Utilization of organic nitrogen by arbuscular mycorrhizal fungi-is there a specific role for protists and ammonia oxidizers?

Arbuscular mycorrhizal (AM) fungi can significantly contribute to plant nitrogen (N) uptake from complex organic sources, most likely in concert with activity of soil saprotrophs and other microbes releasing and transforming the N bound in organic forms. Here, we tested whether AM fungus (Rhizophagus irregularis) extraradical hyphal networks showed any preferences towards certain forms of organic N (chitin of fungal or crustacean origin, DNA, clover biomass, or albumin) administered in spatially discrete patches, and how the presence of AM fungal hyphae affected other microbes. By direct 15N labeling, we also quantified the flux of N to the plants (Andropogon gerardii) through the AM fungal hyphae from fungal chitin and from clover biomass. The AM fungal hyphae colonized patches supplemented with organic N sources significantly more than those receiving only mineral nutrients, organic carbon in form of cellulose, or nothing. Mycorrhizal plants grew 6.4-fold larger and accumulated, on average, 20.3-fold more 15N originating from the labeled organic sources than their nonmycorrhizal counterparts. Whereas the abundance of microbes (bacteria, fungi, or Acanthamoeba sp.) in the different patches was primarily driven by patch quality, we noted a consistent suppression of the microbial abundances by the presence of AM fungal hyphae. This suppression was particularly strong for ammonia oxidizing bacteria. Our results indicate that AM fungi successfully competed with the other microbes for free ammonium ions and suggest an important role for the notoriously understudied soil protists to play in recycling organic N from soil to plants via AM fungal hyphae.

The Uptake and Translocation of 99Tc, 133Cs, 237Np, and 238U Into Andropogon Virginicus With Consideration of Plant Life Stage.

Hydroponic uptake studies were conducted to evaluate the uptake and translocation of Tc, Cs (stable analog for Cs), Np, and U into established and seedling Andropogon virginicus specimens under controlled laboratory conditions. Plant specimens were grown in analyte-spiked Hoagland nutrient solution for 24 h, 3 d, and 5 d. Translocation to shoots was greatest for Tc and Cs, likely due to their analogous nature to plant nutrients, while U (and Np to a lesser extent) predominantly partitioned to root tissue with less extensive translocation to the shoots. Plant age contributed significantly to differences in concentration ratios for all nuclides in shoot tissues (p ≤ 0.024), with higher concentration ratios for seedling specimens. Additionally, duration of exposure was associated with significant differences in concentration ratios of Cs and Tc for seedlings (p = 0.007 and p = 0.030, respectively) while plant part (root or shoot) was associated with significant differences in concentration ratios of established plants (p < 0.001 for both nuclides). Statistically significant increases in radionuclide uptake in seedling specimens relative to established plants under controlled conditions suggests that, in addition to geochemical factors, plant life stage of wild grasses may also be an important factor influencing radionuclide transport in the natural environment.

Utilization of organic nitrogen by arbuscular mycorrhizal fungi-is there a specific role for protists and ammonia oxidizers?

Arbuscular mycorrhizal (AM) fungi can significantly contribute to plant nitrogen (N) uptake from complex organic sources, most likely in concert with activity of soil saprotrophs and other microbes releasing and transforming the N bound in organic forms. Here, we tested whether AM fungus (Rhizophagus irregularis) extraradical hyphal networks showed any preferences towards certain forms of organic N (chitin of fungal or crustacean origin, DNA, clover biomass, or albumin) administered in spatially discrete patches, and how the presence of AM fungal hyphae affected other microbes. By direct 15N labeling, we also quantified the flux of N to the plants (Andropogon gerardii) through the AM fungal hyphae from fungal chitin and from clover biomass. The AM fungal hyphae colonized patches supplemented with organic N sources significantly more than those receiving only mineral nutrients, organic carbon in form of cellulose, or nothing. Mycorrhizal plants grew 6.4-fold larger and accumulated, on average, 20.3-fold more 15N originating from the labeled organic sources than their nonmycorrhizal counterparts. Whereas the abundance of microbes (bacteria, fungi, or Acanthamoeba sp.) in the different patches was primarily driven by patch quality, we noted a consistent suppression of the microbial abundances by the presence of AM fungal hyphae. This suppression was particularly strong for ammonia oxidizing bacteria. Our results indicate that AM fungi successfully competed with the other microbes for free ammonium ions and suggest an important role for the notoriously understudied soil protists to play in recycling organic N from soil to plants via AM fungal hyphae.

