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Bacterial contamination is the most prevalent infectious complication of blood transfusion in the developed world. To mitigate this, several ultraviolet light-based pathogen reduction technologies (PRTs), some of which require photo-chemicals, have been developed to minimize infection transmission. Relative to UV light, visible 405-nm light is safer and has shown potential to be developed as a PRT for the in situ treatment of ex vivo human plasma and platelet concentrates, without the need for photo-chemicals. This study investigates the effect of 405-nm light on human plasma, with focus on the compatibility of antimicrobial light doses with essential plasma clotting factors. To determine an effective antimicrobial dose that is compatible with plasma, prebagged human plasma (up to 300 mL) was seeded with common microbial contaminants and treated with increasing doses of 405-nm light (16 mW cm -2 ; ≤ 403 J cm -2 ). Post-exposure plasma protein integrity was investigated using an AOPP assay, in vitro coagulation tests, and ELISA-based measurement of fibrinogen and Protein S. Microbial contamination in 300 mL prebagged human plasma was significantly reduced (P ≤ 0.05) after exposure to ≤ 288 J cm -2 , with microbial loads reduced by > 96.2%. This dose did not significantly affect the plasma protein quality parameters tested (P > 0.05). Increased doses (≥ 345 J cm -2 ) resulted in a 4.3% increase in clot times with no statistically significant change in protein activity or levels. Overall, this study has demonstrated that the effective microbicidal 405 light dose shows little to no negative effect on plasma quality., (© 2024. The Author(s).)

Chemical and UV light-based pathogen reduction technologies are currently in use for human platelet concentrates (PCs) to enhance safety from transfusion-transmitted infections. Relative to UV light, 405 nm violet-blue light in the visible spectrum is known to be less harmful. Hence, in this report for the first time, we have assessed the global hemostasis activity of PCs stored in plasma and the activities of six plasma coagulation factors (CFs) as a measure of in vitro hemostatic activity following exposure to the microbicidal 405 nm light. Apheresis PC samples collected from each screened human donor (n = 22) were used for testing of PCs and platelet poor plasma (PPP). Both PCs and PPPs were treated for 5 h with 405 nm light to achieve a previously established microbicidal light dose of 270 J/cm 2 . Activated partial thromboplastin time and prothrombin time-based potency assays using a coagulation analyzer and hemostatic capacity via Thromboelastography were analyzed. Thromboelastography analysis of the light-treated PCs and plasma present in the PCs showed little difference between the treated and untreated samples. Further, plasma present in the PCs during the light treatment demonstrated a better stability in potency assays for several coagulation factors compared to the plasma alone prepared from PCs first and subjected to the light treatment separately. Overall, PCs stored in plasma treated with 405 nm violet-blue light retain activity for hemostasis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier B.V.)

Due to its increased safety over ultraviolet light, there is interest in the development of antimicrobial violet-blue light technologies for infection control applications. To ensure compatibility with exposed materials and tissue, the light irradiances and dose regimes used must be suitable for the target application. This study investigates the antimicrobial dose responses and germicidal efficiency of 405 nm violet-blue light when applied at a range of irradiance levels, for inactivation of surface-seeded and suspended bacteria. Bacteria were seeded onto agar surfaces (10 1 -10 8 CFUplate -1 ) or suspended in PBS (10 3 -10 9 CFUmL -1 ) and exposed to increasing doses of 405-nm light (≤ 288 Jcm -2 ) using various irradiances (0.5-150 mWcm -2 ), with susceptibility at equivalent light doses compared. Bacterial reductions ≥ 96% were demonstrated in all cases for lower irradiance (≤ 5 mWcm -2 ) exposures. Comparisons indicated, on a per unit dose basis, that significantly lower doses were required for significant reductions of all species when exposed at lower irradiances: 3-30 Jcm -2 /0.5 mWcm -2  compared to 9-75 Jcm -2 /50 mWcm -2  for low cell density (10 2  CFUplate -1 ) surface exposures and 22.5 Jcm -2 /5 mWcm -2  compared to 67.5 Jcm -2 /150 mWcm -2  for low density (10 3  CFUmL -1 ) liquid exposures (P ≤ 0.05). Similar patterns were observed at higher densities, excluding S. aureus exposed at 10 9  CFUmL -1 , suggesting bacterial density at predictable levels has minimal influence on decontamination efficacy. This study provides fundamental evidence of the greater energy efficacy of 405-nm light for inactivation of clinically-significant pathogens when lower irradiances are employed, further supporting its relevance for practical decontamination applications., (© 2024. The Author(s).)

The highly transmittable nature of SARS-CoV-2 has increased the necessity for novel strategies to safely decontaminate public areas. This study investigates the efficacy of a low irradiance 405-nm light environmental decontamination system for the inactivation of bacteriophage phi6 as a surrogate for SARS-CoV-2. Bacteriophage phi6 was exposed to increasing doses of low irradiance (~0.5 mW cm -2 ) 405-nm light while suspended in SM buffer and artificial human saliva at low (~10 3-4  PFU mL -1 ) and high (~10 7-8  PFU mL -1 ) seeding densities, to determine system efficacy for SARS-CoV-2 inactivation and establish the influence of biologically relevant suspension media on viral susceptibility. Complete/near-complete (≥99.4%) inactivation was demonstrated in all cases, with significantly enhanced reductions observed in biologically relevant media (P < 0.05). Doses of 43.2 and 172.8 J cm -2 were required to achieve ~3 log 10 reductions at low density, and 97.2 and 259.2 J cm -2 achieved ~6 log 10 reductions at high density, in saliva and SM buffer, respectively: 2.6-4 times less dose was required when suspended in saliva compared to SM buffer. Comparative exposure to higher irradiance (~50 mW cm -2 ) 405-nm light indicated that, on a per unit dose basis, 0.5 mW cm -2 treatments were capable of achieving up to 5.8 greater log 10 reductions with up to 28-fold greater germicidal efficiency than that of 50 mW cm -2 treatments. These findings establish the efficacy of low irradiance 405-nm light systems for inactivation of a SARS-CoV-2 surrogate and demonstrate the significant enhancement in susceptibility when suspended in saliva, which is a major vector in COVID-19 transmission., (© 2023 The Authors. Photochemistry and Photobiology published by Wiley Periodicals LLC on behalf of American Society for Photobiology.)

Purpose: Lighting systems which use visible light blended with antimicrobial 405-nm violet-blue light have recently been developed for safe continuous decontamination of occupied healthcare environments. This paper characterises the optical output and antibacterial efficacy of a low irradiance 405-nm light system designed for environmental decontamination applications, under controlled laboratory conditions., Methods: In the current study, the irradiance output of a ceiling-mounted 405-nm light source was profiled within a 3×3×2 m (18 m 3 ) test area; with values ranging from 0.001-2.016 mWcm -2 . To evaluate antibacterial efficacy of the light source for environmental surface decontamination, irradiance levels within this range (0.021-1 mWcm -2 ) at various angular ( Δ ϴ=0-51.3) and linear (∆s=1.6-2.56 m) displacements from the source were used to generate inactivation kinetics, using the model organism, Staphylococcus aureus . Additionally, twelve bacterial species were surface-seeded and light-exposed at a fixed displacement below the source (1.5 m; 0.5 mWcm -2 ) to demonstrate broad-spectrum efficacy at heights typical of high touch surfaces within occupied settings., Results: Results demonstrate that significant (P≤0.05) inactivation was successfully achieved at all irradiance values investigated, with spatial positioning from the source affecting inactivation, with greater times required for inactivation as irradiance decreased. Complete/near-complete (≥93.28%) inactivation of all bacteria was achieved following exposure to 0.5 mWcm -2 within exposure times realistic of those utilised practically for whole-room decontamination (2-16 h)., Conclusion: This study provides fundamental evidence of the efficacy, and energy efficiency, of low irradiance 405-nm light for bacterial inactivation within a controlled laboratory setting, further justifying its benefits for practical infection control applications., Competing Interests: Conflict of interestThe intellectual property rights of the 405 nm light environmental decontamination system belong to the University of Strathclyde, and JGA, MM and SJM are co-inventors., (© The Author(s) 2023.)

