Rejoining of gamma-ray-induced DNA damage in Cryptosporidium parvum measured by the comet assay

Cryptosporidium parvum is a well-known waterborne intracellular protozoan that causes severe diarrheal illness in immunocompromised individuals. This organism is highly resistant to harsh environmental conditions and various disinfectants, and it exhibits one of the highest known resistances to gamma irradiation. We investigated rejoining of gamma-ray-induced DNA damage in C. parvum by neutral comet assay. Oocysts were gamma irradiated at various doses (1, 5, 10, and 25 kGy) and were incubated for various periods (6-96 h) after exposure to 10 kGy. The comet tail moment showed that the number of DNA double-strand breaks increased concomitantly with the gamma irradiation dose. When investigating rejoining after irradiation at 10 kGy, double-strand breaks peaked at 6 h postirradiation, and rejoining was highest at 72 h postirradiation. The observed rejoining pattern suggests that repair process occurs slowly even when complex DNA double-strand breaks in C. parvum were induced by high dose irradiation, 10 kGy.     

Alterations in damage processes in dense cancellous bone following gamma-radiation sterilization

Structurally intact cancellous bone allograft is an attractive tissue form because its high porosity can provide space for delivery of osteogenic factors and also allows for more rapid and complete in-growth of host tissues. Gamma radiation sterilization is commonly used in cancellous bone allograft to prevent disease transmission. Commonly used doses of gamma radiation sterilization (25–35 kGy) have been shown to modify cortical bone post-yield properties and crack propagation but have not been associated with changes in cancellous bone material properties. The purpose of this study was to determine the effects of irradiation on the elastic and yield properties and microscopic tissue damage processes in dense cancellous bone. Cancellous bone specimens (13 control, 14 irradiated to 30 kGy) from bovine proximal tibiae were tested in compression to 1.3% apparent strain and examined for microscopic tissue damage. The yield strain in irradiated specimens (0.93±0.11%, mean±SD) did not differ from that in control specimens (0.90±0.11%, p=0.44). No differences in elastic modulus were observed between groups after accounting for differences in bone volume fraction. However, irradiated specimens showed greater residual strain (p=0.01), increased number of microfractures (p=0.02), and reduced amounts of cross-hatching type damage (p<0.01). Although gamma radiation sterilization at commonly used dosing (30 kGy) does not modify elastic or yield properties of dense cancellous bone, it does cause modifications in damage processes, resulting in increased permanent deformation following isolated overloading.     

Irradiation Sensitivity of Planktonic and Biofilm-Associated Escherichia coli O157:H7 Isolates Is Influenced by Culture Conditions

Ionizing radiation effectively inactivates Escherichia coli O157:H7, but the efficacy of the process against biofilm cells versus that against free-living planktonic cells is not well documented. The radiation sensitivity of planktonic or biofilm cells was determined for three isolates of E. coli O157:H7 (C9490, ATCC 35150, and ATCC 43894). Biofilms were formed on sterile glass slides incubated at 37°C for either 24 h, 48 h, or 72 h. The biofilm and planktonic cultures were gamma irradiated at doses ranging from 0.0 (control) to 1.5 kGy. The dose of radiation value required to reduce the population by 90% (D₁₀) was calculated for each isolate, culture, and maturity based on viable populations at each radiation dose. For each of the times sampled, the D₁₀ values of isolate 43894 planktonic cells (0.454 to 0.479 kGy) were significantly (P < 0.05) higher than those observed for biofilm cells (0.381 to 0.385 kGy), indicating a significantly increased sensitivity to irradiation for cells in the biofilm habitat. At the 24-h sampling time, isolate C9490 showed a similar pattern, in which the D₁₀ values of planktonic cells (0.653 kGy) were significantly higher than those for biofilm cells (0.479 kGy), while isolate 35150 showed the reverse, with D₁₀ values of planktonic cells (0.396 kGy) significantly lower than those for biofilm cells (0.526 kGy). At the 48-h and 72-h sampling times, there were no differences in radiation sensitivities based on biofilm habitat for C9490 or 35150. Biofilm-associated cells, therefore, show a response to irradiation which can differ from that of planktonic counterparts, depending on the isolate and the culture maturity. Culture maturity had a more significant influence on the irradiation efficacy of planktonic cells but not on biofilm-associated cells of E. coli O157:H7.     

