254-nm UVC Research Articles

Effect of intermittent 222 nm krypton-chlorine excilamp irradiation on microbial inactivation in water

In order to assure the microbial safety of drinking water, krypton-chlorine (KrCl) excilamp treatment has emerged as a possible technology to replace the use of conventional 254-nm low-pressure mercury UVC lamps. The aim of this study was to evaluate the disinfection efficiency of the KrCl excilamp emitting power narrow-band UVC radiation at 222-nm against a spectrum of bacteria at different initial populations in model contaminated water and to develop a strategy to effectively sterilize water contaminated with high concentrations of microorganisms. Different inoculum levels (104−5, 105−6, and 106−7 CFU/ml) of Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, and Listeria monocytogenes suspended in sterile distilled water were irradiated with a KrCl excilamp. In water of high inoculum density (106−7 CFU/ml), higher resistance to UVC was observed than in samples of medium and low cell density, as a high concentration of suspended cells lowered the transmittance of UVC rays. However, intermittent application of 222-nm UV irradiation in water containing a high inoculum level showed higher inactivation capacity against all three pathogens than continuous irradiation at the same dose. The results of this study provide an interesting insight into the use of an intermittent 222-nm UVC treatment system in batch-type water reservoir facilities to reduce the risk of waterborne disease.

Disinfection and healing effects of 222-nm UVC light on methicillin-resistant Staphylococcus aureus infection in mouse wounds

UVC radiation is known to be highly germicidal. However, exposure to 254-nm-UVC light causes DNA lesions such as cyclobutane pyrimidine dimers (CPD) in human cells, and can induce skin cancer after long-term repeated exposures. It has been reported that short wavelength UVC is absorbed by proteins in the membrane and cytosol, and fails to reach the nucleus of human cells. Hence, irradiation with 222-nm UVC might be an optimum combination of effective disinfection and biological safety to human cells. In this study, the biological effectiveness of 222-nm UVC was investigated using a mouse model of a skin wound infected with methicillin-resistant Staphylococcus aureus (MRSA). Irradiation with 222-nm UVC significantly reduced bacterial numbers on the skin surface compared with non-irradiated skin. Bacterial counts in wounds evaluated on days 3, 5, 8 and 12 after irradiation demonstrated that the bactericidal effect of 222-nm UVC was equal to or more effective than 254-nm UVC. Histological analysis revealed that migration of keratinocytes which is essential for the wound healing process was impaired in wounds irradiated with 254-nm UVC, but was unaffected in 222-nm UVC irradiated wounds. No CPD-expressing cells were detected in either epidermis or dermis of wounds irradiated with 222-nm UVC, whereas CPD-expressing cells were found in both epidermis and dermis irradiation with 254-nm UVC. These results suggest that 222-nm UVC light may be a safe and effective way to reduce the rate of surgical site and other wound infections.

Wide band gap Ga2O3 as efficient UV-C photocatalyst for gas-phase degradation applications.

α, β, γ, and δ polymorphs of 4.6-4.8eV wide band gap Ga2O3 photocatalysts were prepared via a soft chemistry route. Their photocatalytic activity under 254nm UV-C light in the degradation of gaseous toluene was strongly depending on the polymorph phase. α- and β-Ga2O3 photocatalysts enabled achieving high and stable conversions of toluene with selectivities to CO2 within the 50-90% range, by contrast to conventional TiO2 photocatalysts that fully deactivate very rapidly on stream in similar operating conditions with rather no CO2 production, no matter whether UV-A or UV-C light was used. The highest performances were achieved on the high specific surface area β-Ga2O3 photocatalyst synthesized by adding polyethylene glycol (PEG) as porogen before precipitation, with stable toluene conversion and mineralization rate into CO2 strongly overcoming those obtained on commercial β-Ga2O3. They were attributed to favorable physicochemical properties in terms of high specific surface area, small mean crystallite size, good crystallinity, high pore volume with large size mesopore distribution and appropriate surface acidity, and to the possible existence of a double local internal field within Ga3+ units. In the degradation of hydrogen sulfide, PEG-derived β-Ga2O3 takes advantage from its high specific surface area for storing sulfate, and thus for increasing its resistance to deactivation and the duration at total sulfur removal when compared to other β-Ga2O3 photocatalysts. So, we illustrated the interest of using high surface area β-Ga2O3 in environmental photocatalysis for gas-phase depollution applications.

