Bicarbonate Assimilation Research Articles

Insight into the potential mechanism of bicarbonate assimilation promoted by mixotrophic in CO2 absorption and microalgae conversion system

CO2 absorption-microalgae conversion (CAMC) system is a promising carbon capture and utilization technology. However, the use of HCO3− as a carbon source often led to a slower growth rate of microalgae, which also limited the application of CAMC system. In this study, the assimilation efficiency of HCO3− in CAMC system was improved through mixotrophic, and the potential mechanism was investigated. The HCO3− assimilation efficiency and biomass under mixotrophic were 34.79% and 31.76% higher than that of control. Mixotrophic increased chlorophyll and phycocyanin content, which were beneficial to capture more light energy. The content of ATP and NADPH reached 566.86 μmol/gprot and 672.86 nmol/mgprot, which increased by 31.83% and 27.67% compared to autotrophic. The activity of carbonic anhydrase and Rubisco increased by 18.52% and 22.08%, respectively. Transcriptome showed that genes related to photosynthetic and respiratory electron transport were up-regulated. The synergy of photophosphorylation and oxidative phosphorylation greatly improved energy metabolism efficiency, thus accelerating the assimilation of HCO3−. These results revealed a potential mechanism of promoting the HCO3− assimilation under mixotrophic, it also provided a guidance for using CAMC system to serve carbon neutrality.

An improved carbon fixation management strategy into the crop–soil ecosystem by using biomass ash as the medium

Returning the carbonated biomass ash into the soil can enhance the carbon fixation capacity of crop–soil ecosystem, contributing to the achievement of ‘4 per mille Soils for Food Security and Climate’ goal launched at COP21. The CO 2 sequestration of crop–soil ecosystem induced by biomass ash, however, highly depended on the soil pH and crops growth, which have not​ been elucidated. Here, we explored the effects of different management strategies (watering, chemical fertilizer, and CO 2 -rich biogas slurry on the CO 2 fixation performance of tomato ecosystem (Tom-E) containing one tomato plant, biomass ash, and soil, and evaluated the impact of tomato cultivation concerning carbon fixation. Results showed that when burying biomass ash into the soil, biomass ash did not cause physiological stress on tomato growth, and even improved the tomato qualities. Furthermore, through sequestrating CO 2 from biomass ash, CO 2 fixation capacity of the tomato ecosystem reached about 8.8 ± 1.10, 13.7 ± 0.74, and 20.7 ± 0.53 mmol-CO 2 /Tom-E for watering, chemical fertilizer, and CO 2 -rich biogas slurry management strategies, respectively. However, due to the influence of fertilization management strategy, about 57.4% ± 0.09 and 27.3% ± 0.09 of CO 2 sequestrated in the biomass ash returned to the atmosphere ( i.e , CO 2 loss) for only watering treatment and conventional fertilization (chemical fertilizer) treatment, respectively. Comparatively, as a pH buffering fertilizer, CO 2 -rich biogas slurry application improved the bicarbonate assimilation environment of tomato, which contributed to reducing the CO 2 loss to only 12.5% ± 0.01. Clearly, adopting the carbonated biomass ash and CO 2 -rich biogas slurry as fertilizer provides a suitable and potential strategy for the enhancement of soil carbon sink. • Recycling the biomass ash into soil enhanced the carbon sink of tomato ecosystem. • CO 2 -rich biogas slurry application improved the carbon sink of tomato ecosystem. • DIC assimilation rate of tomato increased by recycling the biomass ash. • Different fertilization management strategies caused different CO 2 sequestration.

N-C27 Predominance and 13C enrichment by bicarbonate assimilation of race A of Botryococcus braunii in Chinese Maoming oilshales

In Maoming oilshales, the presence of races B and L of Botryococcus braunii (B. braunii) was confirmed by the detection of botryococcanes and C40 lycopanes and their derivatives. However, there were no specific biomarkers for race A of B. braunii. The contribution of race A of B. braunii has been implied by the distribution of non-specific n-alkanes with an odd carbon-number preference and a predominance of n-C27. However, confirmation of its contribution to oilshales has been lacking until now. We found that n-C27 is a dominant n-alkane in most Maoming samples which exhibits a positive δ13C excursion (approximately 2–3‰) compared to other homologues. In addition, the extremely heavy δ13C values of botryococcanes (−6.0‰ to −8.5‰) are rationalized to reflect bicarbonate assimilation in race B of B. braunii. The positive excursion of the δ13C values of n-C27 extracted from the same samples could be attributed to bicarbonate assimilation by race A in a similar manner. Bicarbonate assimilation by race A would also explain the isotopic enrichment of n-C14 ~ n-C33 alkanes in the range of −20.1‰ to −26.1‰. They are close to the values of some higher plant sources (C4 plant) and largely different than that of the reported algal n-alkanes (<−30.6‰). The bicarbonate consumption mechanism utilized by both races A and B of B. braunii in Maoming oilshales was triggered by the sharp decrease in pCO2 during the Late Eocene period, recorded globally at the Eocene-Oligocene transition. The presence of race A of B. braunii in most of Maoming sediments indicates that it was wide spread and populated in Maoming Basin over a long period of time, serving as the main biological source material of oilshales on a large scale.

