Effect of gamma irradiation on microbial load,aflatoxins and phytochemicals present in Trigonella foenum-graecum

The effects of gamma irradiation on microbial load, total aflatoxins and phytoconstituents content of Trigonella foenum-graecum have been studied. Gamma irradiation at a dose of 2.5 kGy resulted in 2 log reduction of the total aerobic microbial count. A complete sterilization was, however, observed at 10 kGy. The total aflatoxin level decreased gradually with increase in gamma irradiation dose as compared to its un-irradiated counterparts, whereas the high performance liquid chromatography (HPLC) profile showed no change in the levels of phytochemicals up to the gamma irradiation dose of 10 kGy. HPLC profiles, however, differed in peak areas, and retention times of the components. These results suggest that gamma irradiation at a dose of 5.0 kGy was very effective for microbial decontamination because it did not adversely affect the active components of T. foenum-graecum.     

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 degrees C for 48 h. Resulting biofilms were 18 to 24 microm 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 D(10) value (the dose of radiation required to reduce a population by 1 log(10), or 90%) was calculated for each isolate-culture based on surviving populations at each radiation dose. The D(10) 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 D(10) values of 0.531 and 0.591 kGy, respectively. D(10) 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.     

Radiation inactivation of some food-borne pathogens in fish as influenced by fat levels

The influence of low (0·39–1·1%), medium (4·25%) and high (7·1–32·5%) fat levels in fish on radiation inactivation of four food-borne pathogens was investigated. Cells of Listeria monocytogenes 036, Yersinia enterocolitica F5692, Bacillus cereus and Salmonella typhimurium at logarithmic phase were inoculated in 10% fish homogenates and subjected to gamma irradiation at ice temperature (0–1 °C) with doses ranging from 0·05 to 0·8 kGy. The radiation survival curves of L. monocytogenes and B. cereus were characterized by shoulders, while a tailing effect was depicted by cells of Y. enterocolitica and B. cereus . The D₁₀ values in kGy calculated on the exponential part of the curve ranged from 0·2 to 0·3, 0·15 to 0·25, 0·1 to 0·15 and 0·09 to 0·1 for L. monocytogenes 036, B. cereus, Salm. typhimurium and Y. enterocolitica F5692, respectively. This order (D₁₀) of radiation resistance of each organism was not affected by the fat content of the fish. Inoculated pack studies carried out separately with each pathogen in fatty (Indian sardine, 7·1%) and lean (Golden anchovy, 0·39%) fish showed no difference in their survival after exposure to 1 kGy and 3 kGy doses, which corroborated the above observation. The practical significance of these results in the application of the technology is discussed.     

Effect of gamma irradiation on shelf life and bacterial and viral loads in hard-shelled clams (Mercenaria mercenaria).

The feasibility of using 60Co gamma irradiation to inactivate total coliforms, fecal coliforms, Escherichia coli, Clostridium perfringens, and F-coliphage in hard-shelled clams, Mercenaria mercenaria, was investigated. The results of three trials indicated average D10 values of 1.32 kGy for total coliforms, 1.39 kGy for fecal coliforms, 1.54 kGy for E. coli, 2.71 kGy for C. perfringens, and 13.50 kGy for F-coliphage. Irradiation doses of > 0.5 kGy were significantly lethal to the shellfish.     

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 degrees 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.     

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.     

Effect of monopropylene glycol and gamma irradiation on Yarrowia lipolytica lipase stabilization

This work investigated the effects of monopropylene glycol, protease inhibitor, and gamma irradiation on Yarrowia lipolytica lipase stability during storage. Enzyme liquid stabilization was achieved by addition of monopropylene glycol (MPG) at respective concentrations of 50, 75, and 90%, the protease inhibitors (P2714 and P8215) at 0.1%, and the gamma irradiation with 10kGy, 15kGy, and 25kGy doses. The results showed that monopropylene glycol limited the microorganism growth and decreased the enzymatic activity at high concentration (up to 50%), at two temperatures (20 and 4 degrees C). Enzyme stored at 20 degrees C lost its activity by 80% after two months. This loss was attributed to the protease's effect. At this temperature, the protease's activities have been limited by the specific inhibitors. The gamma irradiations improve microbial safety of liquid enzyme.     

