Microculture model studies on the effect of various gas atmospheres on microbial growth at different temperatures

A microculture technique, employing 96-well tissue culture plates in plastic bags, was used to test the effect of different gas atmospheres (vacuum, air, nitrogen, and carbon dioxide) on the growth of Escherichia coli, Bacillus macerans, Salmonella typhimurium, Candida albicans, Lactobacillus plantarum, Pseudomonas Acinetobacter/Moraxella -group, Brochothrix thermosphacta and Yersinia enterocolitica at 2, 6, and 20°C. In general, carbon dioxide was the most effective inhibitor. The inhibition increased with decreasing temperature. Only the combination of carbon dioxide and 2°C provided complete inhibition of Broch. thermosphacta and Y. enterocolitica.     

Storage of poultry meat under modified atmospheres or vacuum packs: possible role of microbial metabolites as indicator of spoilage

The effect of carbon dioxide (100%), nitrogen (100%), carbon dioxide/oxygen (20% : 80%) or vacuum pack at 3 and 10°C was studied on the microbial flora, in skinless poultry breast fillets or thigh meat. Lactic acid bacteria and Brochothrix thermosphacta were the predominant organisms in samples stored in vacuum packs, carbon dioxide and nitrogen. Pseudomonads grew only in oxygen/carbon dioxide packaging systems. The concentration of lactate diminished in both thigh and breast meat during storage at 3 and 10°C. This decrease was more pronounced in thigh meat stored under 20% : 80% carbon dioxide/oxygen. Acetate increased to varying degrees in all samples regardless of the storage conditions.     

Biocatalytic synthesis of acrylates in organic solvents and supercritical fluids: III. Does carbon dioxide covalently modify enzymes?

We have previously demonstrated that the activity of the lipase (Candida cylindracea) catalyzed transesterification reaction between methylmethacrylate and 2-ethylhexanol in supercritical carbon dioxide is comparatively low. In this article, we have investigated the same reaction in supercritical carbon dioxide with a special emphasis on determining the extent of any interaction between the enzyme and carbon dioxide. Transesterification reaction rates in hexane and supercritical carbon dioxide are compared at different temperatures. In supercritical carbon dioxide, temperature was found to have no significant effect on reaction rate in the range of 40 degrees to 55 degrees C. Above 55 degrees C, however, the reaction rate increased significantly as a function of temperature. It appears that carbon dioxide forms reversible complexes with the free amine groups on the surface of the enzyme. Direct evidence of modification was obtained using mass spectroscopy to detect the extent of modification of a pure protein. The kinetics of the reaction have been studied in hexane, and they obey a ping-pong bi-bi mechanism with inhibition by 2-ethylhexanol. The effect of bubbling carbon dioxide and/or fluoroform on the reaction rate in hexane at different temperatures suggests that the enzyme undergoes shear inactivation in hexane. (c) 1995 John Wiley & Sons, Inc.     

Effect of modified atmosphere composition on the metabolism of glucose by Brochothrix thermosphacta

The influence of atmosphere composition on the metabolism of Brochothrix thermosphacta was studied by analyzing the consumption of glucose and the production of ethanol, acetic and lactic acids, acetaldehyde, and diacetyl-acetoin under atmospheres containing different combinations of carbon dioxide and oxygen. When glucose was metabolized under oxygen-free atmospheres, lactic acid was one of the main end products, while under atmospheres rich in oxygen mainly acetoin-diacetyl was produced. The proportions of the total consumed glucose used for the production of acetoin (aerobic metabolism) and lactic acid (anaerobic metabolism) were used to decide whether aerobic or anaerobic metabolism predominated at a given atmosphere composition. The boundary conditions between dominantly anaerobic and aerobic metabolisms were determined by logistic regression. The metabolism of glucose by B. thermosphacta was influenced not only by the oxygen content of the atmosphere but also by the carbon dioxide content. At high CO(2) percentages, glucose metabolism remained anaerobic under greater oxygen contents.     

