Stable carbon isotope analysis of nucleic acids to trace sources of dissolved substrates used by estuarine bacteria.

The natural abundance of stable carbon isotopes measured in bacterial nucleic acids extracted from estuarine bacterial concentrates was used to trace sources of organic matter for bacteria in aquatic environments. The stable carbon isotope ratios of Pseudomonas aeruginosa and nucleic acids extracted from cultures resembled those of the carbon source on which bacteria were grown. The carbon isotope discrimination between the substrate and total cell carbon from bacterial cultures averaged 2.3% +/- 0.6% (n = 13). Furthermore, the isotope discrimination between the substrate and nucleic acids extracted from bacterial cultures was 2.4% +/- 0.4% (n = 10), not significantly different from the discrimination between bacteria and the substrate. Estuarine water samples were prefiltered through 1-micron-pore-size cartridge filters. Bacterium-sized particles in the filtrates were concentrated with tangential-flow filtration and centrifugation, and nucleic acids were then extracted from these concentrates. Hybridization with 16S rRNA probes showed that approximately 90% of the nucleic acids extracted on two sample dates were of eubacterial origin. Bacteria and nucleic acids from incubation experiments using estuarine water samples enriched with dissolved organic matter from Spartina alterniflora and Cyclotella caspia had stable carbon isotope values similar to those of the substrate sources. In a survey that compared diverse estuarine environments, stable carbon isotopes of bacteria grown in incubation experiments ranged from -31.9 to -20.5%. The range in isotope values of nucleic acids extracted from indigenous bacteria from the same waters was similar, -27.9 to -20.2%. Generally, the lack of isotope discrimination between bacteria and nucleic acids that was noted in the laboratory was observed in the field.(ABSTRACT TRUNCATED AT 250 WORDS)     

Composition of humic acid-degrading estuarine and marine bacterial communities

We examined the bacterial decomposition of humic acids (HA) in two flow-through culture experiments, one inoculated by marine and one by estuarine bacterial communities. In both experiments, the cultures were fed with HA media of salinities of 28 and 14, close to their ambient and a distinctly different, foreign salinity. HA were decomposed to >?60% of the initial concentration within 70?days, and the foreign salinity yielded the highest decomposition. A detrended correspondence analysis of denaturing gradient gel electrophoresis (DGGE) banding patterns showed that during incubation, the bacterial community composition underwent distinct changes. A phylogenetic analysis of DGGE bands excised and bacteria isolated at the end on HA as the sole carbon source showed that Alphaproteobacteria and Gammaproteobacteria largely dominated the communities in the marine flow-through cultures, whereas Gammaproteobacteria, Actinobacteria and Alphaproteobacteria dominated the estuarine communities. Eleven of 13 isolates obtained from both experiments were able to grow on HA as the sole carbon source, seven on phenol and three, affiliated to the Roseobacter clade, on various aromatic acids. The bacteria retrieved from the flow-through cultures were closely (96-99%) affiliated to organisms capable of degrading humic matter, aromatic and aliphatic compounds and also to other bacteria reported previously from the Wadden Sea and Weser estuary.     

Consequences of protist-stimulated bacterial production for estimating protist growth efficiencies

The trophic link between bacteria and bacterivorous protists is a complex interaction that involves feedback of inorganic nutrients and growth substrates that are immeadiately available for prey growth. These interactions were examined in the laboratory and in incubations of concentrated natural assemblages of bacterioplankton. Growth dynamics of estuarine and marine bacterivorous protists were determined in laboratory culture using Vibrio natriegens as prey and were compared to growth of protists on bacterioplankton assemblages concentrated by tangential flow filtration from four northwest Florida Estuaries. Biomass transfers from bacteria to protists were monitored by tracing elemental carbon and nitrogen in particulate fractions of protist added and grazer free controls. Gross growth efficiencies of the protists on naturally occurring bacteria were within the range determined in lab estimates of growth efficiency on cultured bacteria (50%). However, bacterial response to protist excretion products was different in the lab and field incubations, and bacterial growth contributed to the biomass available to protists in the field incubations. As determined by radioisotope-labeled substrate incorporation, a time lag in bacterial reponse to protist excretion products was observed for laboratory batch cultures, allowing accurate estimation of growth efficiency. In incubations with concentrated natural bacterial assemblages, bacterial growth response coincided with protist growth and excretion. The additional bacterial production on protist excretion products reached a maximum of 2–3-fold higher than protist-free controls. In addition, ammonium concentrations increased with protist grazing and growth in lab cultures, but ammonium excreted by protists in concentrates did not accumulate. The C:N values for the bacterial concentrates suggests that these bacteria were nitrogen limited. It is speculated that dissolved organic carbon, concentrated by tangential flow filtration (> 100,000 MW membrane) with the bacterioplankton, was utilized by bacteria when nitrogen was supplied as ammonium and amino acids from protist excretion. Thus, estimates of protist growth efficiency on naturally occurring bacterioplankton, corrected for protist-stimulated bacterial production, were in the range of 13–21%.     