The effect of He-Ne laser radiation on the survival of HeLa cells subjected to ionizing radiation]

A monolayer of HeLa cells, at the stationary phase of growth, exposed to He-Ne laser radiation (632.8 nm; 100 J/m2) either 5 min or 60 min prior to gamma irradiation (0.1-10 Gy; 6.75 Gy/min), or 5 min after irradiation has been investigated. With a 5-min interval between irradiation sessions (both sequences) the survival curves are virtually the same as those for gamma-irradiated cells only. With He-Ne laser radiation delivered 60 min before gamma irradiation with doses exceeding 5 Gy, a fraction of radioresistant cells is identified whose D0 is almost twice as high as D0 of basic cell mass (3.6 and 1.7 Gy respectively. The survival curve becomes a two-component one. A hypothesis is proposed that He-Ne laser radiation activates, in some cells, the processes that promote the repair of radiation damages.     

Microbial decontamination of hospital linens using radiation

The irradiation of hospital linen contaminated with radioresistant microorganisms or hospital microflora with gamma radiation in a dose of 10 kGy ensures the reliable microbial decontamination of such linen. Cotton linen has been found capable of withstanding 15 irradiation cycles in a dose of 10 kGy.     

Role of the competitive microbial flora in the radiation-induced enhancement of ochratoxin production by Aspergillus alutaceus var. alutaceus NRRL 3174.

The radiation sensitivity and the toxigenic potential of conidiospores of the fungus Aspergillus alutaceus var. alutaceus were determined after irradiation with 60Co gamma rays and high-energy electrons. Over the pH range of 3.6 to 8.8, the doses required for a 1 log10 reduction in viability based on the exponential portion of the survival curve ranged from 0.21 to 0.22 kGy, with extrapolation numbers (extrapolation of the exponential portion of the survival curve to zero dose) of 1.01 to 1.33, for electron irradiation, and from 0.24 to 0.27 kGy, with extrapolation numbers of 2.26 to 5.13, for gamma irradiation. Nonsterile barley that was inoculated with conidia of the fungus and then irradiated with either electrons or gamma rays and incubated for prolonged periods at 28 degrees C and at a moisture content of 25% produced less ochratoxin A with increasing doses of radiation. Inoculation of barley following irradiation resulted in enhanced ochratoxin levels compared with unirradiated controls. In these experiments, inoculation with 10(2) spores per g produced greater radiation-induced enhancement than inoculation with 10(5) spores per g. There was no radiation-induced enhancement when the barley was surface sterilized by chemical means prior to irradiation. These results are consistent with the hypothesis that a reduction in the competing microbial flora by irradiation is responsible for the enhanced mycotoxin production observed when nonsterile barley is inoculated with the toxigenic fungus A. alutaceus var. alutaceus after irradiation.     

Role of the competitive microbial flora in the radiation-induced enhancement of ochratoxin production by Aspergillus alutaceus var. alutaceus NRRL 3174.

The radiation sensitivity and the toxigenic potential of conidiospores of the fungus Aspergillus alutaceus var. alutaceus were determined after irradiation with 60Co gamma rays and high-energy electrons. Over the pH range of 3.6 to 8.8, the doses required for a 1 log10 reduction in viability based on the exponential portion of the survival curve ranged from 0.21 to 0.22 kGy, with extrapolation numbers (extrapolation of the exponential portion of the survival curve to zero dose) of 1.01 to 1.33, for electron irradiation, and from 0.24 to 0.27 kGy, with extrapolation numbers of 2.26 to 5.13, for gamma irradiation. Nonsterile barley that was inoculated with conidia of the fungus and then irradiated with either electrons or gamma rays and incubated for prolonged periods at 28 degrees C and at a moisture content of 25% produced less ochratoxin A with increasing doses of radiation. Inoculation of barley following irradiation resulted in enhanced ochratoxin levels compared with unirradiated controls. In these experiments, inoculation with 10(2) spores per g produced greater radiation-induced enhancement than inoculation with 10(5) spores per g. There was no radiation-induced enhancement when the barley was surface sterilized by chemical means prior to irradiation. These results are consistent with the hypothesis that a reduction in the competing microbial flora by irradiation is responsible for the enhanced mycotoxin production observed when nonsterile barley is inoculated with the toxigenic fungus A. alutaceus var. alutaceus after irradiation.     