Antagonism betweenYersinia intermedia andYersinia enterocolitica in water

One strain of Yersinia enterocolitica and one strain of Y. intermedia were grown in peptone water at 25 or 37 degrees C, or in ground water at 15 degrees C. Similar growth rates were observed when these strains were cultivated separately in the same media and at the same temperature. Mixed cultures at 37 degrees C displayed equivalent growth rates. In contrast, mixed cultures incubated at 15 or 25 degrees C were regularly unfavourable to Y. enterocolitica, whereas they did not modify the growth of Y. intermedia. A bacteriophage active on Y. enterocolitica and not on Y. intermedia was characterized from the filtrate of mixed cultures at low temperatures. This phage produced by the lysogenic Y. intermedia strain might be a potential factor responsible for the inhibition of Y. enterocolitica, since no additional antibacterial factor or nutritional competition between Y. intermedia and Y. enterocolitica were found in the mixed cultures.     

Inhibition of Yersinia enterocolitica serotype O3 by natural microflora of pork

Yersinia enterocolitica serotype O3 did not grow but did survive in inoculated raw ground pork kept at 6 and 25 degrees C. The antagonistic effect of microbial flora, especially Hafnia alvei and environmental Yersinia organisms, on the growth of Y. enterocolitica serotype O3 in raw ground pork was evident. These results were supported by evidence of the inhibition of growth of Y. enterocolitica serotype O3 by Enterobacteriaceae, especially H. alvei and environmental Yersinia organisms, in mixed cultures at 6 and 25 degrees C. We suggest that naturally contaminated pork is a source of human infection, since Y. enterocolitica serotype O3 was capable of surviving in the raw pork for a long time.     

Inhibition of Yersinia enterocolitica serotype O3 by natural microflora of pork.

Yersinia enterocolitica serotype O3 did not grow but did survive in inoculated raw ground pork kept at 6 and 25 degrees C. The antagonistic effect of microbial flora, especially Hafnia alvei and environmental Yersinia organisms, on the growth of Y. enterocolitica serotype O3 in raw ground pork was evident. These results were supported by evidence of the inhibition of growth of Y. enterocolitica serotype O3 by Enterobacteriaceae, especially H. alvei and environmental Yersinia organisms, in mixed cultures at 6 and 25 degrees C. We suggest that naturally contaminated pork is a source of human infection, since Y. enterocolitica serotype O3 was capable of surviving in the raw pork for a long time.     

High-temperature controlled atmosphere for post-harvest control of Indian meal moth (Lepidoptera: Pyralidae) on preserved flowers

High carbon dioxide atmospheres combined with high temperature were effective for controlling Indian meal moth, Plodia interpunctella (Hübner) pupae. Pupae were exposed to atmospheres of 60, 80, or 98% carbon dioxide (CO2) in nitrogen (N2), or 60 or 80% CO2 in air at temperatures of 26.7 degrees C or 32.2 degrees C and 60% RH. Controlled atmosphere treatments at 32.2 degrees C controlled pupae faster than the same treatments at the lower temperature. At both temperatures high CO2 concentration treatments combined with nitrogen killed pupae faster than high CO2 concentration treatments combined with air. Exposure to 80% carbon dioxide mixed with nitrogen was the most effective treatment causing 100% mortality in 12 h at 32.2 degrees C and 93.3% mortality in 18 h at 26.6 degrees C. High-temperature controlled atmosphere treatments had no adverse effects on quality of two preserved floral products, Limonium sinuatum (L.) and Gypsophila elegans (Bieb.), tested for 12, 18, and 24 h according to industry standards.     

Factors affecting growth and lipase production by meat lactobacilli strains and Brochothrix thermosphacta

Lipolytic activity of lactobacilli strains and Brochothrix thermosphacta was cell-related; no significant activity was found in the supernatant fluids. Most lipase was produced during the logarithmic phase of growth and was greatly affected by growth conditions. The optimal temperatures for growth and lipase production were respectively 24 degrees C for B. thermosphacta and 30 degrees C for lactobacilli. For all strains, an initial pH of around 7.0 for the medium and low glucose concentration stimulated lipase production. Tributyrin inhibited both growth and lipase production at a concentration of 0.1% for B. thermosphacta or 1% for lactobacilli. Butyric acid (0.1%) and anaerobic culture inhibited lipase production by B. thermosphacta while these two factors had no effect on enzyme production by lactobacilli.     