Seasonal variation and driving factors of carbon and nitrogen stable isotope values of plankton in four lakes of Yunnan Province

浮游生物是湖泊食物网的重要组成，其碳、氮稳定同位素能够反映元素地球化学循环和食物来源的波动，是了解水域生态系统结构变化的重要手段之一.本文选取云南4个不同类型湖泊，开展浮游生物碳、氮稳定同位素组成（δ¹³C、δ¹⁵N）的季节变化与湖泊对比研究.大型深水湖泊（抚仙湖和阳宗海）中，浮游植物δ¹³C值在夏、秋季（-20.34‰±1.98‰）显著高于冬、春季（-28.00‰±2.51‰），反映夏秋季藻类生长速率较高、HCO₃^-无机碳源利用增多等的影响.而小型浅水湖泊（长桥海和大屯海）中浮游植物δ¹³C值在夏季最高（-21.24‰±0.88‰），可能与雨季流域输入增强、陆源有机质占比增加有关.4个湖泊浮游生物δ¹⁵N值具有一致的变化特征，春季显著高于其他季节.分析表明，云南地区雨季以面源污染为主向旱季以点源污染为主的转变，导致氮素营养盐季节性来源差异，并通过生物吸收作用影响了浮游生物δ¹⁵N值的季节变化.在浮游动物与浮游植物的稳定同位素差值（即富集度）方面，营养水平高的小型浅水湖泊中δ¹³C富集度为1.61‰±0.90‰、δ¹⁵N富集度为2.71‰±1.22‰，显著小于营养水平低的大型深水湖泊（分别为2.60‰±0.98‰和4.19‰±1.25‰），表明随着湖泊营养水平的增加，浮游动物更多地以浮游植物为食，导致有机碳在不同营养级之间的传输过程中具有更强的耦合作用，且相邻营养级之间具有更低的δ¹⁵N富集度特征.;Plankton is an important component of lake food webs. Their carbon and nitrogen stable isotope values can reflect the fluctuation of biogeochemical cycling and food sources and can be applied to track the changes of aquatic ecosystem structure. In this paper, four lakes with distinct typology from Yunnan Province were selected to uncover the seasonal changes in carbon and nitrogen stable isotopic compositions of plankton. The carbon stable isotope values of phytoplankton in large deep lakes (Lake Fuxian and Lake Yangzong) are enriched in summer and autumn (mean ±SD: -20.34‰±1.98‰), but relatively depleted in winter and spring (-28.00‰±2.51‰), which may reflect a higher growth rate of plankton and greater assimilation of bicarbonate (HCO₃^-) during the growing season. In small shallow lakes (Lake Changqiao and Lake Datun), carbon stable isotopic values of phytoplankton are mostly enriched in summer (-21.24‰±0.88‰) among the four sampling seasons. This may be related to the increased input of terrigenous organic matter during the rainy season, and a decreased lower water level due to overuse of agricultural irrigation in summer. In contrast to the carbon stable isotope values, the nitrogen stable isotope composition of plankton displayed a similar trend of seasonal variations across lakes where a significantly more positive value was found in spring than in other seasons. Further analyses revealed that that seasonal variation in the nitrogen stable isotope of plankton is related to the seasonal difference of nitrogen sources and their distinct isotopic compositions. Specifically, non-point source pollution, mainly related to agricultural fertilizers with depleted nitrogen stable isotopic value, is often dominant in the rainy season, while point source pollution predominantly derived from sewage input with enriched nitrogen stable isotopic composition is commonly observed in the dry season. In terms of stable isotope fractionation between zooplankton and phytoplankton (i.e. trophic enrichment), small shallow lakes were characterized by smaller carbon (1.61‰±0.90‰) and nitrogen (2.71‰±1.22‰) stable isotopic enrichment than those in large deep lakes (2.60‰±0.98‰ and 4.19‰±1.25‰, respectively) in this study. Zooplankton often displays a higher degree of dependence on autochthonous organic carbon (such as phytoplankton) with increasing nutrient status, reflecting a stronger coupling of organic carbon transfer between trophic levels. Similarly, a smaller degree of nitrogen stable isotope enrichment is evidenced in more eutrophic lakes in this study.

