In Situ Cultivation of Subsurface Microorganisms in a Deep Mafic Sill: Implications for SLiMEs

An in situ culturing device was incubated within a flowing borehole in a mafic sill at 1.474 km depth in Evander Au mine, South Africa. The device was designed to enrich methanogenic, Fe^{3 +}-reducing and SO₄ ^{2 ?}-reducing microorganisms using acetate, formate, methanol, Fe^{3 +}-citrate and SO₄ ^{2 ?} enriched agar and sand cartridges. At the end of the 33 day incubation geochemical analyses detected elevated H₂, acetate, CH₄ and Fe concentrations and depleted SO₄ ^{2 ?} concentrations. 16S rDNA sequences and PLFA analyses revealed that the microbial community composition of the substrate-bearing cartridges were distinct from that of the original borehole water and the non-substrate-bearing control cartridge. 16S rDNA and dissimilatory sulfite reductase, dsrAB, gene sequences indicated the device successfully targeted SO₄ ^{2 ?} reducing bacteria (SRB), which were not detected in the original borehole water. 16S rDNA sequences also revealed a shift in the microbial community from one relying on H₂ based methanogenesis to one suggestive of H₂ based acetogenesis supporting aceticlastic methanogenesis and SO₄ ^{2 ?} reduction compatible with the subsurface lithoautotrophic hypothesis.     

Absorption characteristics and kinetics of CO2 capture into N-methyldiethanolamine aqueous solution catalyzed by the immobilized carbonic anhydrase

Abstract

Carbonic anhydrase (CA) is the most effective CO₂ hydratase catalyst, but the poor storage stability and repeatability of CA limit its development. Therefore, CA was immobilized on the epoxy magnetic composite microspheres to enhance the CO₂ absorption into N-methyldiethanolamine (MDEA) aqueous solution in this work. In the presence of immobilized CA, the CO₂ absorption rate of MDEA solution (10?wt%) (0.63?mmol·min^?1) was greatly improved by almost 40%, and their reaction equilibrium time was shortened from 150?min to 90?min compared with that into MDEA solution. The results indicated that the absorption of CO₂ into MDEA solution had been significantly enhanced by using CA. After the 7th reuse recycle, the activity of the immobilized CA was still closed to its initial value at 313.15?K. Moreover, enzyme catalytic kinetics of immobilized CA was investigated using the p-nitrophenyl acetate (p-NPA) as substrate. The values of Michaelis–Menten constant (K_m) and the maximum velocity (V_max) of the immobilized CA were calculated to be 27.61?mmol/L and 20.14?×?10^?3?mmol·min^?1·mL^?1, respectively. Besides, the kinetics of CO₂ reaction into MDEA with or without CA were also compared. The results showed that CO₂ absorption into CA/MDEA aqueous solution obeyed the pseudo first order regime and the second order kinetics rate constant (k₂) was calculated to be 929?m³·kmol^?1·s^?1, which was twice higher than that of MDEA aqueous solution without immobilized CA (k₂=414 m³·kmol^?1·s^?1) at 313.15?K.     

Non-phototrophic CO 2 fixation by soil microorganisms

Although soils are generally known to be a net source of CO₂ due to microbial respiration, CO₂ fixation may also be an important process. The non-phototrophic fixation of CO₂ was investigated in a tracer experiment with ¹⁴CO₂ in order to obtain information about the extent and the mechanisms of this process. Soils were incubated for up to 91 days in the dark. In three independent incubation experiments, a significant transfer of radioactivity from ¹⁴CO₂ to soil organic matter was observed. The process was related to microbial activity and could be enhanced by the addition of readily available substrates such as acetate. CO₂ fixation exhibited biphasic kinetics and was linearly related to respiration during the first phase of incubation (about 20–40 days). The fixation amounted to 3–5% of the net respiration. After this phase, the CO₂ fixation decreased to 1–2% of the respiration. The amount of carbon fixed by an agricultural soil corresponded to 0.05% of the organic carbon present in the soil at the beginning of the experiment, and virtually all of the fixed CO₂ was converted to organic compounds. Many autotrophic and heterotrophic biochemical processes result in the fixation of CO₂. However, the enhancement of the fixation by addition of readily available substrates and the linear correlation with respiration suggested that the process is mainly driven by aerobic heterotrophic microorganisms. We conclude that heterotrophic CO₂ fixation represents a significant factor of microbial activity in soils.     