Inactivation of Escherichia coli O157:H7 in Apple Juice by Irradiation

Three strains (932, Ent-C9490, and SEA13B88) of Escherichia coli O157:H7 were used to determine the effectiveness of low-dose gamma irradiation for eliminating E. coli O157:H7 from apple juice or cider and to characterize the effect of inducing pH-dependent, stationary-phase acid resistance on radiation resistance. The strains were grown in tryptic soy broth with or without 1% dextrose for 18 h to produce cells that were or were not induced to pH-dependent stationary-phase acid resistance. The bacteria were then transferred to clarified apple juice and irradiated at 2°C with a cesium-137 irradiator. Non-acid-adapted cells had radiation D values (radiation doses needed to decrease a microbial population by 90%) ranging from 0.12 to 0.21 kGy. D values increased to 0.22 to 0.31 kGy for acid-adapted cells. When acid-adapted SEA13B88 cells were tested in five apple juice brands having different levels of suspended solids (absorbances ranging from 0.04 to 2.01 at 550 nm), radiation resistance increased with increasing levels of suspended solids, with D values ranging from 0.26 to 0.35 kGy. Based on these results, a dose of 1.8 kGy should be sufficient to achieve the 5D inactivation of E. coli recommended by the National Advisory Committee for Microbiological Criteria for Foods.     

Sensitivity of Planktonic and Biofilm-Associated Salmonella spp. to Ionizing Radiation

Salmonella enterica forms biofilms that are relatively resistant to chemical sanitizing treatments. Ionizing radiation has been used to inactivate Salmonella on a variety of foods and contact surfaces, but the relative efficacy of the process against biofilm-associated cells versus free-living planktonic cells is not well documented. The radiation sensitivity of planktonic or biofilm-associated cells was determined for three food-borne-illness-associated isolates of Salmonella. Biofilms were formed on sterile glass slides in a coincubation apparatus, using inoculated tryptic soy broth, incubated at 37°C for 48 h. Resulting biofilms were 18 to 24 μm in height as determined by confocal scanning laser microscopy. The planktonic and biofilm cultures were gamma irradiated to doses of 0.0 (control), 0.5, 1.0, 1.5, 2.0 and 2.5 kGy. The D₁₀ value (the dose of radiation required to reduce a population by 1 log₁₀, or 90%) was calculated for each isolate-culture based on surviving populations at each radiation dose. The D₁₀ values of S. enterica serovar Anatum were not significantly (P < 0.05) different for biofilm-associated (0.645 kGy) and planktonic (0.677 kGy) cells. In contrast, the biofilm-associated cells of S. enterica serovar Stanley were significantly more sensitive to ionizing radiation than the respective planktonic cells, with D₁₀ values of 0.531 and 0.591 kGy, respectively. D₁₀ values of S. enterica serovar Enteritidis were similarly reduced for biofilm-associated (0.436 kGy) versus planktonic (0.535 kGy) cells. The antimicrobial efficacy of ionizing radiation is therefore preserved or enhanced in treatment of biofilm-associated bacteria.     

Effect of gamma-ray irradiation on Escherichia coli motility

The effects of ionizing radiation on bacteria are generally evaluated from the dose-dependent survival ratio, which is determined by colony-forming ability and mutation rate. The mutagenic damage to cellular DNA induced by radiation has been extensively investigated; however, the effects of irradiation on the cellular machinery in situ remain unclear. In the present work, we irradiated Escherichia coli cells in liquid media with gamma rays from ⁶⁰Co (in doses up to 8 kGy). The swimming speeds of the cells were measured using a microscope. We found that the swimming speed was unaltered in cells irradiated with a lethal dose of gamma rays. However, the fraction of motile cells decreased in a dose-dependent manner. Similar results were observed when protein synthesis was inhibited by treatment with kanamycin. Evaluation of bacterial swimming speed and the motile fraction after irradiation revealed that some E. coli cells without the potential of cell growth and division remained motile for several hours after irradiation.     

