Phototherapy of Pseudomonas aeruginosa-Infected Wounds: Preclinical Evaluation of Antimicrobial Blue Light (450-460 nm) Using In Vitro Assays and a Human Wound Skin Model.

Objective: To determine effective treatment strategies against bacterial infections of burn wounds with Pseudomonas aeruginosa, we tested different treatment regimens with antibacterial blue light (BL). Background: Infections of burn wounds are serious complications and require effective and pathogen-specific therapy. Hereby, infections caused by P. aeruginosa pose a particular challenge in clinical practice due to its resistance to many antibiotics and topical antiseptics. Methods: LED-based light sources (450-460 nm) with different intensities and treatment times were used. Antibacterial effects against P. aeruginosa were determined by colony-forming unit (CFU) assays, human skin wound models, and fluorescence imaging. Results: In suspension assays, BL (2 h, 40 mW/cm2, 288 J/cm2) reduced bacterial number (>5 log10 CFU/mL). Applying 144 J/cm2, using 40 mW/cm2 for 1 h was more effective (>4 log10 CFU) than using 20 mW/cm2 for 2 h (>1.5 log10 CFU). BL with low irradiance (24 h, 3.5 mW/cm2, 300 J/cm2) only revealed bacterial reduction in thin bacteria-containing medium layers. In infected in vitro skin wounds only BL irradiation (2 h, 40 mW/cm2, 288 J/cm2) exerted a significant antimicrobial efficacy (2.94 log10 CFU/mL). Conclusions: BL treatment may be an effective therapy for P. aeruginosa-infected wounds to avoid radical surgical debridement. However, a significant antibacterial efficacy can only be achieved with higher irradiances and longer treatment times (min. 40 mW/cm2; >1 h), which cannot be easily integrated into regular clinical treatment protocols, for example, during a dressing change. Further studies are necessary to establish BL therapy for infected burns among tissue compatibility and interactions with previous therapeutic agents.

Inhibition of bacterial growth through LED (light-emitting diode) 465 and 630 nm: in vitro.

Photobiomodulation has been used to inactivate bacterial growth, in different laser or LED protocols. Thus, the aim of this study was to verify the inhibition of Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli, in ATCC strains and bacteria collected from patients with skin burns, after irradiation with LED; 300 µl of saline solution with bacterial suspension was irradiated at a concentration of 0.5-0.63, by the McFarland scale, after five serial dilutions, with evaluation of pre- and post-irradiation pH and temperature control. The cultures were placed in a bacteriological incubator at 37 °C for 24 h for later counting of colony-forming units (CFU). Data were analyzed by Shapiro-Wilk tests and single-factor ANOVA, with Tukey post hoc (p < 0.05). Both wavelengths and energy densities tested showed inhibition of bacterial growth. The comparison of the irradiated groups (ATCC) with the control group showed the following: S. aureus and P. aeruginosa 465 nm (40 J/cm2) and 630 nm (50 J/cm2) and E. coli 465 nm (40 J/cm2) and 630 nm (30 J/cm2). Among the ATCC S. aureus groups, there was a difference for 630 nm (30 J/cm2) and 465 nm (30, 40, 50 J/cm2). The bacteria from the burned patients were S. aureus (30 and 50 J/cm2) and P. aeruginosa (50 J/cm2). We conclude that different bacterial strains were reduced into colony-forming units after LED irradiation.

The bactericidal efficacy of femtosecond laser-based therapy on the most common infectious bacterial pathogens in chronic wounds: an in vitro study.