In transfusion medicine, bacterial contamination can occur in ex vivo stored blood plasma, and there are continued efforts to improve blood safety and reduce the risk of transfusion-transmitted infections. Visible 405-nm violet-blue light has demonstrated potential for in situ pathogen reduction in ex vivo stored plasma and platelet concentrates. This study investigates the broad-spectrum antibacterial efficacy and compatibility potential of 405-nm light for treatment of blood plasma. Human plasma seeded with bacteria at a range of densities (10 1 -10 3 , 10 4 -10 6 , 10 7 -10 8  CFU mL -1 ) was exposed to 360 J cm -2 405-nm light (1 h at 0.1 W cm -2 ), with this fixed dose selected based on the initial analysis of inactivation kinetics. One-dimensional protein mobility analysis and measurement of advanced oxidation protein products (AOPP) was conducted to evaluate compatibility of the antimicrobial dose with plasma proteins and, identify upper levels at which protein degradation can be detected. Broad-spectrum antibacterial efficacy was observed with a fixed treatment of 360 J cm -2 , with 98.9-100% inactivation achieved across all seeding densities for all organisms, except E. coli, which achieved 95.1-100% inactivation. At this dose (360 J cm -2 ), no signs of protein degradation occurred. Overall, 405-nm light shows promise for broad-spectrum bacterial inactivation in blood plasma, while preserving plasma protein integrity., (© 2021 The Authors. Photochemistry and Photobiology published by Wiley Periodicals LLC on behalf of American Society for Photobiology.)

The introduction of pathogen reduction technologies (PRTs) to inactivate bacteria, viruses and parasites in donated blood components stored for transfusion adds to the existing arsenal toward reducing the risk of transfusion-transmitted infectious diseases (TTIDs). We have previously demonstrated that 405 nm violet-blue light effectively reduces blood-borne bacteria in stored human plasma and platelet concentrates. In this report, we investigated the microbicidal effect of 405 nm light on one important bloodborne parasite Trypanosoma cruzi that causes Chagas disease in humans. Our results demonstrated that a light irradiance at 15 mWcm -2 for 5 h, equivalent to 270 Jcm -2 , effectively inactivated T. cruzi by over 9.0 Log 10 , in plasma and platelets that were evaluated by a MK2 cell infectivity assay. Giemsa stained T. cruzi infected MK2 cells showed that the light-treated parasites in plasma and platelets were deficient in infecting MK2 cells and did not differentiate further into intracellular amastigotes unlike the untreated parasites. The light-treated and untreated parasite samples were then evaluated for any residual infectivity by injecting the treated parasites into Swiss Webster mice, which did not develop infection even after the animals were immunosuppressed, further demonstrating that the light treatment was completely effective for inactivation of the parasite; the light-treated platelets had similar in vitro metabolic and biochemical indices to that of untreated platelets. Overall, these results provide a proof of concept toward developing 405 nm light treatment as a pathogen reduction technology (PRT) to enhance the safety of stored human plasma and platelet concentrates from bloodborne T. cruzi , which causes Chagas disease., (Copyright © 2020 Jankowska, Nagarkatti, Acharyya, Dahiya, Stewart, Macpherson, Wilson, Anderson, MacGregor, Maclean, Dey, Debrabant and Atreya.)

Bacterial contamination of ex vivo stored platelets is a cause of transfusion-transmitted infection. Violet-blue 405 nm light has recently demonstrated efficacy in reducing the bacterial burden in blood plasma, and its operational benefits such as non-ionizing nature, penetrability, and non-requirement for photosensitizing agents, provide a unique opportunity to develop this treatment for in situ treatment of ex vivo stored platelets as a tool for bacterial reduction. Sealed bags of platelet concentrates, seeded with low-level Staphylococcus aureus contamination, were 405 nm light-treated (3-10 mWcm -2 ) up to 8 h. Antimicrobial efficacy and dose efficiency was evaluated by quantification of the post-treatment surviving bacterial contamination levels. Platelets treated with 10 mWcm -2 for 8 h were further evaluated for survival and recovery in severe combined immunodeficient (SCID) mice. Significant inactivation of bacteria in platelet concentrates was achieved using all irradiance levels, with 99.6-100% inactivation achieved by 8 h ( P < 0.05). Analysis of applied dose demonstrated that lower irradiance levels generally resulted in significant decontamination at lower doses: 180 Jcm -2 /10 mWcm -2 ( P = 0.008) compared to 43.2 Jcm -2 /3 mWcm -2 ( P = 0.002). Additionally, the recovery of light-treated platelets, compared to non-treated platelets, in the murine model showed no significant differences ( P = >0.05). This report paves the way for further comprehensive studies to test 405 nm light treatment as a bactericidal technology for stored platelets., (Copyright © 2020 Maclean, Gelderman, Kulkarni, Tomb, Stewart, Anderson, MacGregor and Atreya.)

Background: The spread of pathogens via the airborne route is often underestimated, and little is known about the extent to which airborne microbial contamination levels vary throughout the day and night in hospital facilities., Aims: To evaluate airborne contamination levels within intensive care unit (ICU) isolation rooms over 10-24-h periods in order to improve understanding of the variability of environmental aerial bioburden, and the extent to which ward activities may contribute., Methods: Environmental air monitoring was conducted within occupied and vacant inpatient isolation rooms. A sieve impactor sampler was used to collect 500-L air samples every 15 min over 10-h (08:00-18:00 h) and 24-h (08:00-08:00 h) periods. Samples were collected, room activity was logged, and bacterial contamination levels were recorded as colony-forming units (cfu)/m 3 air., Findings: A high degree of variability in levels of airborne contamination was observed across all scenarios in the studied isolation rooms. Air bioburden increased as room occupancy increased, with air contamination levels highest in rooms occupied for the longest time during the study (10 days) (mean 104.4 cfu/m 3 , range 12-510 cfu/m 3 ). Counts were lowest in unoccupied rooms (mean 20 cfu/m 3 ) and during the night., Conclusion: Peaks in airborne contamination were directly associated with an increase in activity levels. This study provides the first clear evidence of the extent of variability in microbial airborne levels over 24-h periods in ICU isolation rooms, and found direct correlation between microbial load and ward activity., (Copyright © 2019 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Antimicrobial violet-blue light is an emerging technology designed for enhanced clinical decontamination and treatment applications, due to its safety, efficacy and ease of use. This systematized review was designed to compile the current knowledge on the antimicrobial efficacy of 380-480 nm light on a range of health care and food-related pathogens including vegetative bacteria, bacterial endospores, fungi and viruses. Data were compiled from 79 studies, with the majority focussing on wavelengths in the region of 405 nm. Analysis indicated that Gram-positive and Gram-negative vegetative bacteria are the most susceptible organisms, while bacterial endospores, viruses and bacteriophage are the least. Evaluation of the dose required for a 1 log 10 reduction of key bacteria compared to population, irradiance and wavelength indicated that microbial titer and light intensity had little effect on the dose of 405 nm light required; however, linear analysis indicated organisms exposed to longer wavelengths of violet-blue light may require greater doses for inactivation. Additional research is required to ensure this technology can be used effectively, including: investigating inactivation of multidrug-resistant organisms, fungi, viruses and protozoa; further knowledge about the photodynamic inactivation mechanism of action; the potential for microbial resistance; and the establishment of a standardized exposure methodology., (© 2018 The Authors. Photochemistry and Photobiology published by Wiley Periodicals, Inc. on behalf of American Society for Photobiology.)