Fungal community composition and metabolism under elevated CO2 and O3

Atmospheric CO₂ and O₃ concentrations are increasing due to human activity and both trace gases have the potential to alter C cycling in forest ecosystems. Because soil microorganisms depend on plant litter as a source of energy for metabolism, changes in the amount or the biochemistry of plant litter produced under elevated CO₂ and O₃ could alter microbial community function and composition. Previously, we have observed that elevated CO₂ increased the microbial metabolism of cellulose and chitin, whereas elevated O₃ dampened this response. We hypothesized that this change in metabolism under CO₂ and O₃ enrichment would be accompanied by a concomitant change in fungal community composition. We tested our hypothesis at the free-air CO₂ and O₃ enrichment (FACE) experiment at Rhinelander, Wisconsin, in which Populus tremuloides, Betula papyrifera, and Acer saccharum were grown under factorial CO₂ and O₃ treatments. We employed extracellular enzyme analysis to assay microbial metabolism, phospholipid fatty acid (PLFA) analysis to determine changes in microbial community composition, and polymerase chain reaction–denaturing gradient gel electrophoresis (PCR–DGGE) to analyze the fungal community composition. The activities of 1,4-β-glucosidase (+37%) and 1,4,-β-N-acetylglucosaminidase (+84%) were significantly increased under elevated CO₂, whereas 1,4-β-glucosidase activity (−25%) was significantly suppressed by elevated O₃. There was no significant main effect of elevated CO₂ or O₃ on fungal relative abundance, as measured by PLFA. We identified 39 fungal taxonomic units from soil using DGGE, and found that O₃ enrichment significantly altered fungal community composition. We conclude that fungal metabolism is altered under elevated CO₂ and O₃, and that there was a concomitant change in fungal community composition under elevated O₃. Thus, changes in plant inputs to soil under elevated CO₂ and O₃ can propagate through the microbial food web to alter the cycling of C in soil.     

Kinetics and mechanisms of the CO2 and SO2 uptake by coordinate ion, cis-[Cr(C2O4)(L-L)(OH2)2] {(L-L) = methyl 3-amino-2,3-dideoxy-α-d-arabino-hexopyranoside} as studied by stopped-flow spectrophotometry

The kinetics of CO₂ and SO₂ uptake by a coordinate ion, cis-[Cr(C₂O₄)(L-L)(OH₂)₂]⁺, where L-L stands for a bidentate sugar ligand, methyl 3-amino-2,3-dideoxy-α-d-arabino-hexopyranoside has been studied, over temperature ranges of 5 - 25 and 5 - 20 °C for CO₂ and SO₂, respectively. Investigations were carried out using stopped-flow spectrophotometry in the range of 340-700 nm. Results of the kinetic measurements obtained for both gases were compared. The kinetics and mechanisms of the reactions were suggested and ΔH^‡ values for both processes were determined.     

Geochemical and Microbiological Studies of Nitrous Oxide Variations within the New NEEM Greenland Ice Core during the Last Glacial Period

Deep polar ice cores provide atmospheric records of nitrous oxide (N₂O) and other trace gases reflecting climate history along with a parallel archive of microbial cells transported with mineral dust, marine and volcanic aerosols from around the globe. Our interdisciplinary study of 32 samples from different depths of the recently drilled NEEM Greenland ice core addressed the question whether the identified microorganisms were capable of post-depositional biological production of N₂O in situ. We used high-resolution geochemical and microbiological approaches to examine the N₂O concentrations, the quantitative distributions of dust, Ca⁺², NH₄⁺ and NO₃^? ions related to N cycle pathways, the microbial abundance and diversity at specific NEEM core depths from 1758 m to 1867.8 m. Results showed varying concentrations of N₂O (220–271.5 ppb). Microbial abundance fluctuated between 3.3 × 10⁴ and 3.3 × 10⁶ cells mL^?1 in direct correlation with dust and Ca²⁺ concentrations with higher cell numbers deposited during colder periods. The average values of NH₄⁺ and NO₃^? indicated that substrates were available for the microorganisms capable of utilizing them. PCR amplification of selected functional genes involved in bacterial and archaeal nitrification and denitrification was not successful. Sanger and Illumina MiSeq sequence analyses of SSU rRNA genes showed variable representation of Alpha-, Beta- and Gammaproteobacteria, Firmicutes, Actinobacteria, chloroplasts and fungi. The metabolic potential of the dominant genera of Proteobacteria and Firmicutes as possible N₂O producers suggested that denitrification activity may have led to in-situ production and accumulation of N₂O.     