We investigated the influence of femtosecond laser irradiation on the growth of the two most common infectious bacterial pathogens in wounds; Staphylococcus aureus and Pseudomonas aeruginosa as an attempt to validate optimum parameters for a laser-based bactericidal modality to be used clinically. Bacterial cultures were exposed to femtosecond laser irradiation at different wavelengths, exposure times, and laser powers. The source of femtosecond laser was INSPIRE HF100 laser system, Spectra-Physics, which is pumped by a mode-locked femtosecond Ti: sapphire laser MAI TAI HP, Spectra-Physics. After irradiation, bacterial cells' survival was monitored by observing the clear zones of inhibition in cultured agar plates. Results for all strains indicated that the exposure to femtosecond laser irradiation with a wavelength ranging from ultraviolet (λ > 350 nm) to blue laser light (λ < 480 nm), for a period above 20 min and with a power density of ≈ 0.063 W/cm2, was enough to inhibit both bacterial pathogens with the results maintained for 1 week following irradiation.

Quinine Enhances Photo-Inactivation of Gram-Negative Bacteria.

BACKGROUND: Antimicrobial resistance is a significant concern to public health, and there is a pressing need to develop novel antimicrobial therapeutic modalities. METHODS: In this study, we investigated the capacity for quinine hydrochloride (Q-HCL) to enhance the antimicrobial effects of antimicrobial blue light ([aBL] 405 nm wavelength) against multidrug-resistant (MDR) Gram-negative bacteria in vitro and in vivo. RESULTS: Our findings demonstrated the significant improvement in the inactivation of MDR Pseudomonas aeruginosa and Acinetobacter baumannii (planktonic cells and biofilms) when aBL was illuminated during Q-HCL exposure. Furthermore, the addition of Q-HCL significantly potentiated the antimicrobial effects of aBL in a mouse skin abrasion infection model. In addition, combined exposure of aBL and Q-HCL did not result in any significant apoptosis when exposed to uninfected mouse skin. CONCLUSIONS: In conclusion, aBL in combination with Q-HCL may offer a novel approach for the treatment of infections caused by MDR bacteria.

Antimicrobial Blue Light Inactivation of Microbial Isolates in Biofilms.

BACKGROUND AND OBJECTIVES: Biofilms cause more than 80% of infections in humans, including more than 90% of all chronic wound infections and are extremely resistant to antimicrobials and the immune system. The situation is exacerbated by the fast spreading of antimicrobial resistance, which has become one of the biggest threats to current public health. There is consequently a critical need for the development of alternative therapeutics. Antimicrobial blue light (aBL) is a light-based approach that exhibits intrinsic antimicrobial effect without the involvement of exogenous photosensitizers. In this study, we investigated the antimicrobial effect of this non-antibiotic approach against biofilms formed by microbial isolates of multidrug-resistant bacteria. STUDY DESIGN/MATERIALS AND METHODS: Microbial isolates of Acinetobacter baumannii, Candida albicans, Escherichia coli, Enterococcus faecalis, MRSA, Neisseria gonorrhoeae, Pseudomonas aeruginosa, and Proteus mirabilis were studied. Biofilms were grown in microtiter plates for 24 or 48 hours or in the CDC biofilm reactor for 48 hours and exposed to aBL at 405 nm (60 mW/cm2 , 60 or 30 minutes). The anti-biofilm activity of aBL was measured by viable counts. RESULTS: The biofilms of A. baumannii, N. gonorrhoeae, and P. aeruginosa were the most susceptible to aBL with between 4 and 8 log10 inactivation after 108 J/cm2 (60 mW/cm2 , 30 minutes) or 216 J/cm2 (60 mW/cm2 , 60 minutes) aBL were delivered in the microplates. On the contrary, the biofilms of C. albicans, E. coli, E. faecalis, and P. mirabilis were the least susceptible to aBL inactivation (-0.30, -0.24, -0.84, and -0.68 log10 inactivation, respectively). The same aBL treatment in biofilms developed in the CDC biofilm reactor, caused -1.68 log10 inactivation in A. baumannii and -1.74 and -1.65 log10 inactivation in two different strains of P. aeruginosa. CONCLUSIONS: aBL exhibits potential against pathogenic microorganisms and could help with the significant need for new antimicrobials in clinical practice to manage multidrug-resistant infections. Lasers Surg. Med. © 2019 Wiley Periodicals, Inc.