Quantifying the ultrastructure changes of air-dried and irradiated human amniotic membrane using atomic force microscopy: a preliminary study

Air-dried and sterilized amnion has been widely used as a dressing to treat burn and partial thickness wounds. Sterilisation at the standard dose of 25 kGy was reported to cause changes in the morphological structure as observed under the scanning electron microscope. This study aimed to quantify the changes in the ultrastructure of the air-dried amnion after gamma-irradiated at several doses by using atomic force microscope. Human placentae were retrieved from mothers who had undergone cesarean elective surgery. Amnion separated from chorion was processed and air-dried for 16 h. It was cut into 10?×?10 mm, individually packed and exposed to gamma irradiation at 5, 15, 25 and 35 kGy. Changes in the ultrastructural images of the amnion were quantified in term of diameter of the epithelial cells, size of the intercellular gap and membrane surface roughness. The longest diameter of the amnion cells reduced significantly after radiation (p?<?0.01) however the effect was not dose dependent. No significant changes in the shortest diameter after radiation, except at 35 kGy which decreased significantly when compared to 5 kGy (p?<?0.01). The size of the irradiated air-dried amnion cells reduced in the same direction without affecting the gross ultrastructure. At 15 kGy the intercellular gap decreased significantly (p?<?0.01) with Ra and Rq, values reflecting surface roughness, were significantly the highest (p?<?0.01). Changes in the ultrastructure quantified by using atomic force microscope could complement results from other microscopic techniques.     

Effect of Gamma-irradiation and Heat on Root-knot Nematode,Meloidogyne javanica

Effects of gamma-irradiation on the root-knot nematode Meloidogyne javanica were investigated. A dose of 7.5 kGy killed all second-stage juveniles (J2) within 1 day after treatment. Egg hatch was completely inhibited at 6.25 kGy. A bioassay on tomato measuring galling and egg production was used to determine the infectivity of irradiated J2 and J2 hatched from irradiated eggs. The J2 and eggs irradiated with a dose of 4.25 kGy did not induce galls or reproduce on tomato plants. When nematodes were exposed to combined irradiation and heat treatment, no synergistic effect on J2 or eggs was measured. Heat treatment at 49° C for 10 minutes or 20 minutes without irradiation immobilized J2 and prevented egg development. Irradiation rates needed to kill or incapacitate M. javanica were high and may be impractical as a quarantine measure.     

Roots of the xerophyte Panicum turgidum host a cohort of ionizing-radiation-resistant biotechnologically-valuable bacteria

Bacterial communities associated with roots of Panicum turgidum, exposed to arid conditions, were investigated with a combination of cultural and metataxonomic approaches. Traditional culture-based techniques were used and 32 isolates from the irradiated roots were identified as belonging to Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria phyla. Four actinobacterial strains were shown to be ionizing-radiation (IR)-resistant: Microbacterium sp. PT8 (4.8 kGy (kGy)), Micrococcus sp. PT11 (4.4 kGy), Kocuria rhizophila PT10 (2.9 kGy) and Promicromonospora panici PT9^T (2.6 kGy), based on the D₁₀ dose necessary for a 90% reduction in colony forming units (CFU). Concerning the investigation of microbial communities in situ, metataxonomic analyses of the diversity of IR-resistant microorganisms associated with irradiated roots revealed a marked dominance of Actinobacteria (46.6%) and Proteobacteria (31.5%) compared to Bacteroidetes (4.6%) and Firmicutes (3.2%). Gamma irradiation not only changed the structure of bacterial communities, but also affected their functional properties. Comparative analyses of metabolic profiles indicated the induction of several pathways related to adaptation to oxidative stress in irradiated roots, such as DNA repair, secondary metabolites synthesis, reactive oxygen species (ROS)-mitigating enzymes, etc. P. turgidum is emblematic of desert-adapted plants. Until now, there is no other work that has focused on the microbial profile of irradiated roots of this xerophyte.     

