Microbial activities in soil near natural gas leaks

Summary From the present experiments it may be concluded that in the surroundings of natural gas leaks, methane, ethane and possibly some other components of the natural gas are oxidized by microbial activities as long as oxygen is available. This is demonstrated by an increased oxygen consumption and carbon dioxide production, as well as by increased numbers of different types of bacteria. The resulting deficiency of oxygen, the excess of carbon dioxide, and perhaps the formation of inhibitory amounts of ethylene, are considered to be mainly responsible for the death of trees near natural gas leaks. Also the long period of time needed by the soil to recover, may be due to prolonged microbial activities, as well as to the presence of e. g. ethylene.The present experiments suggest that especially methane-oxidizing bacteria of the Methylosinus trichosporium type were present in predominating numbers and consequently have mainly been responsible for the increased oxygen consumption. However, some fungi oxidizing components of natural gas, including methane and ethane may also have contributed to the increased microbial activities in the soil. The same will be true of a possible secondary microflora on products derived from microorganisms oxidizing natural gas components.     

Some aspects on the role of bacteria in deep water ecosystems

Summary Some remarks on the role of bacteria in deep lakes will be presented with an emphasis on their functioning in the carbon cycle in such ecosystems.The metabolic intensity of most lakes is regulated to a large extent at the primary producer level. Aerobic and anaerobic decomposition by heterotrophic bacteria of the unstable ultimate products of photosynthesis results in the production of carbon dioxide and methane. Bacterial growth occurs at the expense of energy released by the flow of electrons from donors to acceptors. Typical electron acceptors for bacterially mediated reactions are oxygen, nitrate, sulphate and carbon dioxide. When oxygen is used as electron acceptor, the highest amount of energy is released, while the lowest is released when carbon dioxide is used. These reactions are mediated biologically, and the chemical reaction sequence is paralleled by an ecological succession of microorganisms: aerobic heterotrophs, denitrifiers, fermenting bacteria, sulphate reducers, and methane producers. The presence of oxygen is inhibitory to the organisms mediating the last reactions (STUMM, 1966; McCARTY, 1972), and this explains the succession of micro-organisms concomittant with the decrease in redox potential.Both labile and refractory fractions of the pelagial dissolved organic matter can directly be utilized largely by heterotrophic bacteria. At the onset of the summer stratification of Lake Vechten a high concentration (numbers 10⁹ bact/L) of heterotrophic bacteria has been detected in the lower water layers (CAPPENBERG, 1972). The reason for this increase may be a release of nutrients from the mud. During stratification the water layers above the mud become anaerobic owing to the metabolic activity of the bacterial flora and the chemical oxygen demand of the mud. In summer time the highest numbers of heterotrophic bacteria are found in the metalimnion, where organic matter accumulates due to a lower relative rate of sedimentation caused by an increase in viscosity and density.Furthermore it was found that sulphate-reducing bacteria which are capable of reducing sulphates to sulphides using sulphate as terminal electron acceptor, were observed in the hypolimnion only at the time of maximal stratification. During stagnation, gradually decreasing sulphate concentrations are found in the hypolimnion, correlated with increaseing cell numbers of sulphate reducers. As no sulphate could be detected in the mud of Lake Vechten as well, we may conclude that the sulphate concentration limits the number of these bacteria, as can be predicted for similar aquatic environments.Usually the sulphate reducers are found in bottom deposits and are an important group of bacteria of the non-methanogenic populations in mud. Summarizing the biological methane production and its subsequent oxidation by methane-oxidizing bacteria, we may conclude that these processes can be important factors functioning in the carbon cycle in deep fresh-water ecosystems.     