Blue laser light inhibits biofilm formation in vitro and in vivo by inducing oxidative stress.

Resolution of bacterial infections is often hampered by both resistance to conventional antibiotic therapy and hiding of bacterial cells inside biofilms, warranting the development of innovative therapeutic strategies. Here, we report the efficacy of blue laser light in eradicating Pseudomonas aeruginosa cells, grown in planktonic state, agar plates and mature biofilms, both in vitro and in vivo, with minimal toxicity to mammalian cells and tissues. Results obtained using knock-out mutants point to oxidative stress as a relevant mechanism by which blue laser light exerts its anti-microbial effect. Finally, the therapeutic potential is confirmed in a mouse model of skin wound infection. Collectively, these data set blue laser phototherapy as an innovative approach to inhibit bacterial growth and biofilm formation, and thus as a realistic treatment option for superinfected wounds.

Violet-Blue Light Arrays at 405 Nanometers Exert Enhanced Antimicrobial Activity for Photodisinfection of Monomicrobial Nosocomial Biofilms.

Light-emitting diodes (LEDs) demonstrate therapeutic effects for a range of biomedical applications, including photodisinfection. Bands of specific wavelengths (centered at 405 nm) are reported to be the most antimicrobial; however, there remains no consensus on the most effective irradiation parameters for optimal photodisinfection. The aim of this study was to assess decontamination efficiency by direct photodisinfection of monomicrobial biofilms using a violet-blue light (VBL) single-wavelength array (SWA) and multiwavelength array (MWA). Mature biofilms of nosocomial bacteria (Acinetobacter baumannii, Pseudomonas aeruginosa, Escherichia coli, and Staphylococcus aureus) were grown on 96-well polypropylene PCR plates. The biofilms were then exposed to VBL for 2,700 s (SWA) and 1,170 s (MWA) to deliver 0 to 670 J/cm2, and the antibacterial activity of VBL was assessed by comparing the seeding of the irradiated and the nonirradiated biofilms. Nonirradiated groups were used as controls. The VBL arrays were characterized optically (spectral irradiance and beam profile) and thermally. The SWA delivered 401-nm VBL and the MWA delivered between 379-nm and 452-nm VBL, albeit at different irradiances and with different beam profiles. In both arrays, the irradiated groups were exposed to increased temperatures compared to the nonirradiated controls. All bacterial isolates were susceptible to VBL and demonstrated reductions in the seeding of exposed biofilms compared with the nonirradiated controls. VBL at 405 nm exerted the most antimicrobial activity, exhibiting reductions in seeding of up to 94%. Decontamination efficiency is dependent on the irradiation parameters, bacterial species and strain, and experimental conditions. Controlled experiments that ameliorate the heating effects and improve the optical properties are required to optimize the dosing parameters to advance the successful clinical translation of this technology.IMPORTANCE This study reports the efficacy of VBL and blue light (BL) and their antimicrobial activity against mature biofilms of a range of important nosocomial pathogens. While this study investigated the antibacterial activity of a range of wavelengths of between 375 and 450 nm and identified a specific wavelength region (∼405 nm) with increased antibacterial activity, decontamination was dependent on the bacterial species, strain, irradiation parameters, and experimental conditions. Further research with controlled experiments that ameliorate the heating effects and improve the optical properties are required to optimize the dosing parameters to advance the successful clinical translation of this technology.

Photothermal-Induced Antibacterial Activity of Gold Nanorods Loaded into Polymeric Hydrogel against Pseudomonas aeruginosa Biofilm.