Application of a 222-nm krypton-chlorine excilamp to control foodborne pathogens on sliced cheese surfaces and characterization of the bactericidal mechanisms

This study was conducted to investigate the basic spectral properties of a 222-nm krypton-chlorine (KrCl) excilamp and its inactivation efficacy against major foodborne pathogens on solid media, as well as on sliced cheese compared to a conventional 254-nm low-pressure mercury (LP Hg) lamp. Selective media and sliced cheese inoculated with Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, and Listeria monocytogenes were irradiated with a KrCl excilamp and a LP Hg lamp at the same dose. The KrCl excilamp showed full radiant intensity from the outset at a wide range of working temperatures, especially at low temperatures of around 0 to 10°C. Irradiation with 222nm UV-C showed significantly (P<0.05) higher inactivation capacity against all three pathogens than 254-nm radiation on both media and sliced cheese surfaces without generating many sublethally injured cells which potentially could recover. The underlying inactivation mechanisms of 222-nm KrCl excilamp treatment were evaluated by fluorescent staining methods and damage to cellular membranes and intracellular enzyme inactivation were the primary factors contributing to the enhanced bactericidal effect. The results of this study suggest that a 222-nm UV-C surface disinfecting system can be applied as an alternative to conventional LP Hg lamp treatment by the dairy industry.

Norovirus and MS2 inactivation kinetics of UV-A and UV-B with and without TiO2

Germicidal ultraviolet, such as 254-nm UV-C, is a common method of disinfection of pathogenic enteric viruses. However, the disinfection efficacies of UV-A or -B in terms of inactivating waterborne viruses such as norovirus have not been characterized. We evaluated the inactivation kinetics of MS2 bacteriophage and murine norovirus (MNV), a surrogate of human norovirus (NoV), by UV-A and -B. In addition to UV disinfection, we further investigated whether the presence of TiO2 could enhance the virus inactivation kinetics of UV-A and -B. Both MS2 and MNV were highly resistant to UV-A. However, the addition of TiO2 enhanced the efficacy of UV-A for inactivating these viruses. UV-A dose of 1379 mJ/cm2 resulted in a 4 log10 reduction. In comparison, UV-B alone effectively inactivated both MS2 and MNV, as evidenced by the 4 log10 reduction by 367 mJ/cm2 of UV-B. The addition of TiO2 increased the inactivation of MS2; however, it did not significantly increase the efficacy of UV-B disinfection for inactivating MNV. When these treatments were applied to field water such as groundwater, the results were generally consistent with the laboratory findings. Our results clearly indicated that UV-B is useful for the disinfection of waterborne norovirus. However, MNV was quite resistant to UV-A, and UV-A effectively inactivated the tested viruses only when used in combination with TiO2.

Protective effect of nicotinic acid on human albumin during UV-C irradiation

The potential of nicotinic acid (NiAc), a vitamin B3 compound with antioxidant activity, to protect human albumin against the damage caused by exposure to 254-nm UV-C radiation was investigated. Experiments performed at irradiance levels of 3.5–15 W m−2 and application times between 5 and 30 min showed that albumin undergoes rapid dose-dependent aggregation when subjected to irradiation. Addition of 0.2 mM NiAc to the albumin solution resulted in a remarkable reduction of protein damage. Protection provided by NiAc was effective up to radiation doses of about 30 kJ m−2, which are significantly higher than those required for complete microbial and viral inactivation.

