Microbial Flora of Irradiated Dungeness Crabmeat and Pacific Oysters

The microorganisms in Dungeness crabmeat (Cancer magister) and Pacific oysters (Crassostrea gigas) were identified by the replica-plating and computer analysis method. The initial flora of the shellfish and the flora change during storage at 7 C were determined. The microbial flora shifts in both shellfish were also determined after irradiation at 0.1 and 0.4 Mrad and during subsequent storage at 7 C. The Achromobacter species predominated in the initial flora of crabmeat (77.0%). The predominant position of this group increased to 99.2% after 0.1 Mrad and 100% after 0.4 Mrad. A large percentage of Lactobacillus was detected in oysters (55.0%). The Lactobacillus species were the predominant survivors after 0.1 Mrad (92.4%) but the predominant survivors after 0.4 Mrad were Achromobacter species (99.3%).     

Effect of γ Irradiation on the Microflora of Freshwater Fish: III. Spoilage Patterns and Extension of Refrigerated Storage Life of Yellow Perch Fillets Irradiated to 0.1 and 0.2 Megarad1

Maximal shelf life was determined and microbial flora were compared for irradiated (0.1 and 0.2 Mrad) and nonirradiated yellow perch fillets stored at 1 C. Shelf life was estimated by organoleptic determinations. Microbiological studies included determination of the effects of irradiation on the total aerobic microbial population, lag phase, and rate of growth. Genera of organisms isolated from fillets through the course of microbial spoilage were identified, and the proteolytic activity of the organisms was determined. Plate counts for fish prior to irradiation showed the presence of approximately 10(6) organisms per g of sample. Irradiation to 0.1 and 0.2 Mrad produced 1.4 and 3 logarithm reductions of the initial count, respectively. Irradiation to 0.1 and 0.2 Mrad approximately doubled the product's shelf life. Organisms initially isolated from the nonirradiated fillets, in order of decreasing number, consisted of Flavobacterium, Micrococcus-Sarcina, Achromobacter-Alcaligenes-Mima, Pseudomonas, Microbacterium, Vibrio, Bacillus, Corynebacterium, Lactobacillus, Brevibacterium, and Aeromonas. By the 6th and 9th days of fillet storage, Pseudomonas and the Achromobacter group were the predominant organisms. All members of the genus Flavobacterium, but not all members of the genus Pseudomonas, were proteolytically active on raw fish juice-agar and skim milk-agar media. The Achromobacter group was found to be nonproteolytic on both media. Residual flora of fillets irradiated to 0.1 and 0.2 Mrad consisted of the Achromobacter group, Lactobacillus, Micrococcus-Sarcina, and Bacillus. Their sequence in predominance, however, varied with dose level. Not all proteolytic bacteria in the fillets were eliminated by 0.1 and 0.2 Mrad; proteolytic Micrococcus-Sarcina survived these treatments.     

Preparation of immobilized enzymes from acrylic monomers under γ-ray irradiation

Immobilized glucoamylase, invertase, and β-galactosidase were prepared by using 2-hydroxyethyl acrylate and dimethylacrylamide under γ-ray irradiation. In the case of 2-hydroxyethyl acrylate, the monomer-enzyme solution was changed to the gel by irradiation of less than 1.0 Mrad, but it was difficult to eliminate enzyme leakage from the gel. When leakage was eliminated by increased irradiation, the activities of the gels were very low. In the case of dimethylacrylamide, the monomer–enzyme solution was changed to a gel by irradiation of 1.0 Mrad; leakage could be eliminated by irradiation of 2.0 Mrad. This gel possessed very high activity. In the case of acrylic acid-sodium acrylate, the monomer–enzyme solution could not be changed to a gel. In preparing gels, high concentrations of enzyme protein had a tendency to obstruct gelation.     

Citrus compost and its water extract for cultivation of melon plants in greenhouse nurseries. Evaluation of nutriactive and biocontrol effects

Two different types of citrus composts, and their water extracts, were tested with regard to their utilisations as partial substitutes for peat in growing media for melon seedlings in greenhouse nurseries. Both compost showed higher plant growth than peat. Compost composed by citrus waste and green residue (C2) showed greater plant growth than compost obtained from the same organic matrices mentioned above further the addition of sludge obtained from citrus industry (C1). Compost C2 showed a greater auxinic effect than C1 and it was the only one that showed cytokinic effect. Both composts also demonstrated a biocontrol effect against Fusarium oxysporum for melon plants: the effects were also higher in C2 than in C1. Higher number of isolated fungi was active against F. oxysporum in compost C2, than compost C1. No different bacterial biocontrol efficacy was observed between both composts. The water extracts of both composts gave lower plant yields than their solid matrices, their relative effects being similar to those of the solid composts (C2 extract gave higher plant yields than the extract from C1). The biocontrol effects of compost water extracts followed the same trend.     

