Pulsed UV-light inactivation of poliovirus and adenovirus

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.     

Studies on the inactivation of medically important Candida species on agar surfaces using pulsed light

Development of a pulsed-light (PL) approach to inanimate surface decontamination is timely, as the incidence of yeast-related infections in healthcare remains unacceptably high. Critical electrical and biological factors governing the efficacy of PL for the in vitro inactivation of medically important yeast were established in this study. Predetermined cell numbers of yeast were inoculated separately on agar plates and were flashed with ≤90 pulses of broad-spectrum light under varying operating conditions, and their inactivation was measured. Significant differences in inactivation among different yeasts occurred depending on the intensity of the applied lamp discharge energy and the amount of pulsing applied. Levels of yeast sensitivity also varied depending on the distance between the light source and the treatment surface used, and the population size, type and age of cultures treated. Yeast strains were shown to be significantly more resistant to PL irradiation compared with similarly treated bacterial control cultures. A clear relationship was observed between the concentration of eluted proteins from treated yeast and the severity of PL conditions, with scanning electron micrographs showing irreversible cellular damage. Therefore, the findings from this study will enable further development and optimization of PL as a method of decontaminating surfaces in healthcare setting.     

Investigation of critical inter‐related factors affecting the efficacy of pulsed light for inactivating clinically relevant bacterial pathogens

Aims: To investigate critical electrical and biological factors governing the efficacy of pulsed light (PL) for the in vitro inactivation of bacteria isolated from the clinical environment. Development of this alternative PL decontamination approach is timely, as the incidence of health care–related infections remains unacceptably high. Methods and Results: Predetermined cell numbers of clinically relevant Gram‐positive and Gram‐negative bacteria were inoculated separately on agar plates and were flashed with ≤60 pulses of broad‐spectrum light under varying operating conditions, and their inactivation measured. Significant differences in inactivation largely occurred depending on the level of the applied lamp discharge energy (range 3·2–20 J per pulse), the amount of pulsing applied (range 0–60 pulses) and the distance between light source and treatment surface (range 8–20 cm) used. Greater decontamination levels were achieved using a combination of higher lamp discharge energies, increased number of pulses and shorter distances between treatment surface and the xenon light source. Levels of microbial sensitivity also varied depending on the population type, size and age of cultures treated. Production of pigment pyocynanin and alginate slime in mucoid strains of Pseudomonas aeruginosa afforded some protection against lethal action of PL; however, this was evident only by using a combination of reduced amount of pulsing at the lower lamp discharge energies tested. A clear pattern was observed where Gram‐positive bacterial pathogens were more resistant to cidal effects of PL compared to Gram negatives. While negligible photoreactivation of PL‐treated bacterial strains occurred after full pulsing regimes at the different lamp discharge energies tested, some repair was evident when using a combination of reduced pulsing at the lower lamp discharge energies. Strains harbouring genes for multiple resistances to antibiotics were not significantly more resistant to PL treatments. Slight temperature rises (≤4·2°C) were measured on agar surfaces after extended pulsing at higher lamp discharge energies. Presence of organic matter on treatment surface did not significantly affect PL decontamination efficacy, nor did growth of PL‐treated bacteria on selective agar diminish survival compared to similarly treated bacteria inoculated and enumerated on nonselective agar plates. Conclusions: Critical inter‐related factors affecting the effective and repeatable in vitro decontamination performance of PL were identified during this study that will aid further development of this athermal process technology for applications in health care and in industry. Very rapid reductions (c. 7 log₁₀ CFU cm^?2 within ≤10 pulses) occurred using discharge energy of 20 J for all tested clinically relevant bacteria under study when treated at 8 cm distance from xenon light source. While no resistant flora is expected to develop for treatment of microbial pathogens on two‐dimensional surfaces, careful consideration of scale up factors such as design and operational usage of this PL technique will be required to assure operator safety. Significance and Impact of the Study: Findings and conclusions derived from this study will enable further development and optimization of this decontamination technique in health care and in food preparation settings, and will advance the field of nonthermal processing technologies.     