Effect of gamma radiation on native endolithic microorganisms from a radioactive waste deposit site.

A time-course experiment was conducted to evaluate the effects of gamma radiation on the indigenous microbiota present in rock obtained from Yucca Mountain, Nevada Test Site. Microcosms were constructed by placing pulverized Yucca Mountain rock in polystyrene cylinders. Continuous exposure (96 h) at a dose rate of 1.63 Gy/min was used to mimic the near-field environment surrounding waste canisters. The expected maximum surface dose rate from one unbreached canister designed to contain spent nuclear fuels is 0.06 Gy/min. Considering the current repository packing design, multiple canisters within one vault, the cumulative dose rate may well approach that used in this experiment. The microbial communities were characterized after receiving cumulative doses of 0, 0.098, 0. 58, 2.33, 4.67, 7.01 and 9.34 kGy. Radiation-resistant microorganisms in the pulverized rock became viable but nonculturable (VBNC) after a cumulative dose of 2.33 kGy. VBNC microorganisms lose the ability to grow on media on which they have routinely been cultured in response to the environmental stress imposed (i.e. radiation) but can be detected throughout the time course using direct fluorescence microscopy techniques. Two representative exopolysaccharide-producing isolates from Yucca Mountain were exposed to the same radiation regimen in sand microcosms. One isolate was much more radiation-resistant than the other, but both had greater resistance than the general microbial community based on culturable counts. However, when respiring cell counts (VBNC) were compared after irradiation, the results would indicate much more radiation resistance of the individual isolates and the microbial community in general. These results have significant implications for underground storage of nuclear waste as they indicate that indigenous microorganisms are capable of surviving gamma irradiation in a VBNC state.     

Stimulation of macrophages and antitumor activity of radiodetoxified endotoxin

Lipopolysaccharide of Salmonella typhimurium was irradiated with gamma radiation at 10, 15, and 30 kGy doses. A dose of 30 kGy significantly detoxified the LPS (180 times). Mice were injected intraperitoneally with the radiodetoxified LPS, and it was found that it stimulated peritoneal macrophages as was evident from the enhancement of their acid hydrolases and cellular RNA content. Both LPS and radiodetoxified LPS exhibited antitumor activity against S180 cells in Swiss mice. Treatment with 20 micrograms/mouse of either LPS or 30 kGy LPS gave maximum survival of the mice (90%). These mice were found to resist the challenge of S180 cells (1 X 10(6)).     

Exposure to low dose of gamma radiation enhances the excision repair in Saccharomyces cerevisiae

The effect of low doses of ionizing and nonionizing radiation on the radiation response of yeast Saccharomyces cerevisiae toward ionizing and nonionizing radiation was studied. The wild-type strain D273-10B on exposure to 54 Gy gamma radiation (resulting in about 10% cell killing) showed enhanced resistance to subsequent exposure to UV radiation. This induced UV resistance increased with the incubation time between the initial gamma radiation stress and the UV irradiation. Exposure to low doses of UV light on the other hand showed no change in gamma or UV radiation response of this strain. The strains carrying a mutation at rad52 behaved in a way similar to the wild type, but with slightly reduced induced response. In contrast to this, the rad3 mutants, defective in excision repair, showed no induced UV resistance. Removal of UV-induced pyrimidine dimers in wild-type yeast DNA after UV irradiation was examined by analyzing the sites recognized by UV endonuclease from Micrococcus luteus. The samples that were exposed to low doses of gamma radiation before UV irradiation were able to repair the pyrimidine dimers more efficiently than the samples in which low gamma irradiation was omitted. The nature of enhanced repair was studied by scoring the frequency of induced gene conversion and reverse mutation at trp and ilv loci respectively in strain D7, which showed similar enhanced UV resistance induced by low-dose gamma irradiation. The induced repair was found to be essentially error-free. These results suggest that irradiation of strain D273-10B with low doses of gamma radiation enhances its capability for excision repair of UV-induced pyrimidine dimers.     