Innovative large-scale photobioreactor for coal propelled power plant effluents treatment

This paper presents a phycoremediation technology application for effective wastewater treatment of two Israeli coal propelled power plants, one in Ashkelon (Rothenberg) and the second in Hadera (Orot Rabin). The technology is based on a new photobioreactor (PBR) at the heart of a biological integrative pollution prevention and control (Bio-IPPC) system. Unlike other known tubular and flat plate PBRs that are applicable in small Wastewater Treatment Plants (WWTP), this special PBR can be applied in large-scale domestic and industrial WWTPs. The unique PBR interior configuration, which includes multiple high porous polypropylene beds, enables the system to handle pollutants from both gaseous and aquatic effluents. Furthermore, while most existing algal-based technologies preserve cultured strains, this setup relies on WWTP habitat endemic flora. In the pilots of both power plants, the native biomass was single cyanobacteria Synechocystis salina, although the PBR photosynthetic composition can include multiple species of mixotrophic and/or autotrophic microorganisms. The results show that the photosynthetic biomass that grew on the PBR PP beds effectively assimilated the biomass nutrients such as nitrogen, phosphorous, magnesium, boron and calcium, and the toxic elements strontium and fluoride. Moreover, the system also mitigated greenhouse gas emission by the assimilation of bicarbonate (HCO3−) from the aquatic phase and CO2 from the gas phase.

"Candidatus Nitrosotenuis aquarius," an Ammonia-Oxidizing Archaeon from a Freshwater Aquarium Biofilter.

Ammonia is a metabolic waste product excreted by aquatic organisms that causes toxicity when it accumulates. Aquaria and aquaculture systems therefore use biological filters that promote the growth of nitrifiers to convert ammonia to nitrate. Ammonia-oxidizing bacteria (AOB) have been isolated from aquarium biofilters and are available as commercial supplements, but recent evidence suggests that ammonia-oxidizing archaea (AOA) are abundant in aquarium biofilters. In this study, we report the cultivation and closed genome sequence of the novel AOA representative "Candidatus Nitrosotenuis aquarius," which was enriched from a freshwater aquarium biofilter. "Ca Nitrosotenuis aquarius" oxidizes ammonia stoichiometrically to nitrite with a concomitant increase in thaumarchaeotal cells and a generation time of 34.9 h. "Ca Nitrosotenuis aquarius" has an optimal growth temperature of 33°C, tolerates up to 3 mM NH4Cl, and grows optimally at 0.05% salinity. Transmission electron microscopy revealed that "Ca Nitrosotenuis aquarius" cells are rod shaped, with a diameter of ∼0.4 μm and length ranging from 0.6 to 3.6 μm. In addition, these cells possess surface layers (S-layers) and multiple proteinaceous appendages. Phylogenetically, "Ca Nitrosotenuis aquarius" belongs to the group I.1a Thaumarchaeota, clustering with environmental sequences from freshwater aquarium biofilters, aquaculture systems, and wastewater treatment plants. The complete 1.70-Mbp genome contains genes involved in ammonia oxidation, bicarbonate assimilation, flagellum synthesis, chemotaxis, S-layer production, defense, and protein glycosylation. Incubations with differential inhibitors indicate that "Ca Nitrosotenuis aquarius"-like AOA contribute to ammonia oxidation within the aquarium biofilter from which it originated.IMPORTANCE Nitrification is a critical process for preventing ammonia toxicity in engineered biofilter environments. This work describes the cultivation and complete genome sequence of a novel AOA representative enriched from a freshwater aquarium biofilter. In addition, despite the common belief in the aquarium industry that AOB mediate ammonia oxidation, the present study suggests an in situ role for "Ca Nitrosotenuis aquarius"-like AOA in freshwater aquarium biofilters.