Invasive Andropogon gayanus (Gamba grass) alters litter decomposition and nitrogen fluxes in an Australian tropical savanna.

The African grass Andropogon gayanus Kunth. is invading Australian savannas, altering their ecological and biogeochemical function. To assess impacts on nitrogen (N) cycling, we quantified litter decomposition and N dynamics of grass litter in native grass and A. gayanus invaded savanna using destructive in situ grass litter harvests and litterbag incubations (soil surface and aerial position). Only 30% of the A. gayanus in situ litter decomposed, compared to 61% of the native grass litter, due to the former being largely comprised of highly resistant A. gayanus stem. In contrast to the stem, A. gayanus leaf decomposition was approximately 3- and 2-times higher than the dominant native grass, Alloteropsis semilata at the surface and aerial position, respectively. Lower initial lignin concentrations, and higher consumption by termites, accounted for the greater surface decomposition rate of A. gayanus. N flux estimates suggest the N release of A. gayanus litter is insufficient to compensate for increased N uptake and N loss via fire in invaded plots. Annually burnt invaded savanna may lose up to 8.2% of the upper soil N pool over a decade. Without additional inputs via biological N fixation, A. gayanus invasion is likely to diminish the N capital of Australia's frequently burnt savannas.

Uptake of C14-atrazine by prairie grasses in a phytoremediation setting.

Agrochemicals significantly contribute to environmental pollution. In the USA, atrazine is a widely used pesticide and commonly found in rivers, water systems, and rural wells. Phytoremediation can be a cost-effective means of removing pesticides from soil. The objective of this project was to investigate the ability of prairie grasses to remove atrazine. 14C-labeled atrazine was added to sterilized sand and water/nutrient cultures, and the analysis was performed after 21 days. Switchgrass and big bluestem were promising species for phytoremediation, taking up about 40% of the applied [14C] in liquid hydroponic cultures, and between 20% and 33% in sand cultures. Yellow Indiangrass showed low resistance to atrazine toxicity and low uptake of [14C] atrazine in liquid hydroponic cultures. Atrazine degradation increased progressively from sand to roots and leaves. Most atrazine taken up by prairie grasses from sand culture was degraded to metabolites, which accounted for 60-80% of [14C] detected in leaves. Deisopropylatrazine (DIA) was the main metabolite detected in sand and roots, whereas in leaves further metabolism took place, forming increased amounts of didealkylatrazine (DDA) and an unidentified metabolite. In conclusion, prairie grasses achieved high atrazine removal and degradation, showing a high potential for phytoremediation.

NMR-based Metabolomics to Study the Cold-acclimation Strategy of Two Miscanthus Genotypes.

INTRODUCTION: Abiotic stress is a major cause of yield loss in plant culture. Miscanthus, a perennial C4 grass, is now considered a major source of renewable energy, especially for biofuel production. During the first year of planting in Northern Europe, Miscanthus was exposed to frost temperature, which generated high mortality in young plants and large loss of yield. One strategy to avoid such loss is to apply cold-acclimation, which confers on plants a better resistance to low temperature. OBJECTIVES: The aim of this study is to describe the effect of a cold-acclimation period on the metabolome of two Miscanthus genotypes that vary in their frost sensitivity at the juvenile stage. Miscanthus × giganteus (GIG) is particularly sensitive to frost, whereas Miscanthus sinensis August Feder (AUG) is tolerant. MATERIALS AND METHODS: Polar metabolite extraction was performed on Miscanthus, grown in non-acclimated or cold-acclimated conditions. Extracts were analysed by 1 H-NMR followed by multivariate statistical analysis. Discriminant metabolites were identified. RESULTS: More than 40 metabolites were identified in the two Miscanthus genotypes. GIG and AUG showed a different metabolic background before cold treatment, probably related to their genetic background. After cold-acclimation, GIG and AUG metabolomes remained different. The tolerant genotype showed notably higher levels of accumulation in proline, sucrose and maltose when subjected to cold. CONCLUSION: These two genotypes seem to have a different adaptation strategy in cold conditions. The studied change in the metabolome concerns different types of molecules related to the cold-tolerant behaviour of Miscanthus. Copyright © 2016 John Wiley & Sons, Ltd.

A Genomic Resource for the Development, Improvement, and Exploitation of Sorghum for Bioenergy.