Inactivation of Escherichia coli O157:H7, salmonellae, and Campylobacter jejuni in raw ground beef by gamma irradiation.

Raw ground beef patties inoculated with stationary-phase cells of Escherichia coli O157:H7, salmonellae, or Campylobacter jejuni were subjected to gamma irradiation (60Co) treatment, with doses ranging from 0 to 2.52 kGy. The influence of two levels of fat (8 to 14% [low fat] and 27 to 28% [high fat]) and temperature (frozen [-17 to -15 degrees C] and refrigerated [3 to 5 degrees C]) on the inactivation of each pathogen by irradiation was investigated. In ascending order of irradiation resistance, the D10 values ranged from 0.175 to 0.235 kGy (C. jejuni), from 0.241 to 0.307 kGy (E. coli O157:H7), and from 0.618 to 0.800 kGy (salmonellae). Statistical analysis revealed that E. coli O157:H7 had a significantly (P < 0.05) higher D10 value when irradiated at -17 to -15 degrees C than when irradiated at 3 to 5 degrees C. Regardless of the temperature during irradiation, the level of fat did not have a significant effect on the D10 value. Salmonellae behaved like E. coli O157:H7 in low-fat beef, but temperature did not have a significant effect when the pathogen was irradiated in high-fat ground beef. Significantly higher D10 values were calculated for C. jejuni irradiated in frozen than in refrigerated low-fat beef. C. jejuni was more resistant to irradiation in low-fat beef than in high-fat beef when treatment was at -17 to -15 degrees C. Regardless of the fat level and temperature during inactivation, these pathogens were highly sensitive to gamma irradiation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dominant lethal effect of 60Co gamma radiation in Biomphalaria glabrata (SAY, 1818)

The dominant lethal effects of gamma radiation of 60Co in the snail Biomphalaria glabrata were studied. Three groups of 13 wild-type snails were irradiated with single doses of 2.5; 10 and 20 Gy. Crossings were carried out at intervals of 7, 17, 23, 30 and 36 days after irradiation. The dominant lethal effect was observed only at the first crossing occurring 7 days after irradiation with 2.5 Gy. With 10 and 20 Gy, the induction of lethal mutations was detected at 7, 17 and 23 days after irradiation; a dose-response effect was observed. The effect was stronger 7 days after irradiation, decreasing in the succeeding crossings up to 30 days. Cell-killing effects on germ cells were detected in the crossings at 23 days and 30 days after irradiation with 20 Gy. After 36 days, frequencies of malformations resumed background levels; crossing rates partially recovered. These results show that gamma radiation affected all the stages of spermatogenesis. Germ cells at later phases were more sensitive to the mutagenic effect of radiation and the cell killing effects were observed on the youngest cells. This response was similar to the highly homogeneous pattern observed in widely different species and allowed us to estimate some parameters of spermatogenesis in B. glabrata.     

Gamma radiation resistance of Bacillus anthracis spores

The aim of the presented study was determined the effectiveness of action the gamma radiation on water suspension B. anthracis spores. The irradiation was performed using a Cobalt 60 (Co 60) source, by using single and fractionary irradiation doses. In the investigations was used B. anthracis stain "Sterne" 34F2. The obtained results show, that gamma radiation effectively inactivates B. anthracis spores. On the efficiency of sterilization process influence the irradiation's method and the number of spores in 1 ml suspension. In the suspension 1.5 x 10(9) spore in 1 ml, sporicidal doses gamma radiation amount to 25.0 kGy (single dose) or 41.5 kGy (fractionary dose). The volume suspension about definite inoculum of spores, subjected working the gamma rays has not influence on sporicidal effectiveness of radiation sterilization.     

