Aerobic Incubation Conditions Research Articles

Acetylene Tunes Microbial Growth During Aerobic Cometabolism of Trichloroethene.

Microbial aerobic cometabolism is a possible treatment approach for large, dilute trichloroethene (TCE) plumes at groundwater contaminated sites. Rapid microbial growth and bioclogging pose a persistent problem in bioremediation schemes. Bioclogging reduces soil porosity and permeability, which negatively affects substrate distribution and contaminant treatment efficacy while also increasing the operation and maintenance costs of bioremediation. In this study, we evaluated the ability of acetylene, an oxygenase enzyme-specific inhibitor, to decrease biomass production while maintaining aerobic TCE cometabolism capacity upon removal of acetylene. We first exposed propane-metabolizing cultures (pure and mixed) to 5% acetylene (v v-1) for 1, 2, 4, and 8 d and we then verified TCE aerobic cometabolic activity. Exposure to acetylene overall decreased biomass production and TCE degradation rates while retaining the TCE degradation capacity. In the mixed culture, exposure to acetylene for 1-8 d showed minimal effects on the composition and relative abundance of TCE cometabolizing bacterial taxa. TCE aerobic cometabolism and incubation conditions exerted more notable effects on microbial ecology than did acetylene. Acetylene appears to be a viable approach to control biomass production that may lessen the likelihood of bioclogging during TCE cometabolism. The findings from this study may lead to advancements in aerobic cometabolism remediation technologies for dilute plumes.

Kinetics of antimony biogeochemical processes under pre-definite anaerobic and aerobic conditions in a paddy soil

While the transformation of antimony (Sb) in paddy soil has been previously investigated, the biogeochemical processes of highly chemical active Sb in the soil remain poorly understood. In addition, there is a lack of quantitative understanding of Sb transformation in soil. Therefore, in this study, the kinetics of exogenous Sb in paddy soils were investigated under anaerobic and aerobic incubation conditions. The dissolved Sb(V) and the Sb(V) extracted by diffusive gradient technique decreased under anaerobic conditions and then increased under aerobic conditions. The redox reaction of Sb occurred, and Sb bioavailability significantly decreased after 55 days of incubation. The kinetics of Fe and the scanning transmission electron microscopy analysis revealed that the Fe oxides were reduced and became dispersed under anaerobic conditions, whereas they were oxidized and re-aggregated during the aerobic stage. In addition, the redox processes of sulfur and nitrogen were detected under both anaerobic and aerobic conditions. Based on these observations, a simplified kinetic model was established to distinguish the relative contributions of the transformation processes. The bioavailability of Sb was controlled by immobilization as a result of S reduction and by mobilization as a result of Fe reductive dissolution and S oxidation, rather than the pH. These processes coupled with the redox reaction of Sb jointly resulted in the complex behavior of Sb transformation under anaerobic and aerobic conditions. The model-based method and findings of this study provide a comprehensive understanding of the Sb transformation in a complex soil biogeochemical system under changing redox conditions.

Evaluation of a commercial loop-mediated isothermal amplification assay, 3MTM Molecular Detection Assay 2 – Campylobacter, for the detection of Campylobacter from poultry matrices

ABSTRACT 1. The objective of this study was to evaluate performance of a commercial loop-mediated isothermal amplification (LAMP) method as an alternative method for the detection of Campylobacter spp. in primary production samples, poultry rinses and raw poultry products, as compared to the US Department of Agriculture Food Inspection Service Microbiology Laboratory Guide Book PCR reference method, MLG 41A. 2. The Campylobacter spp. LAMP was used in conjunction with a ready-to-use enrichment broth that does not require microaerophilic incubation. After enrichment, boot swabs from poultry farms, carcase rinses and raw poultry products were tested by the LAMP method and the MLG 41A PCR method. 3. The ready-to-use enrichment broth enabled the growth of Campylobacter spp. within 22 to 28 hours under aerobic incubation conditions. The LAMP method enabled Campylobacter detection in the enriched samples of various poultry matrices and had equivalent sensitivity and specificity to the MLG 41A PCR method. 4. No significant difference (95% confidence interval) was found between the alternative and the MLG 41A PCR method, as determined by probability of detection analysis, except for neutralising buffered peptone water post-chill rinsates. For the post-chill neutralising buffered peptone water rinsates, the LAMP method had significantly higher confirmed portions.