In this study, the photothermal-induced bactericidal activity of phospholipid-decorated gold nanorods (DSPE-AuNR) suspension against Pseudomonas aeruginosa planktonic and biofilm cultures was investigated. We found that the treatment of planktonic culture of Pseudomonas aeruginosa with DSPE-AuNR suspension (0.25-0.03 nM) followed by a continuous laser beam exposure resulted in ~6 log cycle reduction of the bacterial viable count in comparison to the control. The percentage reduction of Pseudomonas aeruginosa biofilm viable count was ~2.5-6.0 log cycle upon laser excitation with different concentrations of DSPE-AuNR as compared to the control. The photothermal ablation activity of DSPE-AuNR (0.125 nM) loaded into poloxamer 407 hydrogel against Pseudomonas aeruginosa biofilm resulted in ~4.5-5 log cycle reduction in the biofilm viable count compared to the control. Moreover, transmission electron microscope (TEM) images of the photothermally-treated bacteria revealed a significant change in the bacterial shape and lysis of the bacterial cell membrane in comparison to the untreated bacteria. Furthermore, the results revealed that continuous and pulse laser beam modes effected a comparable photothermal-induced bactericidal activity. Therefore, it can be concluded that phospholipid-coated gold nanorods present a promising nanoplatform to eradicate Pseudomonas aeruginosa biofilm responsible for common skin diseases.

Carbon@polypyrrole nanotubes as a photosensitizer in laser phototherapy of Pseudomonas aeruginosa.

Phototherapy has been offered as an alternative and promising antibacterial strategy to overcome the antibiotic resistance problem. This study evaluated the antibacterial and phototherapy effects of carbon nanotubes with a polypyrrole coating in a core@shell structure (CNTs@PPy) on Pseudomonas aeruginosa (P. aeruginosa). P. aeruginosa was treated with CNTs@PPy at different concentrations (50-500 µg mL-1) in dark or laser light irradiation with a wavelength of 808 nm, a power density of 1000 mW cm-2 for 20 min. Temperature increment, cell viability, formation of reactive oxygen species (ROS) and protein/nucleic acid leakage subsequent the P. aeruginosa treatment were evaluated. The results showed that near-infrared laser irradiation of CNTs@PPy caused to a temperature increment confirming the ability of powerful photokilling of P. aeruginosa in a photothermal route. On the other hand, while CNTs@PPy represented just a 30-50% P. aeruginosa killing rate in dark, laser irradiation of 250 and 500 µg mL-1 concentrations of CNTs@PPy resulted in a ˜70% P. aeruginosa killing rate, along with significant ROS production into the medium and protein and nucleic acid leakage from P. aeruginosa. These later effects were assigned to a photodynamic route activity of CNTs@PPy upon laser irradiation. Therefore, CNTs@PPy acted as a photosensitizer in both photothermal and photodynamic therapies to present an enhanced bactericidal activity to annihilate and destroyed the gram-negative bacteria P. aeruginosa, a cause of many infectious diseases.

Enhanced sterilization and healing of cutaneous pseudomonas infection using 5-aminolevulinic acid as a photosensitizer with 410-nm LED light.

BACKGROUND: Pseudomonas aeruginosa (PA) frequently develops antibiotic-resistant characteristics, which is clinically problematic. The main reason behind the rise of antibiotic-resistant PA is the extensive use of antibiotics. Therefore, a novel technique is needed to treat PA infections. Photodynamic therapy (PDT) is thought to have the potential to be a non-antibiotic treatment for infections. 5-Aminolevulinic acid (ALA), which works as a photosensitizer after being metabolized into protoporphyrin IX (PpIX) in the heme synthetic pathway, is used for PDT. Thus far, the in vivo effectiveness of PDT using ALA against PA is unknown. OBJECTIVE: In this study, we investigated PDT using ALA both in vitro and in vivo. METHODS AND RESULTS: Although PDT with ALA alone did not show a bactericidal effect on PA, PDT with both ALA and EDTA-2Na had a bactericidal effect in vitro. In in vivo experiments, wounds healed faster in PA-infected mice treated with PDT using both EDTA-2Na and ALA compared to non-PDT. CONCLUSION: These results suggest that PDT with EDTA-2Na and ALA is a potential novel treatment option for PA-infected wounds.

