Phylogenetic organization in the assimilation of chemically distinct substrates by soil bacteria

Soils are among the most biodiverse habitats on earth and while the species composition of microbial communities can influence decomposition rates and pathways, the functional significance of many microbial species and phylogenetic groups remains unknown. If bacteria exhibit phylogenetic organization in their function, this could enable ecologically meaningful classification of bacterial clades. Here, we show non-random phylogenetic organization in the rates of relative carbon assimilation for both rapidly mineralized substrates (amino acids and glucose) assimilated by many microbial taxa and slowly mineralized substrates (lipids and cellulose) assimilated by relatively few microbial taxa. When mapped onto bacterial phylogeny using ancestral character estimation this phylogenetic organization enabled the identification of clades involved in the decomposition of specific soil organic matter substrates. Phylogenetic organization in substrate assimilation could provide a basis for predicting the functional attributes of uncharacterized microbial taxa and understanding the significance of microbial community composition for soil organic matter decomposition.     

Dynamics of amino acids in the conditioning film developed on glass panels immersed in the surface seawaters of Dona Paula Bay

The conditioning film developed on glass panels immersed in surface seawater over a period of 24 h was analysed for total organic carbon (OC), total organic nitrogen (ON), and total hydrolyzable amino acid (THAA) concentrations and composition. The concentrations of C and N and THAA increased, whereas the C/N ratio decreased over the period of immersion. The amino acid-C and N accounted for 3.7-6.7% and 10.3-65.3% of OC and ON, respectively. The relative contribution of glycine plus threonine and serine to the total amino acids decreased while that of valine, phenylalanine, isoleucine and leucine increased over the period of immersion. Principal component analysis (PCA) based on mole% amino acid composition showed that the degradation indices (DI) for the conditioning film organic matter increased over the period of immersion. A high C/N ratio, a low %THAA-C, % THAA-N and DI values and the abundance of glycine plus threonine and serine in the conditioning film organic matter during the first few hours following immersion imply that the adsorbed organic matter was mostly derived from degraded organic matter.     

Dynamics of amino acids in the conditioning film developed on glass panels immersed in the surface seawaters of Dona Paula Bay

The conditioning film developed on glass panels immersed in surface seawater over a period of 24?h was analysed for total organic carbon (OC), total organic nitrogen (ON), and total hydrolyzable amino acid (THAA) concentrations and composition. The concentrations of C and N and THAA increased, whereas the C/N ratio decreased over the period of immersion. The amino acid-C and N accounted for 3.7?–?6.7% and 10.3?–?65.3% of OC and ON, respectively. The relative contribution of glycine plus threonine and serine to the total amino acids decreased while that of valine, phenylalanine, isoleucine and leucine increased over the period of immersion. Principal component analysis (PCA) based on mole% amino acid composition showed that the degradation indices (DI) for the conditioning film organic matter increased over the period of immersion. A high C/N ratio, a low %THAA-C, % THAA-N and DI values and the abundance of glycine plus threonine and serine in the conditioning film organic matter during the first few hours following immersion imply that the adsorbed organic matter was mostly derived from degraded organic matter.     

Changes in intracellular amino acids and organic acids induced by nitrogen starvation and nitrate or ammonium resupply in the cyanobacterium Phormidium laminosum