Impact of Different Parameters upon the Production of Virulence Factors in Escherichia coli Strains Isolated from Marine Water

This study aimed to look into the expression of 10 virulence factors (VF) in 100 Escherichia coli strains harvested from the Black Sea Coast. E. coli strains were grown under different conditions, simulating various environmental stressors. The VF production [esculinase, amylase, pore-forming enzymes (hemolysin - spot and CAMP-like hemolysis; lipase; lecithinase)], lysine-decarboxylase, proteases (caseinase, gelatinase) and DNase was investigated at variable temperatures (4°C, 22°C, 37°C, 44°C and 56°C), NaCl (from 0 to 10%) and glucose concentrations (1.5% and 3%), different pH values (5.0, 7.2 and 9.6) and also in aerobic or anaerobic incubation conditions. The investigation of E. coli strains unraveled their ability to grow at 22°C, 37°C, 44°C, regardless of the salinity, pH, and glucose concentration, both in aerobic and anaerobic incubation conditions. The VF were better expressed at 37°C, followed by 22°C, especially siderophores, amylase, and caseinase production. The expression of different VF was variable at a certain salinity, i.e., at 0% NaCl, only amylase and siderophores production was observed. At 2% and 3%, the amylase was better expressed. The best expression of siderophores and caseinase was at 6% NaCl. At higher salinity, the expression of VF started to decrease. The amylase and caseinase were better expressed at pH 9.6 and siderophores at pH 7.2. Higher glucose concentrations (3%) proved to have an inhibitory effect on amylase expression and caseinase. The aerobic/anaerobic incubation conditions exhibited no significant differences in the VF expression. In conclusion, these outcomes reveal the ability of enterobacterial aquatic strains to survive in the presence of different stressors and maintain the expression of potential VF expression even in extreme environmental conditions.

Effect of combination of Oxyrase and sodium thioglycolate on growth of Clostridium perfringens from spores under aerobic incubation

Clostridium perfringens is a strictly anaerobic pathogen that requires absence of oxygen for its growth in laboratory experiments, which is usually attained by using an anaerobic chamber or anaerobic jars. However, it has been demonstrated that C. perfringens may survive for short periods of times due to its adaptive response to O2. Therefore, the objective of this study was to explore the application of Oxyrase (OX) and sodium thioglycolate (ST) as oxygen scavengers, used alone or in combination, for observation of the growth of C. perfringens under aerobic incubation.The growth of C. perfringens from spores in Schaedler Anaerobe Agar containing different levels and combinations of OX and ST was observed at temperatures between 20 and 50 °C. The kinetic parameters, including lag time, specific growth rate, and maximum cell concentrations in the stationary phase, were determined.The results indicated that ST at concentrations of 0.025 and 0.05% (w/w), although allowing eventual growth of C. perfringens, prolonged its lag times, while OX at 1.5% only allowed growth at a lower growth rate in comparison to anaerobic incubation. OX at 3% enhanced the growth of C. perfringens at temperatures between 30 and 50 °C, while higher levels of OX were needed in the medium to support the growth of C. perfringens during storage at 25 °C (>6% OX) and 20 °C (>9% OX), due to the effect of temperature on enzyme activity. No significant difference was found in the kinetic parameters of C. perfringens incubated aerobically with OX and the control (without OX or ST) in an anaerobic chamber. Therefore, OX at appropriate concentrations may allow the observation of the growth of C. perfringens under aerobic incubation conditions without the need of an anaerobic device.