温带针阔混交林土壤碳氮气体通量的主控因子与耦合关系

中高纬度森林地区由于气候条件变化剧烈,土壤温室气体排放量的估算存在很大的不确定性,并且不同碳氮气体通量的主控因子与耦合关系尚不明确。以长白山温带针阔混交林为研究对象,采用静态箱-气相色谱法连续4a(2005—2009年)测定土壤二氧化碳(CO2)、甲烷(CH4)和氧化亚氮(N2O)净交换通量以及温度、水分等相关环境因子。研究结果表明:温带针阔混交林土壤整体上表现为CO2和N2O的排放源和CH4的吸收汇。土壤CH4、CO2和N2O通量的年均值分别为-1.3 kg CH4hm-2a-1、15102.2 kg CO2hm-2a-1和6.13 kg N2O hm-2a-1。土壤CO2通量呈现明显的季节性规律,主要受土壤温度的影响,水分次之;土壤CH4通量的季节变化不明显,与土壤水分显著正相关;土壤N2O通量季节变化与土壤CO2通量相似,与土壤水分、温度显著正相关。土壤CO2通量和CH4通量不存在任何类型的耦合关系,与N2O通量也不存在耦合关系;土壤CH4和N2O通量之间表现为消长型耦合关系。这项研究显示温带针阔混交林土壤碳氮气体通量主要受环境因子驱动,不同气体通量产生与消耗之间存在复杂的耦合关系,下一步研究需要深入探讨环境变化对其耦合关系的影响以及内在的生物驱动机制。     

Growth,14CO2 fixation,activites of photosystems,ribulose 1,5-bisphosphate carboxylase and nitrate reductase in trees as affected by simulated acid rain

In seedlings of the tropical tree speciesErythrina variegata Lam. andHardwickia binata Roxb. exposed to different acidic mist (H₂SO₄, pH 5, 3 and 2) for 5 d significant reduction in seedling growth, biomass accumulation and¹⁴CO₂ fixation were determined. In isolated chloroplasts a decrease in the activities of photosystem 2 and whole electron transport chain was observed only at pH 3 and 2, but no significant change in photosystem 1 activity was observed. SDS-PAGE analysis of crude leaf extracts of ribulose 1,5-bisphosphate carboxylase (RuBPC) indicated a significant loss of 55 and 15 kDa polypeptides at pH 2 inErythrina. The reduction in the RuBPC activity in seedlings grown under acidic mists correlated well with CO₂ fixation.     

Enhanced methane production in a two-phase anaerobic digestion plant, after CO2 capture and addition to organic wastes

Cost-effective technologies are needed to reach the international greenhouse gas (GHG) reduction targets in many fields, including waste and biomass treatment. This work reports the effects of CO₂ capture from a combustion flue gas and its use in a newly-patented, two-phase anaerobic digestion (TPAD) process, to improve energy recovery and to reduce CO₂ emissions. A TPAD process, fed with urban wastewater sludge, was successfully established and maintained for several months at pilot scale. The TPAD process with injection of CO₂ exhibits efficient biomass degradation (58% VSS reduction), increased VFA production during the acidogenic phase (leading to VFA concentration of 8.4 g/L) and high biomethane production (0.350 Sm³/kg_SSV; 0.363 Sm³/m³_react·d). Moreover, CO₂ intake in the acid phase has a positive impact on the overall GHG balance associated to biomethane production, and suggests an improved solution for both emission reduction and biomass conversion into biomethane.     

Physiological variables determined under laboratory conditions may explain the bloom of Aphanizomenon ovalisporum in Lake Kinneret

Response of Aphanizomenon ovalisporum to certain environmental parameters was studied to gain a better understanding of the conditions which may have stimulated its autumnal bloom in Lake Kinneret. Optimal temperature for A. ovalisporum growth was 26–30?°C, resulting in growth rates of 0.2–0.3?day^?1, similar to those observed in the lake. Maximal rate of CO₂ fixation (assimilation numbers of 6–8?μg?C?μg^?1?Chl?h^?1) was obtained at low irradiances (I _k of 40–100?μmol?photons?m^?2?s^?1), 200?μM Pi and low N:Pi ratios. Growth was strongly affected by phosphorus availability, reaching a maximum at Pi concentrations above 40?μM. The high demand for phosphorus was indicated by an increase in alkaline phosphatase activity. The relative abundance of Pi in the cells increased by 4-fold in Pi-rich compared with Pi-limited cultures. Uptake of Pi was faster in Pi-depleted compared with Pi-sufficient cells. Maximal photosynthetic rates and K_1/2(HCO₃ ^?) were 140–220?μmol?O₂?mg^?1?Chl?h^?1 and 10–24?μM, respectively. At pH 7.0 the K _1/2(CO₂) was 2.2 and fell to 0.04?μM at pH 9.0. These data indicated that A. ovalisporum is a HCO₃ ^? user, and can explain its high photosynthetic rates during the bloom, under high pH and low dissolved CO₂ conditions. Na⁺ concentrations of about 5?mM were essential for A. ovalisporum growth at high pH approaching values in the lake.     