In Phormidium laminosum cells, nitrogen starvation caused a decrease in the intracellular levels of all amino acids, except glutamate, and an increase in the total level of the analyzed organic acids. The addition of nitrate or ammonium to N-starved cells resulted in substantial increases in the pool size of most amino acids. Upon addition of ammonium the total level of organic acids diminished, whereas it increased upon addition of nitrate, after a transient decay during the first minutes. Nitrogen resupply stimulated amino acid synthesis, the effect being faster and higher when ammonium was assimilated. The data indicate that nitrate and ammonium assimilation induced an enhancement of carbon flow through the glycolytic and the tricarboxylic-acid pathways to amino acid biosynthesis, with a concurrent decrease in the carbohydrate reserves. The results suggest that the availability of carbon skeletons limited the rate of ammonium assimilation, whereas the availability of reducing equivalents limited the rate of nitrate assimilation.Abbreviations Chl chlorophyll - GOGAT ferredoxin-dependent glutamate synthase (EC 1.4.7.1) - GS glutamine synthetase (EC 6.3.1.2) This work has been supported by grants from the Spanish Ministry of Education and Science (DGICYT and PB92-0464) and the University of the Basque Country (042.310-EC203/94) M.I.T. and J.A.G. were the recipients of fellowships from the Basque Government.     

土壤有机质和外源有机物对甲烷产生的影响

对土壤有机质含量及组分、外源有机物和根系分泌对甲烷产生的影响作了综述。土壤产甲烷量和甲烷排放量随有机质含量增加而提高,与土壤中易矿化有机碳或沸水浸提有机碳含量呈显著相关。外源有机碳加入促进了土壤排放甲,刺激效果与外源有机碳的用量和组成有关。还原力强的有机物如纤维素和半纤维素较还原力弱的有机物如类脂和多糖能够产生更多的甲烷。甲醇、甲基化氨基酸等无其它微生物竞争利用的有机物能被产甲烷菌更多地转化成甲烷。植物根系分泌物也促进甲烷的产生,促进作用大小与植物种类及分泌物的数量和质量有关。外源有机物通过3种方式促进土壤甲烷产生;提高土壤的甲烷底物供应量,降低土壤氧化还原电位,刺激土壤原有有机碳的转化。     

Formation and distribution of free amino acids in Cunninghamella elegans mycelium during growth on media with glucose and dodecane]

The growth, assimilation of organic nitrogen compounds, and production and distribution of free amino acids were studied during the growth of Cunninghamella elegans (--) 1204 on the defined medium 12 containing glucose and dodecane and without an additional carbon source (control). The culture utilized all leucine, glycine, and asparagine after three days of the growth, irrespective of the source of carbon in the medium. Glutamic acid was assimilated only in the presence of glucose. Some 15 free amino acids and 2 amides were detected in the cells, and also some correlations in the composition of the pool in the presence of different sources of carbon in the medium. The substitution of dodecane for glucose decelerates metabolism of free intracellular amino acids and stimulates their accumulation in the cells, especially if there is a deficiency of carbon sources: the overall content of amino acids and amides on the medium with glucose is from 2.2 to 4.1 mg/g; with dodecane, from 9.8 to 16.5 mg/g; in the control, from 16.4 to 25.8 mg/g.     

Comparison of Assimilatory Organic Nitrogen, Sulfur, and Carbon Sources for Growth of Methanobacterium Species

Experiments document the ability of two species of autotrophic methanogens to assimilate and utilize organic substrates as the nutrient sulfur or nitrogen source and as a carbon source during growth on H₂-CO₂. Methanobacterium thermoautotrophicum strain ΔH and the mesophilic species Methanobacterium sp. strain Ivanov grew with glutamine as the nitrogen source or cysteine as the sulfur source. M. thermoautotrophicum also utilized urea as the nitrogen source and as a carbon precursor for methane and cell synthesis. Methanobacterium sp. strain Ivanov grew with methionine as the sulfur source. The growth rate of two different Methanobacterium species was lower on an organic N or S source than on ammonium or sulfide. ³⁵S and ¹⁴C tracer studies demonstrated that amino acid or urea assimilation correlated with time and amount of growth. The rate of [³⁵S]cysteine incorporation was similar in strain ΔH (34 nmol h⁻¹ mg of cells⁻¹) and strain Ivanov (23 nmol h⁻¹ mg of cells⁻¹). However, the rate of [¹⁴C]acetate incorporation was dramatically different (17 versus 208 nmol h⁻¹ mg of cells⁻¹ in strains ΔH and Ivanov, respectively). [¹⁴C]acetate accounted for 1.3 and 21.2% of the total cell carbon synthesized by strains ΔH and Ivanov, respectively. Amino acids and urea were mainly assimilated into the cell protein fraction, but accounted for less than 2.0% of the total cell carbon synthesized. The data suggest that a biochemical-genetic approach to understanding cell carbon synthesis in methanogens is feasible; mutants that are auxotrophic for either acetate, glutamine, cysteine, or methionine are suggested as future targets for genetic studies.     