Suppression of Spoilage Rates of Bombay Duck (Harpodon nehereus) Homogenates by Gamma Radiation

The spoilage rates of unirradiated and irradiated (0.1 Mrad) Bombay duck (Harpodon nehereus) homogenates stored at 0°C, 10°C and 30°C in terms of organoleptic score (OS) total bacterial count (TBC), trimethylamine content (TMA), and total volatile basic nitrogen (TVBN) followed a linear response with increase in temperature. Unirradiated Bombay duck stored at 0°C were found to spoil faster than those irradiated at 0.1 Mrad; the spoilage rates derived for TBC, TMA, TVBN and OS were 0.42, 0.67, 1.93 and ?0.86 respectively for unirradiated samples as compared with corresponding values of 0.31, 0.55, 1.19 and ?0.19 for the irradiated samples. The spoilage rates of unirradiated Bombay duck stored at 10°C and 30°C measured as a function of OS, TMA and TVBN were consistently higher than their irradiated (0.1 Mrad) counterparts. In the case of irradiated samples, TBC was not a satisfactory index of spoilage.     

Effect of incremental doses of radiation on viability of the microbial population on synthetic operating room gowns.

A total of 700 25-cm2 samples of surgical gown material were exposed to doses of cobalt-60 radiation of 0.0 to 0.6 Mrad in 0.1-Mrad increments. Pour plates were made, and the microbial colonies that arose were enumerated, isolated, and identified as to species. The death rate of the microbial population was calculated, and the mean D10 value of 0.269 Mrad was obtained. Analysis showed that the initial population on unirradiated material had been underestimated; when the counts obtained by homogenization of unirradiated material were substituted, a corrected mean D10 value of 0.249 Mrad was obtained. The isolates obtained were identified, and 70.7% were found to be Bacillus spp. with 12 different species identified, 16.2% were Micrococcus spp. with 6 different species identified, and 8.2% were fungi with 10 different species identified. Calculations were made for appropriate doses of radiation to sterilize gowns with this contaminating microbial population. These calculations gave an estimated dose of radiation of 1.98 to 1.81 Mrad to reduce the observed population to 0.001, a standard where 1 gown in 1,000 might contain a living organism. Comparison of the radiation resistance of this population with that of others reported in the literature showed good agreement.     

Effect of Temperature of Liquid Nitrogen on Radiation Resistance of Spores of Clostridium botulinum

An apparatus consisting of a Dewar flask and a relay system controlling the flow of liquid nitrogen permitted the irradiation of samples in tin cans or Pyrex tubes at temperatures ranging from 0 ± 1.5 C to -194 ± 2 C. An inoculated pack comprising 320 cans of ground beef containing 5 × 10⁴ spores of Clostridium botulinum 33A per can (10 cans per radiation dose) was irradiated with Co⁶⁰ at 0 and -196 C. Incubation was carried out at 30 C for 6 months. Approximately 0.9 Mrad more radiation was required to inactivate the spores at -196 C than at 0 C. Cans irradiated at -196 C showed partial spoilage at 3.6 Mrad and no spoilage at 3.9 Mrad; the corresponding spoilage-no spoilage doses at 0 C were 2.7 and 3.0, respectively. The majority of positive cans swelled in 2 to 14 days; occasional swelling occurred as late as 20 days. At progressively higher doses, swelling was delayed proportionally to the radiation dose received. The remaining nonswollen cans had no toxin after 6 months of storage, although occasional cans contained very low numbers of viable spores comprising on the average 0.1% of the original spore inoculum. The D₁₀ values in phosphate buffer were 0.290 Mrad for 0 C and 0.396 Mrad for -196 C; in ground beef, the corresponding D₁₀ values were 0.463 Mrad and 0.680 Mrad, respectively. These D₁₀ values indicate that the lethal effect of γ rays decreased at -196 C as compared with 0 C by 13.5% in phosphate buffer, and by 47% in ground beef.     

Effect of Food Additives and Irradiation on Survival of Salmonella in Oysters

Salmonella typhimurium and S. enteritidis were inoculated into blended oysters, both raw and autoclaved. The oysters were also treated with sodium benzoate (0.1%) or potassium sorbate (0.1%), and irradiated (0.1 Mrad). In both non-irradiated and irradiated samples, greater numbers of Salmonella were recovered after storage at 7 C in the presence of sodium benzoate or potassium sorbate. The results of the autoclaved samples and studies in buffer indicated that this effect was not due to the reduction of competition from the natural flora when the additives were present.     