Recent findings in pulsed light disinfection

Nonthermal disinfection technologies are gaining increasing interest in the field of minimally processed food in order to improve the microbial safety or to extend the shelf life. Especially fresh‐cut produce or meat and fish products are vulnerable to microbial spoilage, but, due to their sensitivity, they require gentle preservation measures. The application of intense light pulses of a broad spectral range comprising ultraviolet, visible and near infrared irradiation is currently investigated as a potentially suitable technology to reduce microbial loads on different food surfaces or in beverages. Considerable research has been performed within the last two decades, in which the impact of various process parameters or microbial responses as well as the suitability of pulsed light (PL) for food applications has been examined. This review summarizes the outcome of the latest studies dealing with the treatment of various foods including the impact of PL on food properties as well as recent findings about the microbicidal action and relevant process parameters.     

Advanced high-power pulsed light device to decontaminate food from pathogens: effects on Salmonella typhimurium viability in vitro

AIMS: The aim of this study was to construct an advanced high-power pulsed light device for decontamination of food matrix and to evaluate its antibacterial efficiency. Key parameters of constructed device-emitted light spectrum, pulse duration, pulse power density, frequency of pulses, dependence of emitted spectrum on input voltage, irradiation homogenicity, possible thermal effects as well as antimicrobial efficiency were evaluated. METHODS AND RESULTS: Antimicrobial efficiency of high-power pulsed light technique was demonstrated and evaluated by two independent methods - spread plate and Miles-Misra method. Viability of Salmonella typhimurium as function of a given light dose (number of pulses) and pulse frequency was examined. According to the data obtained, viability of Salmonella typhimurium reduced by 7 log order after 100 light pulses with power density 133 W cm(-2). In addition, data indicate, that the pulse frequency did not influence the outcome of pathogen inactivation in the region 1-5 Hz. Moreover, no hyperthermic effect was detected during irradiation even after 500 pulses on all shelves with different distance from light source and subsequently different pulse power density (0-252 W cm(-2)). CONCLUSION: Newly constructed high-power pulsed light technique is effective nonthermal tool for inactivation of Salmonella typhimurium even by 7 log order in vitro. SIGNIFICANCE AND IMPACT OF THE STUDY: Novel advanced high-power pulsed light device can be a useful tool for development of nonthermal food decontamination technologies.     

Kinetics of UV(254) inactivation of selected viral pathogens in a static system

Aims: The objective of this study was to estimate UV₂₅₄ inactivation constants for four viral pathogens: influenza virus type A, porcine respiratory and reproductive syndrome virus (PRRSV), bovine viral diarrhoea virus (BVDV) and reovirus. Methods and Results: Viruses in culture medium were exposed to one of nine doses of UV₂₅₄ and then titrated for infectious virus. Analysis showed that viral inactivation by UV₂₅₄ was more accurately described by a two‐stage inactivation model vs a standard one‐stage inactivation model. Conclusions: The results provided evidence for the existence of two heterogeneous viral subpopulations among the viruses tested, one highly susceptible to UV₂₅₄ inactivation and the other more resistant. Importantly, inactivation constants based on the one‐stage inactivation model would have underestimated the UV₂₅₄ dose required for the inactivation of these viruses under the conditions of the experiment. Significance and Impact of the Study: To improve the accuracy of estimates, it is recommended that research involving the inactivation of micro‐organisms evaluates inactivation kinetics using both one‐stage and two‐stage models. These results will be of interest to persons responsible for microbial agents under laboratory or field conditions.     

The effect of repeated pulses of light at the same time on period responses of the rat circadian pacemaker

A consequence of simple velocity-based models is that, in response to light pulses, the circadian period should adjust inversely to phase. In addition, because of the interaction of circadian period and phase response, earlier circadian period changes should modify later circadian period changes. The literature contains few mentions of response curves of circadian period responses following light pulses. Rats were exposed to four pulses of light (60 minutes, 1000 lux) at the same circadian time, a minimum of 26 days apart; we assessed period responses and possible bias in the period-response curve. Modulation of circadian period following light-induced phase responses was examined by assessing the period of running wheel activity onset. Phase and circadian period were not consistently found to share an inverse relationship. Moreover, biases in initial period tended to be increased by the experimental protocol regardless of circadian time of pulse. Rats with a short initial (high-velocity) period had a lengthened period, while rats with a long initial period (low velocity) tended to have a reduce period. However, rats with a long initial period were phase delay biased, not phase advance biased. These results do not support a simple velocity model of the pacemaker. (Chronobiology International, 18(2), 187-201, 2001)     

High-intensity narrow-spectrum light inactivation and wavelength sensitivity of Staphylococcus aureus

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.     