Role of the Nfo and ExoA Apurinic/Apyrimidinic Endonucleases in Radiation Resistance and Radiation-Induced Mutagenesis of Bacillus subtilis Spores

The roles of DNA repair by apurinic/apyrimidinic (AP) endonucleases alone, and together with DNA protection by α/β-type small acid-soluble spore proteins (SASP), in Bacillus subtilis spore resistance to different types of radiation have been studied. Spores lacking both AP endonucleases (Nfo and ExoA) and major SASP were significantly more sensitive to 254-nm UV-C, environmental UV (>280 nm), X-ray exposure, and high-energy charged (HZE)-particle bombardment and had elevated mutation frequencies compared to those of wild-type spores and spores lacking only one or both AP endonucleases or major SASP. These findings further implicate AP endonucleases and α/β-type SASP in repair and protection, respectively, of spore DNA against effects of UV and ionizing radiation.

Roles of Small, Acid-Soluble Spore Proteins and Core Water Content in Survival of Bacillus subtilis Spores Exposed to Environmental Solar UV Radiation

Spores of Bacillus subtilis contain a number of small, acid-soluble spore proteins (SASP) which comprise up to 20% of total spore core protein. The multiple alpha/beta-type SASP have been shown to confer resistance to UV radiation, heat, peroxides, and other sporicidal treatments. In this study, SASP-defective mutants of B. subtilis and spores deficient in dacB, a mutation leading to an increased core water content, were used to study the relative contributions of SASP and increased core water content to spore resistance to germicidal 254-nm and simulated environmental UV exposure (280 to 400 nm, 290 to 400 nm, and 320 to 400 nm). Spores of strains carrying mutations in sspA, sspB, and both sspA and sspB (lacking the major SASP-alpha and/or SASP-beta) were significantly more sensitive to 254-nm and all polychromatic UV exposures, whereas the UV resistance of spores of the sspE strain (lacking SASP-gamma) was essentially identical to that of the wild type. Spores of the dacB-defective strain were as resistant to 254-nm UV-C radiation as wild-type spores. However, spores of the dacB strain were significantly more sensitive than wild-type spores to environmental UV treatments of >280 nm. Air-dried spores of the dacB mutant strain had a significantly higher water content than air-dried wild-type spores. Our results indicate that alpha/beta-type SASP and decreased spore core water content play an essential role in spore resistance to environmentally relevant UV wavelengths whereas SASP-gamma does not.

Antigenotoxic effects of three essential oils in diploid yeast ( Saccharomyces cerevisiae) after treatments with UVC radiation, 8-MOP plus UVA and MMS

Essential oils (EOs) extracted from medicinal plants such as Origanum compactum, Artemisia herba alba and Cinnamomum camphora are known for their beneficial effects in humans. The present study was undertaken to investigate their possible antigenotoxic effects in an eukaryotic cell system, the yeast Saccharomyces cerevisiae. The EOs alone showed some cytotoxicity and cytoplasmic petite mutations, i.e. mitochondrial damage, but they were unable to induce nuclear genetic events. In combination with exposures to nuclear mutagens such as 254-nm UVC radiation, 8-methoxypsoralen (8-MOP) plus UVA radiation and methylmethane sulfonate (MMS), treatments with these EOs produced a striking increase in the amount of cytoplasmic petite mutations but caused a significant reduction in revertants and mitotic gene convertants induced among survivors of the diploid tester strain D7. In a corresponding rho 0 strain, the level of nuclear genetic events induced by the nuclear mutagens UVC and 8-MOP plus UVA resulted in the same reduced level as the combined treatments with the EOs. This clearly suggests a close relationship between the enhancement of cytoplasmic petites (mitochondrial damage) in the presence of the EOs and the reduction of nuclear genetic events induced by UVC or 8-MOP plus UVA. After MMS plus EO treatment, induction of these latter events was comparable at least per surviving fraction in wildtype and rho 0 cells, and apparently less dependent on cytoplasmic petite induction. Combined treatments with MMS and EOs clearly triggered switching towards late apoptosis/necrosis indicating an involvement of this phenomenon in EO-induced cell killing and concomitant decreases in nuclear genetic events. After UVC and 8-MOP plus UVA plus EO treatments, little apoptosis and necrosis were observed. The antigenotoxic effects of the EOs appeared to be predominantly linked to the induction of mitochondrial dysfunction.