Background: Previous work has shown that a ceiling-mounted, 405 nm high-intensity narrow-spectrum light environmental decontamination system (HINS-light EDS) reduces bacterial contamination of environmental surfaces in a burns unit by between 27% and 75%. Examination of the efficacy of the light over extended exposure times and its probable mode of action was performed., Aim: To ascertain the correlation between bacterial kill achieved on sampled surface sites around the burns unit and both irradiance levels of the 405 nm light, and exposure time., Methods: Seventy samples were taken using contact agar plates from surfaces within an occupied side-room in the burns unit before, during, and after a seven-day use of the HINS-light EDS. This was repeated in three separate studies. Statistical analysis determined whether there was significant decrease in environmental contamination during prolonged periods of HINS-light treatment, and whether there was an association between irradiance and bacterial kill., Findings: A decrease of between 22% and 86% in the mean number of surface bacteria was shown during the use of the HINS-light EDS. When the light ceased to be used, increases of between 78% and 309% occurred. There was no correlation between bacterial kill and irradiance levels at each sampling site but strong correlation between bacterial kill and exposure time., Conclusion: Prolonged exposure to the HINS-light EDS causes a cumulative decontamination of the surfaces within a burns unit. The importance of exposure time and possible airborne effect over irradiance levels is emphasized., (Copyright © 2017 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Background: Antimicrobial violet-blue light in the region of 405 nm is emerging as an alternative technology for hospital decontamination and clinical treatment. The mechanism of action is the excitation of endogenous porphyrins within exposed microorganisms, resulting in ROS generation, oxidative damage and cell death. Although resistance to 405 nm light is not thought likely, little evidence has been published to support this. This study was designed to establish if there is potential for tolerance development, using the nosocomial pathogen Staphylococcus aureus as the model organism., Methods: The first stage of this study investigated the potential for S. aureus to develop tolerance to high-intensity 405 nm light if pre-cultured in low-level stress violet-blue light (≤1 mW/cm 2 ) conditions. Secondly, the potential for tolerance development in bacteria subjected to repeated sub-lethal exposure was compared by carrying out 15 cycles of exposure to high-intensity 405 nm light, using a sub-lethal dose of 108 J/cm 2 . Inactivation kinetics and antibiotic susceptibility were also compared., Results: When cultured in low-level violet-blue light conditions, S. aureus required a greater dose of high-intensity 405 nm light for complete inactivation, however this did not increase with multiple (3) low-stress cultivations. Repeated sub-lethal exposures indicated no evidence of bacterial tolerance to 405 nm light. After 15 sub-lethal exposures 1.2 and 1.4 log 10 reductions were achieved for MSSA and MRSA respectively, which were not significantly different to the initial 1.3 log 10 reductions achieved ( P =  0.242 & 0.116, respectively). Antibiotic susceptibility was unaffected, with the maximum change in zone of inhibition being ± 2 mm., Conclusions: Repeated sub-lethal exposure of non-proliferating S. aureus populations did not affect the susceptibility of the organism to 405 nm light, nor to antibiotics. Culture in low-level violet-blue light prior to 405 nm light exposure may increase oxidative stress responses in S. aureus , however, inactivation still occurs and results demonstrate that this is unlikely to be a selective process. These results demonstrate that tolerance from repeated exposure is unlikely to occur, and further supports the potential development of 405 nm light for clinical decontamination and treatment applications.

The requirement for novel decontamination technologies for use in hospitals is ever present. One such system uses 405 nm visible light to inactivate microorganisms via ROS-generated oxidative damage. Although effective for bacterial and fungal inactivation, little is known about the virucidal effects of 405 nm light. Norovirus (NoV) gastroenteritis outbreaks often occur in the clinical setting, and this study was designed to investigate potential inactivation effects of 405 nm light on the NoV surrogate, feline calicivirus (FCV). FCV was exposed to 405 nm light whilst suspended in minimal and organically-rich media to establish the virucidal efficacy and the effect biologically-relevant material may play in viral susceptibility. Antiviral activity was successfully demonstrated with a 4 Log 10 (99.99%) reduction in infectivity when suspended in minimal media evident after a dose of 2.8 kJ cm -2 . FCV exposed in artificial faeces, artificial saliva, blood plasma and other organically rich media exhibited an equivalent level of inactivation using between 50-85% less dose of the light, indicating enhanced inactivation when the virus is present in organically-rich biologically-relevant media. Further research in this area could aid in the development of 405 nm light technology for effective NoV decontamination within the hospital environment.

Objective: This study investigates possible advantages in pulsed over continuous 405-nm light-emitting diode (LED) light for bacterial inactivation and energy efficiency., Background: Alternative nonantibiotic methods of disinfection and infection control have become of significant interest. Recent studies have demonstrated the application of systems using 405-nm LEDs for continuous disinfection of the clinical environment, and also for potential treatment of contaminated wounds., Methods: Liquid suspensions of 10 3 colony-forming units/mL populations of Staphylococcus aureus were subject to pulsed 405-nm light of different frequencies, duty cycles, and intensities and for different lengths of time., Results: Pulsed exposures with the same average irradiance of 16 mW/cm 2 and varying duty cycle (25%, 50%, 75%) showed very similar performance compared with continuous exposures, with 95-98% reduction of S. aureus achieved for all duty cycles. The pulsing frequency was varied in intervals from 100 Hz to 10 kHz and appeared to have little effect on antimicrobial efficacy. However, when comparing pulsed with continuous exposure, an improvement in inactivation per unit optical energy was achieved, with results showing an increase of approximately 83% in optical efficiency., Conclusions: These results suggest that under pulsed conditions, a lower energy consumption and lower perceived brightness could be achieved, thus potentially providing improved operating conditions for medical/infection control applications without compromising antimicrobial efficacy.

Bacterial inactivation by 405 nm light is accredited to the photoexcitation of intracellular porphyrin molecules resulting in energy transfer and the generation of reactive oxygen species that impart cellular oxidative damage. The specific mechanism of cellular damage, however, is not fully understood. Previous work has suggested that destruction of nucleic acids may be responsible for inactivation; however, microscopic imaging has suggested membrane damage as a major constituent of cellular inactivation. This study investigates the membrane integrity of Escherichia coli and Staphylococcus aureus exposed to 405 nm light. Results indicated membrane damage to both species, with loss of salt and bile tolerance by S. aureus and E. coli, respectively, consistent with reduced membrane integrity. Increased nucleic acid release was also demonstrated in 405 nm light-exposed cells, with up to 50 % increase in DNA concentration into the extracellular media in the case of both organisms. SYTOX green fluorometric analysis, however, demonstrated contradictory results between the two test species. With E. coli, increasing permeation of SYTOX green was observed following increased exposure, with >500 % increase in fluorescence, whereas no increase was observed with S. aureus. Overall, this study has provided good evidence that 405 nm light exposure causes loss of bacterial membrane integrity in E. coli, but the results with S. aureus are more difficult to explain. Further work is required to gain greater understanding of the inactivation mechanism in different bacterial species, as there are likely to be other targets within the cell that are also impaired by the oxidative damage from photo-generated reactive oxygen species.

