Molecular Mechanism For The Eliminating Effect Of 460 Nm Visible Light On Methicillin-Resistant Staphylococcus Aureus Infection

BACKGROUND AND OBJECTIVES:Treatment of MRS A infection is intractable and the mechanisms of antibacterial effect of 400-500 nm VL are still unclear.In this study we evaluate the outcomes after irradiation of planktonic and biofilm state MRSA,and wound infection with MRSA via 460 nm VL illuminating technique(VLIT),and further explore the mechanisms.METHODS:Effects of 460 nm VL on both planktonic and biofilm state MRSA were evaluated.Bacteria were treated by exposure to 460 nm radiation(12,60,120,240,or 360 J/cm2,;60 mW/cm2)for VLIT.Other samples were exposed to 630 nm(240 J/cm2),or not treated.The microbe inhibiting rate,live/dead stain and microstructure observation were evaluated to assess the outcomes.Model of wound infection with MRSA was created on dorsal skin of the C57BL/6 mouse.120 J/cm2 illumination per day was conducted on treatment group.Outcomes were survival rate,wound healing rate,inflammatory infiltrate at 0,1,3,and 14 days(histological assessment).RNA sequencing technique was applied to analyze the whole genome variety of MRSA after 460 nm VL exposure and verified by qPCR.GS-11P was used as a phage inhibitor to evaluate the effect of prophage induction in the process of 460 nm VLIT on MRSA and phage particles were observed by SEM.Assessment of the therapeutic effect of 460 nm VLIT on human skin wounds infection with MRSA was conducted by scoring the severity of swelling,secretion and pain of wounds.120 J/cm2 was delivered once a day on wounds for 5 to 14 days.RESULTS:460 nm VLIT showed great capability of killing both planktonic and biofilm state MRSA.60 J/cm2 and higher fluencies produced efficient inhibiting effect on planktonic MRSA and over 96%bacteria was killed by enhancing energy to 240 J/cm2.For biofilm state MRSA,240 and 360 J/cm2 produced 84.67%and 93.34%inhibiting rate respectively.With much better survival rate(P<0.05),wound healing rate(85.27±2.49 VS 15.78±0.98,P<0.001)and less inflammatory cells infiltration(P<0.05 on day 14)compared to control group,460 nm VLIT showed strong antibacterial effect on wound infection with MRSA.Results of RNA-seq and qPCR have revealed that prophage induction occurred in MRSA genome after irradiated by 460 nm VL,and 20 μg/mL GS-11P can significantly repress the temperate phage activation.Phage particles were clearly found by SEM.Wounds infection with MRSA in patients’ were greatly inhibited by illuminating with 460 nm VL once per day.During the whole illuminating process,the temperature measured at every minute was not above 37 ℃ which indicated no thermal effect involved in the antibacterial processCONCLUSION:460 nm VILT produces an strong antibacterial effect on MRSA infection and prophage induction play a critical role in it.460 nm VILT is potentially used to against other multi-resistant microbes and further to instead of UV as routine method for sterilization.