Analysis of bacteriocinogenic properties of Yersinia enterocolitica strains

The aim of the study was the investigation of bacteriocinogenic properties of 102 Yersinia enterocolitica strains. The influence of selected factors on the production of bacteriocins by Y. enterocolitica and properties of jersiniacin 44JPSBKOH were also investigated. Bacteriocinogenic properties of Y. enterocolitica strains were tested by using the delayed cross-streaking method. It was found that the production of bacteriocins by Y. enterocolitica depended on the type of media on which the producer and indicator strains were grown. It turned out that some strains of Y. enterocolitica showed bacteriocinogenic properties at 25 degrees C, 30 degrees C and 37 degrees C irrespective of the presence of manganese ions in medium. In the presence of iron ions these strains showed bacteriocinogenic properties only at 25 degrees C. Y. enterocolitica strains which required Mn2+ or Mn7+ ions for bacteriocins production showed this activity only at 25 degrees C but in presence of Fe3+ ions they had no bacteriocinogenic properties. The partially purified jersiniacin 44JPSBKOH is a protein, its molecular weight was estimated to be 40 kDa. Yersiniacin 44JPSBKOH was active in the pH range of 3 to 9. Its bactericidal activity was rapidly lost when heated to 100 degrees C and treated with proteolytic enzymes. Yersiniacin 44JPSBKOH showed bactericidal activity against other Y. enterocolitica strains and some strains of Pseudomonas aeruginosa isolated from humans.     

Supercritical carbon dioxide extraction of chamomile flowers: extraction efficiency, stability, and in-line inclusion of chamomile-carbon dioxide extract in beta-cyclodextrin

The extraction of chamomile flowers using supercritical carbon dioxide was investigated with respect to extraction efficiency and compared with solvent extraction. The stability of matricine, a sensitive constituent of the essential oil of chamomile, in these extracts was studied during storage at different temperatures over 6 months. Matricine was stable at -30 degrees C. A slight decrease (80-90% recovery) occurred at +5 degrees C, whereas complete decomposition of matricine took place within 3-4 months at room temperature and at +30 degrees C, respectively. An in-line inclusion of chamomile constituents in beta-cyclodextrin (beta-CD) during the extraction process was assessed and inclusion rates between 40 and 95% were obtained depending on the amount of beta-CD and the type of chamomile constituent. No further stabilization of matricine in the carbon dioxide extract/beta-CD complexes was achieved. High residual water contents in the complexes even after freeze-drying were identified as accelerating the decomposition. In addition, the extractability of flavonoids, such as apigenin and apigenin-7-glucoside, was determined. Apigenin-7-glucoside, the more hydrophilic substance, was not extractable with pure carbon dioxide and showed a recovery of 11% using methanol modified carbon dioxide (18%, w/w) at 60 degrees C and 380 bar. Extraction conditions in the two-phase region of the binary mixture carbon dioxide-methanol (70 degrees C, 100 bar) led to a drastic change in fluid polarity and hence extractability increased to 92-95%.     

Inhibition of Listeria innocua and Brochothrix thermosphacta in vacuum-packaged meat by addition of bacteriocinogenic Lactobacillus curvatus CRL705 and its bacteriocins

AIMS: To evaluate the inhibition effectiveness of Lactobacillus curvatus CRL705 used as a bioprotective culture and of its bacteriocins, lactocin 705 and lactocin AL705, against Listeria innocua, Brochothrix thermosphacta and indigenous lactic acid bacteria (LAB) in vacuum-packaged meat stored at 2 degrees C. METHODS AND RESULTS: The live culture of Lact. curvatus CRL705 as well as synthetic lactocin 705 and purified lactocin AL705 were shown to be similarly effective in preventing the growth of B. thermosphacta and L. innocua in meat discs in contrast to control samples in which these micro-organisms grew rapidly, their numbers increasing by 3.0- and 2.1-log cycles respectively. In addition, indigenous LAB population showed a lower growth rate in the presence of lactocin 705. Bacteriocin activity was detected in the meat discs during 36 days at 2 degrees C irrespective of the biopreservation strategy applied. Changes in pH were not significantly different in meat discs treated with the protective culture when compared with control samples. CONCLUSIONS: Lactobacillus curvatus CRL705 and the produced bacteriocins, lactocin 705 and lactocin AL 705, were effective in inhibiting L. innocua and B. thermosphacta. The use of the bioprotective culture in refrigerated vacuum-packaged fresh meat would be more feasible from an economic and legal point of view. SIGNIFICANCE AND IMPACT OF THE STUDY: Establishment of biopreservation as a method to ensure the microbiological safety of vacuum-packaged fresh meat at 2 degrees C.     