A Comparison of the Sensitivity of Photodamage Assays in Rat Basophilic Leukemia Cells¶

Many aspects of cellular function or physiology can be used to indicate the level of damage resulting from the application of potentially deleterious agents such as drugs, solvents or even light. The dose required to reach a specific biological endpoint will necessarily depend on the characteristics of the damage induced by the agent. By using multiple biological probes, it is possible to get a more complete description of the type of damage induced. Photodamage was induced in rat basophilic leukemia cells by either 254-nm UVC light exposure or rose bengal photosensitization. Damage was measured by three quantitative assays employing fluorescent probes: calcein, to measure nonspecific esterase activity, propidium iodide (PI), to measure loss of plasma membrane integrity, rhodamine 123 (R123) to measure mitochondrial depolarization, and the incorporation of 5′-bromodeoxyuridine (BrdU), to measure the progress of cell replication. BrdU incorporation was found to be the most sensitive indicator for both forms of photodamage. For UVC photodamage, the BrdU assay was 330 times more sensitive than the other two assays. For rose bengal photosensitization, the BrdU assay was 48 or 62 times more sensitive than either the R123 or calcein/PI assays, respectively.

A Comparison of the Sensitivity of Photodamage Assays in Rat Basophilic Leukemia Cells

Many aspects of cellular function or physiology can be used to indicate the level of damage resulting from the application of potentially deleterious agents such as drugs, solvents or even light. The dose required to reach a specific biological endpoint will necessarily depend on the characteristics of the damage induced by the agent. By using multiple biological probes, it is possible to get a more complete description of the type of damage induced. Photodamage was induced in rat basophilic leukemia cells by either 254-nm UVC light exposure or rose bengal photosensitization. Damage was measured by three quantitative assays employing fluorescent probes: calcein, to measure nonspecific esterase activity, propidium iodide (PI), to measure loss of plasma membrane integrity, rhodamine 123 (R123) to measure mitochondrial depolarization, and the incorporation of 5'-bromodeoxyuridine (BrdU), to measure the progress of cell replication. BrdU incorporation was found to be the most sensitive indicator for both forms of photodamage. For UVC photodamage, the BrdU assay was 330 times more sensitive than the other two assays. For rose bengal photosensitization, the BrdU assay was 48 or 62 times more sensitive than either the R123 or calcein/PI assays, respectively.

Role of the spore coat layers in Bacillus subtilis spore resistance to hydrogen peroxide, artificial UV-C, UV-B, and solar UV radiation.

Spores of Bacillus subtilis possess a thick protein coat that consists of an electron-dense outer coat layer and a lamellalike inner coat layer. The spore coat has been shown to confer resistance to lysozyme and other sporicidal substances. In this study, spore coat-defective mutants of B. subtilis (containing the gerE36 and/or cotE::cat mutation) were used to study the relative contributions of spore coat layers to spore resistance to hydrogen peroxide (H(2)O(2)) and various artificial and solar UV treatments. Spores of strains carrying mutations in gerE and/or cotE were very sensitive to lysozyme and to 5% H(2)O(2), as were chemically decoated spores of the wild-type parental strain. Spores of all coat-defective strains were as resistant to 254-nm UV-C radiation as wild-type spores were. Spores possessing the gerE36 mutation were significantly more sensitive to artificial UV-B and solar UV radiation than wild-type spores were. In contrast, spores of strains possessing the cotE::cat mutation were significantly more resistant to all of the UV treatments used than wild-type spores were. Spores of strains carrying both the gerE36 and cotE::cat mutations behaved like gerE36 mutant spores. Our results indicate that the spore coat, particularly the inner coat layer, plays a role in spore resistance to environmentally relevant UV wavelengths.

Artificial and solar UV radiation induces strand breaks and cyclobutane pyrimidine dimers in Bacillus subtilis spore DNA.