With high productivity and stress tolerance, numerous grass genera of the Andropogoneae have emerged as candidates for bioenergy production. To optimize these candidates, research examining the genetic architecture of yield, carbon partitioning, and composition is required to advance breeding objectives. Significant progress has been made developing genetic and genomic resources for Andropogoneae, and advances in comparative and computational genomics have enabled research examining the genetic basis of photosynthesis, carbon partitioning, composition, and sink strength. To provide a pivotal resource aimed at developing a comparative understanding of key bioenergy traits in the Andropogoneae, we have established and characterized an association panel of 390 racially, geographically, and phenotypically diverse Sorghum bicolor accessions with 232,303 genetic markers. Sorghum bicolor was selected because of its genomic simplicity, phenotypic diversity, significant genomic tools, and its agricultural productivity and resilience. We have demonstrated the value of sorghum as a functional model for candidate gene discovery for bioenergy Andropogoneae by performing genome-wide association analysis for two contrasting phenotypes representing key components of structural and non-structural carbohydrates. We identified potential genes, including a cellulase enzyme and a vacuolar transporter, associated with increased non-structural carbohydrates that could lead to bioenergy sorghum improvement. Although our analysis identified genes with potentially clear functions, other candidates did not have assigned functions, suggesting novel molecular mechanisms for carbon partitioning traits. These results, combined with our characterization of phenotypic and genetic diversity and the public accessibility of each accession and genomic data, demonstrate the value of this resource and provide a foundation for future improvement of sorghum and related grasses for bioenergy production.

Common mycorrhizal networks amplify competition by preferential mineral nutrient allocation to large host plants.

Arbuscular mycorrhizal (AM) fungi interconnect plants in common mycorrhizal networks (CMNs) which can amplify competition among neighbors. Amplified competition might result from the fungi supplying mineral nutrients preferentially to hosts that abundantly provide fixed carbon, as suggested by research with organ-cultured roots. We examined whether CMNs supplied (15) N preferentially to large, nonshaded, whole plants. We conducted an intraspecific target-neighbor pot experiment with Andropogon gerardii and several AM fungi in intact, severed or prevented CMNs. Neighbors were supplied (15) N, and half of the target plants were shaded. Intact CMNs increased target dry weight (DW), intensified competition and increased size inequality. Shading decreased target weight, but shaded plants in intact CMNs had mycorrhizal colonization similar to that of sunlit plants. AM fungi in intact CMNs acquired (15) N from the substrate of neighbors and preferentially allocated it to sunlit, large, target plants. Sunlit, intact CMN, target plants acquired as much as 27% of their nitrogen from the vicinity of their neighbors, but shaded targets did not. These results suggest that AM fungi in CMNs preferentially provide mineral nutrients to those conspecific host individuals best able to provide them with fixed carbon or representing the strongest sinks, thereby potentially amplifying asymmetric competition below ground.

Effect of fibrolytic enzymes added to a Andropogon gayanus grass silage-concentrate diet on rumen fermentation in batch cultures and the artificial rumen (Rusitec).

In vitro batch cultures were used to screen four fibrolytic enzyme mixtures at two dosages added to a 60 : 40 silage : concentrate diet containing the C(4) tropical grass Andropogon gayanus grass ensiled at two maturities - vegetative stage (VS) and flowering stage (FS). Based on these studies, one enzyme mixture was selected to treat the same diets and evaluate its impact on fermentation using an artificial rumen (Rusitec). In vitro batch cultures were conducted as a completely randomized design with two runs, four replicates per run and 12 treatments in a factorial arrangement (four enzyme mixtures×three doses). Enzyme additives (E1, E2, E3 and E4) were commercial products and contained a range of endoglucanase, exoglucanase and xylanase activities. Enzymes were added to the complete diet 2 h before incubation at 0, 2 and 4 µl/g of dry matter (DM). Gas production (GP) was measured after 3, 6, 12, 24 and 48 h of incubation. Disappearance of DM (DMD), NDF (NDFD) and ADF (ADFD) were determined after 24 and 48 h. For all four enzyme mixtures, a dosage effect (P>0.05) DM, N, NDF or ADF disappearance after 48 h of incubation nor daily ammonia-N, volatile fatty acids or CH(4) production. However, enzyme application increased (P<0.05) microbial N production in feed particle-associated (loosely-associated) and silage feed particle-bound (firmly associated) fractions. With A. gayanus silage diets, degradation may not be limited by microbial colonization, but rather by the ability of fibrolytic enzymes to degrade plant cell walls within this recalcitrant forage.