Physiological Profiling of Soil Microbial Communities in a Florida Scrub-Oak Ecosystem: Spatial Distribution and Nutrient Limitations

Rapid physiological profiling of heterotrophic microbial communities enables intensive analysis of the factors affecting activity in aerobic habitats, such as soil. Previous methods for performing such profiling were severely limited due to enrichment bias and inflexibility in incubation conditions. We tested a new physiological profiling approach based on a microtiter plate oxygen sensor system (Becton Dickinson Oxygen Biosensor System (BDOBS)), which allows for testing of lower substrate addition (i.e., lower enrichment potential) and manipulation of physiochemical assay conditions, such as pH and nutrients. Soil microbial communities associated with a scrub-oak forest ecosystem on Merritt Island Wildlife Refuge in central Florida, USA, were studied in order to evaluate microbial activity in a nutrient poor soil and to provide baseline data on the site for subsequent evaluation of the effects of elevated CO₂ on ecosystem function. The spatial variation in physiological activity amongst different habitats (litter, bulk soil, and rhizosphere) was examined as a function of adaptation to local resources (i.e., water soluble extracts of roots and leaf litter) and the degree of N and P limitation. All the communities were primarily N-limited, with a secondary P limitation, which was greater in the rhizosphere and bulk soil. The litter community showed greater overall oxygen consumption when exposed to litter extracts relative to the rhizosphere or soil, suggesting acclimation toward greater use of the mixed substrates in the extract. Root extracts were readily used by communities from all the habitats with no habitat specific acclimation observed. A priming effect was detected in all habitats; addition of glucose caused a significant increase in the use of soil organic carbon. Response to added glucose was only observed with N and P addition, suggesting that C may be lost to the groundwater from these porous soils because nutrient limitation prevents C immobilization.     

THE RESPIRATION OF LUMINOUS BACTERIA AND THE EFFECT OF OXYGEN TENSION UPON OXYGEN CONSUMPTION

1. The respiration of luminous bacteria has been studied by colorimetric and manometric methods. 2. Limulus oxyhaemocyanin has been used as a colorimetric indicator of oxygen consumption and indicator dyes were used for colorimetric determination of carbon dioxide production. 3. The Thunberg-Winterstein microrespirometer has been used for the measurement of the rate of oxygen consumption by luminous bacteria at different partial pressures of oxygen. 4. The effect of oxygen concentration upon oxygen consumption has been followed from equilibrium with air to low pressures of oxygen. 5. Luminous bacteria consume oxygen and produce carbon dioxide independent of oxygen pressures from equilibrium with air (152 mm.) to approximately 22.80 mm. oxygen or 0.03 atmosphere. 6. Dimming of a suspension of luminous bacteria occurs when oxygen tension is lowered to approximately 2 mm. Hg (0.0026 atmosphere) and when the rate of respiration becomes diminished one-half. 7. Pure nitrogen stops respiratory activity and pure oxygen irreversibly inhibits oxygen consumption. 8. The curve for rate of oxygen consumption with oxygen concentration is similar to curves for adsorption of gasses at catalytic surfaces, and agrees with the Langmuir equation for the expression of the amount of gas adsorbed in unimolecular layer at catalytic surfaces with gas pressure. 9. A constant and maximum rate of oxygen consumption occurs in small cells when oxygen concentration becomes sufficient to entirely saturate the surface of the oxidative catalyst of the cell.     

Identification of trehalose as a compatible solute in different species of acidophilic bacteria

The major industrial heap bioleaching processes are located in desert regions (mainly Chile and Australia) where fresh water is scarce and the use of resources with low water activity becomes an attractive alternative. However, in spite of the importance of the microbial populations involved in these processes, little is known about their response or adaptation to osmotic stress. In order to investigate the response to osmotic stress in these microorganisms, six species of acidophilic bacteria were grown at elevated osmotic strength in liquid media, and the compatible solutes synthesised were identified using ion chromatography and MALDI-TOF mass spectrometry. Trehalose was identified as one of, or the sole, compatible solute in all species and strains, apart from Acidithiobacillus thiooxidans where glucose and proline levels increased at elevated osmotic potentials. Several other potential compatible solutes were tentatively identified by MALDITOF analysis. The same compatible solutes were produced by these bacteria regardless of the salt used to produce the osmotic stress. The results correlate with data from sequenced genomes which confirm that many chemolithotrophic and heterotrophic acidophiles possess genes for trehalose synthesis. This is the first report to identify and quantify compatible solutes in acidophilic bacteria that have important roles in biomining technologies.     