Utilization of Dissolved Nitrogen by Heterotrophic Bacterioplankton: a Comparison of Three Ecosystems

The contributions of different organic and inorganic nitrogen and organic carbon sources to heterotrophic bacterioplankton in batch cultures of oceanic, estuarine, and eutrophic riverine environments were compared. The importance of the studied compounds was surprisingly similar among the three ecosystems. Dissolved combined amino acids (DCAA) were most significant, sustaining from 10 to 45% of the bacterial carbon demands and from 42 to 112% of the bacterial nitrogen demands. Dissolved free amino acids (DFAA) supplied 2 to 7% of the carbon and 6 to 24% of the nitrogen incorporated into the bacterial biomass, while dissolved DNA (D-DNA) sustained less than 5 and 12% of the carbon and nitrogen requirements, respectively. Ammonium was the second most important source of nitrogen, meeting from 13 to 45% of the bacterial demand in the oceanic and estuarine cultures and up to 270% of the demand in riverine cultures. Nitrate was taken up in the oceanic cultures (uptake equaled up to 46% of the nitrogen demand) but was released in the two others. Assimilation of DCAA, DFAA, and D-DNA combined supplied 43% of the carbon demand of the bacteria in the oceanic cultures, while approximately 25% of the carbon requirements were met by the three substrates at the two other sites. Assimilation of nitrogen from DCAA, DFAA, D-DNA, NH₄⁺, and NO₃^-, on the other hand, exceeded production of particulate organic nitrogen in one culture at 27 h and in all cultures over the entire incubation period (50 h). These results suggest that the studied nutrient sources may fully support the nitrogen needs but only partially support the carbon needs of microbial communities of geographically different ecosystems. Furthermore, a comparison of the initial concentrations of the different substrates indicated that relative pool sizes of the substrates seemed to influence which substrates were primarily being utilized by the bacteria.     

Compositions isotopiques naturelles des bactéries hétérotrophes et détermination de l'origine du carbone organique dissous biodisponible

Several studies of salt marsh systems have attempted to quantify the flow of organic matter between the land and coastal waters. However, the techniques used could not identify sources of dissolved organic carbon (DOC) rapidly assimilated by heterothrophic bacteria. Recently, the assay of carbon isotope ratios has allowed characterization of the different sources of organic matter in salt marshes. In this study, we wanted to find out if the natural isotopic composition assayed in heterotrophic bacteria distinguished the origin of bioavailable DOC. We determined the δ¹³C values for 1) three bacterial strains and their nucleic acids cultured on glucose and tryptose substrates, respectively, and 2) naturally occurring bacteria recovered from seawater in which salt marsh vegetation had been immersed. First, we showed that the isotopic fractionation was the same for the three bacterial strains cultured on the same synthetic substrate, but could vary depending on the nature of DOC. There was no significant difference between the δ¹³ C values of bacteria and their nucleic acids. Second, natural bacteria were grown in a medium enriched in DOC from halophytic plants. The δ¹³C values of this community were close to those of dissolved organic carbon from plant leachates. The comparison between the isotopic ratios of natural bacteria in Vibrio alginolyticus showed that the heterogeneity of the bacterial community averaged the isotopic fractionation from the preferential assimilation of organic compounds in the medium by each bacterial strain. The δ¹³ C values recorded for the bacterial community in the field and their nucleic acids made it possible to identify the source of organic matter readily accessible to microorganisms in a coastal ecosystem.     