The effect of low intensity ultraviolet‐C light on monoclonal antibodies

As part of an investigation to identify potential new viral reduction strategies, ultraviolet‐C (UV‐C) light was examined. Although this technology has been known for decades to possess excellent virus inactivation capabilities, UV‐C light can also introduce significant unwanted damage to proteins. To study the effect on monoclonal antibodies, three different antibodies were subjected to varying levels of UV‐C light using a novel dosing device from Bayer Technology Services GmbH. The range of fluencies (or doses) covered was between 0 and 300 J/m² at a wavelength of 254 nm. Product quality data generated from the processed pools showed only minimal damage done to the antibodies. Aggregate formation was low for two of the three antibodies tested. Acidic and basic variants increased for all three antibodies, with the basic species increasing more than the acidic species. Peptide maps made for the three sets of pools showed no damage to two of the three antibody backbones, whereas the third antibody had very low levels of methionine oxidation evident. Samples held at 2–8°C for 33 days showed no increase in aggregates or charge variants, indicating that the proteins did not degrade and were not damaged further by reactive or catalytic species that may have been created on exposure to UV‐C light. Overall, UV‐C light was shown to induce very little damage to monoclonal antibodies at lower fluencies and appears to be a viable option for viral inactivation in biotechnology applications. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009     

Optimization of the photodynamic inactivation of prions by a phthalocyanine photosensitizer: The crucial involvement of singlet oxygen

Prion disorders are fatal neurodegenerative diseases caused by the autocatalytic conversion of a natively occurring prion protein (PrP^C) into its misfolded infectious form (PrP^TSE). The proven resistance of PrP^TSE to common disinfection procedures increases the risk of prion transmission in medical settings. Herein, we present the effective photodynamic inactivation (PDI) of prions by disulfonated hydroxyaluminum phthalocyanine (AlPcOH(SO₃)₂) utilizing two custom‐built red light sources. The treatment eliminates PrP^TSE signal in infectious mouse brain homogenate with efficiency that depends on light intensity but has a low effect on the overall protein content. Importantly, singlet oxygen (O₂(¹Δ_g)) is the only species significantly photogenerated by AlPcOH(SO₃)₂, and it is responsible for the PDI of prions. More intensive light conditions show not only higher O₂(¹Δ_g) production but also decreases in AlPcOH(SO₃)₂ photostability. Our findings suggest that PDI by AlPcOH(SO₃)₂‐generated O₂(¹Δ_g) represents a promising approach for prion inactivation that may be useful in future decontamination strategies for delicate medical tools.     

A study on the inactivation of micro-organisms and enzymes by high pressure CO2

This study addresses some microbial inactivation phenomena induced by high pressure CO2 over micro-organisms and enzymes. The activity of four selected enzymes was measured before and after treatment with CO2 under pressure in both buffer solutions and natural cellular environment (E. coli cells and tomato paste). Results are reported for acid phosphatase, alkaline phosphatase, ATPase, and pectinase at different conditions of temperature, CO2 pressure, and treatment time (32-40 degrees C, 85-150 bar, 30-70 min). The results obtained show that the high pressure CO2 treatment induces an inactivation of cellular enzymatic activity higher than the one caused on the same enzymes in solution. However, the measured activity difference is not caused by a damage at the enzymes molecular level but is a consequence of the permeabilization of the cellular envelopes which leads to a release of unmodified enzymes from the cells with simultaneous drop of enzymatic cellular activity. The reported data suggest that the bacterial cell death is probably due not to a selective effect of high pressure CO2 treatment but to simultaneous detrimental action of CO2 on cellular membrane and cell wall.     