Engineering analysis of the high-density heterotrophic cultivation of mung bean sprouts

This study investigated the heterotrophic growth behavior of mung beans cultivated in an individual bed under water supply. The fresh weight of mung beans in the bed was estimated, and changes in temperature, and oxygen and carbon dioxide concentrations were recorded during the cultivation period. The specific growth rate, oxygen uptake rate, and carbon dioxide evolution rate, based on the fresh weight in the bed, were calculated. Growth under heterotrophic cultivation can be classified into the following three stages. Reductions in specific oxygen uptake rate, specific carbon dioxide evolution rate, and specific energy production rate corresponded to that of specific growth rate. Indicators of biological activity related to oxygen and carbon dioxide were evaluated quantitatively for beds under high-density heterotrophic cultivation. Moreover, the results obtained from this study successfully demonstrate that there is a relationship between the growth of mung beans and indicators of biological activity.     

Influence of respirometry methods on intraspecific variation in standard metabolic rates in newts

Standard metabolic rate (SMR) is both a highly informative and variable trait. Variation in SMR stems not only from diverse intrinsic and extrinsic factors, but also from the use of diverse methods for metabolic measurements. We measured CO(2) production (VCO(2)) and oxygen consumption rates (VO(2)) using two flow-through respirometry modes, continuous and intermittent (stop-flow), to evaluate their potential contribution to SMR variation in Alpine newts, Ichthyosaura alpestris. Both respirometry modes yielded similar and repeatable VCO(2) values. Although VO(2) was highly repeatable, continuous respirometry produced lower VO(2) than the intermittent method. During intermittent measurements, the total number of activity bouts was higher than during continuous respirometry trials. Statistical correction for disparate activity levels minimized variation in oxygen consumption between respirometry modes. We conclude that use of either method of flow-through respirometry, if properly applied, introduced less noise to SMR estimates than a variation in activity levels.     

Assessment of in-situ bioremediation at a refinery waste-contaminated site and an aviation gasoline contaminated site

A combination of geochemical, microbiological and isotopic methods were used to evaluate in-situ bioremediation of petroleum hydrocarbons at one site contaminated with refinery waste and a second site contaminated with aviation gasoline at Alameda Point, California. At each site, geochemical and microbiological characteristics from four locations in the contaminated zone were compared to those from two uncontaminated background locations. At both sites, the geochemical indicators of in-situbiodegradation includeddepleted soil gas and groundwater oxygen, elevated groundwater alkalinity, and elevated soil gas carbon dioxide and methane in the contaminated zone relative to the background. Radiocarbon content of methane and carbon dioxide measured in soil gas at both sites indicated that they were derived from hydrocarbon contaminant degradation. Direct microscopy of soil core samples using cell wall stains and activity stains, revealed elevated microbial numbers and enhanced microbial activities in contaminated areas relative to background areas, corroborating geochemical findings. While microbial plate counts and microcosm studies using soil core samples provided laboratory evidence for the presence of some microbial activity and contaminant degradation abilities, they did not correlate well with either contaminant location, geochemical, isotopic, or direct microscopy data.     

Column Bioleaching of Low-Grade Chalcopyritic Ore Using Moderate Thermophile Bacteria

Bacterially assisted heap leaching is an economical technology for treating low grade copper sulphides. In the present research, bioleaching of low grade chalcopyrite ore (1% chalcopyrite, and 3% pyrite) have been investigated using moderate thermophilic bacteria. The ore sample has low solubility in acid solution (about 5%). Experiments were carried out using column reactors and the effect of particle size (?12.7, ?19.07 and ?25.04 mm) and external addition of carbon dioxide to the induced air (10% v/v) have been investigated. Results have shown that the copper recovery increased with reducing particle size, and carbon dioxide addition improved bacterial activity and copper dissolution. In the optimum condition, i.e., particles finer than 12.07 mm and 10% (v/v) carbon dioxide addition, 69.68% of copper were extracted.     