Utilization of Dissolved Nitrogen by Heterotrophic Bacterioplankton: Effect of Substrate C/N Ratio

The significance of dissolved combined amino acids (DCAA), dissolved free amino acids (DFAA), and dissolved DNA (D-DNA) as sources of C and N for marine bacteria in batch cultures with variable substrate C/N ratios was studied. Glucose, ammonium, alanine, and phosphate were added to the cultures to produce C/N ratios of 5, 10, and 15 and to ensure that phosphorus was not limiting. Maximum bacterial particulate organic carbon production (after 25 h of incubation) was inversely correlated with the C/N ratio: with the addition of identical amounts of carbon, the levels of production were 9.0-, 10.0-, and 11.1-fold higher at C/N ratios of 15, 10, and 5, respectively, relative to an unamended control. The bacterial growth efficiency increased from 22% (control cultures) to 44 to 53% in the cultures with manipulated C/N ratios (C/N-manipulated cultures). Net carbon incorporation from DCAA, DFAA, and D-DNA supported on average 19, 4, and 3% (control cultures and cultures to which only phosphate was added [+P cultures]) and 5, 4, and 0.3% of the particulate organic carbon production (C/N-manipulated cultures), respectively. In the C/N-manipulated cultures, a 2.6- to 3.4-fold-higher level of incorporation of DCAA, relative to that in the control cultures, occurred. Incorporation of D-DNA increased with the substrate C/N ratio, suggesting that D-DNA mainly was a source of N to the bacteria. Organic N (DCAA, DFAA, and D-DNA) sustained 14 to 49% of the net bacterial N production. NH₄⁺ was the dominant N source and constituted 55 to 99% of the total N uptake. NO₃^- contributed up to 23% to the total N uptake but was released in two cultures. The studied N compounds sustained nearly all of the bacterial N demand. Our results show that the C/N ratio of dissolved organic matter available to bacteria has a significant influence on the incorporation of individual compounds like DCAA and D-DNA.     

Effects of elemental composition on the incorporation of dietary nitrogen and carbon isotopic signatures in an omnivorous songbird

The use of stable isotopes to infer diet requires quantifying the relationship between diet and tissues and, in particular, knowing of how quickly isotopes turnover in different tissues and how isotopic concentrations of different food components change (discriminate) when incorporated into consumer tissues. We used feeding trials with wild-caught yellow-rumped warblers (Dendroica coronata) to determine delta15N and delta13C turnover rates for blood, delta15N and delta13C diet-tissue discrimination factors, and diet-tissue relationships for blood and feathers. After 3 weeks on a common diet, 36 warblers were assigned to one of four diets differing in the relative proportion of fruit and insects. Plasma half-life estimates ranged from 0.4 to 0.7 days for delta13C and from 0.5 to 1.7 days for delta15N . Half-life did not differ among diets. Whole blood half-life for delta13C ranged from 3.9 to 6.1 days. Yellow-rumped warbler tissues were enriched relative to diet by 1.7-3.6% for nitrogen isotopes and by -1.2 to 4.3% for carbon isotopes, depending on tissue and diet. Consistent with previous studies, feathers were the most enriched and whole blood and plasma were the least enriched or, in the case of carbon, slightly depleted relative to diet. In general, tissues were more enriched relative to diet for birds on diets with high percentages of insects. For all tissues, carbon and nitrogen isotope discrimination factors increased with carbon and nitrogen concentrations of diets. The isotopic signature of plasma increased linearly with the sum of the isotopic signature of the diet and the discrimination factor. Because the isotopic signature of tissues depends on both elemental concentration and isotopic signature of the diet, attempts to reconstruct diet from stable isotope signatures require use of mixing models that incorporate elemental concentration.     