How the environmental fate of clomazone in rice fields is influenced by amendment with olive-mill waste under different regimes of irrigation and tillage.

Irrigation and tillage systems alone or in combination with organic amendments can strongly influence soil properties, which in turn may also modify the environmental fate of any pesticides applied. This study was aimed at determining how amendment with composted olive-mill waste (W) influenced leaching, sorption, and persistence of the herbicide clomazone in rice field soils under different tillage and irrigation management practices. The field trial conducted covered 3 years in succession, with six treatments: irrigation by sprinkler and conventional tillage without (ST) or with W application (80 Mg ha-1 ) (STW), irrigation by sprinkler but no tillage (SNT), irrigation by sprinkler but no tillage with W application (SNTW), and continuous flooding irrigation and tillage without (FT) and with W application (FTW). Application of W significantly increased the adsorption of clomazone to soil in the first and third years. In the first year, the persistence of clomazone under aerobic (t1/2 =33.1-36.3 days) and anaerobic incubation conditions (t1/2 =3.43-10.8 days) decreased after W application to t1/2 values in the ranges 18.1-29.7 and 3.06-5.44 days, respectively. However, in the third year, although clomazone persistence increased significantly in SNT and ST when W was applied under both incubation conditions, it decreased significantly in FT management under anaerobic incubation conditions. The addition of W led to less leaching of clomazone, particularly for the FT case where the herbicide leaching losses were 2.8 and 2.6 times lower in the first and third years after W addition, respectively. Using W as an organic amendment could be regarded as an invaluable strategy to reduce water contamination by clomazone in rice production, especially under traditional tillage and flooding management. © 2019 Society of Chemical Industry.

Olive mill sludge may reduce water contamination by 4-chloro-2-methylphenoxyacetic acid (MCPA) in non-flooding but enhance it in flooding rice cropping agroecosystems

A field experiment covering three years was conducted to evaluate how composted olive mill sludge (OS) influenced MCPA's environmental fate in rice soils under six combinations of tillage and irrigation cultivation techniques: tillage and sprinkler irrigation without (TS) or with (TSOS) the addition of OS (80 Mg ha−1), no-tillage and sprinkler irrigation without (NTS) or with (NTSOS) OS, and tillage and continuous flooding without (TF) or with (TFOS) OS. The measurements made in the first and third years after OS application were taken to constitute the “direct” and “residual” effects, respectively. After OS amendment, Kd (partition coefficients) values in the direct year were lower by factors of 1.1, 1.3, and 1.9 in TSOS, NTSOS, and TFOS, respectively, relative to the corresponding unamended soils, and in the residual year by factors of 1.1 and 1.5 in TSOS and NTOS, but greater by a factor of 1.5 in TFOS, than in the corresponding unamended soils, respectively. The dissipation of MCPA was very fast under both anaerobic (t1/2 = 1.80–5.29 d) and aerobic (t1/2 = 2.23–9.42 d) incubation conditions. The field application of OS led to a decrease in MCPA persistence under both incubation conditions, especially in the TF case. However, while under aerobic conditions the half-life (t1/2) decreased after OS addition in the direct and residual years, under anaerobic condition it only decreased in the direct year. While the application of OS in TS and NTS led to less leaching of MCPA, in TF it led to 1.4 and 1.2 times more leaching losses of the herbicide for the direct and residual years, respectively. Therefore, the use of OS in rice production could be considered an effective strategy for reducing water contamination by MCPA in at least the short- and medium-terms after its application, but only under non-flooding crop management regimes irrespective of the tillage practice implemented.