Properties of halogenated and sulfonated porphyrins relevant for the selection of photosensitizers in anticancer and antimicrobial therapies.

The impact of substituents on the photochemical and biological properties of tetraphenylporphyrin-based photosensitizers for photodynamic therapy of cancer (PDT) as well as photodynamic inactivation of microorganisms (PDI) was examined. Spectroscopic and physicochemical properties were related with therapeutic efficacy in PDT of cancer and PDI of microbial cells in vitro. Less polar halogenated, sulfonamide porphyrins were most readily taken up by cells compared to hydrophilic and anionic porphyrins. The uptake and PDT of a hydrophilic porphyrin was significantly enhanced with incorporation in polymeric micelles (Pluronic L121). Photodynamic inactivation studies were performed against Gram-positive (S. aureus, E. faecalis), Gram-negative bacteria (E. coli, P. aeruginosa, S. marcescens) and fungal yeast (C. albicans). We observed a 6 logs reduction of S. aureus after irradiation (10 J/cm2) in the presence of 20 µM of hydrophilic porphyrin, but this was not improved with incorporation in Pluronic L121. A 2-3 logs reduction was obtained for E. coli using similar doses, and a decrease of 3-4 logs was achieved for C. albicans. Rational substitution of tetraphenylporphyrins improves their photodynamic properties and informs on strategies to obtain photosensitizers for efficient PDT and PDI. However, the design of the photosensitizers must be accompanied by the development of tailored drug formulations.

Catalytic reduction of 4-nitrophenol and photo inhibition of Pseudomonas aeruginosa using gold nanoparticles as photocatalyst.

A simple, green method is described for the synthesis of Gold (Au) nanoparticles (NPs) using Cotoneaster horizontalis extract as a phyto-reducer and capping agent with superior photo inhibition activity against Pseudomonas aeruginosa. Different from the other methods used elevated temperatures for nanoparticles synthesis, the novelty of our method lies in its energy saving process and fast synthesis rates (~5min for AuNPs), and its potential to tune the nanoparticles size and afterward their catalytic activity. The starch, fatty acid and reducing sugars present in the extract are mostly responsible for repaid reduction rate Au+3 ions to AuNPs. Strong Plasmon resonance (SPR) of AuNPs was observed at 560nm, which indicates the formation of gold nanoparticles. Uv-visible spectroscopy, high resolution transmission electron microscope (HRTEM) and energy dispersion X-ray diffraction (XRD) were preformed to find out the formation of AuNPs. Proficient reduction of 4-nitrophenol (4-NP) into 4-aminophenol (4-AP) in the presence of AuNPs and NaBH4 was observed and was found to depend upon the nanoparticle size or the extract concentration. The AuNPs was also evaluated for antibacterial against P. aeruginosa. Before transferred it into antibacterial activity, it placed under visible light for 120min. The same experiment was performed in dark as control medium. The photo irradiated AuNPs was observed to be more effective against P. aeruginosa. The result showed that diameter of zone of inhibition of visible light irradiated AuNPs against P. aeruginosa was 17 (±0.5) and in dark was 8 (±0.4) mm.

In vitro antimicrobial activity of LED irradiation on Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an opportunistic pathogen responsible of many deaths due to nosocomial pneumonia each year. It is particularly resistant to many different classes of antibiotics and disinfectants. For all these reasons, there is the necessity to find novel approaches of treatment. The aim of this study was to evaluate the effect of 880nm light emitting diodes (LED) irradiation on P. aeruginosa, in vitro. Different LED irradiation parameters (time, energy output and the addition of methylene blue and chlorhexidine) have been tested in order to evaluate the effects on this bacterium. After treatment, the colony forming units per milliliter (CFU mL-1) were recorded and the data were submitted to ANOVA and Bonferroni post hoc tests at a level of significance of 5%. A statistical significant reduction of bacterial count has been registered after 5min of LED irradiation. The antibacterial effect was directly proportional to irradiation time and the output energy. The pre-treatment with methylene blue, seems to be not effective against P. aeruginosa, independently from irradiation parameters. On the contrary, the contemporary action of LED and chlorhexidine has shown a great reduction of bacterial count that was statistical significant respect chlorhexidine and LED alone. The effect of LED irradiation was visible also after 24h, when a lower bacterial count characterized all irradiated samples respect controls.