Light at wavelength 405 nm is an effective bactericide. Previous studies showed that exposing mammalian cells to 405 nm light at 36 J/cm(2) (a bactericidal dose) had no significant effect on normal cell function, although at higher doses (54 J/cm(2)), mammalian cell death became evident. This research demonstrates that mammalian and bacterial cell toxicity induced by 405 nm light exposure is accompanied by reactive oxygen species production, as detected by generation of fluorescence from 6-carboxy-2',7'-dichlorodihydrofluorescein diacetate. As indicators of the resulting oxidative stress in mammalian cells, a decrease in intracellular reduced glutathione content and a corresponding increase in the efflux of oxidised glutathione were observed from 405 nm light treated cells. The mammalian cells were significantly protected from dying at 54 J/cm(2) in the presence of catalase, which detoxifies H2O2. Bacterial cells were significantly protected by sodium pyruvate (H2O2 scavenger) and by a combination of free radical scavengers (sodium pyruvate, dimethyl thiourea (OH scavenger) and catalase) at 162 and 324 J/cm(2). Results therefore suggested that the cytotoxic mechanism of 405 nm light in mammalian cells and bacteria could be oxidative stress involving predominantly H2O2 generation, with other ROS contributing to the damage., (Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The ability of Clostridium difficile to form highly resilient spores which can survive in the environment for prolonged periods causes major contamination problems. Antimicrobial 405 nm light is being developed for environmental decontamination within hospitals, however further information relating to its sporicidal efficacy is required. This study aims to establish the efficacy of 405 nm light for inactivation of C. difficile vegetative cells and spores, and to establish whether spore susceptibility can be enhanced by the combined use of 405 nm light with low concentration chlorinated disinfectants. Vegetative cells and spore suspensions were exposed to increasing doses of 405 nm light (at 70-225 mW/cm(2)) to establish sensitivity. A 99.9% reduction in vegetative cell population was demonstrated with a dose of 252 J/cm(2), however spores demonstrated higher resilience, with a 10-fold increase in required dose. Exposures were repeated with spores suspended in the hospital disinfectants sodium hypochlorite, Actichlor and Tristel at non-lethal concentrations (0.1%, 0.001% and 0.0001%, respectively). Enhanced sporicidal activity was achieved when spores were exposed to 405 nm light in the presence of the disinfectants, with a 99.9% reduction achieved following exposure to 33% less light dose than required when exposed to 405 nm light alone. In conclusion, C. difficile vegetative cells and spores can be successfully inactivated using 405 nm light, the sporicidal efficacy can be significantly enhanced when exposed in the presence of low concentration chlorinated disinfectants. Further research may lead to the potential use of 405 nm light decontamination in combination with selected hospital disinfectants to enhance C. difficile cleaning and infection control procedures., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Objective: To investigate the use of 405 nm light for inhibiting the growth of selected species of dermatophytic and saprophytic fungi., Background Data: The increasing incidence and resilience of dermatophytic fungal infections is a major issue, and alternative treatment methods are being sought., Methods: The sensitivity of the dermatophytic fungi Trichophyton rubrum and Trichophyton mentagrophytes to 405 nm violet-blue light exposure was investigated, and the results compared with those obtained with the saprophytic fungus Aspergillus niger. Microconidia of T. rubrum and T. mentagrophytes and conidia of A. niger were seeded onto Sabauroud dextrose agar plates and irradiated with 405 nm light from an indium-gallium-nitride 99-DIE light-emitting diode (LED) array and the extent of inhibition was measured., Results: Germination of the microconidia of the Trichophyton species was completely inhibited using an irradiance of 35 mW/cm(2) for 4 h (dose of 504 J/cm(2)). A. niger conidia showed greater resistance, and colonial growth developed after light exposure. In liquid suspension tests, 405 nm light dose levels of 360, 720, and 1440 J/cm(2) resulted in complete inactivation of T. rubrum microconidia, whereas A. niger showed greater resistance, and at the highest dose level applied (1440 J/cm(2)) although A niger hyphae were completely inactivated, only a 3-log10 reduction of a 5-log10 conidial suspension was achieved., Conclusions: The study results demonstrate the relatively high sensitivity of Trichophyton microconidia to 405 nm violet-blue light, and this is may be of potential interest regarding the control and treatment of dermatophyte infections.

Bacterial contamination of injectable stored biological fluids such as blood plasma and platelet concentrates preserved in plasma at room temperature is a major health risk. Current pathogen reduction technologies (PRT) rely on the use of chemicals and/or ultraviolet light, which affects product quality and can be associated with adverse events in recipients. 405 nm violet-blue light is antibacterial without the use of photosensitizers and can be applied at levels safe for human exposure, making it of potential interest for decontamination of biological fluids such as plasma. As a pilot study to test whether 405 nm light is capable of inactivating bacteria in biological fluids, rabbit plasma and human plasma were seeded with bacteria and treated with a 405 nm light emitting diode (LED) exposure system (patent pending). Inactivation was achieved in all tested samples, ranging from low volumes to prebagged plasma. 99.9% reduction of low density bacterial populations (≤10 3  CFU mL -1 ), selected to represent typical "natural" contamination levels, was achieved using doses of 144 Jcm -2 . The penetrability of 405 nm light, permitting decontamination of prebagged plasma, and the nonrequirement for photosensitizing agents provide a new proof of concept in bacterial reduction in biological fluids, especially injectable fluids relevant to transfusion medicine.

High-intensity narrow-spectrum (HINS) light is a novel violet-blue light inactivation technology which kills bacteria through a photodynamic process, and has been shown to have bactericidal activity against a wide range of species. Specimens from patients with infected hip and knee arthroplasties were collected over a one-year period (1 May 2009 to 30 April 2010). A range of these microbial isolates were tested for sensitivity to HINS-light. During testing, suspensions of the pathogens were exposed to increasing doses of HINS-light (of 123mW/cm(2) irradiance). Non-light exposed control samples were also used. The samples were then plated onto agar plates and incubated at 37°C for 24 hours before enumeration. Complete inactivation (greater than 4-log10 reduction) was achieved for all of the isolates. The typical inactivation curve showed a slow initial reaction followed by a rapid period of inactivation. The doses of HINS-light required ranged between 118 and 2214 J/cm(2). Gram-positive bacteria were generally found to be more susceptible than Gram-negative. As HINS-light uses visible wavelengths, it can be safely used in the presence of patients and staff. This unique feature could lead to its possible use in the prevention of infection during surgery and post-operative dressing changes. Cite this article: Bone Joint J 2015;97-B:283-8., (©2015 The British Editorial Society of Bone & Joint Surgery.)

Background: It is acknowledged that activities such as dressing changes and bed sheet changes are high-risk events; creating surges in levels of airborne bacteria. Burns patients are particularly high dispersers of pathogens; due to their large, often contaminated, wound areas. Prevention of nosocomial cross-contamination is therefore one of the major challenges faced by the burns team. In order to assess the contribution of airborne spread of bacteria, air samples were taken repeatedly throughout and following these events, to quantify levels of airborne bacteria., Methods: Air samples were taken at 3-min intervals before, during and after a dressing and bed change on a burns patient using a sieve impaction method. Following incubation, bacterial colonies were enumerated to calculate bacterial colony forming units per m(3) (cfu/m(3)) at each time point. Statistical analysis was performed, whereby the period before the high-risk event took place acted as a control period. The periods during and after the dressing and bed sheet changes were examined for significant differences in airborne bacterial levels relative to the control period. The study was carried out four times, on three patients with burns between 35% total burn surface area (TBSA) and 51% TBSA., Results: There were significant increases in airborne bacteria levels, regardless of whether the dressing change or bed sheet change took place first. Of particular note, is the finding that significantly high levels (up to 2614cfu/m(3)) of airborne bacteria were shown to persist for up to approximately 1h after these activities ended., Discussion: This is the most accurate picture to date of the rapidly changing levels of airborne bacteria within the room of a burns patient undergoing a dressing change and bed change. The novel demonstration of a significant increase in the airborne bacterial load during these events has implications for infection control on burns units. Furthermore, as these increased levels remained for approximately 1h afterwards, persons entering the room both during and after such events may act as vectors of transmission of infection. It is suggested that appropriate personal protective equipment should be worn by anyone entering the room, and that rooms should be quarantined for a period of time following these events., Conclusion: Airborne bacteria significantly increase during dressing and sheet changes on moderate size burns, and remain elevated for up to an hour following their cessation., (Copyright © 2014 Elsevier Ltd and ISBI. All rights reserved.)