Effects of ionizing radiation and preservation on biomechanical properties of human costal cartilage

Tissue banks around the world store human cartilage obtained from cadaveric donors for use in diverse reconstructive surgical procedures. To ensure this tissue is sterile at the time of distribution, tissues may be sterilized by ionizing radiation. In this work, we evaluate the physical changes in deep frozen costal cartilage (?70 °C) or costal cartilage preserved in high concentrations of glycerol (>98 %) followed by a terminal sterilization process using ionizing radiation, at 3 different doses (15, 25 and 50 kGy). Tension and compression tests were carried out to determine the mechanical changes related both to the different preservation methods and irradiation doses. For both methods of preservation, tension strength was increased by about 24 %, when cartilage tissue was irradiated with 15 kGy. Deep frozen samples, when irradiated with 25 or 50 kGy, had a decrease in their mechanical performance, albeit to a lesser extent than when tissues were preserved in high concentration of glycerol and equally irradiated. In conclusion, processing in high concentration of glycerol did not increase tissue protection against radiation damage; while cartilage preserved in high concentrations of glycerol withstands radiation up to 25 kGy, deep frozen human costal cartilage may be sterilized with a doses up to 50 kGy without significant mechanical impact.     

Combining the Sterile Insect Technique with Wolbachia-Based Approaches: II- A Safer Approach to Aedes albopictus Population Suppression Programmes,Designed to Minimize the Consequences of Inadvertent Female Release

Due to the absence of a perfect method for mosquito sex separation, the combination of the sterile insect technique and the incompatible insect technique is now being considered as a potentially effective method to control Aedes albopictus. In this present study first we examine the minimum pupal irradiation dose required to induce complete sterility in Wolbachia triple-infected (HC), double-infected (GUA) and uninfected (GT) female Ae. albopictus. The HC line is a candidate for Ae. albopictus population suppression programmes, but due to the risk of population replacement which characterizes this triple infected line, the individuals to be released need to be additionally irradiated. After determining the minimum irradiation dose required for complete female sterility, we test whether sterilization is sufficient to prevent invasion of the triple infection from the HC females into double-infected (GUA) populations. Our results indicate that irradiated Ae. albopictus HC, GUA and GT strain females have decreased fecundity and egg hatch rate when irradiated, inversely proportional to the dose, and the complete sterilization of females can be acquired by pupal irradiation with doses above 28 Gy. PCR-based analysis of F₁ and F₂ progeny indicate that the irradiated HC females, cannot spread the new Wolbachia wPip strain into a small cage GUA population, released at a 1:5 ratio. Considering the above results, we conclude that irradiation can be used to reduce the risk of population replacement caused by an unintentional release of Wolbachia triple-infected Ae. albopictus HC strain females during male release for population suppression.     

Optimization of exopolysaccharide welan gum production by Alcaligenes sp. CGMCC2428 with Tween-40 using response surface methodology

The effect of additives on welan gum production produced by fermentation with Alcaligenes sp. CGMCC2428 was studied. Tween-40 was the best additive for improving welan gum production and welan gum displayed better rheological properties than that obtained by control fermentation without additives. Response surface methodology was employed to optimize the culture conditions for welan gum production in the shake flask culture, including Tween-40 concentration, pH and culture temperature. The optimal conditions were determined as follows: Tween-40 concentration 0.94 g/l, pH 6.9 and temperature 29.6 °C. The corresponding experimental concentration of welan gum was 23.62 ± 0.60 g/l, which was agreed closely with the predicted value (23.48 g/l). Validation experiments were also carried out to prove the adequacy and the accuracy of the model obtained. The welan gum fermentation in a 7.5 l bioreactor reached 24.90 ± 0.68 g/l.     

Gamma radiation effects on seed germination, growth and pigment content, and ESR study of induced free radicals in maize (Zea mays)

The effects of gamma radiation are investigated by studying plant germination, growth and development, and biochemical characteristics of maize. Maize dry seeds are exposed to a gamma source at doses ranging from 0.1 to 1 kGy. Our results show that the germination potential, expressed through the final germination percentage and the germination index, as well as the physiological parameters of maize seedlings (root and shoot lengths) decreased by increasing the irradiation dose. Moreover, plants derived from seeds exposed at higher doses (≤0.5 kGy) did not survive more than 10 days. Biochemical differences based on photosynthetic pigment (chlorophyll a, chlorophyll b, carotenoids) content revealed an inversely proportional relationship to doses of exposure. Furthermore, the concentration of chlorophyll a was higher than chlorophyll b in both irradiated and non-irradiated seedlings. Electron spin resonance spectroscopy used to evaluate the amount of free radicals induced by gamma ray treatment demonstrates that the relative concentration of radiation-induced free radicals depends linearly on the absorbed doses.     