Carbon Isotopic Fractionation Does Not Occur during Dark Respiration in C3 and C4 Plants

The magnitude of possible carbon isotopic fractionation during dark respiration was investigated with isolated mesophyll cells from mature leaves of common bean (Phaseolus vulgaris L.), a C3 plant, and corn (Zea mays L.), a C4 plant. Mesophyll protoplasts were extracted from greenhouse-grown leaves and incubated in culture solutions containing different carbohydrate substrates (fructose, glucose, and sucrose) with known [delta]13C values. The CO2 produced by protoplasts after incubation in the dark was collected, purified, and analyzed for its carbon isotope ratio. From observations of the isotope ratios of the substrate and respired CO2, we calculated the carbon isotope discrimination associated with metabolism of each of these substrates. In eight of the 10 treatment combinations, the carbon isotope ratio discrimination was not significantly different from 0. In the remaining two treatment combinations, the carbon isotope ratio discrimination was 1[per mille (thousand) sign]. From these results, we conclude that there is no significant carbon isotopic discrimination during mitochondrial dark respiration when fructase, glucose, or sucrose are used as respiratory substrates.     

Microbial Utilization of Estuarine Dissolved Organic Carbon: a Stable Isotope Tracer Approach Tested by Mass Balance

The natural stable isotope values of different plants have been used to trace the fate of organic carbon that enters estuarine ecosystems. Experiments were designed to determine the magnitude of (delta) (sup13)C changes of dissolved organic carbon (DOC) derived from tidal marsh vegetation that occurred during bacterial decomposition. Bacteria were grown on DOC leached from estuarine Spartina alterniflora and Typhus angustifolia plants. In four experiments, 25 to 80% of the initial carbon (2.6 to 9.1 mM organic C) was converted to bacterial biomass and CO(inf2). Mass balance calculations showed good recovery of total C and (sup13)C at the end of these experiments (100% (plusmn) 14% total C; (plusmn) 1(permil) (delta) (sup13)C). The (delta) (sup13)C values of DOC, bacterial biomass, and respired CO(inf2) changed only slightly in the four experiments by average values of -0.6, +1.4, and +0.5(permil), respectively. These changes are small relative to the range of (delta) (sup13)C values represented by different organic carbon sources to estuaries. Thus, microbial (delta) (sup13)C values determined in the field helped to identify the source of the carbon assimilated by bacteria.     

Incorporation of [h]leucine and [h]valine into protein of freshwater bacteria: uptake kinetics and intracellular isotope dilution

Incorporation of [H]leucine and [H]valine into proteins of freshwater bacteria was studied in two eutrophic lakes. Incorporation of both amino acids had a saturation level of about 50 nM external concentration. Only a fraction of the two amino acids taken up was used in protein synthesis. At 100 nM, the bacteria respired 91 and 78% of leucine and valine taken up, respectively. Respiration of H and C isotopes of leucine gave similar results. Most of the nonrespired leucine was recovered in bacterial proteins, while only up to one-half of the nonrespired valine occurred in proteins. In intracellular pools of the bacteria, [H]leucine reached an isotope saturation of 88 to 100% at concentrations of >40 nM. For [H]valine, an isotope equilibrium of about 90% was obtained at concentrations of >80 nM. Within an incubation period of typically 1 h, tritiated leucine and valine incorporated into proteins of the bacteria reached an isotope saturation of 2 to 6%. In a 99-h batch experiment, bacterial protein synthesis calculated from incorporation of leucine and valine corresponded to 31 and 51% (10 nM) and 89 and 97% (100 nM), respectively, of the chemically determined protein production. Measured conversion factors of 100 nM leucine and valine were 6.4 x 10 and 6.6 x 10 cells per mol, respectively, and fell within the expected theoretical values. The present study demonstrates that incorporation of both valine and leucine produces realistic measurements of protein synthesis in freshwater bacteria and that the incorporation can be used as a measure of bacterial production.     

稳定性同位素探测技术在微生物生态学研究中的应用

稳定性同位素标记技术同分子生物学技术相结合而发展起来的稳定性同位素探测技术（stableisotope probing,SIP）,在对各种环境中微生物群落组成进行遗传分类学鉴定的同时,可确定其在环境过程中的功能,提供复杂群落中微生物相互作用及其代谢功能的大量信息,具有广阔的应用前景.其基本原理是：将原位或微宇宙（microcosm）的环境样品暴露于稳定性同位素富集的基质中,这些样品中存在的某些微生物能够以基质中的稳定（性同位素为碳源或氮源进行物质代谢并满足其自身生长需要,基质中的稳定性同位素被吸收同化进入微生物体内,参与各类物质如核酸（DNA和RNA）及磷脂脂肪酸（PLFA）等的生物合成,通过提取、分离、纯化、分析这些微生物体内稳定性同位素标记的生物标志物,从而将微生物的组成与其功能联系起来.在介绍稳定性同位素培养基质的选择及标记方法、合适的生物标志物的选择及提取分离方法的基础上,举例阐述了此项技术在甲基营养菌、有机污染物降解菌、根际微生物生态、互营微生物、宏基因组学等方面的应用.     