Background: Although the germicidal properties of ultraviolet (UV) light have long been known, it is only comparatively recently that the antimicrobial properties of visible violet-blue 405 nm light have been discovered and used for environmental disinfection and infection control applications., Aim: To review the antimicrobial properties of 405 nm light and to describe its application as an environmental decontamination technology with particular reference to disinfection of the hospital environment., Methods: Extensive literature searches for relevant scientific papers and reports., Findings: A large body of scientific evidence is now available that provides underpinning knowledge of the 405 nm light-induced photodynamic inactivation process involved in the destruction of a wide range of prokaryotic and eukaryotic microbial species, including resistant forms such as bacterial and fungal spores. For practical application, a high-intensity narrow-spectrum light environmental disinfection system (HINS-light EDS) has been developed and tested in hospital isolation rooms. The trial results have demonstrated that this 405 nm light system can provide continuous disinfection of air and exposed surfaces in occupied areas of the hospital, thereby substantially enhancing standard cleaning and infection control procedures., Conclusion: Violet-blue light, particularly 405 nm light, has significant antimicrobial properties against a wide range of bacterial and fungal pathogens and, although germicidal efficacy is lower than UV light, this limitation is offset by its facility for safe, continuous use in occupied environments. Promising results on disinfection efficacy have been obtained in hospital trials but the full impact of this technology on reduction of healthcare-associated infection has yet to be determined., (Copyright © 2014 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Exposure to narrowband violet-blue light around 405 nm wavelength can induce lethal oxidative damage to bacteria and fungi, however effects on viruses are unknown. As photosensitive porphyrin molecules are involved in the microbicidal inactivation mechanism, and since porphyrins are absent in viruses, then any damaging effects of 405 nm light on viruses might appear unlikely. This study used the bacteriophage ɸC31, as a surrogate for non-enveloped double-stranded DNA viruses, to establish whether 405 nm light can induce virucidal effects. Exposure of ɸC31 suspended in minimal media, nutrient-rich media, and porphyrin solution, demonstrated differing sensitivity of the phage. Significant reductions in phage titer occurred when exposed in nutrient-rich media, with ~3-, 5- and 7-log 10 reductions achieved after exposure to doses of 0.3, 0.5 and 1.4 kJ/cm 2 , respectively. When suspended in minimal media a 0.3-log 10 reduction ( P = 0.012) occurred after exposure to 306 J/cm 2 : much lower than the 2.7- and > 2.5-log 10 reductions achieved with the same dose in nutrient-rich, and porphyrin-supplemented media, suggesting inactivation is accelerated by the photo-activation of light-sensitive components in the media. This study provides the first evidence of the interaction of narrowband 405 nm light with viruses, and demonstrates that viral susceptibility to 405 nm light can be significantly enhanced by involvement of exogenous photosensitive components. The reduced susceptibility of viruses in minimal media, compared with that of other microorganisms, provides further evidence that the antimicrobial action of 405 nm light is predominantly due to the photo-excitation of endogenous photosensitive molecules such as porphyrins within susceptible microorganisms.

The antimicrobial effects of 405 nm light have generated interest in its use as an emerging disinfection technology with potential food-related applications. The aim of this study was to assess the bactericidal efficacy of 405 nm light for inactivation of Escherichia coli and Listeria monocytogenes under sub-lethally stressed environmental conditions. Bacteria were exposed to 405 nm light from a light emitting diode (LED) array under various temperature, salt (NaCl) and acid conditions to determine if bacterial susceptibility to 405 nm light inactivation is affected when exposed under these conditions. Non-stressed bacterial populations (10(5) CFU/mL) were exposed to increasing doses of 405 nm light (~70 mW/cm(2)) and the inactivation results were compared with those generated under stress conditions. Bacteria were held at various temperatures (4°C, 22°C and 45°C), acid concentrations (pH3, 3.5 and 7) and salt concentrations (0%, 0.8%, 10% and 15% NaCl), and simultaneously exposed to 405 nm light. Enhanced inactivation of both E. coli and L. monocytogenes was achieved when light exposure was combined with each of the sub-lethal stresses, with significantly increased inactivation rates compared to non-stressed populations (P≤0.05). One exception was with L. monocytogenes when light-exposed in the presence of 15% salt, as this combination reduced bacterial inactivation. The greatest enhancement of 405 nm light inactivation for both bacterial species was achieved when light exposure was combined with sub-lethal acid stress conditions at pH3. This was demonstrated by a 5-log10 reduction of E. coli following a 405 nm light dose of 84 J/cm(2) compared to 378 J/cm(2) for non-stressed populations (77% reduction in dose) and by a 5-log10 reduction of L. monocytogenes achieved with a dose of 42 J/cm(2) which corresponded to 50% of the dose required for the equivalent reduction of non-stressed populations. This acid-enhanced 405 nm light inactivation effect was demonstrated with E. coli and L. monocytogenes when dispersed in liquid suspension and when deposited on a test surface. Overall, results from this study have shown that sub-lethally stressed bacteria have increased susceptibility to 405 nm light inactivation, thereby providing a synergistic inactivation effect, findings which increase the potential of this new light-based decontamination technology for food related applications., (© 2013.)

Healthcare associated infections pose a major threat to patients admitted to hospitals and infection rates following orthopedic arthroplasty surgery are as high as 4%. A 405-nm high-intensity narrow spectrum light has been proven to reduce environmental contamination in hospital isolation rooms, and there is potential to develop this technology for application in arthroplasty surgery. Cultured rat osteoblasts were exposed to varying light intensities and it was found that exposures of up to a dose of 36 J/cm2 had no significant effect on cell viability [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay], function (alkaline phosphatase activity), and proliferation rate (BrdU cell proliferation assay). High irradiance exposures (54 J/cm2) significantly affected the cell viability indicating that the effects of 405-nm light on osteoblasts are dose dependent. Additionally, exposure of a variety of clinically related bacteria to a dose of 36 J/cm2 resulted in up to 100% kill. These results demonstrating the differential sensitivity of osteoblasts and bacteria to 405-nm light are an essential step toward developing the technique for decontamination in orthopedic surgery.

This study assessed the effects of high-intensity violet light on selected yeast and mould fungi. Cell suspensions of Saccharomyces cerevisiae, Candida albicans, and dormant and germinating spores (conidia) of the mould Aspergillus niger were exposed to high-intensity narrow band violet light with peak output at 405 nm generated from a light-emitting diode (LED) array. All three fungal species were inactivated by the 405-nm light without a requirement for addition of exogenous photosensitiser chemicals. Of the fungal species tested, S. cerevisiae was most sensitive and dormant conidia of A. niger were most resistant to 405-nm light exposure. Five-log10 colony forming units per millilitre (CFU ml(-1)) reductions of the tested species required exposure doses of 288 J cm(-2) for S. cerevisiae, 576 J cm(-2) for C. albicans, and a much higher value of 2.3 kJ cm(-2) for dormant conidia of A. niger. During germination, A. niger conidia became more sensitive to 405-nm light exposure and sensitivity increased as germination progressed over an 8 h test period. Light exposure under aerobic and anaerobic conditions, together with results obtained using ascorbic acid as a scavenger of reactive oxygen species, revealed that 405-nm light inactivation in fungi involved an oxygen-dependent mechanism, as previously described in bacteria. The inactivation results achieved with yeast cells and fungal spores together with operational advantages associated with the use of a visible (nonultraviolet (UV)) light source highlight the potential of 405-nm light for fungal decontamination applications., (Copyright © 2013 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.)

Attachment of bacteria to surfaces and subsequent biofilm formation remains a major cause of cross-contamination capable of inducing both food-related illness and nosocomial infections. Resistance to many current disinfection technologies means facilitating their removal is often difficult. The aim of this study was to investigate the efficacy of 405 nm light for inactivation of bacterial attached as biofilms to glass and acrylic. Escherichia coli biofilms (10(3)-10(8) CFU mL(-1)) were generated on glass and acrylic surfaces and exposed for increasing times to 405 nm light (5-60 min) at ca 140 mW cm(-2). Successful inactivation of biofilms has been demonstrated, with results highlighting complete/near-complete inactivation (up to 5 log10 reduction on acrylic and 7 log10 on glass). Results also highlight that inactivation of bacterial biofilms could be achieved whether the biofilm was on the upper "directly exposed" surface or "indirectly exposed" underside surface. Statistically significant inactivation was also shown with a range of other microorganisms associated with biofilm formation (Staphylococcus aureus, Pseudomonas aeruginosa and Listeria monocytogenes). Results from this study have demonstrated significant inactivation of bacteria ranging from monolayers to densely populated biofilms using 405 nm light, highlighting that with further development this technology may have potential applications for biofilm decontamination in food and clinical settings., (© 2013 Wiley Periodicals, Inc. Photochemistry and Photobiology © 2013 The American Society of Photobiology.)