A novel intramolecular G-quartet-containing fold of single-stranded d(GT)8 and d(GT)16 oligonucleotides

Human genome is shown to be enriched with (GT)_n stretches of lengths from 8 to 20 dinucleotides. Low temperature (T ≤ 10 °C) conformations of d(GT)_n oligonucleotides (n = 7, 8, 12, 16, 20) were studied by means of circular dichroism (CD), thermal melting, ethidium bromide (EtBr) probing and single nucleotide substitutions. Rotational relaxation times for EtBr:d(GT)_n complexes confirmed a monomolecular state of the oligonucleotides. CD spectra indicated involvement of all guanines of d(GT)₈ and d(GT)₁₆ in G-quartets, while dT(GT)₇, d(GT)₁₂ and d(GT)₂₀ were shown to be only partially ordered. The schemes of the d(GT)₈ and d(GT)₁₆ folds are suggested.     

Using gamma irradiation and low temperature on microbial decontamination of red meat in Iran

Gamma irradiation can be used as one of the most efficient methods to reduce microorganisms in food. The irradiation of food is used for a number of purposes, including microbiological control, insects control and inhibition of sprouting and delay of senescence of living food. The aim of this study was to study effects of gamma irradiation, refrigeration and frozen storage as the combination process for improvement of red meat shelf-life. The bovine meat samples were treated with 0, 0.5, 1, 2 and 3 kGy of gamma irradiation and kept in refrigerator for 3 weeks and in freezer for 8 months. The control and irradiated samples were stored at 4–7°C and at −18°C for refrigeration and frozen storage, respectively; and microbial and chemical analyze was done at 1 week and 2 months intervals. In this study the optimum dose of gamma radiation in order to decrease the total count of Mesophilic bacteria, Coliforms, Staphylococcus aureus and especially for elimination of Salmonella was obtained at 3 kGy. Microbial analysis indicated that irradiation and storage at low temperature had a significant effect on the reduction of microbial loads. There was no significant difference in chemical characteristics during freezing storage in bovine meat. Also, irradiated meat samples (3 kGy) were stored in 4–7°C for 14 days, compared to 3 days for non irradiated samples.     

Ultrastructural changes of bovine pulmonary artery endothelial cells irradiated in vitro with a 137Cs source

Bovine pulmonary artery endothelial cells in culture were evaluated by phase-contrast and electron microscopy at various times after being irradiated with ¹³⁷Cs in vitro. Cells irradiated prior to reaching confluency showed vacuolization and increased numbers of lysosomes beginning at 48 hr after irradiation with 300–500 rad and at 24 hr after irradiation with 1500–5000 rad. After 7 days the morphological changes appeared to be reversible for cells receiving the lower doses, but were progressive for higher doses of radiation. The same qualitative changes, with a delayed onset, were observed for cells irradiated at confluency. An observed decrease in the endoplasmic reticulum and polysomes occurred only late in the course of radiation injury. There was no observable structural alteration of mitochondria even when there was evidence of otherwise marked cytoplasmic injury. We conclude that structural changes of the lysosomes constitute an early phase of injury by irradiation of the endothelial cell in culture, while decreases in endoplasmic reticulum and polysomcs occur relatively late. The mitochondrial structure of the endothelial cell appears to be relatively resistant to radiation. All morphological changes occur subsequent to impaired transport of α-aminoisobutyric acid, which is observed within 6 hr as previously reported (Kwock et al., 1982).     

Ionizing-Radiation Resistance in the Desiccation-Tolerant Cyanobacterium Chroococcidiopsis

The effect of X-ray irradiation on cell survival, induction, and repair of DNA damage was studied by using 10 Chroococcidiopsis strains isolated from desert and hypersaline environments. After exposure to 2.5 kGy, the percentages of survival for the strains ranged from 80 to 35%. In the four most resistant strains, the levels of survival were reduced by 1 or 2 orders of magnitude after irradiation with 5 kGy; viable cells were recovered after exposure to 15 kGy but not after exposure to 20 kGy. The severe DNA damage evident after exposure to 2.5 kGy was repaired within 3 h, and the severe DNA damage evident after exposure to 5 kGy was repaired within 24 h. The increase in trichloroacetic acid-precipitable radioactivity in the culture supernatant after irradiation with 2.5 kGy might have been due to cell lysis and/or an excision process involved in DNA repair. The radiation resistance of Chroococcidiopsis strains may reflect the ability of these cyanobacteria to survive prolonged desiccation through efficient repair of the DNA damage that accumulates during dehydration.     