Blue light treatment of Pseudomonas aeruginosa: Strong bactericidal activity, synergism with antibiotics and inactivation of virulence factors.

Pseudomonas aeruginosa is among the most common pathogens responsible for both acute and chronic infections of high incidence and severity. Additionally, P. aeruginosa resistance to conventional antimicrobials has increased rapidly over the past decade. Therefore, it is crucial to explore new therapeutic options, particularly options that specifically target the pathogenic mechanisms of this microbe. The ability of a pathogenic bacterium to cause disease is dependent upon the production of agents termed 'virulence factors', and approaches to mitigate these agents have gained increasing attention as new antibacterial strategies. Although blue light irradiation is a promising alternative approach, only limited and preliminary studies have described its effect on virulence factors. The current study aimed to investigate the effects of lethal and sub-lethal doses of blue light treatment (BLT) on P. aeruginosa virulence factors. We analyzed the inhibitory effects of blue light irradiation on the production/activity of several virulence factors. Lethal BLT inhibited the activity of pyocyanin, staphylolysin, pseudolysin and other proteases, but sub-lethal BLT did not affect the production/expression of proteases, phospholipases, and flagella- or type IV pili-associated motility. Moreover, a eukaryotic cytotoxicity test confirmed the decreased toxicity of blue light-treated extracellular P. aeruginosa fractions. Finally, the increased antimicrobial susceptibility of P. aeruginosa treated with sequential doses of sub-lethal BLT was demonstrated with a checkerboard test. Thus, this work provides evidence-based proof of the susceptibility of drug-resistant P. aeruginosa to BLT-mediated killing, accompanied by virulence factor reduction, and describes the synergy between antibiotics and sub-lethal BLT.

Violet 405-nm light: a novel therapeutic agent against common pathogenic bacteria.

BACKGROUND: The increasing incidence of healthcare-associated infections (HAIs) and multidrug-resistant organisms demonstrate the need for innovative technological solutions. Staphylococcus aureus, Streptococcus pneumonia, Escherichia coli, and Pseudomonas aeruginosa in particular are common pathogens responsible for a large percentage of indwelling medical device-associated clinical infections. The bactericidal effects of visible light sterilization (VLS) using 405-nm is one potential therapeutic under investigation. MATERIALS AND METHODS: Light-emitting diodes of 405-nm were used to treat varying concentrations of S aureus, S pneumonia, E coli, and P aeruginosa. Irradiance levels between 2.71 ± 0.20 to 9.27 ± 0.36 mW/cm2 and radiant exposure levels up to 132.98 ± 6.68 J/cm2 were assessed. RESULTS: Dose-dependent effects were observed in all species. Statistically significant reductions were seen in both Gram-positive and Gram-negative bacteria. At the highest radiant exposure levels, bacterial log10 reductions were E coli-6.27 ± 0.54, S aureus-6.10 ± 0.60, P aeruginosa-5.20 ± 0.84, and S pneumoniae-6.01 ± 0.59. Statistically significant results (<0.001*) were found at each time point. CONCLUSIONS: We have successfully demonstrated high-efficacy bacterial reduction using 405-nm light sterilization. The VLS showed statistical significance against both Gram-positive and Gram-negative species with the given treatment times. The ß-lactam antibiotic-resistant E coli was the most sensitive to VLS, suggesting light therapy could a suitable option for sterilization in drug-resistant bacterial species. This research illustrates the potential of using VLS in treating clinically relevant bacterial infections.

