic reduction factor varied between ≥ 5.29 and ≥ 5.31, (0.001% MB sensitizer) and ≥ 4.6 and ≥ 5.31 (0.0001% MB) respectively. Extending the LED irradiation time from 2 to 5 and 10 minutes did not change these results. In contrast approaches of BCoV-infected cells in the dark, treated with 0.001% and 0.0001% MB sensitizer alone, a lot of residual viruses could be detected after 10 minutes of incubation (RF 0.9 and RF 1.23 for 0.001% MB and 0.0001% MB respectively) In our SARS-CoV-2 experiments with VERO E6 infected cells the irradiation time was reduced to 1, 2 and 3 minutes for both concentrations with increasing broadband LED radiation intensity from 20 to 50 and 100.000 lx. (RF 4.67 for 0.001% and 0.0001% respectively). This showed a minimum concentration of 0.0001%MB and a minimum radiation intensity of 20,000 lx leads to a 99.99% reduction of intracellular and extracellular viruses after one minute exposure.

Keywords: Methylene blue; PDI (photodynamic inactivation); Phenothiazine dye in aPDT; SARS-CoV-2 inactivation; aPDT (antimicrobial photodynamic therapy).