Ways to Improve Quality Control of Culture Medium Used for Mycoplasma Detection

An urgent safety concern associated with biological products is contamination with mycoplasmas, which may originate from donor tissues and organs, virus harvests, culture medium components, trypsin, animal blood serum, as well as be transmitted by personnel involved in the manufacture of medicines. Currently, due to an increase in the range of biologicals available, there is a need for more sensitive and specific test methods. In the Russian practice, microbiological (culture-based) testing of finished pharmaceutical products for mycoplasma contamination is performed using complex culture media whose sensitivity depends on the quality of proteins, ingredients, and reagents used. Growth promotion properties of the media are determined according to the State Pharmacopoeia of the Russian Federation, 14th ed., using a single test strain — Mycoplasma arginini G230 (M. arginini G230 industry reference material). The aim of the study was to analyse current Russian and foreign requirements for the quality control of culture media that are used for mycoplasma detection, in order to update and improve the quality control procedure in Russia. It was demonstrated that a compelling advantage of the State Pharmacopoeia of the Russian Federation is the possibility of using a semi-liquid culture medium which does not require special aerobic or anaerobic incubation conditions and allows for quantification of mycoplasma colonies and determination of mycoplasma titre in culture medium while testing its growth promotion properties using reference М. arginini G230 test strain. The analysis revealed some differences in Russian and foreign requirements for quality evaluation of culture media. These differences were taken into account when developing recommendations for improvement of the Russian test procedure, i.e. enlarging the range of test strains used and development of respective reference standards.

Behaviour of bispyribac-sodium in aerobic and anaerobic rice-growing conditions with and without olive-mill waste amendment

Water and tillage management practices alone or in combination with organic amendments can alter the behaviour of pesticides. This study evaluates the sorption, persistence, and leaching of bispyribac-sodium (BYS) under anaerobic and aerobic rice-growing conditions with and without soil amendment by composted olive mill waste (OW). For this purpose, field experiments were conducted over three years. They involved six treatments: no-tillage and sprinkler irrigation (aerobic) without (NTA) and with OW application (NTAOW), tillage and aerobic without (TA) and with OW application (TAOW), and tillage and flooding without (TF) and with OW application (TFOW). In all three OW treatments, the amendment was applied in the first year. The Kf values were significantly greater in TF than in [NTA and TA] for the first and third years of the experiment by factors of [1.5 and 2.3] and [1.6 and 1.2], respectively. While in the first year Kf was 1.5 times greater in TF than TFOW, at the end of the experiment it was 1.6 times lower. In the non-amended treatments, the persistence of BYS was up to 1.9 times greater under anaerobic than under aerobic incubation conditions. Except for the NTA treatment in the third year, the persistence of BYS under aerobic incubation conditions (t1/2 = 35.5–53.7 d) was enhanced when OW was applied to the soil (t1/2 = 44.5–80.1 d). Under anaerobic incubation conditions, in the first year the BYS persistence was significantly reduced in NTA and TA when OW was added to the soil, but in TF it was enhanced. Conversely, in the third year the persistence of BYS was significantly increased in NTA and TA by OW addition, but decreased under TF management. BYS leaching was similar under the aerobic (TA and NTA) and anaerobic (TF) treatments in the first year, but was less under the TF treatment at the end of the experiment. Independently of the water and tillage management, the addition of OW significantly influenced BYS sorption and persistence, reducing the amount of the herbicide leached, although the magnitude of the effect depended on the time elapsed after its application to the soil. The use of OW could be considered as a strategy with which to reduce water contamination by BYS under aerobic and anaerobic rice-growing conditions, especially in the short term after application of this organic amendment.

Nitrogen mineralization simulation dynamic in tobacco soil

Laboratory and field culture experiments have been carried out at two sites which show that temperature and soil moisture significantly affected nitrogen mineralization in the soil in a tobacco field. Intermittent leaching under aerobic incubation conditions was adopted to develop dynamic simulation models of organic nitrogen mineralization under different temperature regimes (10, 15, 20, 25, 30, 35, 30, 25, and 20 °C) and soil gravimetric moisture content (0.1, 7%, 13%, 20%, 27%, 33%, 40%, 47%, and 53%). The results show that the highest N mineralization rate appeared at 35°C and the lowest was 10°C. Soil N mineralization rate, as affected by soil moisture, is a single peak curve with a peak at 40%. The interaction of temperature and moisture on soil N mineralization is significant. Based on the results of a field trial, the combined model given by governing equation:

Behaviour of bentazon as influenced by water and tillage management in rice-growing conditions.