Infection rates after arthroplasty surgery are between 1-4 %, rising significantly after revision procedures. To reduce the associated costs of treating these infections, and the patients' post-operative discomfort and trauma, a new preventative method is required. High intensity narrow spectrum (HINS) 405 nm light has bactericidal effects on a wide range of medically important bacteria, and it reduced bacterial bioburden when used as an environmental disinfection method in a Medical Burns Unit. To prove its safety for use for environmental disinfection in orthopaedic theatres during surgery, cultured osteoblasts were exposed to HINS-light of intensities up to 15 mW/cm2 for 1 h (54 J/cm2). Intensities of up to 5 mW/cm2 for 1 h had no effect on cell morphology, activity of alkaline phosphatase, synthesis of collagen or osteocalcin expression, demonstrating that under these conditions this dose is the maximum safe exposure for osteoblasts; after exposure to 15 mW/cm2 all parameters of osteoblast function were significantly decreased. Viability (measured by protein content and Crystal Violet staining) of the osteoblasts was not influenced by exposure to 5 mW/cm2 for at least 2 h. At 5 mW/cm2 HINS-light is an effective bactericide. It killed 98.1 % of Staphylococcus aureus and 83.2 % Staphylococcus epidermis populations seeded on agar surfaces, and is active against both laboratory strains and clinical isolates from infected hip and knee arthroplasties. HINS-light could have potential for development as a method of disinfection to reduce transmission of bacteria during arthroplasty, with wider applications in diverse surgical procedures involving implantation of a medical device.

Routine nursing activities such as dressing/bed changes increase bacterial dispersal from burns patients, potentially contaminating healthcare workers (HCW) carrying out these tasks. HCW thus become vectors for transmission of nosocomial infection between patients. The suspected relationship between %total body surface area (%TBSA) of burn and levels of bacterial release has never been fully established. Bacterial contamination of HCW was assessed by contact plate samples (n=20) from initially sterile gowns worn by the HCW during burns patient dressing/bed changes. Analysis of 24 gowns was undertaken and examined for relationships between %TBSA, time taken for activity, and contamination received by the HCW. Relationships between size of burn and levels of HCW contamination, and time taken for the dressing/bed change and levels of HCW contamination were best described by exponential models. Burn size correlated more strongly (R(2)=0.82, p<0.001) than time taken (R(2)=0.52, p<0.001), with levels of contamination received by the HCW. Contamination doubled with every 6-9% TBSA increase in burn size. Burn size was used to create a model to predict bacterial contamination received by a HCW carrying out bed/dressing changes. This may help with the creation of burn-specific guidelines on protective clothing worn by HCW caring for burns patients., (Copyright © 2012 Elsevier Ltd and ISBI. All rights reserved.)

Resistance of bacterial endospores to treatments, including biocides, heat and radiation is a persistent problem. This study investigates the susceptibility of Bacillus and Clostridium endospores to 405 nm visible light, wavelengths which have been shown to induce inactivation of vegetative bacterial cells. Suspensions of B. cereus endospores were exposed to high-intensity 405 nm light generated from a light-emitting diode array and results demonstrate the induction of a sporicidal effect. Up to a 4-log(10) CFU mL(-1) reduction in spore population was achieved after exposure to a dose of 1.73 kJ cm(-2). Similar inactivation kinetics were demonstrated with B. subtilis, B. megaterium and C. difficile endospores. The doses required for inactivation of endospores were significantly higher than those required for inactivation of B. cereus and C. difficile vegetative cells, where ca 4-log(10) CFU mL(-1) reductions were achieved after exposure to doses of 108 and 48 J cm(-2), respectively. The significant increase in dose required for inactivation of endospores compared with vegetative cells is unsurprising due to the notorious resilience of these microbial structures. However, the demonstration that visible light of 405 nm can induce a bactericidal effect against endospores is significant, and could have potential for incorporation into decontamination methods for the removal of bacterial contamination including endospores., (© 2012 Wiley Periodicals, Inc. Photochemistry and Photobiology © 2012 The American Society of Photobiology.)

The antimicrobial properties of light is an area of increasing interest. This study investigates the sensitivity of the significant foodborne pathogen Listeria monocytogenes to selected wavelengths of visible light. Results demonstrate that exposure to wavelength region 400-450 nm, at sufficiently high dose levels (750 J cm(-2)), induced complete inactivation of a 5 log(10) population. Exposure to wavelengths longer than 450 nm did not cause significant inactivation. Analysis of 10 nm bandwidths between 400 and 450 nm confirmed 405(± 5) nm light to be most effective for the inactivation of L. monocytogenes, with a lesser bactericidal effect also evident at other wavelengths between 400 and 440 nm. Identification of the optimum bactericidal wavelength enabled the comparison of inactivation using 405(± 5) nm filtered light and a 405 nm light-emitting diode (LED) array (14 nm FWHM). Results demonstrate similar inactivation kinetics, indicating that the applied dose of 405 nm light is the important factor. Use of the 405 nm LED array for the inactivation of L. monocytogenes and other Listeria species resulted in similar kinetics, with up to 5 log(10) reductions with a dose of 185 J cm(-2). Comparative data for the 405 nm light inactivation of L. monocytogenes and other important foodborne pathogens, Escherichia coli, Salmonella enteritidis and Shigella sonnei, are also presented, with L. monocytogenes showing higher susceptibility to inactivation through 405 nm light exposure., (© 2012 Wiley Periodicals, Inc. Photochemistry and Photobiology © 2012 The American Society of Photobiology.)

Pulsed electric field (PEF) treatment was examined as a potential decontamination method for tissue engineering biomatrices by determining the susceptibility of a range of microorganisms whilst within a collagen gel. High intensity pulsed electric fields were applied to collagen gel biomatrices containing either Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, Candida albicans, Saccharomyces cerevisiae or the spores of Aspergillus niger. The results established varying degrees of microbial PEF susceptibility. When high initial cell densities (10(6)-10(7) CFU ml(-1)) were PEF treated with 100 pulses at 45 kV cm(-1), the greatest log reduction was achieved with S. cerevisiae (~6.5 log(10) CFU ml(-1)) and the lowest reduction achieved with S. epidermidis (~0.5 log(10) CFU ml(-1)). The results demonstrate that inactivation is influenced by the intrinsic properties of the microorganism treated. Further investigations are required to optimise the microbial inactivation kinetics associated with PEF treatment of collagen gel biomatrices.

Infections are the leading cause of morbidity and mortality in burn patients and prevention of contamination from exogenous sources including the hospital environment is becoming increasingly emphasised. The High-Intensity Narrow-Spectrum light Environmental Decontamination System (HINS-light EDS) is bactericidal yet safe for humans, allowing continuous disinfection of the environment surrounding burn patients. Environmental samples were collected from inpatient isolation rooms and the outpatient clinic in the burn unit, and comparisons were then made between the bacterial contamination levels observed with and without use of the HINS-light EDS. Over 1000 samples were taken. Inpatient studies, with sampling carried out at 0800 h, demonstrated a significant reduction in the average number of bacterial colonies following HINS-light EDS use of between 27% and 75%, (p<0.05). There was more variation when samples were taken at times of increased activity in the room. Outpatient studies during clinics demonstrated a 61% efficacy in the reduction of bacterial contamination on surfaces throughout the room during the course of a clinic (p=0.02). The results demonstrate that use of the HINS-light EDS allows efficacious bacterial reductions over and above that achieved by standard cleaning and infection control measures in both inpatient and outpatient settings in the burn unit., (Copyright © 2011 Elsevier Ltd and ISBI. All rights reserved.)

The bactericidal effect of 405 nm light was investigated on taxonomically diverse bacterial pathogens from the genera Salmonella, Shigella, Escherichia, Listeria, and Mycobacterium. High-intensity 405 nm light, generated from an array of 405-nm light-emitting diodes (LEDs), was used to inactivate bacteria in liquid suspension and on exposed surfaces. L. monocytogenes was most readily inactivated in suspension, whereas S. enterica was most resistant. In surface exposure tests, L. monocytogenes was more susceptible than Gram-negative enteric bacteria to 405 nm light when exposed on an agar surface but interestingly less susceptible than S. enterica after drying onto PVC and acrylic surfaces. The study findings, that 405 nm light inactivates diverse types of bacteria in liquids and on surfaces, in addition to the safety advantages of this visible (non-UV wavelength) light, indicate the potential of this technology for a range of decontamination applications.