Markers and signals associated with nitrogen assimilation in higher plants

A key concept underpinning current understanding of the carbon/nitrogen (C/N) interaction in plants is that the capacity for N assimilation is aligned to nutrient availability and requirements by the integrated perception of signals from hormones, nitrate, sugars, organic acids, and amino acids. Studies on the nature and integration of these signals over the last ten years has revealed a complex network of controls brokered by an interplay of C and N signals. These controls not only act to orchestrate the relative rates of C and N assimilation and carbohydrate and amino acid production, but they also have a significant influence on plant development. Amino acids are the hub around which the processes of N assimilation, associated C metabolism, photorespiration, export of organic N from the leaf, and the synthesis of nitrogenous end-products revolve. Since specific major amino acids or their relative ratios are modulated differentially by photorespiration and N assimilation, even though these processes are tightly intermeshed, they are potentially powerful markers for metabolite profiling and metabolomics approaches to the study of plant biology. Moreover, while minor amino acids show marked diurnal rhythms, their contents fluctuate in a co-ordinated manner. It is probable that factors associated with early events and processes in C and N assimilation influence the relative composition of minor amino acids.     

Shift in carbon flow and stimulation of amino-acid turnover induced by nitrate and ammonium assimilation in Anacystis nidulans

The influence of nitrate and ammonium assimilation on the flow of recently fixed carbon has been determined in intact Anacystis nidulans cells actively fixing CO₂. Assimilation of nitrate or ammonium resulted in substantial increases in the incorporation of carbon into acid-soluble metabolites, the magnitude of the effect being dependent on the irradiance. The radiolabel in sugar phosphate was virtually unaffected by nitrogen assimilation, whereas that in organic acids and, in particular, in amino acids was markedly increased. Enhancement of carbon incorporation into amino acids induced by nitrogen assimilation was not accompanied by parallel increases in the size of the amino acid pools. This resulted in an appreciable increase of the specific radioactivity of most amino acids under conditions of nitrogen assimilation. The data indicate that nitrate and ammonium assimilation induce an enhancement of carbon flow through the glycolytic and the tricarboxylic-acid pathways to oxaloacetate and α-ketoglutarate, as well as a stimulation of amino-acid turnover. These effects were more pronounced at saturating irradiance. We thank the Dirección General de Investigación Científica y Técnica, Spain (research grant PB88-0019) and the Plan Andaluz de Investigación (grupo 3101) for financial support, and P. Pérez de León for excellent secretarial assistance.     

Does elevated atmospheric [CO2] alter diurnal C uptake and the balance of C and N metabolites in growing and fully expanded soybean leaves?

Increases in growth at elevated [CO2] may be constrained by a plant's ability to assimilate the nutrients needed for new tissue in sufficient quantity to match the increase in carbon fixation and/or the ability to transport those nutrients and carbon in sufficient quantity to growing organs and tissues. Analysis of metabolites provides an indication of shifts in carbon and nitrogen partitioning due to rising atmospheric [CO2] and can help identify where bottlenecks in carbon utilization occur. In this study, the carbon and nitrogen balance was investigated in growing and fully expanded soybean leaves exposed to elevated [CO2] in a free air CO2 enrichment experiment. Diurnal photosynthesis and diurnal profiles of carbon and nitrogen metabolites were measured during two different crop growth stages. Diurnal carbon gain was increased by c. 20% in elevated [CO2] in fully expanded leaves, which led to significant increases in leaf hexose, sucrose, and starch contents. However, there was no detectable difference in nitrogen-rich amino acids and ureides in mature leaves. By contrast to mature leaves, developing leaves had high concentrations of ureides and amino acids relative to low concentrations of carbohydrates. Developing leaves at elevated [CO2] had smaller pools of ureides compared with developing leaves at ambient [CO2], which suggests N assimilation in young leaves was improved by elevated [CO2]. This work shows that elevated [CO2] alters the balance of carbon and nitrogen pools in both mature and growing soybean leaves, which could have down-stream impacts on growth and productivity.     