Eastern Mediterranean hydroclimate reconstruction over the last 3600 years based on sedimentary n-alkanes, their carbon and hydrogen isotope composition and XRF data from the Gialova Lagoon, SW Greece

Understanding past hydroclimate variability and related drivers is essential to improve climate forecasting capabilities especially in areas with high climatic sensitivity, such as the Mediterranean. This can be achieved by using a broad spectrum of high resolution, multiple proxy records which can also allow us to assess linkages between regional hydroclimate variability and shifts in the large-scale atmospheric patterns. Here, we present a multiproxy reconstruction of the central-eastern Mediterranean hydroclimate changes over the last 3600 years based on a sediment core from the Gialova Lagoon, a shallow coastal ecosystem in SW Peloponnese, Greece. Our combined dataset consists of the distribution and compound-specific carbon and hydrogen isotope (δ13C and δD) composition of n-alkanes, bulk organic matter properties and X-ray fluorescence (XRF) core scanning data. This approach was complemented with a semi-quantitative analysis of plant remains in the core. The results indicate a high contribution of local aquatic vegetation to organic matter. Large δ13C variations in predominantly aquatic plant-derived mid-chain alkanes (C23-25) mainly reflect changes in the aquatic plant abundance and their carbon source. Our data suggest that higher δ13C23-25 values (up to −19‰) largely correspond to expansion of aquatic vegetation during wet and/or cold periods causing carbon-limiting conditions in the water and assimilation of isotopically-enriched bicarbonate by the plants. The δD records of the individual n-alkanes (C17 to C31) exhibit a nearly identical pattern to each other, which implies that they all reflect changes in the source water isotope composition, driven by hydroclimate variability. In addition, the δD profiles are consistent with the XRF data with both proxies being driven by a common hydroclimate signal. We observe two major shifts from dry and/or warm periods at ca 3600-3000 cal BP and ca 1700-1300 cal BP to wet and/or cold episodes at ca 3000-2700 cal BP and ca 1300-900 cal BP. The period ca 700-200 cal BP is the wettest and/or coldest in our record and coeval with the Little Ice Age. The climatic fluctuation reported in this study can be explained by the relative dominance of high-latitude (e.g. North Atlantic Oscillation during winters) and the low-latitude atmospheric patterns (Intertropical convergence zone, Subtropical High and the effects of Asian monsoons during summers) which suggests an Atlantic-Mediterranean-Monsoon climate link in this area for the late Holocene.

Augmenting the Calvin\u2013Benson\u2013Bassham cycle by a synthetic malyl-CoA-glycerate carbon fixation pathway

The Calvin–Benson–Bassham (CBB) cycle is presumably evolved for optimal synthesis of C3 sugars, but not for the production of C2 metabolite acetyl-CoA. The carbon loss in producing acetyl-CoA from decarboxylation of C3 sugar limits the maximum carbon yield of photosynthesis. Here we design a synthetic malyl-CoA-glycerate (MCG) pathway to augment the CBB cycle for efficient acetyl-CoA synthesis. This pathway converts a C3 metabolite to two acetyl-CoA by fixation of one additional CO2 equivalent, or assimilates glyoxylate, a photorespiration intermediate, to produce acetyl-CoA without net carbon loss. We first functionally demonstrate the design of the MCG pathway in vitro and in Escherichia coli. We then implement the pathway in a photosynthetic organism Synechococcus elongates PCC7942, and show that it increases the intracellular acetyl-CoA pool and enhances bicarbonate assimilation by roughly 2-fold. This work provides a strategy to improve carbon fixation efficiency in photosynthetic organisms.

Insights into the Fundamental Physiology of the Uncultured Fe-Oxidizing Bacterium Leptothrix ochracea.

Leptothrix ochracea is known for producing large volumes of iron oxyhydroxide sheaths that alter wetland biogeochemistry. For over a century, these delicate structures have fascinated microbiologists and geoscientists. Because L. ochracea still resists long-term in vitro culture, the debate regarding its metabolic classification dates back to 1885. We developed a novel culturing technique for L. ochracea using in situ natural waters and coupled this with single-cell genomics and nanoscale secondary-ion mass spectrophotometry (nanoSIMS) to probe L. ochracea's physiology. In microslide cultures L. ochracea doubled every 5.7 h and had an absolute growth requirement for ferrous iron, the genomic capacity for iron oxidation, and a branched electron transport chain with cytochromes putatively involved in lithotrophic iron oxidation. Additionally, its genome encoded several electron transport chain proteins, including a molybdopterin alternative complex III (ACIII), a cytochrome bd oxidase reductase, and several terminal oxidase genes. L. ochracea contained two key autotrophic proteins in the Calvin-Benson-Bassham cycle, a form II ribulose bisphosphate carboxylase, and a phosphoribulose kinase. L. ochracea also assimilated bicarbonate, although calculations suggest that bicarbonate assimilation is a small fraction of its total carbon assimilation. Finally, L. ochracea's fundamental physiology is a hybrid of those of the chemolithotrophic Gallionella-type iron-oxidizing bacteria and the sheathed, heterotrophic filamentous metal-oxidizing bacteria of the Leptothrix-Sphaerotilus genera. This allows L. ochracea to inhabit a unique niche within the neutrophilic iron seeps.IMPORTANCELeptothrix ochracea was one of three groups of organisms that Sergei Winogradsky used in the 1880s to develop his hypothesis on chemolithotrophy. L. ochracea continues to resist cultivation and appears to have an absolute requirement for organic-rich waters, suggesting that its true physiology remains unknown. Further, L. ochracea is an ecological engineer; a few L. ochracea cells can generate prodigious volumes of iron oxyhydroxides, changing the ecosystem's geochemistry and ecology. Therefore, to determine L. ochracea's basic physiology, we employed new single-cell techniques to demonstrate that L. ochracea oxidizes iron to generate energy and, despite having predicted genes for autotrophic growth, assimilates a fraction of the total CO2 that autotrophs do. Although not a true chemolithoautotroph, L. ochracea's physiological strategy allows it to be flexible and to extensively colonize iron-rich wetlands.