DNA damage and biological expression of adenovirus: a comparison of liquid versus frozen conditions of exposure to gamma rays

Human adenovirus type 2 (Ad 2) was irradiated with 137Cs gamma rays in the liquid state at 0 degree C. DNA breaks were correlated with the inactivation of several viral functions and compared to results obtained previously for irradiation of Ad 2 under frozen conditions at -75 degrees C. Irradiation at 0 degree C induced 170 +/- 20 single-strand breaks and 2.6 +/- 0.4 double-strand breaks/Gy/10(12) Da in the viral DNA. Viral adsorption to human KB cells was inactivated with a D0 of 9.72 +/- 1.18 kGy, whereas the inactivation of Ad 2 plaque formation had a D0 of 0.99 +/- 0.14 or 1.1 +/- 0.29 kGy when corrected for the effect of radiation on virus adsorption. For the adsorbed virus, an average of 4.3 +/- 1.7 single-strand and 0.065 +/- 0.02 double-strand breaks were induced in the viral DNA per lethal hit. In contrast, irradiation of Ad 2 at -75 degrees C results in 2.6- to 3.4-fold less DNA breakage per Gy and a 5.6-fold increase in D0 for plaque formation of the adsorbed virus. Furthermore, although host cell reactivation (HCR) of Ad 2 viral structural antigen production for irradiated virus was substantially reduced in the xeroderma pigmentosum fibroblast strain (XP25RO) compared to normal strains for irradiation at -75 degrees C (57% HCR), it was only slightly reduced compared to normal for irradiation at 0 degree C (88% HCR). These results indicate that the spectrum of DNA damage is both quantitatively and qualitatively different for the two conditions of irradiation.     

Evaluation of delayed apoptotic response in lethally irradiated human melanoma cell lines

To assess the lethal doses of gamma radiation and corresponding apoptotic response in new established human melanoma cell lines we exposed exponentially growing cultures to 8-100 Gy gamma radiation. The apoptosis and cell survival were determined by trypan blue exclusion, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) reaction, agarose gel electrophoresis, colony forming assay, and long-term survival assay. The maximal DNA fragmentation 3 days after irradiation was observed in cultures irradiated with 20 Gy (36.9% TUNEL positive cells). The cultures irradiated with 50 and 100 Gy contained 18.7% and 16.4% TUNEL positive cells, respectively. Cultures exposed to 8 and 20 Gy gamma radiation recovered by week 3-4. Lethally irradiated (50 and 100 Gy) cultures which contained less apoptotic cells by day 3 died by week 5. A detectable increase in melanoma cell pigmentation after irradiation was also observed. The survival of human melanoma cell cultures after exposure to gamma radiation does not correlate with the level of apoptotic cells by day 3. At high radiation doses (> 50 Gy) when the radiation induced cell pigmentation is not inhibited the processes of apoptotic DNA fragmentation might be preferentially inactivated.     

Shifts in microbial community composition in tannery-contaminated soil in response to increased gamma radiation

Contaminated sites from man-made activities such as old-fashioned tanneries are inhabited by virulent microorganisms that exhibit more resistance against extreme and toxic environmental conditions. We investigated the effect of different Gamma radiation doses on microbial community composition in the sediment of an old-fashioned tannery. Seven samples collected from the contaminated sites received different gamma radiation doses (I = 0.0, II = 5, III = 10, VI = 15, V = 20, VI = 25, and VII = 30 kGy) as an acute exposure. The shift in microbial community structure was assessed using the high throughput 454 pyrosequencing. Variations in diversity, richness, and the shift in operational taxonomic units (OTUs) were investigated using statistical analysis. Our results showed that the control sample (I) had the highest diversity, richness, and OTUs when compared with the irradiated samples. Species of Halocella, Parasporobacterium, and Anaerosporobacter had the highest relative abundance at the highest radiation dose of 30 kGy. Members of the Firmicutes also increased by 20% at the highest radiation dose when compared with the control sample (0.0 kGy). Representatives of Synergistetes decreased by 25% while Bacteroidetes retained a steady distribution across the range of gamma radiation intensities. This study provides information about potential “radioresistant” and/or “radiotolerant” microbial species that are adapted to elevated level of chemical toxicity such as Cr and Sr in tannery. These species can be of a high biotechnological and environmental importance.