Total and hydrolizable particulate organic matter (carbohydrates, proteins and lipids) at a coastal station in Terra Nova Bay (Ross Sea, Antarctica)

We analysed quantity and quality of particulate organic matter during the austral summer 1994/1995 at a coastal station in Terra Nova Bay (Ross Sea, Antarctica). Our main aims were to investigate the origin and biochemical composition of particulate organic matter (POM), to measure its availability for consumers through the study of its digestible fraction (measured by using different enzymes separately) and to highlight the role of hydrolizable compounds in the organic matter diagenesis in the coastal waters at Terra Nova Bay. Temporal and spatial patterns of chlorophyll-a concentrations were reflected by the particulate organic carbon, nitrogen and total biopolymeric carbon concentrations, suggesting that most POM originated directly from phytoplankton. The most evident feature of POM in the coastal waters at Terra Nova Bay was the dominance of proteins (on average 57% of total biopolymeric particulate carbon), followed by carbohydrates (25%) and lipids (18%). We found that about 30% of the refractory particulate organic carbon (assumed to be present only after the complete exploitation of particulate organic nitrogen) did not originate from biopolymeric carbon (as sum of carbohydrate, protein and lipid carbon). This allows us to suggest the use of the digestible fraction of particulate biopolymeric carbon as a more accurate measure of the food availability of POM for consumers. In Terra Nova Bay coastal waters, most of the particulate protein pool was associated with large phytoplankton cells or phytodetritus. As a result, the protein pool appeared less available (i.e. less digestible) than the one present in oligotrophic waters where, conversely, most particulate organic nitrogen is sequestered into bacteria. The relative low availability of the protein pool, together with the rapid sinking of POM and the low remineralization rates of benthic heterotrophic microbes, are suggested as possible factors in determining the “inefficiency” in organic matter recycling of coastal waters at Terra Nova Bay, which behaves as a “loss type” system. Received: 17 June 1997 / Accepted 25 September 1997     

Extracellular enzyme kinetics scale with resource availability

Microbial community metabolism relies on external digestion, mediated by extracellular enzymes that break down complex organic matter into molecules small enough for cells to assimilate. We analyzed the kinetics of 40 extracellular enzymes that mediate the degradation and assimilation of carbon, nitrogen and phosphorus by diverse aquatic and terrestrial microbial communities (1160 cases). Regression analyses were conducted by habitat (aquatic and terrestrial), enzyme class (hydrolases and oxidoreductases) and assay methodology (low affinity and high affinity substrates) to relate potential reaction rates to substrate availability. Across enzyme classes and habitats, the scaling relationships between apparent V_max and apparent K_m followed similar power laws with exponents of 0.44 to 0.67. These exponents, called elasticities, were not statistically distinct from a central value of 0.50, which occurs when the K_m of an enzyme equals substrate concentration, a condition optimal for maintenance of steady state. We also conducted an ecosystem scale analysis of ten extracellular hydrolase activities in relation to soil and sediment organic carbon (2,000–5,000 cases/enzyme) that yielded elasticities near 1.0 (0.9 ± 0.2, n = 36). At the metabolomic scale, the elasticity of extracellular enzymatic reactions is the proportionality constant that connects the C:N:P stoichiometries of organic matter and ecoenzymatic activities. At the ecosystem scale, the elasticity of extracellular enzymatic reactions shows that organic matter ultimately limits effective enzyme binding sites. Our findings suggest that one mechanism by which microbial communities maintain homeostasis is regulating extracellular enzyme expression to optimize the short-term responsiveness of substrate acquisition. The analyses also show that, like elemental stoichiometry, the fundamental attributes of enzymatic reactions can be extrapolated from biochemical to community and ecosystem scales.     