Superoxide dismutase in the anal gills of the mosquito larvae of Aedes aegypti: its inhibition by alpha-terthienyl.

The anal gills of the mosquito larvae of Aedes aegypti were shown to possess superoxide dismutase (EC 1.15.1.1) activity, which increased with the maturation of the larvae from instar 1 to instar 4. This enzyme was highly inhibited upon treatment of the larvae with alpha-terthienyl (2,2':5,2"-terthiophene) and subsequent exposure to long-wave ultraviolet light. Inhibition also occurred with treatment of the crude enzyme extract in a similar fashion. Exposure of the enzyme to the ultraviolet light alone or the treatment of the enzyme with alpha-terthienyl in darkness could not manifest this inhibition. This finding adds a new dimension to the complex mechanism(s) proposed for the photodynamic toxicity of alpha-terthienyl.     

Application of pulsed light technology for fruits and vegetables disinfection: A review

Non-thermal technologies can maintain fruit and vegetable products quality better than traditional thermal processing. Pulsed light (PL) is a non-thermal method for microbial inactivation (vegetative cells and spores) in fruits and vegetables. The PL treatment involves the application of intense and short-duration pulses of broad spectrum wavelengths ranging from UV to near-infrared (100–1100 nm). This review summarized application of PL technology to control microbial contamination and increasing shelf-life of some fruits and vegetables including apple, blueberries, grape, orange, strawberries, carrot, lettuce, spinach, and tomato. The microbial inactivation in very short treatment times, low energy used by this system, flexibility for solid or liquid samples, few residual compounds and no synthetic chemicals that cause environmental pollution or harm humans, is benefits of PL technique. The efficiency of PL disinfection is closely associated with the input voltage, fluence (energy dose), composition of the emitted light spectrum, number of lamps, the distance between samples and light source, and frequency and number of applied pulses. The PL treatments control pathogenic and spoilage microorganisms, so it facilitates the growth and development of the starter microorganisms affecting product quality.     

Modelling and optimization of inactivation of Lactobacillus plantarum by pulsed electric field treatment

AIMS: The effect of critical pulsed electric field (PEF) process parameters, such as electric field strength, pulse length and number of pulses, on inactivation of Lactobacillus plantarum was investigated. METHODS AND RESULTS: Experiments were performed in a pH 4.5 sodium phosphate buffer having a conductivity of 0.1 S m-1, using a laboratory-scale continuous PEF apparatus with a co-linear treatment chamber. An inactivation model was developed as a function of field strength, pulse length and number of pulses. Based on this inactivation model, the conditions for a PEF treatment were optimized with respect to the minimum energy required to obtain a certain level of inactivation. It was shown that the least efficient process parameter in the range investigated was the number of pulses. The most efficient way to optimize inactivation of Lact. plantarum was to increase the field strength up to 25.7 kV cm-1, at the shortest pulse length investigated, 0.85 micros, and using a minimum number of pulses. The highest inactivation of Lact. plantarum at the lowest energy costs is obtained by using the equation: E=26.7tau0.23, in which E is the field strength and tau the pulse length. An optimum is reached by substituting tau with 5.1. CONCLUSIONS: This study demonstrates that the correct choice of parameters, as predicted by the model described here, can considerably improve the PEF process. SIGNIFICANCE AND IMPACT OF THE STUDY: The knowledge gained in this study improves the understanding of the limitations and opportunities of the PEF process. Consequently, the advantage of the PEF process as a new option for non-thermal decontamination can be better utilized.     