Mycorrhizal phenotypes and the Law of the Minimum.

Mycorrhizal phenotypes arise from interactions among plant and fungal genotypes and the environment. Differences in the stoichiometry and uptake capacity of fungi and plants make arbuscular mycorrhizal (AM) fungi inherently more nitrogen (N) limited and less phosphorus (P) limited than their host plants. Mutualistic phenotypes are most likely in P-limited systems and commensal or parasitic phenotypes in N-limited systems. Carbon (C) limitation is expected to cause phenotypes to shift from mutualism to commensalism and even parasitism. Two experiments compared the influence of fertilizer and shade on mycorrhizas in Andropogon gerardii across three naturally N-limited or P-limited grasslands. A third experiment examined the interactive effects of N and P enrichment and shade on A. gerardii mycorrhizas. Our experiments generated the full spectrum of mycorrhizal phenotypes. These findings support the hypothesis that mutualism is likely in P-limited systems and commensalism or parasitism is likely in N-limited systems. Furthermore, shade decreased C-assimilation and generated less mutualistic mycorrhizal phenotypes with reduced plant and fungal biomass. Soil fertility is a key controller of mycorrhizal costs and benefits and the Law of the Minimum is a useful predictor of mycorrhizal phenotype. In our experimental grasslands arbuscular mycorrhizas can ameliorate P-limitation but not N-limitation.

Design and operation of a continuous 13C and 15N labeling chamber for uniform or differential, metabolic and structural, plant isotope labeling.

Tracing rare stable isotopes from plant material through the ecosystem provides the most sensitive information about ecosystem processes; from CO2 fluxes and soil organic matter formation to small-scale stable-isotope biomarker probing. Coupling multiple stable isotopes such as (13)C with (15)N, (18)O or (2)H has the potential to reveal even more information about complex stoichiometric relationships during biogeochemical transformations. Isotope labeled plant material has been used in various studies of litter decomposition and soil organic matter formation(1-4). From these and other studies, however, it has become apparent that structural components of plant material behave differently than metabolic components (i.e. leachable low molecular weight compounds) in terms of microbial utilization and long-term carbon storage(5-7). The ability to study structural and metabolic components separately provides a powerful new tool for advancing the forefront of ecosystem biogeochemical studies. Here we describe a method for producing (13)C and (15)N labeled plant material that is either uniformly labeled throughout the plant or differentially labeled in structural and metabolic plant components. Here, we present the construction and operation of a continuous (13)C and (15)N labeling chamber that can be modified to meet various research needs. Uniformly labeled plant material is produced by continuous labeling from seedling to harvest, while differential labeling is achieved by removing the growing plants from the chamber weeks prior to harvest. Representative results from growing Andropogon gerardii Kaw demonstrate the system's ability to efficiently label plant material at the targeted levels. Through this method we have produced plant material with a 4.4 atom%(13)C and 6.7 atom%(15)N uniform plant label, or material that is differentially labeled by up to 1.29 atom%(13)C and 0.56 atom%(15)N in its metabolic and structural components (hot water extractable and hot water residual components, respectively). Challenges lie in maintaining proper temperature, humidity, CO2 concentration, and light levels in an airtight (13)C-CO2 atmosphere for successful plant production. This chamber description represents a useful research tool to effectively produce uniformly or differentially multi-isotope labeled plant material for use in experiments on ecosystem biogeochemical cycling.

Plant tissue analysis for explosive compounds in phytoremediation and phytoforensics.