Bentazon is a widely used herbicide in rice agroecosystems that has commonly been found in water resources. To assess how tillage and water regimes affect sorption/desorption, dissipation and leaching of bentazon in Mediterranean rice-growing conditions, field experiments were carried out using tillage and flooding (TF), tillage and sprinkler irrigation (TS), no-tillage and sprinkler irrigation (NTS) and long-term no-tillage and sprinkler irrigation (NTS7). After 3 years, the Kd values in TS were 2.3, 1.6 and 1.7 times lower than the values in NTS7, NTS and TF respectively. Greater sorption of bentazon was related to higher contents in total organic carbon and, although to a lesser extent, in humic acids and dissolved organic carbon. The persistence of bentazon was significantly greater under anaerobic (half-life DT50 = 94.1-135 days) than under aerobic (DT50 = 42.4-91.3 days) incubation conditions for all management regimes. Leaching losses of bentazon were reduced from 78 and 74% in TS and TF to 61 and 62% in NTS7 and NTS respectively. The mid- and long-term implementation of sprinkler irrigation in combination with no-tillage could be considered a management system that is effective at reducing water contamination by bentazon in Mediterranean rice-growing agroecosystems. © 2017 Society of Chemical Industry.

Potential carbon emissions dominated by carbon dioxide from thawed permafrost soils

A meta-analysis of soil incubation studies from the permafrost zone suggests that thawing under aerobic conditions, which releases CO2, will strengthen the permafrost carbon feedback more than waterlogged systems, which releases CO2 and CH4. Increasing temperatures in northern high latitudes are causing permafrost to thaw1, making large amounts of previously frozen organic matter vulnerable to microbial decomposition2. Permafrost thaw also creates a fragmented landscape of drier and wetter soil conditions3,4 that determine the amount and form (carbon dioxide (CO2), or methane (CH4)) of carbon (C) released to the atmosphere. The rate and form of C release control the magnitude of the permafrost C feedback, so their relative contribution with a warming climate remains unclear5,6. We quantified the effect of increasing temperature and changes from aerobic to anaerobic soil conditions using 25 soil incubation studies from the permafrost zone. Here we show, using two separate meta-analyses, that a 10 °C increase in incubation temperature increased C release by a factor of 2.0 (95% confidence interval (CI), 1.8 to 2.2). Under aerobic incubation conditions, soils released 3.4 (95% CI, 2.2 to 5.2) times more C than under anaerobic conditions. Even when accounting for the higher heat trapping capacity of CH4, soils released 2.3 (95% CI, 1.5 to 3.4) times more C under aerobic conditions. These results imply that permafrost ecosystems thawing under aerobic conditions and releasing CO2 will strengthen the permafrost C feedback more than waterlogged systems releasing CO2 and CH4 for a given amount of C.

Ten years of elevated atmospheric CO2 doesn't alter soil nitrogen availability in a rice paddy

Elevated atmospheric carbon dioxide (CO2) concentration can affect soil nitrogen (N) cycling in natural and seminatural ecosystems, but the response of soil N cycling to elevated atmospheric CO2 in intensively managed agricultural ecosystems characterized by large N fertilizer inputs remains poorly understood. Here, we investigated the effects of seven and 10 years of elevated CO2 levels on soil gross N transformation rates using the 15N dilution technique at the Rice Free Air CO2 Enrichment (Rice-FACE) experiment in China. Our results show that under aerobic incubation conditions after the first seven years of CO2 enrichment, gross rates of N mineralization, NH4+ immobilization, nitrification, and NO3− immobilization were not significantly different between N application rates or between CO2 treatments. None of the four rates were affected by elevated CO2 levels under both aerobic and water-logged incubation for a further three years of CO2 enrichment. As a result, elevated CO2 levels did not result in changes in available N for plants and soil microbes, and thus did not increase the potential risks of N losses through leaching and runoff. These results are probably associated with the lack of changes in soil organic C and N concentrations due to elevated CO2. In contrast, elevated CO2 levels significantly increased N2O emissions for both incubation conditions. In general, our results suggest that 10 years of elevated atmospheric CO2 concentration had negligible effects on soil N availability in a rice paddy field.