Contribution of Bicarbonate Assimilation to Carbon Pool Dynamics in the Deep Mediterranean Sea and Cultivation of Actively Nitrifying and CO2-Fixing Bathypelagic Prokaryotic Consortia.

Covering two-thirds of our planet, the global deep ocean plays a central role in supporting life on Earth. Among other processes, this biggest ecosystem buffers the rise of atmospheric CO2. Despite carbon sequestration in the deep ocean has been known for a long time, microbial activity in the meso- and bathypelagic realm via the “assimilation of bicarbonate in the dark” (ABD) has only recently been described in more details. Based on recent findings, this process seems primarily the result of chemosynthetic and anaplerotic reactions driven by different groups of deep-sea prokaryoplankton. We quantified bicarbonate assimilation in relation to total prokaryotic abundance, prokaryotic heterotrophic production and respiration in the meso- and bathypelagic Mediterranean Sea. The measured ABD values, ranging from 133 to 370 μg C m−3 d−1, were among the highest ones reported worldwide for similar depths, likely due to the elevated temperature of the deep Mediterranean Sea (13–14°C also at abyssal depths). Integrated over the dark water column (≥200 m depth), bicarbonate assimilation in the deep-sea ranged from 396 to 873 mg C m−2 d−1. This quantity of produced de novo organic carbon amounts to about 85–424% of the phytoplankton primary production and covers up to 62% of deep-sea prokaryotic total carbon demand. Hence, the ABD process in the meso- and bathypelagic Mediterranean Sea might substantially contribute to the inorganic and organic pool and significantly sustain the deep-sea microbial food web. To elucidate the ABD key-players, we established three actively nitrifying and CO2-fixing prokaryotic enrichments. Consortia were characterized by the co-occurrence of chemolithoautotrophic Thaumarchaeota and chemoheterotrophic proteobacteria. One of the enrichments, originated from Ionian bathypelagic waters (3,000 m depth) and supplemented with low concentrations of ammonia, was dominated by the Thaumarchaeota “low-ammonia-concentration” deep-sea ecotype, an enigmatic and ecologically important group of organisms, uncultured until this study.

Root-derived bicarbonate assimilation in response to variable water deficit in Camptotheca acuminate seedlings.

Water deficit is one of the key factors that limits the carbon (C) assimilation and productivity of plants. The effect of variable water deficit on recently root-derived bicarbonate assimilation in Camptotheca acuminate seedlings was investigated. Three-month-old seedlings were subjected to three water regimes, well-watered (WW), moderate stress (MS), and severe stress (SS) induced by polyethyleneglycol, in conjunction with relatively high (H) and low (L) natural 13C-abundance of NaHCO3-labeled treatments in hydroponics for 14 days. The δ13C of the newly expanded leaves in H were generally more enriched in heavy isotopes than were those in L, indicative of the involvement of bicarbonate in aboveground tissues. The C isotope fractionation of newly expanded leaves relative to air (∆13Cair-leaves) ranged from 17.78 to 21.78‰ among the treatments. The ∆13Cair-leaves under the MS and SS treatments in H were both more negative than was that in L. A linear regression between Ci/Ca and ∆13Cair-leaves in both L and H were different from the theoretical regression. On the basis of the two end-member mixing model, the proportion of fixed CO2 supplied from bicarbonate contributing to the total photosynthetically inorganic C assimilation were 10.34, 20.05 and 16.60% under the WW, MS, and SS treatments, respectively. These results indicated that the increase in water deficit decreased the atmospheric CO2 gain but triggered a compensatory use of bicarbonate in C. acuminate seedlings.