High-intensity narrow-spectrum (HINS) 405-nm light is a novel technology developed to address the significant problem of health-care associated infection. Its potential for wound-decontamination applications is assessed on mammalian cells and bacteria. The fibroblast-populated collagen lattice (FPCL) is used as an in vitro model of wound healing, and the effect of HINS light on contraction is examined. Effects on cell proliferation, morphological changes, and α-smooth muscle actin (α-SMA) expression are investigated. Bactericidal effects are assessed using the bacterium Staphylococcus epidermidis. Low doses of HINS light were found to have no significant inhibitory effects on FPCL contraction, cell proliferation, or α-SMA expression. Doses of up to 18 Jcm(-2) had no significant inhibitory effects on FPCL cell numbers, and this dose was shown to cause almost complete inactivation of bacteria. These results show that HINS light has potential for disinfection applications without adversely influencing wound healing.

Pulsed electric field (PEF) treatment has been proposed as a decontamination method for labile matrices used in tissue engineering applications. Through the application of PEF, a non-thermal treatment that causes bacterial inactivation through the irreversible rupture of microbial cell membranes, inactivation is achieved without loss of scaffold structure and function. However, some microorganisms are less susceptible to PEF treatment. This study shows that treatment with PEF and nisin, a food preservative bacteriocin, has a synergistic effect on the inactivation of Staphylococcus epidermidis in collagen gels. Almost complete inactivation of a 10(3) -10(4) CFU/mL S. epidermidis population was achieved when treated with a combination of PEF and 500 IU/mL nisin, with results demonstrating a 3.4 log(10) reduction, compared with 0.66 log(10) reduction with PEF alone. Nisin, at concentrations up to 3000 IU/mL, had no discernable toxicity to mammalian 3T3 cells when added to the culture medium or incorporated into the collagen gels. This combined decontamination method, involving PEF plus nisin, may provide a non-destructive process for inactivation of PEF-resistant bacteria in labile tissue engineering scaffolds., (Copyright © 2010 Wiley Periodicals, Inc.)

The performance of a new decontamination technology, referred to as 'high-intensity narrow-spectrum light environmental decontamination system' (HINS-light EDS) was evaluated by a series of three studies carried out in a hospital isolation room used to treat burns patients. The ceiling-mounted HINS-light EDS emits high-intensity 405nm light which, although bactericidal, is harmless to patients and staff thereby permitting continuous environmental disinfection throughout the day. Performance efficacy was assessed by contact agar plate sampling and enumeration of staphylococcal bacteria on environmental surfaces within the room before, during and after HINS-light EDS treatment. When the room was unoccupied, use of HINS-light EDS resulted in ∼90% reduction of surface bacterial levels and when the room was occupied by an MRSA-infected burns patient, reductions between 56% and 86% were achieved, with the highest reduction (86%) measured following an extended period of HINS-light EDS operation. In an on/off intervention study, surface bacterial levels were reduced by 62% by HINS-light EDS treatment and returned to normal contamination levels two days after the system was switched off. These reductions of staphylococci, including Staphylococcus aureus and meticillin-resistant S. aureus, by HINS-light EDS treatment were greater than the reductions achieved by normal infection control and cleaning activities alone. The findings provide strong evidence that HINS-light EDS, used as a supplementary procedure, can make a significant contribution to bacterial decontamination in clinical environments., (Copyright © 2010 The Hospital Infection Society. Published by Elsevier Ltd. All rights reserved.)

Although considerable research has been carried out on a range of environmental factors that impact on the survival of Campylobacter jejuni, there is limited information on the effects of violet/blue light on this pathogen. This investigation was carried out to determine the effects of high-intensity 405-nm light on C. jejuni and to compare this with the effects on two other important Gram-negative enteric pathogens, Salmonella enteritidis and Escherichia coli O157:H7. High-intensity 405-nm light generated from an array of 405-nm light-emitting diodes was used to inactivate the test bacteria. The results demonstrated that while all three tested species were susceptible to 405-nm light inactivation, C. jejuni was by far the most sensitive organism, requiring a total dose of 18 J cm⁻² of 405-nm light to achieve a 5-log₁₀ reduction. This study has established that C. jejuni is particularly susceptible to violet/blue light at a wavelength of 405 nm. This finding, coupled with the safety-in-use advantages of this visible (non-ultraviolet wavelength) light, suggests that high-intensity 405-nm light may have applications for control of C. jejuni contamination levels in situations where this type of illumination can be effectively applied.

This study demonstrates the susceptibility of a variety of medically important bacteria to inactivation by 405-nm light from an array of light-emitting diodes (LEDs), without the application of exogenous photosensitizer molecules. Selected bacterial pathogens, all commonly associated with hospital-acquired infections, were exposed to the 405-nm LED array, and the results show that both gram-positive and gram-negative species were successfully inactivated, with the general trend showing gram-positive species to be more susceptible than gram-negative bacteria. Detailed investigation of the bactericidal effect of the blue-light treatment on Staphylococcus aureus suspensions, for a range of different population densities, demonstrated that 405-nm LED array illumination can cause complete inactivation at high population densities: inactivation levels corresponding to a 9-log(10) reduction were achieved. The results, which show the inactivation of a wide range of medically important bacteria including methicillin-resistant Staphylococcus aureus, demonstrate that, with further development, narrow-spectrum 405-nm visible-light illumination from an LED source has the potential to provide a novel decontamination method with a wide range of potential applications.

Aims: To investigate the effectiveness of pulsed electric field (PEF) treatment as a new method for inactivation of micro-organisms in complex biomatrices and to assess this by quantifying the inactivation of Escherichia coli seeded in collagen gels., Methods and Results: PEF was applied to E. coli seeded collagen gels in static (nonflowing) chambers. The influence of electric field strength, pulse number and seeded cell densities were investigated. The highest level of inactivation was obtained at the maximum field strength of 45 kV cm(-1). For low levels of E. coli contamination (10(3) CFU ml(-1)), PEF treatment resulted in no viable E. coli being recovered from the gels. However, PEF treatment of gels containing higher cell densities (>or=10(4) CFU ml(-1)) did not achieve complete inactivation of E. coli., Conclusions: PEF treatment successfully inactivated E. coli seeded in collagen gels by 3 log(10) CFU ml(-1). Complete inactivation was hindered at high cell densities by the tailing effect observed., Significance and Impact of the Study: PEF shows potential as a novel, nondestructive method for decontamination of collagen-based matrices. Further investigation is required to ensure its compatibility with other proteins and therapeutic drugs for tissue engineering and drug delivery applications.

Exposure to visible-light causes the photoinactivation of certain bacteria by a process that is believed to involve the photo-stimulation of endogenous intracellular porphyrins. Studies with some bacterial species have reported that this process is oxygen-dependent. This study examines the role of oxygen in the visible-light inactivation of Staphylococcus aureus. Suspensions of S. aureus were exposed to broadband visible-light under both oxygen depletion and oxygen enhancement conditions to determine whether these environmental modifications had any effect on the staphylococcal inactivation rate. Oxygen enhancement was achieved by flowing oxygen over the surface of the bacterial sample during light inactivation and results demonstrated an increased rate of staphylococcal inactivation, with approximately 3.5 times less specific dose being required for inactivation compared to that for a non-enhanced control. Oxygen depletion, achieved through the addition of oxygen scavengers to the S. aureus suspension, further demonstrated the essential role of oxygen in the light inactivation process, with significantly reduced staphylococcal inactivation being observed in the presence of oxygen scavengers. The results of the present study demonstrate that the presence of oxygen is important for the visible-light inactivation of S. aureus, thus providing supporting evidence that the nature of the mechanism occurring within the visible-light-exposed staphylococci is photodynamic inactivation through the photo-excitation of intracellular porphyrins.