In vitro and in vivo antimicrobial activity of combined therapy of silver nanoparticles and visible blue light against Pseudomonas aeruginosa.

Silver nanoparticles (AgNPs) have been used as potential antimicrobial agents against resistant pathogens. We investigated the possible therapeutic use of AgNPs in combination with visible blue light against a multidrug resistant clinical isolate of Pseudomonas aeruginosa in vitro and in vivo. The antibacterial activity of AgNPs against P. aeruginosa (1×10(5) colony forming unit/mL) was investigated at its minimal inhibitory concentration (MIC) and sub-MIC, alone and in combination with blue light at 460 nm and 250 mW for 2 hours. The effect of this combined therapy on the treated bacteria was then visualized using transmission electron microscope. The therapy was also assessed in the prevention of biofilm formation by P. aeruginosa on AgNP-impregnated gelatin biopolymer discs. Further, in vivo investigations were performed to evaluate the efficacy of the combined therapy to prevent burn-wound colonization and sepsis in mice and, finally, to treat a real infected horse with antibiotic-unresponsive chronic wound. The antimicrobial activity of AgNPs and visible blue light was significantly enhanced (P<0.001) when both agents were combined compared to each agent alone when AgNPs were tested at MIC, 1/2, or 1/4 MIC. Transmission electron microscope showed significant damage to the cells that were treated with the combined therapy compared to other cells that received either the AgNPs or blue light. In addition, the combined treatment significantly (P<0.001) inhibited biofilm formation by P. aeruginosa on gelatin discs compared to each agent individually. Finally, the combined therapy effectively treated a horse suffering from a chronic wound caused by mixed infection, where signs of improvement were observed after 1 week, and the wound completely healed after 4 weeks. To our knowledge, this combinatorial therapy has not been investigated before. It was proved efficient and promising in managing infections caused by multidrug resistant bacteria and could be used as an alternative to conventional antibiotic therapy.

Antimicrobial blue light inactivation of Pseudomonas aeruginosa by photo-excitation of endogenous porphyrins: In vitro and in vivo studies.

Pseudomonas aeruginosa is among the most common pathogens that cause nosocomial infections and is responsible for about 10% of all hospital-acquired infections. In the present study, we investigated the potential development of tolerance of P. aeruginosa to antimicrobial blue light by carrying 10 successive cycles of sublethal blue light inactivation. The high-performance liquid chromatographic (HPLC) analysis was performed to identify endogenous porphyrins in P. aeruginosa cells. In addition, we tested the effectiveness of antimicrobial blue light in a mouse model of nonlethal skin abrasion infection by using a bioluminescent strain of P. aeruginosa. The results demonstrated that no tolerance was developed to antimicrobial blue light in P. aeruginosa after 10 cycles of sub-lethal inactivation. HPLC analysis showed that P. aeruginosa is capable of producing endogenous porphyrins in particularly, coproporphyrin III, which are assumed to be responsible for the photodynamic effects of blue light alone. P. aeruginosa infection was eradicated by antimicrobial blue light alone (48 J/cm(2) ) without any added photosensitizer molecules in the mouse model. In conclusion, endogenous photosensitization using blue light should gain considerable attention as an effective and safe alternative antimicrobial therapy for skin infections. Lasers Surg. Med. 48:562-568, 2016. © 2016 Wiley Periodicals, Inc.

Red and infrared laser therapy inhibits in vitro growth of major bacterial species that commonly colonize skin ulcers.