The effect of carbon dioxide on the growth of Yersinia enterocolitica in a simulated milk medium

Carbon dioxide (30 mmol/l) was shown to inhibit the growth of four strains of Yersinia enterocolitica grown separately in a simulated milk medium at 7°C and 60 rev/min for 4 d. This indicates that addition of CO₂ to refrigerated raw milk supplies is likely to be a safe process with respect to that organism.     

Antagonism by gram-negative bacteria to growth of Yersinia enterocolitica in mixed cultures

The inhibition of the growth of Yersinia enterocolitica by other gram-negative bacteria in mixed cultures at 32 degrees C was not the consequence of a depletion in essential nutrients, an unfavorable change in pH or oxygen tension or the production of toxic metabolic products. The inability of Y. enterocolitica to attain its potential maximum population in mixed cultures appeared instead to result from "metabolic crowding," which occurred when the faster-growing antagonistic organism reached stationary-phase density. Lowering the incubation temperature, a technique commonly used in "cold" enrichment for isolation of Y. enterocolitica, tended to equalize growth rates and thereby allowed Y. enterocolitica to achieve a higher population.     

A note on the microbiology of retail packs of prepared salad vegetables

Retail packs of mixed, prepared salad vegetables from two different manufacturers were stored at 7 degrees C until the end of storage-life (sell-by date plus 1 d), when the microbial flora was examined. The quality of the salads was acceptable at the end of storage life. The oxygen concentrations in packs were lower, and the carbon dioxide concentrations were higher, than those in air. High numbers of bacteria were present, with Pseudomonas spp. and Enterobacter agglomerans predominating in packs of both salads, together with lactic acid bacteria in one of the salads. Significant numbers of pectolytic bacteria including Pseudomonas spp. and Erwinia carotovora were detected. Despite the presence of high numbers of coliform bacteria, Escherichia coli was not detected in batches of one salad, and was detected in relatively low numbers in batches of the other. Yersinia spp., predominantly environmental strains of Yersinia enterocolitica, were isolated by enrichment from all samples tested; Staphylococcus aureus and enterococci were not detected.     

Growth and end-product formation in fermenter cultures of Brochothrix thermosphacta ATCC 11509T and two psychrotrophic Lactobacillus spp. in different gaseous atmospheres

The effects of different gaseous atmospheres were determined on the maximum specific growth rate (μ_max) and end-product formation by Brochothrix thermosphacta ATCC 11509^T, Lactobacillus viridescens SMRICC 174 and Lactobacillus sp. SMRICC 173 (homofermentative). The highest μ_max-values for Lact. viridescens (0.47/h) and Broc. thermosphacta (0.49/h) were obtained in air. Under anaerobic conditions μ_max was reduced, an atmosphere containing CO₂ alone giving the greatest reduction. Lactobacillus sp. 173 did not grow in air or N₂. Aerobic growth was obtained by adding peroxidase while anaerobic growth occurred in the presence of 5–20% CO₂. Carbon dioxide alone reduced the growth rate. All test organisms produced mainly lactic acid anaerobically. Lactobacillus viridescens also produced ethanol while Broc. thermosphacta produced small amounts of ethanol and formic acid. With O₂ present, the number of end-products increased for all organisms. Lactobacillus sp. 173 produced small amounts of acetic acid and acetoin together with lactic acid. Oxygen induced acetic acid production in Lact. viridescens and Broc. thermosphacta . Aerobically, Broc. thermosphacta also produced a large amount of acetoin and smaller amounts of 2,3-butanediol, iso -valeric acid and iso -butyric acid. The production of lactic acid by Broc. thermosphacta was completely prevented under strictly aerobic conditions. All test organisms consumed O₂ during aerobic growth. Hydrogen peroxide was produced by Lact. viridescens and Lactobacillus sp. 173.     

Growth and end-product formation in fermenter cultures of Brochothrix thermosphacta ATCC 11509T and two psychrotrophic Lactobacillus spp. in different gaseous atmospheres

