Nutrient Constraints on Metabolism Affect the Temperature Regulation of Aquatic Bacterial Growth Efficiency

Inorganic nutrient availability and temperature are recognized as major regulators of organic carbon processing by aquatic bacteria, but little is known about how these two factors interact to control bacterial metabolic processes. We manipulated the temperature of boreal humic stream water samples within 0–25°C and measured bacterial production (BP) and respiration (BR) with and without inorganic nitrogen?+?phosphorus addition. Both BP and BR increased exponentially with temperature in all experiments, with Q ₁₀ values varying between 1.2 and 2.4. The bacterial growth efficiency (BGE) showed strong negative relationships with temperature in nutrient-enriched samples and in natural stream water where community-level BP and BR were not limited by nutrients. However, there were no relationships between BGE and temperature in samples where BP and BR were significantly constrained by the inorganic nutrient availability. The results suggest that metabolic responses of aquatic bacterial communities to temperature variations can be strongly dependent on whether the bacterial metabolism is limited by inorganic nutrients or not. Such responses can have consequences for both the carbon flux through aquatic food webs and for the flux of CO₂ from aquatic systems to the atmosphere.     

碳源与氮源限制下细菌代谢调节研究进展

营养限制是微生物最常面临的环境胁迫之一。除了在营养物质匮乏的海洋、冰川、沙漠、深层地表等自然环境中,越来越多的人工环境也出现了营养限制的特征,例如各类微污染水体、提标改造的废水生物处理系统等。基质浓度极大地影响着包括细菌在内的许多微生物的生长、代谢及群落结构,最终导致其功能的改变。为了在营养限制条件下维持生存,微生物首先需感知营养供给的减少,其后通过基因、蛋白质、信号分子、代谢产物等对各代谢过程进行全局调控,最后改变基质亲和力、生长速率、运动能力、形态等以适应营养不足。胞内各种信号物质及其触发的响应是微生物应对营养胁迫的关键。本文分别梳理了以细菌为代表的微生物应对碳源、氮源限制时的关键信号物质、受体蛋白/调控过程及响应结果,并分析了碳氮限制响应过程中的相互作用,以期为极端环境微生物的认识、营养限制条件下微生物的应用,尤其是低浓度污染物生物处理、生物监测等领域提供理论基础。     

微生物对分解底物碳氮磷化学计量的响应和调节机制

微生物分解者的生存和生长策略、群落结构和功能会随着底物化学计量特征而改变, 从而强烈地影响底物的分解速度和元素的周转速率, 进而影响生态系统的功能过程。深入理解微生物生理代谢活动和群落结构与陆地生态系统过程之间的联系及其对全球变化的响应和反馈机理是生态学和全球变化生物学领域值得关注的重大科学命题。该文基于生态化学计量学理论和代谢理论, 首先介绍微生物在陆地生态系统碳氮磷循环中的作用; 然后综述微生物对分解底物化学计量变异性的响应和调节的4种主要机制: (1)调整微生物自身化学计量特征; (2)调整微生物群落结构; (3)产生特定的胞外酶以获取受限制的资源; (4)改变碳氮磷元素利用效率。最后, 通过分析当前研究不足, 提出该领域亟需关注的科学问题有: (1)综合阐明微生物对底物化学计量变化响应的各种机制及其相对重要性; (2)探索全球变化对微生物驱动的碳氮磷循环的影响; (3)探索微生物对底物化学计量变化适应对策的时空变化。     

好氧反硝化细菌碳氮代谢特点及途径的研究进展

好氧反硝化作用的发现打破了反硝化只能在严格厌氧条件下进行的传统认知,为生物脱氮提供了一条新的途径,已成为近些年的研究热点。碳源可为好氧反硝化过程提供能量和电子供体,其代谢难易程度直接影响着好氧反硝化细菌的脱氮效率,因此有必要明确碳源在好氧反硝化脱氮过程中的代谢机理。基于此,本文阐述了好氧反硝化细菌的种类及其对硝态氮与亚硝态氮的代谢途径;系统分析了不同好氧反硝化细菌对碳氮源代谢的差异与代谢机理;综合分析了碳代谢差异对好氧反硝化脱氮过程的影响,并对未来的研究方向进行了展望,旨在深入理解好氧反硝化细菌同时去除碳氮的机理,为提高废水生物脱氮除碳效率提供理论依据。     

Responses and regulation mechanisms of microbial decomposers to substrate carbon,nitrogen, and phosphorus stoichiometry