Quantification of photosynthetic inorganic carbon utilisation via a bidirectional stable carbon isotope tracer

The amount of bicarbonate utilised by plants is usually ignored because of limited measurement methods. Accordingly, this study quantified the photosynthetic assimilation of inorganic carbon (CO2 and HCO3−) by plants. The net photosynthetic CO2 assimilation (PN), the photosynthetic assimilation of CO2 and bicarbonate (PN’), the proportion of increased leaf area (fLA) and the stable carbon isotope composition (δ13C) of Orychophragmus violaceus (Ov) and Brassica juncea (Bj) under three bicarbonate levels (5, 10 and 15 mm NaHCO3) were examined to determine the relationship among PN, PN’ and fLA. PN’, not PN, changed synchronously with fLA. Moreover, the proportions of exogenous bicarbonate and total bicarbonate (including exogenous bicarbonate and dissolved CO2-generated bicarbonate) utilised by Ov were 2.27 % and 5.28 % at 5 mm bicarbonate, 7.06 % and 13.28 % at 10 mm bicarbonate, and 8.55 % and 17.31 % at 15 mm bicarbonate, respectively. Meanwhile, the proportions of exogenous bicarbonate and total bicarbonate utilised by Bj were 1.77 % and 3.28 % at 5 mm bicarbonate, 2.11 % and 3.10 % at 10 mm bicarbonate, and 2.36 % and 3.09 % at 15 mm bicarbonate, respectively. Therefore, the dissolved CO2-generated bicarbonate and exogenous bicarbonate are important sources of inorganic carbon for plants.

Elemental sulfur and acetate can support life of a novel strictly anaerobic haloarchaeon.

Archaea domain is comprised of many versatile taxa that often colonize extreme habitats. Here, we report the discovery of strictly anaerobic extremely halophilic euryarchaeon, capable of obtaining energy by dissimilatory reduction of elemental sulfur using acetate as the only electron donor and forming sulfide and CO2 as the only products. This type of respiration has never been observed in hypersaline anoxic habitats and is the first example of such metabolic capability in the entire Archaea domain. We isolated and cultivated these unusual organisms, selecting one representative strain, HSR2, for detailed characterization. Our studies including physiological tests, genome sequencing, gene expression, metabolomics and [(14)C]-bicarbonate assimilation assays revealed that HSR2 oxidized acetate completely via the tricarboxylic acid cycle. Anabolic assimilation of acetate occurred via activated glyoxylate bypass and anaplerotic carboxylation. HSR2 possessed sulfurtransferase and an array of membrane-bound polysulfide reductase genes, all of which were expressed during the growth. Our findings suggest the biogeochemical contribution of haloarchaea in hypersaline anoxic environments must be reconsidered.

Acetate and bicarbonate assimilation and metabolite formation in Chlamydomonas reinhardtii: a 13C-NMR study.

Cellular metabolite analyses by 13C-NMR showed that C. reinhardtii cells assimilate acetate at a faster rate in heterotrophy than in mixotrophy. While heterotrophic cells produced bicarbonate and CO2 aq, mixotrophy cells produced bicarbonate alone as predominant metabolite. Experiments with singly 13C-labelled acetate (13CH3-COOH or CH3-13COOH) supported that both the 13C nuclei give rise to bicarbonate and CO2 aq. The observed metabolite(s) upon further incubation led to the production of starch and triacylglycerol (TAG) in mixotrophy, whereas in heterotrophy the TAG production was minimal with substantial accumulation of glycerol and starch. Prolonged incubation up to eight days, without the addition of fresh acetate, led to an increased TAG production at the expense of bicarbonate, akin to that of nitrogen-starvation. However, such TAG production was substantially high in mixotrophy as compared to that in heterotrophy. Addition of mitochondrial un-coupler blocked the formation of bicarbonate and CO2 aq in heterotrophic cells, even though acetate uptake ensued. Addition of PSII-inhibitor to mixotrophic cells resulted in partial conversion of bicarbonate into CO2 aq, which were found to be in equilibrium. In an independent experiment, we have monitored assimilation of bicarbonate via photoautotrophy and found that the cells indeed produce starch and TAG at a much faster rate as compared to that in mixotrophy and heterotrophy. Further, we noticed that the accumulation of starch is relatively more as compared to TAG. Based on these observations, we suggest that acetate assimilation in C. reinhardtii does not directly lead to TAG formation but via bicarbonate/CO2 aq pathways. Photoautotrophic mode is found to be the best growth condition for the production of starch and TAG and starch in C. reinhardtii.