Detoxification of Salmonella typhimurium lipopolysaccharide by ionizing radiation

The efficiency of ionizing radiation in detoxifying the lethal determinant(s) of the lipopolysaccharide (LPS) of Salmonella typhimurium, S. enteritidis, and Escherichia coli in aqueous solution and associated with heat-killed S. typhimurium cells in suspension decreased with doses above 1 Mrad. The 50% end point of inactivation was more than 7.0 Mrad for heat-killed salmonellae and 4.8, 4.5, and 1.0 Mrad for the LPS of S. typhimurium, S. enteritidis, and E. coli, respectively. After exposure to 20 Mrad, S. typhimurium LPS retained a small portion of its lethal properties although the ld(50) was much greater than 9.5 mg per 20-g mouse. However, at -184 C, no inactivation of the lethal determinant(s) occurred after exposure to as much as 20 Mrad. This demonstrated the significance of the indirect effect and the mobility and formation of free radicals. At 22 C, the optical density at 400 mmu increased and the pH decreased with increasing radiation dose, but no qualitative changes were observed in the infrared spectrum. No change was observed in the pyrogenicity of S. typhimurium LPS; a slight decrease in antigenicity was revealed when 6 days, but not when 1 day, elapsed between vaccination and challenge in the mouse protection test. The results were interpreted as evidence of the existence of two or more lethal and antigenic determinants. The differential effect of radiation on these properties and on the pyrogenic component(s) probably are indicative of separate functional sites for lethal, antigenic, and pyrogenic activities.     

Deletion of mouse rad9 causes abnormal cellular responses to DNA damage, genomic instability, and embryonic lethality

The fission yeast Schizosaccharomyces pombe rad9 gene promotes cell survival through activation of cell cycle checkpoints induced by DNA damage. Mouse embryonic stem cells with a targeted deletion of Mrad9, the mouse ortholog of this gene, were created to evaluate its function in mammals. Mrad9(-/-) cells demonstrated a marked increase in spontaneous chromosome aberrations and HPRT mutations, indicating a role in the maintenance of genomic integrity. These cells were also extremely sensitive to UV light, gamma rays, and hydroxyurea, and heterozygotes were somewhat sensitive to the last two agents relative to Mrad9(+/+) controls. Mrad9(-/-) cells could initiate but not maintain gamma-ray-induced G(2) delay and retained the ability to delay DNA synthesis rapidly after UV irradiation, suggesting that checkpoint abnormalities contribute little to the radiosensitivity observed. Ectopic expression of Mrad9 or human HRAD9 complemented Mrad9(-/-) cell defects, indicating that the gene has radioresponse and genomic maintenance functions that are evolutionarily conserved. Mrad9(+/-) mice were generated, but heterozygous intercrosses failed to yield Mrad9(-/-) pups, since embryos died at midgestation. Furthermore, Mrad9(-/-) mouse embryo fibroblasts were not viable. These investigations establish Mrad9 as a key mammalian genetic element of pathways that regulate the cellular response to DNA damage, maintenance of genomic integrity, and proper embryonic development.     

Application of Replica Plating and Computer Analysis for Rapid Identification of Bacteria in Some Foods: II. Analysis of Microbial Flora in Irradiated Dover Sole (Microstomus pacificus)

This investigation was carried out to determine the nature of the microbial flora shifts in dover sole as a result of irradiation and storage at 6 C. The relationship was determined between the microorganisms which initially survive irradiation and those making up the final spoilage flora. A total of 2,723 isolates were examined by use of the replica-plating and computer analysis method. The spoilage of the unirradiated control samples during storage at 6 C was almost entirely due to the growth of Pseudomonas. This group, which occupied 25% of the fresh flora, grew to nearly 100% in 2 days of storage. In contrast, irradiation doses of 0.1, 0.2, 0.3, and 0.4 Mrad favored the growth of Achromobacter and yeasts. The Micrococcus, which survived radiation, did not grow at 6 C. At 0.5 Mrad, spoilage of fish samples was due entirely to the growth of yeasts.     