The loss of stratospheric ozone and the accompanying increase in solar UV flux have led to concerns regarding decreases in global microbial productivity. Central to understanding this process is determining the types and amounts of DNA damage in microbes caused by solar UV irradiation. While UV irradiation of dormant Bacillus subtilis endospores results mainly in formation of the "spore photoproduct" 5-thyminyl-5,6-dihydrothymine, genetic evidence indicates that an additional DNA photoproduct(s) may be formed in spores exposed to solar UV-B and UV-A radiation (Y. Xue and W. L. Nicholson, Appl. Environ. Microbiol. 62:2221-2227, 1996). We examined the occurrence of double-strand breaks, single-strand breaks, cyclobutane pyrimidine dimers, and apurinic-apyrimidinic sites in spore DNA under several UV irradiation conditions by using enzymatic probes and neutral or alkaline agarose gel electrophoresis. DNA from spores irradiated with artificial 254-nm UV-C radiation accumulated single-strand breaks, double-strand breaks, and cyclobutane pyrimidine dimers, while DNA from spores exposed to artificial UV-B radiation (wavelengths, 290 to 310 nm) accumulated only cyclobutane pyrimidine dimers. DNA from spores exposed to full-spectrum sunlight (UV-B and UV-A radiation) accumulated single-strand breaks, double-strand breaks, and cyclobutane pyrimidine dimers, whereas DNA from spores exposed to sunlight from which the UV-B component had been removed with a filter ("UV-A sunlight") accumulated only single-strand breaks and double-strand breaks. Apurinic-apyrimidinic sites were not detected in spore DNA under any of the irradiation conditions used. Our data indicate that there is a complex spectrum of UV photoproducts in DNA of bacterial spores exposed to solar UV irradiation in the environment.

Mutagenic specificity of solar UV light in nucleotide excision repair-deficient rodent cells.

To investigate the role of nucleotide excision repair (NER) in the cellular processing of carcinogenic DNA photoproducts induced by defined, environmentally relevant portions of the solar wavelength spectrum, we have determined the mutagenic specificity of simulated sunlight (310-1100 nm), UVA (350-400 nm), and UVB (290-320 nm), as well as of the "nonsolar" model mutagen 254-nm UVC, at the adenine phosphoribosyltransferase (aprt) locus in NER-deficient (ERCC1) Chinese hamster ovary (CHO) cells. The frequency distributions of mutational classes induced by UVB and by simulated sunlight in repair-deficient CHO cells were virtually identical, each showing a marked increase in tandem CC-->TT transitions relative to NER-proficient cells. A striking increase in CC-->TT events was also previously documented for mutated p53 tumor-suppressor genes from nonmelanoma tumors of NER-deficient, skin cancer-prone xeroderma pigmentosum patients, compared to normal individuals. The data therefore indicate that the aprt gene in NER-deficient cultured rodent cells irradiated with artificial solar light generates the same distinctive "fingerprint" for sunlight mutagenesis as the p53 locus in NER-deficient humans exposed to natural sunlight in vivo. Moreover, in strong contrast to the situation for repair-component CHO cells, where a significant role for UVA was previously noted, the mutagenic specificity of simulated sunlight in NER-deficient CHO cells and of natural sunlight in humans afflicted with xeroderma pigmentosum can be entirely accounted for by the UVB portion of the solar wavelength spectrum.

Site and strand specificity of UVB mutagenesis in the SUP4-o gene of yeast.

DNA sequencing was used to characterize 208 mutations induced in the SUP4-o tRNA gene of the yeast Saccharomyces cerevisiae by UVB (285-320 nm) radiation. The results were compared to those for an analysis of 211 SUP4-o mutations induced by 254-nm UVC light. In each case, greater than 90% of the mutations were single base-pair changes but G.C----A.T transitions predominated and accounted for more of the mutations induced by UVB than UVC. Double substitutions, single base-pair deletions, and more complex events were also recovered. However, UVB induced 3-fold more tandem substitutions than UVC and nontandem double events were detected only after irradiation with UVC. Virtually all induced substitutions occurred at sites where the pyrimidine of the base pair was part of a dipyrimidine sequence. Although the site specificities were consistent with roles for cyclobutane dimers and pyrimidine-pyrimidone(6-4) lesions in mutation induction, preliminary photoreactivation data implicated cyclobutane dimers as the major form of premutational DNA damage for both agents. Intriguingly, there was a preference for both UVB- and UVC-induced mutations to occur at sites where the dipyrimidine was on the transcribed strand.