Substrate Use of Pseudovibrio sp. Growing in Ultra-Oligotrophic Seawater

Marine planktonic bacteria often live in habitats with extremely low concentrations of dissolved organic matter (DOM). To study the use of trace amounts of DOM by the facultatively oligotrophic Pseudovibrio sp. FO-BEG1, we investigated the composition of artificial and natural seawater before and after growth. We determined the concentrations of dissolved organic carbon (DOC), total dissolved nitrogen (TDN), free and hydrolysable amino acids, and the molecular composition of DOM by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI FT-ICR-MS). The DOC concentration of the artificial seawater we used for cultivation was 4.4 μmol C L^-1, which was eight times lower compared to the natural oligotrophic seawater we used for parallel experiments (36 μmol C L ^-1). During the three-week duration of the experiment, cell numbers increased from 40 cells mL^-1 to 2x10⁴ cells mL ^-1 in artificial and to 3x10⁵ cells mL ^-1 in natural seawater. No nitrogen fixation and minor CO₂ fixation (< 1% of cellular carbon) was observed. Our data show that in both media, amino acids were not the main substrate for growth. Instead, FT-ICR-MS analysis revealed usage of a variety of different dissolved organic molecules, belonging to a wide range of chemical compound groups, also containing nitrogen. The present study shows that marine heterotrophic bacteria are able to proliferate with even lower DOC concentrations than available in natural ultra-oligotrophic seawater, using unexpected organic compounds to fuel their energy, carbon and nitrogen requirements.     

Irrigation as a fuel pump to freshwater ecosystems

We generated a detailed time series of total dissolved hydrolyzable amino acids (DHAA) in a watershed dominated by irrigated agriculture in northern California, USA to investigate the roles of hydrologic and seasonal changes on the composition of dissolved organic matter (DOM). DHAA are sensitive indicators of the degradation state and reactivity of DOM. DHAA concentrations ranged from 0.55 to 9.96 μM (median 3.51 ± 1.80 μM), with expected peaks during high-discharge storms and unexpected high values throughout the low-discharge irrigation season. Overall, summer irrigation was a critical hydrologic regime for DOM cycling since it mobilized DOM similar in concentration and reactivity to DOM released during storms. Together, irrigation and storm flows exported DOM with (1) the largest DHAA contributions to the dissolved organic carbon and the dissolved organic nitrogen pools, (2) the largest proportion of basic amino acids, and (3) the lowest degradation extent based on multiple indices. In this highly disturbed terrestrial system, UV–vis absorbance did not correlate with DHAA concentrations, while classic interpretations of common amino acid indicators (e.g., proportion of basic amino acids, degradation index, percent of non-protein amino acids) were prone to conflicting characterizations of DOM reactivity. Therefore, a new parameter (processing ratio, PR) derived from individual amino acid concentrations was developed that demonstrated a strong potential for mechanistic-driven characterization of the extent of DOM diagenesis in freshwaters. Irrigated agriculture altered stream biogeochemistry by releasing a continuous supply of reactive DOM (lowest PR values), thereby providing an additional energy source to downstream ecosystems.     

Organic acid accumulation may inhibit N2 fixation in phosphorus-stressed lupin nodules.