Microbial Properties of Composts That Suppress Damping-Off and Root Rot of Creeping Bentgrass Caused by Pythium graminicola

Composts prepared from a variety of feedstocks were tested for their ability to suppress seedling and root diseases of creeping bentgrass caused by Pythium graminicola. Among the most suppressive materials in laboratory experiments were different batches of a brewery sludge compost and a biosolids compost from Endicott, N.Y. Batches of these composts that were initially not suppressive to Pythium damping-off became more suppressive with increasing compost age. Leaf, yard waste, food, and spent mushroom composts as well as certain biosolids, cow manure, chicken-cow manure, and leaf-chicken manure composts were not suppressive to Pythium damping-off. In some cases, turkey litter, chicken manure, chicken-leaf, and food waste composts were inhibitory to creeping bentgrass seed germination in laboratory experiments. Microbial populations varied among all of the composts tested. Bacterial populations were high in all composts except the turkey litter compost, in which populations were 1,000- to 10,000-fold lower than in the other composts tested. Among the highest populations of heterotrophic fungi and antibiotic-producing actinomycetes were those found in all batches of the brewery sludge compost, whereas the lowest populations were found in turkey litter, chicken manure, and food waste composts. Heat treatment of suppressive composts reduced populations of bacteria, fungi, and actinomycetes in all composts tested. Disease suppressiveness was also reduced or eliminated in heated composts. Amending heated composts with small amounts of nonheated compost restored suppressive properties and partially restored microbial populations to wild-type levels. A strong negative relationship between compost microbial activity (as measured by the hydrolysis of fluorescein diacetate) and Pythium damping-off severity was observed. When composts were applied to creeping bentgrass in field experiments, a significant level of suppressiveness was evident with some composts when disease pressure was high (i.e., disease ratings high in uninoculated plots). A 1991 batch of turkey litter compost and the 1990 batch of Endicott biosolids were consistently suppressive to foliar symptoms of Pythium root rot on creeping bentgrass. This study indicates that suppression of Pythium diseases of creeping bentgrass in batches of brewery sludge and Endicott biosolids composts, and possibly in other suppressive composts examined in less detail in this study, is related directly to the microbial activities in the composts. On the other hand, the mechanisms of Pythium suppression in turkey litter and perhaps other poultry-based composts is not related directly to the compost microbial activity. Although turkey litter showed a lack of suppressiveness in laboratory bioassays and low microbial populations and activity, it resulted in a significant and consistent level of suppressiveness in field experiments. Therefore, the microbiological properties of Pythium-suppressive composts may differ substantially, and measurements of microbial populations and activity may not be predictive of the level of disease suppression in all composts.     

Sensitivity of aflatoxin b1 to ionizing radiation.

Aflatoxin B1 in a 5-micrograms/ml water solution was sensitive to ionizing radiation. Inactivation was assayed by the Ames microsome mutagenicity test and confirmed by thin-layer chromatography. Destruction of aflatoxin B1 had already begun at 2.5 kilograms (kGy; 1 kGy = 0.1 Mrad), but a dose exceeding 10 kGy was necessary for total destruction.     

Stimulation of environmentally controlled mushroom composting by polysaccharidases

Polysaccharidases adsorbed on commercial amylodextrins were added to environmentally controlled composts of straw plus poultry manure. After 5 days of composting at 48°C, microbial enzyme activities and numbers of bacteria were higher in the treated compost than in the control. During the next phase at 80°C, between days 5 and 6, more C and N were solubilized in the treated compost. After introducing a microbial inoculum on day 6, and maintaining the substrate at 48°C, colonization by bacteria was faster in the treated compost and consequently, more fibre was degraded. Differences between composts in yields of Agaricus bisporus after 5 weeks of cropping were not significant (P=0.05).The authors are with INRA, Station de Recherches sur les Champignons, BP 81, 33883 Villenave d'Ornon, France     

Sensitivity of aflatoxin b1 to ionizing radiation

Aflatoxin B1 in a 5-micrograms/ml water solution was sensitive to ionizing radiation. Inactivation was assayed by the Ames microsome mutagenicity test and confirmed by thin-layer chromatography. Destruction of aflatoxin B1 had already begun at 2.5 kilograms (kGy; 1 kGy = 0.1 Mrad), but a dose exceeding 10 kGy was necessary for total destruction.     

Antimicrobial activity of hyaluronic acid

In this work the biological activity of hyaluronic acid (HA), isolated from fowl crests, is evaluated. The data on the physico-chemical analysis of HA are presented. The preparation obtained is characterized by a high content of the main substance and high relative viscosity. Sterilization conditions for HA, depending on the degree of microbial contamination of the preparation, were carried out. As revealed in this study, irradiation with a dose of 1.0 Mrad is sufficient for obtaining a sterile preparation. HA has been found to possess inhibiting activity with respect to Pseudomonas.     