Plant tissue analysis methods were evaluated for six explosive compounds to assess uptake and phytoforensic methods development to quantify explosives in plant to obtain the plant data for the evaluation of explosive contamination in soil and groundwater. Four different solvent mixtures containing acetonitrile or methanol were tested at variable extraction ratios to compare the extraction efficiency for six explosive compounds: 2,4,6-trinitrotoluene (TNT), pentaerythritoltetranitrate (PETN), hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX), 2-amino-4,6-dinitrotoluene (2ADNT), and 2,4-Dinitroanisole (DNAN), in Laurel Willow (Salix pentandra) stem and range grass Big Bluestem (Andropogon gerardii) using LC-MS/MS. Plant tissues were spiked with 500 ng/g of explosives and extracted using ultrasonically-assisted solvent extraction. With the ratio of fresh plant mass to solvent volume of 1:20 for willow and 1:40 for big bluestem grass, results indicated that all explosives in willow except HMX were extracted at higher than 73.3% by using 20 mL of methanol, 50:50 (v/v) methanol:water, or acetonitrile, whereas HMX was extracted with the highest recovery of 61.3% by 20 mL of acetonitrile. In big bluestem grass, the most effective solvents were 20 mL of either methanol or 50:50 (v/v) methanol:water for PETN extraction with a recovery of higher than 101.2% and 20 mL of 50:50 (v/v) methanol:water for HMX, RDX, TNT, 2ADNT, and DNAN extraction with a recovery of 83.8%, 104.4%, 97.5%, 80.7%, and 108.2%, respectively. However, unlike methanol and acetonitrile, 50:50 (v/v) methanol:water provided no problem of leading or split peak in chromatogram; therefore, it was preferred in the test and performed a method validation. Results indicated that 50:50 (v/v) methanol:water provided good repeatability and recovery and method detection limits at 0.5-20 ng/g fresh weight or 8.8-61.3 ng/g dry weight. Overall, results suggested that solvent extraction efficiency of explosives in plant was influenced by plant species and solvent used, and method presented here was believed to provide the preliminary data with respect to the analysis of simultaneous explosives in plants with LC-MS/MS.

Invasive Andropogon gayanus (gamba grass) is an ecosystem transformer of nitrogen relations in Australian savanna.

Invasion by the African grass Andropogon gayanus is drastically altering the understory structure of oligotrophic savannas in tropical Australia. We compared nitrogen (N) relations and phenology of A. gayanus and native grasses to examine the impact of invasion on N cycling and to determine possible reasons for invasiveness of A. gayanus. Andropogon gayanus produced up to 10 and four times more shoot phytomass and root biomass, with up to seven and 2.5 times greater shoot and root N pools than native grass understory. These pronounced differences in phytomass and N pools between A. gayanus and native grasses were associated with an altered N cycle. Most growth occurs in the wet season when, compared with native grasses, dominance of A. gayanus was associated with significantly lower total soil N pools, lower nitrification rates, up to three times lower soil nitrate availability, and up to three times higher soil ammonium availability. Uptake kinetics for different N sources were studied with excised roots of three grass species ex situ. Excised roots of A. gayanus had an over six times higher-uptake rate of ammonium than roots of native grasses, while native grass Eriachne triseta had a three times higher uptake rate of nitrate than A. gayanus. We hypothesize that A. gayanus stimulates ammonification but inhibits nitrification, as was shown to occur in its native range in Africa, and that this modification of the soil N cycle is linked to the species' preference for ammonium as an N source. This mechanism could result in altered soil N relations and could enhance the competitive superiority and persistence of A. gayanus in Australian savannas.

Effect of elevated CO2 and drought on soil microbial communities associated with Andropogon gerardii.

Our understanding of the effects of elevated atmospheric CO2, singly and in combination with other environmental changes,on plant-soil interactions is incomplete. Elevated CO2 effects on C4 plants, though smaller than on C3 species, are mediated mostly via decreased stomatal conductance and thus water loss. Therefore, we characterized the interactive effect of elevated CO2 and drought on soil microbial communities associated with a dominant C4 prairie grass, Andropogon gerardii Vitman. Elevated CO2 and drought both affected resources available to the soil microbial community. For example, elevated CO2 increased the soil C:N ratio and water content during drought, whereas drought alone decreased both. Drought significantly decreased soil microbial biomass. In contrast, elevated CO2 increased biomass while ameliorating biomass decreases that were induced under drought. Total and active direct bacterial counts and carbon substrate use (overall use and number of used sources) increased significantly under elevated CO2. Denaturing gradient gel electrophoresis analysis revealed that drought and elevated CO2, singly and combined, did not affect the soil bacteria community structure.We conclude that elevated CO2 alone increased bacterial abundance and microbial activity and carbon use, probably in response to increased root exudation. Elevated CO2 also limited drought-related impacts on microbial activity and biomass,which likely resulted from decreased plant water use under elevated CO2. These are among the first results showing that elevated CO2 and drought work in opposition to modulate plant-associated soil-bacteria responses, which should then influence soil resources and plant and ecosystem function.

Effects of N on plant response to heat-wave: a field study with prairie vegetation.