Chemolithotrophic primary production in a subglacial ecosystem.

Glacial comminution of bedrock generates fresh mineral surfaces capable of sustaining chemotrophic microbial communities under the dark conditions that pervade subglacial habitats. Geochemical and isotopic evidence suggests that pyrite oxidation is a dominant weathering process generating protons that drive mineral dissolution in many subglacial systems. Here, we provide evidence correlating pyrite oxidation with chemosynthetic primary productivity and carbonate dissolution in subglacial sediments sampled from Robertson Glacier (RG), Alberta, Canada. Quantification and sequencing of ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) transcripts suggest that populations closely affiliated with Sideroxydans lithotrophicus, an iron sulfide-oxidizing autotrophic bacterium, are abundant constituents of microbial communities at RG. Microcosm experiments indicate sulfate production during biological assimilation of radiolabeled bicarbonate. Geochemical analyses of subglacial meltwater indicate that increases in sulfate levels are associated with increased calcite and dolomite dissolution. Collectively, these data suggest a role for biological pyrite oxidation in driving primary productivity and mineral dissolution in a subglacial environment and provide the first rate estimate for bicarbonate assimilation in these ecosystems. Evidence for lithotrophic primary production in this contemporary subglacial environment provides a plausible mechanism to explain how subglacial communities could be sustained in near-isolation from the atmosphere during glacial-interglacial cycles.

Heterotrophic bicarbonate assimilation is the main process of de novo organic carbon synthesis in hadal zone of the Hellenic Trench, the deepest part of Mediterranean Sea.

Ammonium-oxidizing chemoautotrophic members of Thaumarchaea are proposed to be the key players in the assimilation of bicarbonate in the dark (ABD). However, this process may also involve heterotrophic metabolic pathways, such as fixation of carbon dioxide (CO2) via various anaplerotic reactions. We collected samples from the depth of 4900 m at the Matapan-Vavilov Deep (MVD) station (Hellenic Trench, Eastern Mediterranean) and used the multiphasic approach to study the ABD mediators in this deep-sea ecosystem. At this depth, our analysis indicated the occurrence of actively CO2-fixing heterotrophic microbial assemblages dominated by Gammaproteobacteria with virtually no Thaumarchaea present. [14C]-bicarbonate incorporation experiments combined with shotgun [14C]-proteomic analysis identified a series of proteins of gammaproteobacterial origin. More than quarter of them were closely related with Alteromonas macleodii ‘deep ecotype’ AltDE, the predominant organism in the microbial community of MVD. The present study demonstrated that in the aphotic/hadal zone of the Mediterranean Sea, the assimilation of bicarbonate is associated with both chemolithoauto- and heterotrophic ABD. In some deep-sea areas, the latter may predominantly contribute to the de novo synthesis of organic carbon which points at the important and yet underestimated role heterotrophic bacterial populations can play the in global carbon cycle/sink in the ocean interior.

Microbial life in the Lake Medee, the largest deep-sea salt-saturated formation

Deep-sea hypersaline anoxic lakes (DHALs) of the Eastern Mediterranean represent some of the most hostile environments on our planet. We investigated microbial life in the recently discovered Lake Medee, the largest DHAL found to-date. Medee has two unique features: a complex geobiochemical stratification and an absence of chemolithoautotrophic Epsilonproteobacteria, which usually play the primary role in dark bicarbonate assimilation in DHALs interfaces. Presumably because of these features, Medee is less productive and exhibits reduced diversity of autochthonous prokaryotes in its interior. Indeed, the brine community almost exclusively consists of the members of euryarchaeal MSBL1 and bacterial KB1 candidate divisions. Our experiments utilizing cultivation and [14C]-assimilation, showed that these organisms at least partially rely on reductive cleavage of osmoprotectant glycine betaine and are engaged in trophic cooperation. These findings provide novel insights into how prokaryotic communities can adapt to salt-saturated conditions and sustain active metabolism at the thermodynamic edge of life.