Organic carbon and mineral nutrient limitation of oxygen consumption, bacterial growth and efficiency in the Norwegian Sea

To evaluate the role of bacteria in the transformation of organic matter in subarctic waters, we investigated the effect of mineral nutrients (ammonia and phosphate) and organic carbon (glucose) enrichment on heterotrophic bacterial processes and community structure. Eight experiments were done in the Norwegian Sea during May and June 2008. The growth-limiting factor (carbon or mineral nutrient) for heterotrophic bacteria was inferred from the combination of nutrient additions that stimulated highest bacterial oxygen consumption, biomass, production, growth rate and bacterial efficiency. We conclude that heterotrophic bacteria were limited by organic carbon and co-limited by mineral nutrients during the prevailing early nano-phytoplankton (1–10 μm) bloom conditions. High nucleic acid (HNA) bacteria became dominant (>80%) only when labile carbon and mineral nutrient sources were available. Changes in bacterial community structure were investigated using denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR)-amplified 16S ribosomal RNA genes. The bacterial community structure changed during incubation time, but neither carbon nor mineral nutrient amendment induced changes at the end of the experiments. The lack of labile organic carbon and the availability of mineral nutrients are key factors controlling bacterial activity and the role of the microbial food web in carbon sequestration.     

Comparison of biooxidation with carbon dioxide assimilation during bacterial growth on ferrous ion or elemental sulfur.

Biomass and oxygen uptake activity profiles of a mixed bioleaching culture were studied and compared at various temperatures. Bacteria were grown on ferrous ion or elemental sulfur in a Micro-Oxymax respirometer apparatus that allowed measurement of both oxygen consumption and carbon dioxide assimilation. Balanced growth was observed between 10 degrees C and 35 degrees C, with an optimum at 30 degrees C, on both energy sources. No significant growth was observed at the lowest temperature used, 5 degrees C, or at the highest temperature used, 40 degrees C. The oxygen to carbon dioxide molar yield was 50:1 when growing on ferrous ion but only 17:1 when growing on elemental sulfur. Upon transfer from a sulfide ore to a new energy source, greater numbers in the inoculum reduced the duration of the lag phase. Lag phase duration was also reduced by proximity to the optimum growth temperature. A longer lag phase decreased the achievable growth rate of the cells exponentially, significantly affecting biooxidation activity.     

Validation of respirometry as a short-term method to assess the efficacy of biocides

This study shows that a short-term respirometric measurement based on the rate of oxygen uptake needed to oxidize glucose is a reliable and fast method to assess biocide efficacy against P. fluorescens cells. Respiratory activity using oxygen consumption rate, the determination of viable and nonviable cells using Live/Dead BacLight kit and colony formation units (CFU), were compared as indicators of the biocidal efficacy of ortho-phthalaldehyde (OPA). The results showed that determining the effect of OPA against P. fluorescens using the different methods leads to different conclusions. The minimum bactericidal concentration (MBC) was 80 mgl(-1), 100 mgl(-1) and 65 mgl(-1) respectively, using respiratory activity, viability using BacLight counts and culturability. The plate count method was shown to underestimate the biocidal action of OPA, whilst data from respirometry and viability using Live/Dead BacLight kit correlated strongly and were not statistically different when yellow cells were considered nonviable. Respirometry therefore represents an expeditious, non-destructive and accurate method to determine the antimicrobial action of biocides against aerobic heterotrophic bacteria.     