More intense, more frequent, and longer heat-waves are expected in the future due to global warming, which could have dramatic ecological impacts. Increasing nitrogen (N) availability and its dynamics will likely impact plant responses to heat stress and carbon (C) sequestration in terrestrial ecosystems. This field study examined the effects of N availability on plant response to heat-stress (HS) treatment in naturally-occurring vegetation. HS (5 d at ambient or 40.5 degrees C) and N treatments (+/-N) were applied to 16 1 m(2) plots in restored prairie vegetation dominated by Andropogon gerardii (warm-season C4 grass) and Solidago canadensis (warm-season C3 forb). Before, during, and after HS, air, canopy, and soil temperature were monitored; net CO2 assimilation (P(n)), quantum yield of photosystem II (Phi(PSII)), stomatal conductance (g(s)), and leaf water potential (Psi(w)) of the dominant species and soil respiration (R(soil)) of each plot were measured daily during HS. One week after HS, plots were harvested, and C% and N% were determined for rhizosphere and bulk soil, and above-ground tissue (green/senescent leaf, stem, and flower). Photosynthetic N-use efficiency (PNUE) and N resorption rate (NRR) were calculated. HS decreased P(n), g(s), Psi(w), and PNUE for both species, and +N treatment generally increased these variables (+/-HS), but often slowed their post-HS recovery. Aboveground biomass tended to decrease with HS in both species (and for green leaf mass in S. canadensis), but decrease with +N for A. gerardii and increase with +N for S. canadensis. For A. gerardii, HS tended to decrease N% in green tissues with +N, whereas in S. canadensis, HS increased N% in green leaves. Added N decreased NRR for A. gerardii and HS increased NRR for S. canadensis. These results suggest that heat waves, though transient, could have significant effects on plants, communities, and ecosystem N cycling, and N can influence the effect of heat waves.

Land-use changes alter CO2 flux patterns of a tall-grass Andropogon field and a savanna-woodland continuum in the Orinoco lowlands.

Land use changes in the savannas of the Orinoco lowlands have resulted in a mosaic of vegetation. To elucidate how these changes have affected carbon exchanges with the atmosphere, we measured CO2 fluxes by eddy covariance and soil CO2 efflux systems along a disturbance gradient beginning with a cultivated tall-grass Andropogon field (S1) and extending over three savanna sites with increasing woody cover growing above native herbaceous vegetation. The savanna sites included a herbaceous savanna (S2), a tree savanna (S3) and a woodland savanna (S4). During the wet season, maximum diurnal net ecosystem exchange (NEE) over the S1-S4 sites was 6.6-9.3, 6.6-7.9, 10.6-11.3 and 9.3-10.6 micromol m(-2) s(-1), respectively. The rate of CO2 uptake over S1 was lower than that for C4 grasses elsewhere because of pasture degradation. Soil respiration and temperature were exponentially related when soil water content (theta) was above 0.083 m(3) m(-3); however, soil respiration declined markedly as theta decreased from 0.083-0.090 to 0.033-0.056 m(3) m(-3). There were bursts of CO2 emission when dry soils were rewetted by rainfall. During the wet season, all sites constituted carbon sinks with maximum net daily ecosystem production (NEP) of 2.1, 1.7, 2.1 and 2.1 g C m(-2) day(-1), respectively. During the dry season, the savanna sites (S2-S4) became carbon sources with maximum emission fluxes of -0.5, -1.4 and -1.6 g C m(-2) day(-1), respectively, whereas the tall-grass field (S1) remained a carbon sink with a maximum NEP of 0.3 g C m(-2) day(-1) at the end of the season. For all measurement periods, annual NEP of sites S1-S4 was 366, 6, 116 and 139 g C m(-2), respectively. Comparisons of carbon source/sink dynamics across a wide range of savannas indicate that savanna carbon budgets can change in sign and magnitude. On an annual basis, gross primary production over the S1-S4 stands was 797, 803, 136 and 1230 g C m(-2), respectively. Net primary productivity (NPP) of the S1-S4 stands, calculated from eddy covariance measurements as the daily sum of NEE and day and night heterotrophic respiration was 498, 169, 181 and 402 g C m-2 year-1, respectively. These values were slightly higher than NPP based on harvest measurements (432, 162, 176 and 386 g C m(-2) year(-1), respectively), presumably because fine roots were incompletely harvested. Soil water content limited carbon uptake at all sites, and water-use efficiency (WUE) was related to rainfall dynamics. During the dry season, all sites except the cultivated tall-grass Andropogon field (S1) had a negative WUE. Although our results are specific to the Orinoco vegetational mosaic, the effects of land-use practices on the controls and physiological functions of the studied ecosystems may be generalized to other savannas.