Low-level laser therapy (LLLT) is used in chronic wounds due to its healing effects. However, bacterial species may colonize these wounds and the optimal parameters for effective bacterial inhibition are not clear. The aim of this study was to analyze the effect of LLLT on bacterial growth in vitro. Bacterial strains including Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa were suspended in saline solution at a concentration of 10(3) cells/ml and exposed to laser irradiation at wavelengths of 660, 830, and 904 nm at fluences of 0 (control), 3, 6, 12, 18, and 24 J/cm(2). An aliquot of the irradiated suspension was spread on the surface of petri plates and incubated at 37 °C for quantification of colony-forming unit after 24, 48, and 72 h. Laser irradiation inhibited the growth of S. aureus at all wavelengths and fluences higher than 12 J/cm(2), showing a strong correlation between increase in fluence and bacterial inhibition. However, for P. aeruginosa, LLLT inhibited growth at all wavelengths only at a fluence of 24 J/cm(2). E. coli had similar growth inhibition at a wavelength of 830 nm at fluences of 3, 6, 12, and 24 J/cm(2). At wavelengths of 660 and 904 nm, growth inhibition was only observed at fluences of 12 and 18 J/cm(2), respectively. LLLT inhibited bacterial growth at all wavelengths, for a maximum of 72 h after irradiation, indicating a correlation between bacterial species, fluence, and wavelength.

Blue Laser Inhibits Bacterial Growth of Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa.

OBJECTIVE: The purpose of this study was to analyze the influence of blue laser on bacterial growth of the main species that usually colonize cutaneous ulcers, as well as its effect over time following irradiation. BACKGROUND DATA: The use of blue laser has been described as an adjuvant therapeutic method to inhibit bacterial growth, but there is no consensus about the best parameters to be used. METHODS: Strains of Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa ATCC 27853, and Escherichia coli ATCC 25922 were suspended in saline solution at a concentration of 1.5×10(3) colony forming units (CFU)/mL. Next, 300 µL of this suspension was transferred to a microtitulation plate and exposed to a single blue laser irradiation (450 nm) at fluences of 0 (control), 3, 6, 12, 18, and 24 J/cm(2). Each suspension was spread over the surface of a Petri plate before being incubated at 37°C, and counts of CFU were determined after 24 and 48 h. RESULTS: Blue laser inhibited the growth of S. aureus and P. aeruginosa at fluences >6 J/cm(2). On the other hand, E. coli was inhibited at all fluences tested, except at 24 J/cm(2). CONCLUSIONS: Blue laser light was capable of inhibiting bacterial growth at low fluences over time, thus presenting no time-dependent effect.

In Vitro Evaluation of Antimicrobial Photodynamic Therapy Associated with Hydroalcoholic Extracts of Schinopsis brasiliensis Engl.: New Therapeutic Perspectives.

OBJECTIVE: The aim of this study was to evaluate the photodynamic potential of extracts of Schinopsis brasiliensis Engl. on bacteria involved in several human infections. BACKGROUND DATA: Photodynamic therapy (PDT) involves the interaction of light with an appropriate and photosensitizer wavelength, and the prospect of existing photosensitive compounds in herbal extracts enhanced by the application of laser diode has been promising. METHODS: The antibacterial activity against Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa and Enterococcus faecalis was obtained by the disk diffusion method on agar. The laser diode InGaAIP was used with 660 nm wavelength, 100 mW, and 4 J/cm(2), and the application was performed in a timely manner for 34 sec on each disk tested. The groups tested were: Laser and bark extract (B+L+); bark extract only (B+L-); Laser and leaf extract (F+L+); leaf extract only (F+L-); Laser and malachite green (M+L+); malachite green only (M+L-); and laser only (L+). RESULTS: There were significant differences between the B+L- and B+L+ groups (p=0.029) and between the L+F- and L+F+ groups (p=0.029) at various concentrations of the nebulized extracts of bark and leaf. Among the tested pathogens, S. aureus showed the highest zone of inhibition, 24.55±0.36 mm in group B+L+, 500 mg.mL(-1). CONCLUSIONS: PDT with malachite green was effective, and groups B+L+ and F+L+ showed excellent activity on the bacteria tested, suggesting the presence of photosensitizers in extracts of S. brasiliensis Engl.