Pericellular oxygen monitoring with integrated sensor chips for reproducible cell culture experiments

Here we present an application, in two tumour cell lines, based on the Sensing Cell Culture Flask system as a cell culture monitoring tool for pericellular oxygen sensing. T-47D (human breast cancer) and T98G (human brain cancer) cells were cultured either in atmospheric air or in a glove-box set at 4% oxygen, in both cases with 5% CO2 in the gas phase. Pericellular oxygen tension was measured with the help of an integrated sensor chip comprising oxygen sensor arrays. Obtained results illustrate variation of pericellular oxygen tension in attached cells covered by stagnant medium. Independent of incubation conditions, low pericellular oxygen concentration levels, usually associated with hypoxia, were found in dense cell cultures. Respiration alone brought pericellular oxygen concentration down to levels which could activate hypoxia-sensing regulatory processes in cultures believed to be aerobic. Cells in culture believed to experience conditions of mild hypoxia may, in reality, experience severe hypoxia. This would lead to incorrect assumptions and suggests that pericellular oxygen concentration readings are of great importance to obtain reproducible results when dealing with hypoxic and normoxic (aerobic) incubation conditions. The Sensing Cell Culture Flask system allows continuous monitoring of pericellular oxygen concentration with outstanding long-term stability and no need for recalibration during cell culture experiments. The sensor is integrated into the flask bottom, thus in direct contact with attached cells. No additional equipment needs to be inserted into the flask during culturing. Transparency of the electrochemical sensor chip allows optical inspection of cells attached on top of the sensor.

Microbial Production of 1,3-Propanediol by a Newly Isolated <i>Citrobacter freundii</i> Strain CF-5

Microorganisms tolerant to 1, 3-propanediol (1, 3-PDO) were screened out from various environmental samples, and isolate CF-5 was found to produce 2.8 g/L 1, 3-PDO with glycerol as carbon source. Based on 16S rRNA gene sequence analysis, CF-5 was identified as aCitrobacter freundiistrain. The fermentation conditions of strain CF-5 were analyzed. Higher biomass production was obtained in the medium supplemented with limited amount of glucose without repression of conversion efficiency of glycerol to 1, 3-PDO. Under the combined aerobic and oxygen limited incubation conditions, higher yield of 1, 3-PDO was reached. The fed-batch fermentation was performed in a 5 L bioreactor, 1, 3-PDO was found mainly produced in exponential growth phase cells ofC. freundiiCF-5 with the highest productivity 0.63 g/L*h, and the overall yield of 1, 3-PDO fermentation was 11.8 g/L within 36 h fermentation.

Higher Water Temperature and Incubation under Aerobic and Microaerobic Conditions Increase the Recovery and Diversity of Arcobacter spp. from Shellfish