Distribution of carbon isotopes ((13)C/(12)C) in cells and temporal organization of cellular processes]

Recent studies on fractionation of carbon isotopes in biological systems are reviewed. It follows that direct experimental proofs have been obtained that 1) basic fractionation of carbon isotopes in the cell is related to isotope effect in pyruvate decarboxylation; 2) fractionation of carbon isotopes in the above reaction in vivo proceeds with exhausting substrate pool. The latter provides natural relationship between metabolites isotope distribution and sequence of their synthesis in the cell cycle, or with the temporal organization of cellular metabolism. The non-steady and periodic pattern of pyruvate decarboxylation due to the exhausting substrate pool well agrees with the existing notions on reciprocal oscillations in the cell glycolytic chain. Experimental data are presented corroborating indirectly the existence of oscillations in bacterial cells. Earlier proposed model of the mechanism of carbon isotope fractionation based on the above principles can be used for analysing changes in isotopic characteristics of the organisms and interpreting their relations with metabolic processes.     

Utilization of Organic Nitrogen Sources by Two Phytoplankton Species and a Bacterial Isolate in Pure and Mixed Cultures

Algal production of dissolved organic carbon and the regeneration of nutrients from dissolved organic carbon by bacteria are important aspects of nutrient cycling in the sea, especially when inorganic nitrogen is limiting. Dissolved free amino acids are a major carbon source for bacteria and can be used by phytoplankton as a nitrogen source. We examined the interactions between the phytoplankton species Emiliania huxleyi and Thalassiosira pseudonana and a bacterial isolate from the North Sea. The organisms were cultured with eight different amino acids and a protein as the only nitrogen sources, in pure and mixed cultures. Of the two algae, only E. huxleyi was able to grow on amino acids. The bacterium MD1 used all substrates supplied, except serine. During growth of MD1 in pure culture, ammonium accumulated in the medium. Contrary to the expectation, the percentage of ammonium regenerated from the amino acids taken up showed no correlation with the substrate C/N ratio. In mixed culture, the algae grew well in those cultures in which the bacteria grew well. The bacterial yields (cell number) were also higher in mixed culture than in pure culture. In the cultures of MD1 and T. pseudonana, the increase in bacterial yield (number of cells) over that of the pure culture was comparable to the bacterial yield in mixed culture on a mineral medium. This result suggests that T. pseudonana excreted a more-or-less-constant amount of carbon. The bacterial yields in mixed cultures with E. huxleyi showed a smaller and less consistent difference than those of the pure cultures of MD1. It is possible that the ability of E. huxleyi to use amino acids influenced the bacterial yield. The results suggest that interactions between algae and bacteria influence the regeneration of nitrogen from organic carbon and that this influence differs from one species to another.     

High-throughput isotopic analysis of RNA microarrays to quantify microbial resource use

Most microorganisms remain uncultivated, and typically their ecological roles must be inferred from diversity and genomic studies. To directly measure functional roles of uncultivated microbes, we developed Chip-stable isotope probing (SIP), a high-sensitivity, high-throughput SIP method performed on a phylogenetic microarray (chip). This approach consists of microbial community incubations with isotopically labeled substrates, hybridization of the extracted community rRNA to a microarray and measurement of isotope incorporation—and therefore substrate use—by secondary ion mass spectrometer imaging (NanoSIMS). Laboratory experiments demonstrated that Chip-SIP can detect isotopic enrichment of 0.5 atom % ¹³C and 0.1 atom % ¹⁵N, thus permitting experiments with short incubation times and low substrate concentrations. We applied Chip-SIP analysis to a natural estuarine community and quantified amino acid, nucleic acid or fatty acid incorporation by 81 distinct microbial taxa, thus demonstrating that resource partitioning occurs with relatively simple organic substrates. The Chip-SIP approach expands the repertoire of stable isotope-enabled methods available to microbial ecologists and provides a means to test genomics-generated hypotheses about biogeochemical function in any natural environment.     