新型强脉冲光治疗雀斑临床疗效观察

目的:观察新型强脉冲光治疗雀斑的疗效。方法:采用飞顿辉煌激光360嫩肤系统(以色列飞顿公司),波长570～950nm,光斑面积10mm×30mm,脉宽10、12、15ms,能量14～19J/cm~2。治疗40例雀斑患者,每三周一次,共治疗5次,末次治疗后评价患者雀斑的疗效。结查:40例患者经过治疗后,18例(45%)基本完全消退,14例(35%)明显消退,8例(20%)好转,总有效率为100%。所有患者面部治疗区域皮肤质地较以前更光滑、细腻,无严重不良反应出现。结论:采用新型强脉冲光治疗雀斑疗效显著、安全,副作用少。     

Effects of light pulses on the locomotor activity rhythm of Orchestia montagui (Amphipoda,Talitridae)

Animals of the amphipod Orchestia montagui are kept in constant darkness with two short light pulses. One pulse is applied at the beginning of subjective night (around the dusk) and the other one at the end of subjective night (around the dawn). The pulse duration is estimated in the order of one or two hours around the dusk as well as the dawn. The locomotor activity rhythm was monitored in individual animals in summer under constant temperature. Results revealed that whatever the experimental conditions, under continuous or interrupted darkness by pulses, two endogenous components have been highlighted. In fact, Periodogram analysis showed the presence of ultradian and circadian periods around 12 and 24 h, respectively. The shortest circadian period and the most important inter-individual variability was observed under pulse of 2 h around the dusk with mean value equal to τ_DD+pulse = 24h38′ ± 4h34′. The activity profiles are in majority unimodal. Moreover, the most activity peak showed a slipping of its location from the middle of subjective night under constant darkness to the middle of subjective day under pulse. Globally, the locomotor activity rhythm of O. montagui was better defined under pulses and specimens were significantly more active under continuous darkness. Moreover, a great variability around the activity time was observed especially with pulse of 1 h.     

非剥脱性嫩肤技术在皮肤光老化中的应用

目的：介绍国内外近10年来非剥脱性嫩肤技术治疗皮肤光老化临床研究概况,以期进一步了解非剥脱性嫩肤技术在皮肤光老化中的应用新进展。方法：应用计算机检索Pubmed和中国生物医学文献数据库1999／2009的相关文章,限定文章语言种类为英文和中文,检索词“photoaging（光老化）,laser（激光）,Intense pulsed light（强光）,radiofrequency（射频）,Light Emitting Diode（发光二极管）”。对资料进行初审,选择临床实验研究文献查找全文。纳入标准：①有明确诊断标准。②随机对照实验或对照实验。结果：共收集到相关文献110篇,按上述标准纳入23篇,其余文献均被排除。结论：非剥脱性嫩肤技术治疗皮肤光老化疗效可靠,具有副作用少,方法简单,安全性高,易被患者接受等优势。但不管单独使用哪种方法治疗皮肤光老化其效果均不是很完美,必需紧密结合患者疾病的具体情况和非剥脱性嫩肤设备的性能进行个性化的治疗。     

Interaction between the effects of light pulses of different chromatic content on Hydra attenuata and H. magnipapillata

The pattern of Hydra periodic shortening-elongation behaviour can be modified by photic stimulation. The response time between the end of a pulse stimulus and the beginning of the next body shortening event is a function of: 1) the polarity of the pulse; 2) the intensity of the pulse, 3) the phase of application of the pulse during a shortening-elongation period, 4) the wavelength of the pulse, 5) the chromatic composition of the background illumination. The combined effects of pulse light stimuli of different wavelength applied either simultaneously or in immediate sequence are reported here. The results give some insight into the possible mechanism of phototransduction.     

Selective fluorophore-assisted light inactivation of voltage-gated calcium channels

Fluorophore-assisted light inactivation (FALI) is an investigative tool to inactivate fluorescently labeled proteins by a mechanism of in situ photodestruction. We found that Cav 1.2 (L-type) and Cav 3.1 (T-type) calcium channels, labeled by genetic fusion with GFP derivatives, show differential sensitivity to FALI. Specifically, FALI silences Cav 1.2 calcium channels containing EYFP-labeled α 1C subunits but does not affect the EYFP-α 1G Cav 3.1 calcium channels or Cav 1.2 channels containing EYFP-labeled β subunits. Our findings limit the applicability of acceptor photobleaching for the measurements of FRET but open an opportunity to combine the fluorescent imaging of the live cell expressing labeled calcium channels with selective functional inactivation of their specific subsets.