Some Arcobacter species are considered emerging food-borne and waterborne pathogens, and shellfish have been suggested as one of their reservoirs. However, only a few studies have investigated the presence of Arcobacter in this kind of food. This study assesses the prevalence and diversity of Arcobacter spp. in shellfish by multiplex PCR (m-PCR) and culturing methods (under different atmospheric conditions) and evaluates the possible influence of environmental parameters (temperature, salinity, and harvesting bay). Arcobacter was detected by m-PCR and/or culturing in 61 (29.9%) of 204 shellfish samples. Of the positive samples by culturing, 41.1% were obtained under only aerobic incubation conditions, while 23.2% were obtained under only microaerobic conditions. Of 476 investigated isolates, 118 belonged to different enterobacterial repetitive intergenic consensus (ERIC)-PCR genotypes (strains) and to 11 different species. This study shows the highest diversity of Arcobacter species ever observed in samples from any origin. The most prevalent species was Arcobacter butzleri (60.2%), followed by Arcobacter molluscorum (21.2%). The prevalence of Arcobacter was significantly higher during the summer than in other seasons, being associated with an increase in water temperature. Results confirm that shellfish are a reservoir for a remarkable diversity of Arcobacter spp.

Effects of land-use type and nitrogen addition on nitrous oxide and carbon dioxide production potentials in Japanese Andosols

Land-use type and nitrogen (N) addition strongly affect nitrous oxide (N2O) and carbon dioxide (CO2) production, but the impacts of their interaction and the controlling factors remain unclear. The aim of this study was to evaluate the effect of both factors simultaneously on N2O and CO2 production and associated soil chemical and biological properties. Surface soils (0–10 cm) from three adjacent lands (apple orchard, grassland and deciduous forest) in central Japan were selected and incubated aerobically for 12 weeks with addition of 0, 30 or 150 kg N ha–1 yr–1. Land-use type had a significant (p < 0.001) impact on the cumulative N2O and CO2 production. Soils from the apple orchard had higher N2O and CO2 production potentials than those from the grassland and forest soils. Soil net N mineralization rate had a positive correlation with both soil N2O and CO2 production rates. Furthermore, the N2O production rate was positively correlated with the CO2 production rate. In the soils with no N addition, the dominant soil properties influencing N2O production were found to be the ammonium-N content and the ratio of soil microbial biomass carbon to nitrogen (MBC/MBN), while those for CO2 production were the content of nitrate-N and soluble organic carbon. N2O production increased with the increase in added N doses for the three land-use types and depended on the status of the initial soil available N. The effect of N addition on CO2 production varied with land use type; with the increase of N addition doses, it decreased for the apple orchard and forest soils but increased for the grassland soils. This difference might be due to the differences in microbial flora as indicated by the MBC/MBN ratio. Soil N mineralization was the major process controlling N2O and CO2 production in the examined soils under aerobic incubation conditions.

Inactivation and sub-lethal injury of salmonella typhi, salmonella typhimurium and vibrio cholerae in copper water storage vessels

BackgroundThis study provides information on the antibacterial effect of copper against the water-borne pathogens Salmonella Typhi, Salmonella Typhimurium and Vibrio cholerae.MethodsSuspensions of each pathogen were kept in water within a traditional copper vessel at 30°C for 24 h. Samples were withdrawn, diluted and plated onto suitable growth media. Conventional enumeration of healthy (uninjured) bacteria was carried out using standard aerobic incubation conditions. Additionally, reactive oxygen species-neutralised (ROS-n) conditions were achieved by adding the peroxide scavenger sodium pyruvate to the medium with anaerobic incubation, to enumerate uninjured (ROS-insensitive) and injured (ROS-sensitive) bacteria. Differences between log-transformed means of conventional (aerobic) and ROS-n counts were statistically evaluated using t tests.ResultsOverall, all three pathogens were inactivated by storage in copper vessels for 24 h. However, for shorter-term incubation (4-12 h), higher counts were observed under ROS-n conditions than under aerobic conditions, which demonstrate the presence of substantial numbers of sub-lethally injured cells prior to their complete inactivation.ConclusionsThe present study has for the first time confirmed that these bacterial pathogens are inactivated by storage in a copper vessel within 24 h. However, it has also demonstrated that it is necessary to account for short-term sub-lethal injury, manifest as ROS-sensitivity, in order to more fully understand the process. This has important practical implications in terms of the time required to store water within a copper vessel to completely inactivate these bacteria and thereby remove the risk of water-borne disease transmission by this route.