Bacterial sulfur reduction in hot vents

Abstract: Elemental sulfur can be reduced through different microbial processes, including catabolically significant sulfur respiration and reduction of sulfur in the course of fermentation. Both of these processes are found in thermophilic microorganisms inhabiting continental and submarine hot vents, where elemental sulfur is one of the most common sulfur species. Among extreme thermophiles, respresented mainly by Archaea, sulfur-respiring bacteria include hydrogen-utilizing lithoautotrophs and heterotrophs, oxidizing complex organic substrates. Some marine heterotrophic sulfur-reducing Archaea were found to ferment peptides and polysaccharides, using elemental sulfur as an electron sink and thus avoiding the formation of molecular hydrogen which is highly inhibiting. Moderately thermophilic communities contain eubacterial sulfur reducers capable of lithotrophic and heterotrophic growth. Total mineralization of organic matter is carried out by a complex microbial system consisting of fermentative heterotrophs, which use elemental sulfur as an electron sink, and sulfur-respiring bacteria of the genus Desulfurella , which oxidize other fermentation products, yielding only CO_f2 and H_f2S. The most remarkable thermophilic microbial community is the thermophilic cyanobacterial mat found in the Uzon caldera, Kamchatka, which contains elemental sulfur among the layers. Organic matter produced by the thermophilic Oscillatoria is completely and rapidly mineralized by means of sulfur reduction.     

Interrelationships between <Emphasis Type="Italic">Dunaliella</Emphasis> and halophilic prokaryotes in saltern crystallizer ponds

Thanks to their often very high population densities and their simple community structure, saltern crystallizer ponds form ideal sites to study the behavior of halophilic microorganisms in their natural environment at saturating salt concentrations. The microbial community is dominated by square red halophilic Archaea, recently isolated and described as Haloquadratum walsbyi, extremely halophilic red rod-shaped Bacteria of the genus Salinibacter, and the unicellular green alga Dunaliella as the primary producer. We review here, the information available on the microbial community structure of the saltern crystallizer brines and the interrelationships between the main components of their biota. As Dunaliella produces massive amounts of glycerol to provide osmotic stabilization, glycerol is often postulated to be the most important source of organic carbon for the heterotrophic prokaryotes in hypersaline ecosystems. We assess here, the current evidence for the possible importance of glycerol and other carbon sources in the nutrition of the Archaea and the Bacteria, the relative contribution of halophilic Bacteria and Archaea to the heterotrophic activity in the brines, and other factors that determine the nature of the microbial communities that thrive in the salt-saturated brines of saltern crystallizer ponds. Three-letter abbreviations for names of genera of Halobacteriaceae conform the recommendations of the ICSP Subcommittee on the Taxonomy of Halobacteriaceae.     

Controlled gas environments in industrial fermentations

The gas environment is of major importance in controlling aerobic fermentation processes for the manufacture of microbial products. Oxygen and carbon dioxide levels in gas-liquid equilibria affect productivity and energy consumption in such processes and appear to be implicated in the regulation of microbial metabolism. Gas-liquid transfer has been intensively studied by many investigators for Newtonian and non-Newtonian fluids, primarily in terms of oxygen-limitation in biomass and product formation. More recentreports show that microbial growth and product formation are affected by levels of oxygen and carbon dioxide in the gas environment, suggesting that microbial metabolism may be directed towards specific products by the control of such environments. High product concentrations may also be obtained by solid substrate fermentations with mycelial organisms cultured on semi-solid agricultural products at low moisture contents. Such methods are commonly used in the Orient for the manufacture of enzymes and traditional fermented foods and could probably be extended to other microbial products. This review covers fundamental aspects of engineering research in microbial processes that suggest applications for controlled gas environments in submerged culture and solid substrate fermentations of potential industrial interest.     

Effect of chemical water pollution on the physiological activity of microorganisms

Anionactive detergents occurring in water in concentrations up to 100 micrograms . ml-1 do not exert inhibitory effects on the present microflora, either in case of heterotrophic bacteria, indicators of fecal pollution, or pathogenic and facultative pathogenic bacteria, including viruses. On the contrary, the studied bacterial species are able to decompose surfactants and utilize them as carbon source. Though, the rate of degradation is different with respect to respective substances, as well as with regard to the species of organisms, and their physiological activity. Only at higher detergent concentrations inhibition of microbial cell growth may be observed.     