This study was conducted to investigate the bactericidal effects of visible light on methicillin-sensitive and methicillin-resistant Staphylococcus aureus (MRSA), and subsequently identify the wavelength sensitivity of S. aureus, in order to establish the wavelengths inducing maximum inactivation. Staphylococcus aureus, including MRSA strains, were shown to be inactivated by exposure to high-intensity visible light, and, more specifically, through a series of studies using a xenon broadband white-light source in conjunction with a selection of optical filters, it was found that inactivation of S. aureus occurs upon exposure to blue light of wavelengths between 400 and 420 nm, with maximum inactivation occurring at 405+/-5 nm. This visible-light inactivation was achieved without the addition of exogenous photosensitisers. The significant safety benefit of these blue-light wavelengths over UV light, in addition to their ability to inactivate medically important microorganisms such as MRSA, emphasises the potential of exploiting these non-UV wavelengths for disinfection applications.

Aims: To apply scanning electron microscopy, image analysis and a fluorescent viability stain to assess lethal and sublethal injury in food-borne bacteria exposed to pulsed-plasma gas discharges (PPGD)., Methods and Results: The fluorescent redox probe 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) was used for enumerating actively respiring cells of Campylobacter jejuni, Escherichia coli, Listeria monocytogenes, Staphylococcus aureus and Salmonella enterica serovar Typhimurium that were suspended in sterile water at 4 degrees C and exposed to separate PPGD and heat treatments. While there was good agreement between use of respiratory staining (RS) and direct-selective agar plate counting (PC) for enumerating untreated bacteria, there were c. 1 and 3 log-unit differences in surviving cell numbers per millilitre for test organisms subjected to PPGD and heat treatments respectively, when enumerated by these different viability indicators. PPGD-treated bacteria were markedly altered at the cellular level when examined by scanning electron microscopy., Conclusions: Use of this RS method revealed that substantial subpopulations of test bacteria rendered incapable of forming colonies by separate PPGD and heat treatments may remain metabolically active., Significance and Impact of the Study: Use of this RS method offers interesting perspectives on assessing established and novel microbial inactivation methods, and may also provide a better understanding of mechanisms involved in microbial inactivation induced by high-intensity PPGD treatments.

A pulsed-plasma gas-discharge (PPGD) system was developed for the novel decontamination of chilled poultry wash water. Treatment of poultry wash water in the plasma generation chamber for up to 24 s at 4 degrees C reduced Escherichia coli NCTC 9001, Campylobacter jejuni ATCC 33560, Campylobacter coli ATCC 33559, Listeria monocytogenes NCTC 9863, Salmonella enterica serovar Enteritidis ATCC 4931, and S. enterica serovar Typhimurium ATCC 14028 populations to non-detectable levels (< or = 8 log CFU/ml). Although similar PPGD treatments at 4 degrees C also produced significant reductions (> or = 3 log CFU/ml) in recalcitrant B. cereus NCTC 11145 endospore numbers within 30 s, the level of endospore reduction was dependent on the nature of the sparged gas used in the plasma treatments. Scanning electron microscopy revealed that significant damage occurred at the cellular level in PPGD-treated test organisms. This electrotechnology delivers energy in intense ultrashort bursts, generating products such as ozone, UV light, acoustic and shock waves, and pulsed electric fields that have multiple bactericidal properties. This technology offers an exciting complementary or alternative approach for treating raw poultry wash water and for preventing cross-contamination in processing environments.

Aims: To study the pulsed ultraviolet (UV) inactivation of poliovirus and adenovirus., Methods and Results: Viral suspensions of 2 ml volume were exposed to varying numbers of polychromatic light pulses emitted from a xenon flashlamp. Ten pulses produced an approximately 4 log(10) reduction in poliovirus titre, and no infectious poliovirus remained after 25 pulses. With adenovirus, 10 pulses resulted in an approximately 1 log(10) reduction in infectivity. Adenovirus required 100 pulses to produce an approximately 3 log(10) reduction in infectivity, and 200 pulses to produce a greater than 4 log(10) reduction., Conclusions: Adenovirus was more resistant to pulsed UV treatment than poliovirus although both viruses showed susceptibility to the treatment., Significance and Impact of the Study: Pulsed UV-light treatment proved successful in the inactivation of poliovirus and adenovirus, and represents an alternative to continuous-wave UV treatment.

Inactivation of Escherichia coli is examined using ultra-violet (UV) radiation from a pulsed xenon flashlamp. The light from the discharge has a broadband emission spectrum extending from the UV to the infrared region with a rich UV content. The flashlamp provides high-energy UV output using a small number of short-duration pulses (30 micros). The flashlamp is used with a monochromator to investigate the wavelength sensitivity of E. coli to inactivation by the pulsed UV light. Using 8 nm wide pulses of UV radiation, the most efficient inactivation is found to occur at around 270 nm and no inactivation is observed above 300 nm. A pyroelectric detector allows the energy dose to be determined at each wavelength, and a peak value for E. coli population reduction of 0.43 log per mJ/cm(2) is measured at 270 nm. The results are compared with the published data available for continuous UV light sources.

Aims: To apply scanning electron microscopy, image analysis and a fluorescent viability stain to assess lethal and sublethal in food-borne bacteria exposed to high-intensity pulsed electric fields (PEF)., Methods and Results: A rapid cellular staining method using the fluorescent redox probes 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) and 4',6-diamidino-2-phylindole was used for enumerating actively respiring cells of Listeria mononcytogenes, Bacillus cereus and Escherichia coli. This respiratory staining (RS) approach provided good agreement with the conventional plate count agar method for enumerating untreated and high-intensity PEF-treated bacteria suspended in 0.1% (w/v) peptone water. However, test organisms subjected to similar levels of lethality by heating at 56 degrees C resulted in ca 3-log-unit difference in surviving cell numbers ml(-1) when enumerated by these different viability indicators. PEF-treated bacteria were markedly altered at the cellular level when examined by scanning electron microscopy., Conclusions: While PEF-treatment did not produce sublethally injured cells (P < 0.05), substantial subpopulations of test bacteria rendered incapable of forming colonies by heating may remain metabolically active., Significance and Impact of the Study: The fluorescent staining method offers interesting perspectives on assessing established and novel microbial inactivation methods. Use of this approach may also provide a better understanding of the mechanisms involved in microbial inactivation induced by PEF.

The influence of treatment temperature and pulsed electric fields (PEF) on the viability of Mycobacterium paratuberculosis cells suspended in 0.1% (wt/vol) peptone water and in sterilized cow's milk was assessed by direct viable counts and by transmission electron microscopy (TEM). PEF treatment at 50 degrees C (2,500 pulses at 30 kV/cm) reduced the level of viable M. paratuberculosis cells by approximately 5.3 and 5.9 log(10) CFU/ml in 0.1% peptone water and in cow's milk, respectively, while PEF treatment of M. paratuberculosis at lower temperatures resulted in less lethality. Heating alone at 50 degrees C for 25 min or at 72 degrees C for 25 s (extended high-temperature, short-time pasteurization) resulted in reductions of M. paratuberculosis of approximately 0.01 and 2.4 log(10) CFU/ml, respectively. TEM studies revealed that exposure to PEF treatment resulted in substantial damage at the cellular level to M. paratuberculosis.

The physical effects of high-intensity pulsed electric fields (PEF) on the inactivation of diarrhoeagenic Bacillus cereus cells suspended in 0.1% peptone water were examined by transmission electron microscopy (TEM). The levels of PEF-induced microbial cell death were determined by enumeration on tryptone soy yeast extract agar and Bacillus cereus-selective agar plates. Following exposure to lethal levels of PEF, TEM investigation revealed irreversible cell membrane rupture at a number of locations, with the apparent leakage of intracellular contents. This study provides a clearer understanding of the mechanism of PEF-induced cellular damage, information that is essential for the further optimization of this emerging food-processing technology.