Pleiotropic effects of a cold shock protein homolog PprM on the proteome of Deinococcus radiodurans

An extremophile D. radiodurans encodes a non-cold shock inducible cold shock protein homolog DR_0907 (also known as PprM). The DR_0907 ORF was deleted by knockout mutagenesis and the resultant deletion mutant (ΔpprM D. radiodurans) displayed growth defect as well as gamma-radiation sensitivity (D₁₀ values = ΔpprM D. radiodurans: 12.1 kGy versus wild type (WT) D. radiodurans: 14 kGy). 2D gel based comparative proteomics revealed a comparable induction of DNA repair proteins in ΔpprM D. radiodurans and WT D. radiodurans recovering from 5 kGy gamma irradiation (⁶⁰Co gamma source, dose rate: 2 kGy/h), suggesting that pprM does not cause radiation sensitivity through modulation of DdrO-regulated DNA repair genes. However, deletion of pprM did result in repression of several proteins that belonged to vital housekeeping pathways such as metabolism and protein homeostasis that might contribute to slow growth phenotype. These deficiencies intrinsic to ΔpprM D. radiodurans might also contribute to its radiation sensitivity.     

Physiological responses of the halophilic archaeon Halobacterium sp. strain NRC1 to desiccation and gamma irradiation

We report that the halophilic archaeon Halobacterium sp. strain NRC-1 is highly resistant to desiccation, high vacuum and ⁶⁰Co gamma irradiation. Halobacterium sp. was able to repair extensive double strand DNA breaks (DSBs) in its genomic DNA, produced both by desiccation and by gamma irradiation, within hours of damage induction. We propose that resistance to high vacuum and ⁶⁰Co gamma irradiation is a consequence of its adaptation to desiccating conditions. Gamma resistance in Halobacterium sp. was dependent on growth stage with cultures in earlier stages exhibiting higher resistance. Membrane pigments, specifically bacterioruberin, offered protection against cellular damages induced by high doses (5 kGy) of gamma irradiation. High-salt conditions were found to create a protective environment against gamma irradiation in vivo by comparing the amount of DSBs induced by ionizing radiation in the chromosomal DNA of Halobacterium sp. to that of the more radiation-sensitive Escherichia coli that grows in lower-salt conditions. No inducible response was observed after exposing Halobacterium sp. to a nonlethal dose (0.5 kGy) of gamma ray and subsequently exposing the cells to either a high dose (5 kGy) of gamma ray or desiccating conditions. We find that the hypersaline environment in which Halobacterium sp. flourishes is a fundamental factor for its resistance to desiccation, damaging radiation and high vacuum.     

The influence of the irradiation regime upon mycotoxins production under experimental conditions

The paper handles the problem of the inactivation of the toxinogenic strain Aspergillus flavus following the application of gamma radiation to wheat. The amount of the applied dose and of the absorbed dose of ionizing radiation upon the inhibition of mycelium growth and toxin production were defined. The aflatoxin B1 was determined by extracting in chloroform and developed on Silufol R within the choroform; aceton system. The applied doses of gamma radiation (3-30 kGy) have show that the absorbed dose does not inhibit aflatoxin production. By combining the action of gamma radiation with humidity of the wheat (humidity 13-15%; 25% irradiation 6 kGy) an inactivation was reached. With the help of toxicologico-genetical tests (the Dominant Lethal Mutations Test, the Three Generations Test) the influence was traced of contaminated, irradiated substrates upon the health of experimental animals. It follows from the results obtained that in long-term feeding with contaminated wheat irradiated by gamma rays no positive mutagenic activity has been recorded. It allows to presume that wheat of humidity of 25% contaminated by a weakly toxigenic strain Aspergillus flavus irradiated by a dose of 6 kGy, and wheat of a humidity of 13-15%, contaminated by a strongly toxinogenic strain of Aspergillus flavus, irradiated by a dose of 6 kGy, are no genetic risk for white rats.