The effects of different gaseous atmospheres were determined on the maximum specific growth rate (mumax) and end-product formation by Brochothrix thermosphacta ATCC 11509T, Lactobacillus viridescens SMRICC 174 and Lactobacillus sp. SMRICC 173 (homofermentative). The highest mumax-values for Lact. viridescens (0.47/h) and Broc. thermosphacta (0.49/h) were obtained in air. Under anaerobic conditions mumax was reduced, an atmosphere containing CO2 alone giving the greatest reduction. Lactobacillus sp. 173 did not grow in air or N2. Aerobic growth was obtained by adding peroxidase while anaerobic growth occurred in the presence of 5-20% CO2. Carbon dioxide alone reduced the growth rate. All test organisms produced mainly lactic acid anaerobically. Lactobacillus viridescens also produced ethanol while Broc. thermosphacta produced small amounts of ethanol and formic acid. With O2 present, the number of end-products increased for all organisms. Lactobacillus sp. 173 produced small amounts of acetic acid and acetoin together with lactic acid. Oxygen induced acetic acid production in Lact. viridescens and Broc. thermosphacta. Aerobically, Broc. thermosphacta also produced a large amount of acetoin and smaller amounts of 2,3-butanediol, iso-valeric acid and iso-butyric acid. The production of lactic acid by Broc. thermosphacta was completely prevented under strictly aerobic conditions. All test organisms consumed O2 during aerobic growth. Hydrogen peroxide was produced by Lact. viridescens and Lactobacillus sp. 173.     

Antagonism by gram-negative bacteria to growth of Yersinia enterocolitica in mixed cultures.

The inhibition of the growth of Yersinia enterocolitica by other gram-negative bacteria in mixed cultures at 32 degrees C was not the consequence of a depletion in essential nutrients, an unfavorable change in pH or oxygen tension or the production of toxic metabolic products. The inability of Y. enterocolitica to attain its potential maximum population in mixed cultures appeared instead to result from "metabolic crowding," which occurred when the faster-growing antagonistic organism reached stationary-phase density. Lowering the incubation temperature, a technique commonly used in "cold" enrichment for isolation of Y. enterocolitica, tended to equalize growth rates and thereby allowed Y. enterocolitica to achieve a higher population.     

Effects of low temperature and freeze-thaw cycles on hydrocarbon biodegradation in Arctic tundra soil.

Degradation of petroleum hydrocarbons was monitored in microcosms with diesel fuel-contaminated Arctic tundra soil incubated for 48 days at low temperatures (-5, 0, and 7 degrees C). An additional treatment was incubation for alternating 24-h periods at 7 and -5 degrees C. Hydrocarbons were biodegraded at or above 0 degrees C, and freeze-thaw cycles may have actually stimulated hydrocarbon biodegradation. Total petroleum hydrocarbon (TPH) removal over 48 days in the 7, 0, and 7 and -5 degrees C treatments, respectively, was 450, 300, and 600 microg/g of soil. No TPH removal was observed at -5 degrees C. Total carbon dioxide production suggested that TPH removal was due to biological mineralization. Bacterial metabolic activity, indicated by RNA/DNA ratios, was higher in the middle of the experiment (day 21) than at the start, in agreement with measured hydrocarbon removal and carbon dioxide production activities. The total numbers of culturable heterotrophs and of hydrocarbon degraders did not change significantly over the 48 days of incubation in any of the treatments. At the end of the experiment, bacterial community structure, evaluated by ribosomal intergenic spacer length analysis, was very similar in all of the treatments but the alternating 7 and -5 degrees C treatment.     

Effects of manufacturing conditions on the adsorption capacity of heavy metal ions by Makino bamboo charcoal

The objective of this study was to investigate the effects of manufacturing conditions on the adsorption capacity of heavy metal ions by Makino bamboo charcoal. Results show that the specific surface area and iodine number of bamboo charcoal activated at 900 degrees C were larger than those of bamboo charcoal activated at 800 degrees C. The specific surface area of bamboo charcoal activated at 800 degrees C by carbon dioxide was larger than that of charcoal activated by steam. However, a contrary result was observed when the activation temperature was 900 degrees C. The total volume and proportion of micropores in bamboo charcoal activated by carbon dioxide were greater than those in the other sample groups. However, the total volume and bulk volume of meso- and macropores, and average pore diameter for bamboo charcoal activated by steam were greater than those in the other sample groups. Using 5g bamboo charcoal (10-30 mesh) with a soaking time of 24h, a better adsorption effect on Pb2+ (100%), Cu2+ (100%), and Cr3+ (88-98%) was found. However, medium frequencies were observed for the adsorption of Cd2+ (40-80%) and Ni2+ (20-60%). Very limited adsorption of As5+ was detected in this study. For the same charcoal grain sizes, the adsorption capacity of 0.5g of charcoal was better than that of 0.1g. The improved adsorption effect of the sample group activated by steam was compared with the sample group activated by carbon dioxide.