Influence of enhanced CO2 on growth and photosynthesis of the red algaeGracilaria sp. andG. chilensis

The influence of elevated CO₂ concentrations on growth and photosynthesis ofGracilaria sp. andG. chilensis was investigated in order to procure information on the effective utilization of CO₂. Growth of both was enhanced by CO₂ enrichment (air + 650 ppm CO₂, air + 1250 ppm CO₂, the enhancement being greater inGracilaria sp. Both species increased uptake of NO₃ ^– with CO₂ enrichment. Photosynthetic inorganic carbon uptake was depressed inG. chilensis by pre-culture (15 days) with CO₂ enrichment, but little affected inGracilaria sp. Mass spectrometric analysis showed that O₂ uptake was higher in the light than in the dark for both species and in both cases was higher inGracilaria sp. The higher growth enhancement inGracilaria sp. was attributed to greater depression of photorespiration by the enrichment of CO₂ in culture.     

Microalgal removal of CO<Subscript>2</Subscript> from flue gases: Changes in medium pH and flue gas composition do not appear to affect the photochemical yield of microalgal cultures

Our research objectives are to determine under what conditions microalgal-based CO₂ capture from flue gases is economically attractive. Specifically, our objective here was to select microalgae that are temperature, pH and flue gas tolerant. Microalgae were grown under five different temperatures, three different pH and five different flue gas mixtures besides 100% CO₂ (gas concentrations that the cells were exposed to ranged 5.7–100% CO₂, 0–3504 ppm SO₂, 0–328 ppm NO, and 0–126 ppm NO₂). Our results indicate that the microalgal strains tested exhibit a substantial ability to withstand a wide range of temperature (54 strains tested), pH (20 strains tested) and flue gas composition (24 strains tested) likely to be encountered in cultures used for carbon sequestration from smoke stack gases. Our results indicate that microalgal photosynthesis is a limited but viable strategy for CO₂ capture from flue gases produced by stationary combustion sources.     

Effects of CO2 enrichment,nutrient addition,and fungal endophyte-infection on the growth of two grasses

Summary Increasing atmospheric carbon dioxide (CO₂) concentration is expected to increase plant productivity and alter plant/plant interactions, but little is known about its effects on symbiotic interactions with microorganisms. Interactions between perennial ryegrass, Lolium perenne (a C3 plant), and purpletop grass, Tridens flavus (a C4 plant), and their clavicipitaceous fungal endophytes (Acremonium lolii and Balansia epichloe, respectively) were investigated by growing the grasses under 350 and 650 l l^– 1 CO₂ at two nutrient levels. Infected and uninfected perennial ryegrass responded with increased growth to both CO₂ enrichment and nutrient addition. Biomass and leaf area of infected and uninfected plants responded similarly to CO₂ enrichment. When growth analysis parameters were calculated, there were significant increases in relative growth rate and net assimilation rate of infected plants compared to uninfected plants, although the differences remained constant across CO₂ and nutrient treatments. Growth of purpletop grass did not increase with CO₂ enrichment or nutrient addition and there were no significant differences between infected and uninfected plants. CO₂ enrichment did not alter the interactions between these two host grasses and their endophytic-fungal symbionts.     