Nodulated lupins (Lupinus angustifolius cv. Wonga) were hydroponically grown under conditions of low phosphate (LP) or adequate phosphate (HP) to assess the effect of phosphoenolpyruvate carboxylase (PEPC)-derived organic acids on nitrogen assimilation in LP nodules. LP conditions are linked to altered organic acid metabolism, by the engagement of PEP metabolism via PEPC. In LP nodules, the enhanced organic acid synthesis may reduce the available organic carbon for nitrogen assimilation. The diversion of carbon between the organic acid- and amino acid pools was assessed through key nodular enzymes and (14)CO(2) metabolism. Under LP conditions, increased rates of organic acid synthesis via PEPC and malate dehydrogenase (MDH), coincided with reduced nitrogen assimilation via aspartate aminotransferase (AAT), aspartate synthetase (AS) and glutamine synthetase (GS)/glutamate synthase (GOGAT) activities. There was a preferential metabolism of nodular (14)CO(2) into organic acids and particularly into malate. High malate levels were associated with reduced N(2) fixation and synthesis of amino acids. These results indicate that phosphorus deficiency can enhance malate synthesis in nodules, but that excessive malate accumulation may inhibit N(2) fixation and nitrogen assimilation.     

The Effects of Soil Bacterial Community Structure on Decomposition in a Tropical Rain Forest

Soil microorganisms are key drivers of terrestrial biogeochemical cycles, yet it is still unclear how variations in soil microbial community composition influence many ecosystem processes. We investigated how shifts in bacterial community composition and diversity resulting from differences in carbon (C) availability affect organic matter decomposition by conducting an in situ litter manipulation experiment in a tropical rain forest in Costa Rica. We used bar-coded pyrosequencing to characterize soil bacterial community composition in litter manipulation plots and performed a series of laboratory incubations to test the potential functional significance of community shifts on organic matter decomposition. Despite clear effects of the litter manipulation on soil bacterial community composition, the treatments had mixed effects on microbial community function. Distinct communities varied in their ability to decompose a wide range of C compounds, and functional differences were related to both the relative abundance of the two most abundant bacterial sub-phyla (Acidobacteria and Alphaproteobacteria) and to variations in bacterial alpha-diversity. However, distinct communities did not differ in their ability to decompose native dissolved organic matter (DOM) substrates that varied in quality and quantity. Our results show that although resource-driven shifts in soil bacterial community composition have the potential to influence decomposition of specific C substrates, those differences may not translate to differences in DOM decomposition rates in situ. Taken together, our results suggest that soil bacterial communities may be either functionally dissimilar or equivalent during decomposition depending on the nature of the organic matter being decomposed.     

Nitrogen metabolism in plant cell suspension cultures: I. Effect of amino acids on growth

Certain amino acids inhibit growth of tobacco (Nicotiana tabacum L. var. xanthi), tomato (Lycopersicon esculentum) carrot (Daucus carota), and soybean (Glycerine max L. co. Mandarin) cell cultures when nitrate or urea are the nitrogen sources but not when ammonia is the nitrogen source. These amino acids also inhibit development of nitrate reductase activity (NADH:nitrate oxidoreductase EC 1.6.6.1) in tobacco and tomato cultures. Threonine, the most inhibitory amino acid, also inhibits nitrate uptake in tobacco cells. Arginine, and some other amino acids, abolish the inhibition effects caused by other amino acids. We suggest that amino acids inhibit assimilation of intracellular ammonium into amino acids in cells grown on nitrate or urea.     

Distribution of particulate amino acids in the Bransfield Strait

Summary The depth distribution and composition of residual amino acids was determined in December 1980 in the Bransfield Strait. The major components were aspartic and glutamic acids together with alanine, glycine and serine. Non-protein amino acids were not found in significant amounts. On the average, the amino acids accounted for 25% of the total organic carbon and 50% of the organic nitrogen. Amino acid nitrogen/chlorophyll ratios indicated that in most euphotic zone samples phytoplankton in good physiological condition was present whereas heavily degraded material was found in deeper water layers. Statistical treatment and cluster analysis show distinct differences between euphotic and aphotic zone samples which could be used to characterize phytoplankton communities.