Effects of key parameters in recycling of metals from petroleum refinery waste catalysts in bioleaching process

Environmental laws concerning spent catalysts disposal have become increasingly more severe in recent years. Due to the toxic nature of spent catalysts, their disposal can pollute the environment. The recovery of heavy metals decreases the environmental impact of the waste catalysts and the recycled product can be further used for industrial purposes. Bio-hydrometallurgical approaches, such as bioleaching, appear to offer good prospects for recovering valuable metals from spent refinery catalysts. Currently, identifying and modifying the parameters that influenced the efficiency of bioleaching is important for industrial sector. The biological system can be further improved through optimizing the bioleaching parameters, such as the nutrient culture media, amount of oxygen and carbon dioxide, pH, temperature, inoculum, metal resistance of microorganisms, chemistry of solid waste, particle size of solid waste, solid liquid ratio, bioleaching period, size of substrate, shaking speed, and also the development of more effective bioleaching microorganisms. In our previous review (Asghari et al. in J Ind Eng Chem 19:1069–1081, 2013), information available in the literature on the bioleaching fundamentals of spent catalysts with a focus on recent developments was reviewed in detail. In this study, the effects of most important factors that influence an efficient bioleaching process of spent refinery catalysts with the hope that these valuable and useful data can help determine the most efficient process will be discussed. The details of metals recovery with a focus on the effects of different variables in the bioleaching such as reaction time, pulp density, initial pH, particle size, nutrient concentration, temperature and buffer will also be presented.     

Mineralization in the Ems-Dollard estuary effects on carbon and oxygen budgets

Summary The Ems estuary is a tidal system, showing typical estuaries characteristics, such as gradients of salinity and suspended matter in the water phase. The tidal amplitude is 2.5–3 m. Tidal flats cover 40% of the total area in the outer part and 75% in the innermost part of the estuary, the Dollard. In the sediment aerobic heterotrophic bacteria are concentrated in the upper 2 cm, the numbers rapidly decreasing with depth.Oxygen production and consumption rates in the sediment, and oxygen consumption in the water are measured, together with environmental parameters and numbers of aerobic heterotrophic bacteria. Using a conversion factor of 12/32, oxygen measurements are translated to organic carbon. Input of organic carbon from external sources (particulate carbon from the river Ems and the North Sea, and organic waste discharge) is calculated for the Dollard. An organic carbon budget for the Dollard, using these calculated figures from one year's measurements, showed that most of the organic carbon entering the systems, originated from external sources.Mineralization was quantitatively more important in the sediment than it was in the water phase. The calculated import and production of organic carbon in the system was larger than the calculated consumption. An explanation for this difference is probably the export of soluble organic carbon, which was not measured, to the adjoining Waddensea, which must be considerable. It was concluded that heterotrophic bacteria must play an important role in carbon fluxes in the Dollard and that studies of growth yield under in situ conditions are necessary for a better understanding of their role in the ecosystem.     

A study on biological activity measurements and heterotrophic bacteria in a small freshwater lake

A 6-m-deep lake has been sampled to measure the temporal and depth-wise distribution of heterotrophic bacteria and biological activity in the water. Surface, mid-depth and bottom waters were analysed at monthly intervals for a period of one year. The coefficient of heterotrophic activity, alkaline phosphatase activity and biological oxygen demand are used as an index of biological activity. The bacterial community was at maximum during spring, coinciding with high values of biological activity. Highest biological activity was observed in the bottom waters. Dissolved organic carbon showed a significant positive correlation with most of the biological activity parameters. This suggests that biological activity, as measured by the coefficient of heterotrophic activity, was more closely related to the concentration of substrates than to population density of heterotrophic bacteria.     

Spatial and seasonal distribution of heterotrophic bacteria in pond water and sediments under different management practices

Population densities and generation time of heterotrophic bacteria were estimated in water and sediments of fish ponds used for polyculture, monoculture and traditional systems of fish farming over a period of two and a half years. Spatial differences in microbial densities in the ponds were related to the organic loadings of the farm site and farm management. A number of environmental factors such as nitrogen, dissolved organic carbon, bicarbonate and phosphate of water were found to be responsible for the seasonal changes of heterotrophic bacteria in these ponds.