地质封存二氧化碳与深地微生物相互作用研究进展

地质封存将工业和能源相关领域生产活动产生的二氧化碳（CO₂）进行捕集并注入到深部地下岩石构造中,以实现长期储存的目标,是降低温室气体排放、实现CO₂长期封存的重要可行性手段之一。向深部地下地质构造中注入大量CO₂会导致深地环境发生显著变化,进而引起原生微生物活性及群落结构发生明显改变。因此,地质封存CO₂能够直接或间接影响深地微生物驱动的生物地球化学过程。同时,微生物在短期和长期的超临界CO₂（scCO₂）胁迫作用下,也会通过不同的适应性进化方式影响CO₂在地下环境中的迁移、转化和赋存形态。本文介绍了国内外二氧化碳捕获与封存发展现状以及地质封存CO₂影响条件下的scCO₂-水-微生物-矿物的相互作用领域的最新科研进展,并展望了利用深地微生物强化CO₂固定以及将其转化为高附加值产物的潜力。     

Catalytic effect of water,water dimer,HCOOH and H2SO4 on the isomerisation of HON(O)NNO2 to ON(OH)NNO2: a mechanism study

ABSTRACT

In this article, we report a theoretical investigation on the role of several catalysts in the isomerisation mechanisms of HON(O)NNO₂ to ON(OH)NNO₂ by theoretical method of CBS-QB3. The isomerisation reactions with catalyst X (X?=?H₂O, (H₂O)₂, HCOOH and H₂SO₄) are multi-hydrogen atom transfer reactions. Compared to the isomerisation mechanisms and rate constant of HON(O)NNO₂ to ON(OH)NNO₂ without catalysts, incorporation of the catalyst X shows different positive catalytic effects on affecting the reaction processes, with the H₂SO₄-assisted reaction being the most favourable. Such different catalytic effects are mainly related to the size of the ring structure in X-assisted transition states and the different values of pK_a and proton affinities for HCOOH and H₂SO₄. Besides, compared with the barrier height of the isomerisation process from HON(O)NNO₂ to ON(OH)NNO₂ with HN(NO₂)₂ and HON(O)NNO₂, the barrier of H₂SO₄-assisted reaction is lower by 9.3 and 4.5?kcal?·mol^?1, meanwhile, the rate constant of H₂SO₄ catalyzed is larger than water and water dimer–assisted by 3–5 and 2–3 orders of magnitude, respectively. So, H₂SO₄-assisted reaction is the most favourable.     

青藏高原高寒草甸土壤CO2排放对模拟氮沉降的早期响应

研究大气氮沉降输入对青藏高原高寒草甸土壤-大气界面CO₂交换通量的影响,对于准确评价全球变化背景下区域碳平衡至关重要。通过构建多形态、低剂量的增氮控制试验,利用静态箱-气相色谱法测定土壤CO₂排放通量,同时测定相关土壤变量和地上生物量,分析高寒草甸土壤CO₂排放特征及其主要驱动因子。研究结果表明:低、高剂量氮输入倾向于消耗土壤水分,而中剂量氮输入有利于土壤水分的保持;施氮初期总体上增加了土壤无机氮含量,铵态氮累积效应更为显著;施氮显著增加地上生物量和土壤CO₂排放通量,铵态氮的促进效应显著高于硝态氮。另外,土壤CO₂排放通量主要受土壤温度驱动,其次为地上生物量和铵态氮储量。上述结果反映了氮沉降输入短期内可能刺激了植物生长和土壤微生物活性,加剧了土壤-大气界面CO₂排放。     

Competitive Formation of Hydroxycarbonate Green Rust 1 versus Hydroxysulphate Green Rust 2 in Shewanella putrefaciens Cultures

The formation of hydroxysulphate green rust 2, a Fe(II-III) compound commonly found during corrosion processes of iron-based materials in seawater, has not yet been reported in bacterial cultures. Here we used Shewanella putrefaciens, a dissimilatory iron-reducing bacterium to anaerobically catalyze the transformation of a ferric oxyhydroxide, lepidocrocite (γ-FeOOH), into Fe(II) in the presence of various sulphate concentrations. Biotransformation assays of γ-FeOOH were performed with formate as the electron donor under a variety of concentrations. The results showed that the competitive formation of hydroxycarbonate green rust 1 (GR1(CO₃ ^2?)) and hydroxysulphate green rust 2 (GR2(SO₄ ^{2 ?})) depended upon the relative ratio (R) of bicarbonate and sulphate concentrations. When R ≥ 0.17, GR1(CO₃ ^{2 ?}) only was formed whereas when R < 0.17, a mixture of GR2(SO₄ ^{2 ?}) and GR1(CO₃ ^{2 ?}) was obtained. These results demonstrated that the hydroxysulphate GR2 can originate from the microbial reduction of γ-FeOOH and confirmed the preference for carbonate over sulphate during green rust precipitation. The solid phases were characterized by X-ray diffraction, transmission Mössbauer spectroscopy and scanning electron microscopy. Diffuse reflectance infrared Fourier transform spectroscopy confirmed the presence of intercalated carbonate and sulphate in green rust's structure. This study sheds light on the influence of dissimilatory iron-reducing bacteria on microbiologically influenced corrosion.     