不同产地三疣梭子蟹的稳定同位素比值特征及原产地溯源

探索稳定同位素对三疣梭子蟹产地溯源的可行性,可为保护地理标志产品、追溯原产地来源提供理论依据。本研究以黄海、渤海和东海3个主产区海域的三疣梭子蟹为对象,分析了其碳、氮稳定同位素值差异性和不同组织中稳定同位素比值的相关性。结果表明: 不同产地的碳、氮稳定同位素比值有显著性差异,不同组织之间有明显的同位素分馏效应。利用建立的判别分析模型,通过三疣梭子蟹不同组织稳定同位素比值进行产地判别分析,采用肌肉和腮中碳、氮稳定同位素的产地判别正确率(>95%)明显高于肝胰性腺,说明肌肉和腮稳定同位素比值可以对不同海域的三疣梭子蟹进行有效区分。本研究填补了目前三疣梭子蟹稳定同位素溯源技术的研究空白。     

Stable carbon isotope fractionation by sulfate-reducing bacteria

Biogeochemical transformations occurring in the anoxic zones of stratified sedimentary microbial communities can profoundly influence the isotopic and organic signatures preserved in the fossil record. Accordingly, we have determined carbon isotope discrimination that is associated with both heterotrophic and lithotrophic growth of pure cultures of sulfate-reducing bacteria (SRB). For heterotrophic-growth experiments, substrate consumption was monitored to completion. Sealed vessels containing SRB cultures were harvested at different time intervals, and delta(13)C values were determined for gaseous CO(2), organic substrates, and products such as biomass. For three of the four SRB, carbon isotope effects between the substrates, acetate or lactate and CO(2), and the cell biomass were small, ranging from 0 to 2 per thousand. However, for Desulfotomaculum acetoxidans, the carbon incorporated into biomass was isotopically heavier than the available substrates by 8 to 9 per thousand. SRB grown lithoautotrophically consumed less than 3% of the available CO(2) and exhibited substantial discrimination (calculated as isotope fractionation factors [alpha]), as follows: for Desulfobacterium autotrophicum, alpha values ranged from 1.0100 to 1.0123; for Desulfobacter hydrogenophilus, the alpha value was 0.0138, and for Desulfotomaculum acetoxidans, the alpha value was 1.0310. Mixotrophic growth of Desulfovibrio desulfuricans on acetate and CO(2) resulted in biomass with a delta(13)C composition intermediate to that of the substrates. The extent of fractionation depended on which enzymatic pathways were used, the direction in which the pathways operated, and the growth rate, but fractionation was not dependent on the growth phase. To the extent that environmental conditions affect the availability of organic substrates (e.g., acetate) and reducing power (e.g., H(2)), ecological forces can also influence carbon isotope discrimination by SRB.     

Lipid biomarkers and carbon isotope signatures of a microbial (Beggiatoa) mat associated with gas hydrates in the gulf of Mexico

White and orange mats are ubiquitous on surface sediments associated with gas hydrates and cold seeps in the Gulf of Mexico. The goal of this study was to determine the predominant pathways for carbon cycling within an orange mat in Green Canyon (GC) block GC 234 in the Gulf of Mexico. Our approach incorporated laser-scanning confocal microscopy, lipid biomarkers, stable carbon isotopes, and 16S rRNA gene sequencing. Confocal microscopy showed the predominance of filamentous microorganisms (4 to 5 mum in diameter) in the mat sample, which are characteristic of Beggiatoa. The phospholipid fatty acids extracted from the mat sample were dominated by 16:1omega7c/t (67%), 18:1omega7c (17%), and 16:0 (8%), which are consistent with lipid profiles of known sulfur-oxidizing bacteria, including Beggiatoa. These results are supported by the 16S rRNA gene analysis of the mat material, which yielded sequences that are all related to the vacuolated sulfur-oxidizing bacteria, including Beggiatoa, Thioploca, and Thiomargarita. The delta13C value of total biomass was -28.6 per thousand; those of individual fatty acids were -29.4 to -33.7 per thousand. These values suggested heterotrophic growth of Beggiatoa on organic substrates that may have delta13C values characteristic of crude oil or on their by-products from microbial degradation. This study demonstrated that integrating lipid biomarkers, stable isotopes, and molecular DNA could enhance our understanding of the metabolic functions of Beggiatoa mats in sulfide-rich marine sediments associated with gas hydrates in the Gulf of Mexico and other locations.     