Intracellular changes of metal elements by fucoidan extracted from brown seaweed (Cladosiphon okamuranus)

This study was undertaken to elucidate the intracellular changes of metal elements after the administration of fucoidan extracted from Cladosiphon okamuranus. TRL1215 cells (normal rat liver cell line) were treated with 0, 0.1, or 1.0 mg/ml fucoidan and incubated in 5% CO2 at 37 degrees C. The cellular levels of Mg, Al, Fe, and Zn were significantly increased in the 1.0 mg/ml fucoidan-treated cells compared to those of the 0.1 mg/ml fucoidan-treated cells and the control. Next, TRL1215 cells were cultured on Mylar film overnight. At 24 h after 5-bromo-2'-deoxyuridine dosing, 0, 0.1, or 1.0 mg/ml fucoidan was treated for 9 h. The cellular distribution of elements was analyzed using in-air micro-micro-particle induced X-ray emission. The X-ray spectra showed that yields of Al, Mg, and Zn were high in order of the 1.0 mg/ml fucoidan-treated sample, the 0.1 mg/ml fucoidan-treated sample, and the control. Fe yield was mildly increased by fucoidan administration. In fucoidan-treated cells, the focal accumulation of Br was correlated spatially with phosphorous-rich region, suggesting that Br was localized within the nucleus. Al distribution provided a spatial association with Br map. These data suggest that fucoidan increases the accumulations of Al, Mg, Fe, and Zn in normal rat hepatocytes, and fucoidan-binding Al is postulated to be transferred into the nucleus.     

Effect of γ-Irradiation on the Microflora of Freshwater Fish: I. Microbial Load, Lag Period, and Rate of Growth on Yellow Perch (Perca flavescens) Fillets

Microbial flora were compared in irradiated and nonirradiated yellow perch fillets. These studies included effects of irradiation on the total microbial population, the lag phase, and rate of growth in this freshwater fishery product. The work was conducted concurrently with sensory and chemical evaluation, and constituted part of an investigation designed to evaluate the effect of substerilization doses (0.3 and 0.6 Mrad) of Co⁶⁰ γ rays on the storage life of yellow perch fillets at 1.0 or 6.0 C. In five storage tests, total plate counts prior to irradiation did not exceed 8.7 × 10⁵ per gram of sample; this count was reduced nearly 100% by irradiation with either 0.3 or 0.6 Mrad. Progressively lower maximal bacterial populations and lengthened lag phases were obtained as more radiation was used. The growth rate of the population did not appear to decrease significantly. Microbial data obtained in these studies confirmed the sensory and chemical studies, by indicating that irradiation can significantly extend the refrigerated shelf life of freshwater fish.     

Effect of γ-Irradiation on the Microflora of Freshwater Fish: I. Microbial Load,Lag Period,and Rate of Growth on Yellow Perch (Perca flavescens) Fillets

Microbial flora were compared in irradiated and nonirradiated yellow perch fillets. These studies included effects of irradiation on the total microbial population, the lag phase, and rate of growth in this freshwater fishery product. The work was conducted concurrently with sensory and chemical evaluation, and constituted part of an investigation designed to evaluate the effect of substerilization doses (0.3 and 0.6 Mrad) of Co⁶⁰ γ rays on the storage life of yellow perch fillets at 1.0 or 6.0 C. In five storage tests, total plate counts prior to irradiation did not exceed 8.7 × 10⁵ per gram of sample; this count was reduced nearly 100% by irradiation with either 0.3 or 0.6 Mrad. Progressively lower maximal bacterial populations and lengthened lag phases were obtained as more radiation was used. The growth rate of the population did not appear to decrease significantly. Microbial data obtained in these studies confirmed the sensory and chemical studies, by indicating that irradiation can significantly extend the refrigerated shelf life of freshwater fish.     

Change in Microflora of Sewage Sludge by Gamma-ray Irradiation

Total bacteria of activated dewatered sludge cake of Takasaki city which amounted to 2 × 10⁹ per gram diminished rapidly with the radiation dose, but slowly after 0.5 Mrad, and 10³ per gram survived even after 10 Mrad irradiation. However, coliforms which amounted to 8 × 10⁷ per gram were inactivated below 0.5 Mrad irradiation. The predominant bacteria in non-irradiated sludge were Pseudomonas cepacia and it mainly survived up to 2 Mrad, but Bacillus were predominant at 0.5 to 0.7 Mrad irradiation. The main residual flora from 2 to 5 Mrad was composed of Pseudomonas soranacearum, P. cepacia and P. delafieldii, and the main residual flora in more than 5 Mrad irradiated sludge was P. flava . These typical strains of Pseudomonas in phosphate buffer were radiation sensitive, and their D₁₀ values were from 0.005 to 0.021 Mrad under aerobic irradiation conditions.