Elevated CO2 alleviates the effects of low P on the growth of N2-fixing Acacia auriculiformis and Acacia mangium

Nodulated seedlings of Acacia auriculiformis Cunn. ex Benth and Acacia mangium Willd were grown with different phosphorus (P) regimes for 90 days, and half of them were exposed to elevated CO₂ (800 μl l⁻¹) during the last 30 days. Under ambient CO₂, plant growth and the amount of N fixed symbiotically in N₂-fixing seedlings decreased with the decrease of supplied P; this relationship did not occur under elevated CO₂. The increase in plant biomass by elevated CO₂ at low P was accompanied by the increase in internal P use efficiency, the amount of N fixed symbiotically and N use efficiency. Elevated CO₂ recovered the low P-induced reduction in leaf dry matter per unit area or unit fresh weight, but it had no effect on the low P-induced increase in partitioning dry matter to roots. These results suggest that elevated CO₂ alleviates the low P effect mainly by increasing the use efficiency of internal P for plant growth and symbiotic N₂ fixation, and the source-sink relationship is possibly an important driving force for this effect of elevated CO₂ in A. auriculiformis and A. mangium.     

Responses of soil microbiota of a late successional alpine grassland to long term CO2 enrichment

We investigated microbial responses in a late successional sedge-dominated alpine grassland to four seasons of CO₂ enrichment. Part of the plots received fertilizer equivalent to 4.5g N m⁻² a⁻¹. Soil basal respiration (R _mic ), the metabolic quotient for CO₂ (qCO₂=R _mic /C _mic ), microbial C and N (C _mic and N _mic ) as well as total soil organic C and N showed no response to CO₂ enrichment alone. However, when the CO₂ treatment was combined with fertilizer addition R _mic and qCO₂ were statistically significantly higher under elevated CO₂ than under ambient conditions (+57% and +71%, respectively). Fertilizer addition increased microbial N pools by 17%, but this was not influenced by elevated CO₂. Microbial C was neither affected by elevated CO₂ nor fertilizer. The lack of a CO₂-effect in unfertilized plots was suprising in the light of our evidence (based on C balance) that enhanced soil C inputs must have occurred under elevated CO₂ regardless of fertilizer treatment. Based on these data and other published work we suggest that microbial responses to elevated CO₂ in such stable, late-successional ecosystems are limited by the availability of mineral nutrients and that results obtained with fertile or heavily disturbed substrates are unsuitable to predict future microbial responses to elevated CO₂ in natural systems. However, when nutrient limitation is removed (e.g. by wet nitrogen deposition) microbes make use of the additional carbon introduced into the soil system. We believe that the response of natural ecosystems to elevated CO₂ must be studied in situ in natural, undisturbed systems.     

Photosynthetic CO<Subscript>2</Subscript> uptake in seedlings of two tropical tree species exposed to oscillating elevated concentrations of CO<Subscript>2</Subscript>

Do short-term fluctuations in CO₂ concentrations at elevated CO₂ levels affect net CO₂ uptake rates of plants? When exposed to 600 μl CO₂ l^?1, net CO₂ uptake rates in shoots or leaves of seedlings of two tropical C₃ tree species, teak (Tectona grandis L. f.) and barrigon [Pseudobombax septenatum (Jacq.) Dug.], increased by 28 and 52% respectively. In the presence of oscillations with half-cycles of 20 s, amplitude of ca. 170 μl CO₂ l^?1 and mean of 600 μl CO₂ l^?1, the stimulation in net CO₂ uptake by the two species was reduced to 19 and 36%, respectively, i.e. the CO₂ stimulation in photosynthesis associated with a change in exposure from 370 to 600 μl CO₂ l^?1 was reduced by a third in both species. Similar reductions in CO₂-stimulated net CO₂ uptake were observed in T. grandis exposed to 40-s oscillations. Rates of CO₂ efflux in the dark by whole shoots of T. grandis decreased by 4.8% upon exposure of plants grown at 370 μl CO₂ l^?1 to 600 μl CO₂ l^?1. The potential implications of the observations on CO₂ oscillations and dark respiration are discussed in the context of free-air CO₂ enrichment (FACE) systems in which short-term fluctuations of CO₂ concentration are a common feature.