Diversity of lactic acid bacteria from modified atmosphere packaged sliced cooked meat products at sell-by date assessed by PCR-denaturing gradient gel electrophoresis

The predominant lactic acid bacteria (LAB) microbiota associated with three types of modified atmosphere packaged (MAP) sliced cooked meat products (i.e. ham, turkey and chicken) was analyzed at sell-by date using a combination of culturing and molecular population fingerprinting. Likewise routine analyses during industrial MAP production, meat samples were plated on the general heterotrophic Plate Count Agar (PCA) and on the LAB-specific de Man, Rogosa, Sharpe (MRS) agar under different temperature and atmosphere conditions. Subsequently, community DNA extracts were prepared from culturable bacterial fractions harvested from both media and used for PCR targeting the V3 hyper-variable region of the 16S rRNA gene followed by denaturing gradient gel electrophoresis (DGGE) of PCR amplicons (PCR-DGGE). Irrespective of aerobic or anaerobic incubation conditions, V3-16S rDNA DGGE fingerprints of culturable fractions from PCA and MRS medium displayed a high level of similarity indicating that LAB constituted the most dominant group in the culturable bacterial community. Comparison of DGGE profiles of fractions grown at 20, 28 or 37 °C indicated that part of the culturable community consisted of psychrotrophs. Four DGGE bands were common among cooked ham, turkey and chicken products, suggesting that these represent the microbiota circulating in the plant where all three MAP product types were sliced and packaged. Based on band sequencing and band position analysis using LAB reference strains, these four bands could be assigned to Lactobacillus sakei and/or the closely related Lactobacillus fuchuensis, Lactobacillus curvatus, Carnobacterium divergens and Leuconostoc carnosum. In conclusion, the PCR-DGGE approach described in this study allows to discriminate, identify and monitor core and occasional LAB microbiota of MAP sliced cooked meat products and provides valuable complementary information to the current plating procedures routinely used in industrial plants.

GROWTH RESPONSE OF LACTOBACILLUS RHAMNOSUS TO MICROAEROPHILIC INCUBATION AND ADDITION OF REDUCTANTS IN A RIBOFLAVIN MICROTITER PLATE ASSAY

ABSTRACT Given the importance of riboflavin in human diets, a rapid riboflavin assay is needed by the food industry to allow simultaneous determination of riboflavin in numerous food sources. The growth responses of Lactobacillus rhamnosus to microaerophilic conditions and three reducing agents were examined as an attempt to reduce riboflavin assay time. Ascorbic acid, dithiothreitol and thioglycollate were applied at three concentrations. None of the concentrations of reducing agents caused a further increase of bacterial optical density (OD) or had a significant effect on the growth rates. More optimal microaerophilic conditions were achieved through the use of an anaerobic chamber. The growth rate of L. rhamnosus under microaerophilic conditions (0.30 ± 0.05/h) was not significantly different (P &gt; 0.05) from the growth rate of the bacteria under aerobic conditions (0.34 ± 0.07/h). However, after 14 h of bacterial growth, the OD of the bacterial culture under anaerobic hood incubation (0.71 ± 0.09) was significantly higher (P &lt; 0.05) than under aerobic incubation conditions (0.58 ± 0.05). Additional research is needed involving more controlled atmospheres for incubation during growth and a wider range of reductants to optimize and shorten the time required for riboflavin microtiter plate assay.PRACTICAL APPLICATIONSQuantifying riboflavin in foods and food ingredients is important to the food industry. Lactobacillus rhamnosus growth response to varying levels of riboflavin is used to estimate the amount of this vitamin. In an effort to optimize growth on microtiter plates and decrease assay time, microaerophilic conditions were examined. Although rate of growth was not altered, cell concentration measured as optical density (OD) did increase. Microaerophilic incubation of microtiter plates represents a potential approach to decrease assay time and provide the food industry with more timely results.