Enrichment and Characterization of an Autotrophic Ammonia-Oxidizing Archaeon of Mesophilic Crenarchaeal Group I.1a from an Agricultural Soil

Soil nitrification is an important process for agricultural productivity and environmental pollution. Though one cultivated representative of ammonia-oxidizing Archaea from soil has been described, additional representatives warrant characterization. We describe an ammonia-oxidizing archaeon (strain MY1) in a highly enriched culture derived from agricultural soil. Fluorescence in situ hybridization microscopy showed that, after 2 years of enrichment, the culture was composed of >90% archaeal cells. Clone libraries of both 16S rRNA and archaeal amoA genes featured a single sequence each. No bacterial amoA genes could be detected by PCR. A [¹³C]bicarbonate assimilation assay showed stoichiometric incorporation of ¹³C into Archaea-specific glycerol dialkyl glycerol tetraethers. Strain MY1 falls phylogenetically within crenarchaeal group I.1a; sequence comparisons to "Candidatus Nitrosopumilus maritimus" revealed 96.9% 16S rRNA and 89.2% amoA gene similarities. Completed growth assays showed strain MY1 to be chemoautotrophic, mesophilic (optimum at 25°C), neutrophilic (optimum at pH 6.5 to 7.0), and nonhalophilic (optimum at 0.2 to 0.4% salinity). Kinetic respirometry assays showed that strain MY1's affinities for ammonia and oxygen were much higher than those of ammonia-oxidizing bacteria (AOB). The yield of the greenhouse gas N₂O in the strain MY1 culture was lower but comparable to that of soil AOB. We propose that this new soil ammonia-oxidizing archaeon be designated "Candidatus Nitrosoarchaeum koreensis."

Assessing the influence of the carbon oxidation-reduction state on organic pollutant biodegradation in algal–bacterial photobioreactors

The influence of the carbon oxidation-reduction state (CORS) of organic pollutants on their biodegradation in enclosed algal-bacterial photobioreactors was evaluated using a consortium of enriched wild-type methanotrophic bacteria and microalgae. Methane, methanol and glucose (with CORS -4, -2 and 0, respectively) were chosen as model organic pollutants. In the absence of external oxygen supply, microalgal photosynthesis was not capable of supporting a significant methane and methanol biodegradation due to their high oxygen demands per carbon unit, while glucose was fully oxidized by photosynthetic oxygenation. When bicarbonate was added, removal efficiencies of 37 ± 4% (20 days), 65 ± 4% (11 days) and 100% (2 days) were recorded for CH(4,) CH(3)OH and C(6)H(12)O(6), respectively due to the additional oxygen generated from photosynthetic bicarbonate assimilation. The use of NO(3)(-) instead of NH(4)(+) as nitrogen source (N oxidation-reduction state of +5 vs. -3) resulted in an increase in CH(4) degradation from 0 to 33 ± 3% in the absence of bicarbonate and from 37 ± 4% to 100% in the presence of bicarbonate, likely due to a decrease in the stoichiometric oxygen requirements and the higher photosynthetic oxygen production. Hypothetically, the CORS of the substrates might affect the CORS of the microalgal biomass composition (higher lipid content). However, the total lipid content of the algal-bacterial biomass was 19 ± 7% in the absence and 16 ± 2% in the presence of bicarbonate.

High bicarbonate assimilation in the dark by Arctic bacteria

Although both autotrophic and heterotrophic microorganisms incorporate CO₂ in the dark through different metabolic pathways, this process has usually been disregarded in oxic marine environments. We studied the significance and mediators of dark bicarbonate assimilation in dilution cultures inoculated with winter Arctic seawater. At stationary phase, bicarbonate incorporation rates were high (0.5-2.5 μg C L⁻¹ d⁻¹) and correlated with rates of bacterial heterotrophic production, suggesting that most of the incorporation was due to heterotrophs. Accordingly, very few typically chemoautotrophic bacteria were detected by 16S rRNA gene cloning. The genetic analysis of the biotin carboxylase gene accC putatively involved in archaeal CO₂ fixation did not yield any archaeal sequence, but amplified a variety of bacterial carboxylases involved in fatty acids biosynthesis, anaplerotic pathways and leucine catabolism. Gammaproteobacteria dominated the seawater cultures (40-70% of cell counts), followed by Betaproteobacteria and Flavobacteria as shown by catalyzed reporter deposition fluorescence in situ hybridization (CARDFISH). Both Beta- and Gammaproteobacteria were active in leucine and bicarbonate uptake, while Flavobacteria did not take up bicarbonate, as measured by microautoradiography combined with CARDFISH. Within Gammaproteobacteria, Pseudoalteromonas-Colwellia and Oleispira were very active in bicarbonate uptake (ca. 30 and 70% of active cells, respectively), while the group Arctic96B-16 did not take up bicarbonate. Our results suggest that, potentially, the incorporation of CO₂ can be relevant for the metabolism of specific Arctic heterotrophic phylotypes, promoting the maintenance of their cell activity and/or longer survival under resource depleted conditions.