稳定同位素技术在植物水分利用研究中的应用

近20a稳定同位素技术在植物生态学研究中的应用得到了长足发展,使得对植物与水分关系也有了更深一步的了解。介绍稳定同位素性碳、氢、氧同位素在研究植物水分关系中的应用及进展,以期能为国内植物水分利用研究提供参考。由于植物根系从土壤中吸收水分时并不发生同位素分馏,对木质部水分同位素分析有助于对植物利用水分来源,生态系统中植物对水分的竞争和利用策略的研究,更好地了解生态系统结构与功能。稳定碳同位素作为植物水分利用效率的一个间接指标,在不同水分梯度环境中,及植物不同代谢产物与水分关系中有着广泛的应用。同位素在土壤-植被-大气连续体水分中的应用,有助于了解生态系统的水分平衡。随着稳定同位素方法的使用,植物与水分关系的研究将取得更大的进展。     

Use of the [(14)C]leucine incorporation technique to measure bacterial production in river sediments and the epiphyton.

Bacterial production is a key parameter for the understanding of carbon cycling in aquatic ecosystems, yet it remains difficult to measure in many aquatic habitats. We therefore tested the applicability of the [(14)C]leucine incorporation technique for the measurement of bulk bacterial production in various habitats of a lowland river ecosystem. To evaluate the method, we determined (i) extraction efficiencies of bacterial protein from the sediments, (ii) substrate saturation of leucine in sediments, the biofilms on aquatic plants (epiphyton), and the pelagic zone, (iii) bacterial activities at different leucine concentrations, (iv) specificity of leucine uptake by bacteria, and (v) the effect of the incubation technique (perfused-core incubation versus slurry incubation) on leucine incorporation into protein. Bacterial protein was best extracted from sediments and precipitated by hot trichloroacetic acid treatment following ultrasonication. For epiphyton, an alkaline-extraction procedure was most efficient. Leucine incorporation saturation occurred at 1 microM in epiphyton and 100 nM in the pelagic zone. Saturation curves in sediments were difficult to model but showed the first level of leucine saturation at 50 microM. Increased uptake at higher leucine concentrations could be partly attributed to eukaryotes. Addition of micromolar concentrations of leucine did not enhance bacterial electron transport activity or DNA replication activity. Similar rates of leucine incorporation into protein calculated for whole sediment cores were observed after slurry and perfused-core incubations, but the rates exhibited strong vertical gradients after the core incubation. We conclude that the leucine incorporation method can measure bacterial production in a wide range of aquatic habitats, including fluvial sediments, if substrate saturation and isotope dilution are determined.     

SHORT‐TERM MEASUREMENT OF CARBON STABLE ISOTOPE DISCRIMINATION IN PHOTOSYNTHESIS AND RESPIRATION BY AQUATIC MACROPHYTES,WITH MARINE MACROALGAL EXAMPLES1

Progress in the study of stable isotope discrimination in carbon assimilation by aquatic macrophytes has been slower than for other groups of primary producers, such as phytoplankton and terrestrial plants. A probable reason has been the methodologies employed for such a study: field collections or long‐term incubations, both relying on the observation of changes in carbon isotope composition of plant tissue. Here, we present a short‐term incubation method based on the change in carbon stable isotope composition in water. Its fundamental advantage over the other approaches is that the change in stable isotope composition in water in a closed system is much faster than in the plant tissue. We applied the method to investigate the relationship between carbon assimilation intensity and isotope discrimination. The results included a relatively small discrimination in respiration, a significant influence of carbon assimilation rate on discrimination, and the suggestion of HCO₃^? or CO₂ uptake in photosynthesis. The information gathered using this method would be difficult to obtain in other ways, and so we believe that it should contribute to a better understanding of the physiology and ecology of aquatic macrophytes.