The survival and growth strategies, community structure and functions of microbial decomposers vary with substrate stoichiometry, which profoundly influences substrate decomposition, turnover, and hence the carbon and nutrient cycles of terrestrial ecosystems. It is crucial to understand the relationships among microbial metabolism, community structure and ecosystem processes of terrestrial ecosystems and their responses and feedbacks to global changes. In this review, we first introduced the significance of microbial decomposers in the carbon, nitrogen, and phosphorus cycles of terrestrial ecosystems from perspectives of ecological stoichiometry and metabolic theories. Then we synthesized four potential mechanisms of microbial response and control on substrate stoichiometric variations, i.e., through (1) modifying microbial stoichiometry, (2) shifting microbial community structure, (3) producing extracellular enzymes to acquire limiting resources, and (4) changing microbial carbon, nitrogen, and phosphor use efficiencies. Finally, we proposed three research directions in this field: (1) to comprehensively explore various microbial mechanisms in response to changes in substrate stoichiometry and the relative importance of these mechanisms; (2) to examine influences of global changes on microbial-driven cycles of carbon, nitrogen, and phosphorus; and (3) to explore spatiotemporal changes in the strategies of microbial adaptation to changes in the substrate stoichiometry.     

施氏假单胞菌氮代谢调控蛋白GlnK与碳信号分子α-酮戊二酸的体外互作研究

细菌中PⅡ蛋白是一类氮代谢调控因子,可通过感知胞内碳氮信号变化调整自身与靶蛋白的相互作用,从而实现对下游基因的级联调控。α-酮戊二酸是胞内碳状态的重要信号分子,前期研究发现PⅡ蛋白可结合α-酮戊二酸感应细胞内的碳状态,但不同菌中二者的结合存在差异。施氏假单胞菌A1501（Pseudomononas stutzeri A1501）只含有1种PⅡ蛋白GlnK,其在A1501固氮调控中发挥重要作用,但具体碳信号感知与转导机制有待进一步探索。基于此,通过大肠杆菌外源表达GlnK蛋白,并通过微量热泳动（microscale thermophoresis,MST）方法研究GlnK蛋白与碳信号分子α-酮戊二酸的体外互作,发现二者可以体外结合,且进一步证实GlnK蛋白的第89位甘氨酸（G89,Gly89）可能与互作相关。研究结果为进一步解析α-酮戊二酸在A1501中的信号转导奠定了基础,也为深入解析固氮菌的碳氮代谢偶联机制提供了理论支持。     

Overlapping binding of PhoP and AfsR to the promoter region of glnR in Streptomyces coelicolor

Growth of soil bacteria is often limited by the availability of essential nutrients such as carbon, nitrogen and phosphate. The reaction to a specific nutrient starvation triggers interconnected responses to equilibrate the metabolism. It is known that PhoP (response regulator involved in phosphate control) specifically binds to several promoters of genes involved in nitrogen metabolism which are also regulated by GlnR (regulator involved in nitrogen control). In this article we report a novel cross-talk between GlnR and the SARP-like regulator, AfsR. AfsR binds to some PhoP-regulated promoters including those of afsS (a small regulatory protein of secondary metabolism), pstS (a component of the phosphate transport system) and phoRP (encoding the two component system itself). We have characterized the regulation exerted upon the nitrogen regulator glnR gene by AfsR, using EMSA and DNase I footprinting assays as well as in vivo expression studies with ΔphoP, ΔafsR and ΔafsR-ΔphoP mutants. Both PhoP and AfsR proteins are able to bind to overlapping regions within the glnR promoter producing different effects. This work demonstrates a cross-talk of three different regulators of both primary and secondary metabolism.     

Microcommunities and microgradients: Linking nutrient regeneration,microbial mutualism,and high sustained aquatic primary production

Nutrient regeneration is essential to sustained primary production in the aquatic environment because of coupled physical and metabolic gradients. The commonly evaluated ecosystem perspective of nutrient regeneration, as is illustrated among planktonic paradigms of lake ecosystems, functions only at macrotemporal and spatial scales. Most inland waters are small and shallow. Consequently, most organic matter of these waters is derived from photosynthesis of emergent, floating-leaved, and submersed higher plants and microflora associated with living substrata and detritus, including sediments, as well as terrestrial sources. The dominant primary productivity of inland aquatic ecosystems is not planktonic, but rather is associated with surfaces. The high sustained rates of primary production among sessile communities are possible because of the intensive internal recycling of nutrients, including carbon. Steep gradients exist within these attached microbial communities that (a) require rapid, intensive recycling of carbon, phosphorus, nitrogen, and other nutrients between producers, particulate and dissolved detritus, and bacteria and protists: (b) augment internal community recycling and losses with small external inputs of carbon and nutrients from the overlying water or from the supporting substrata; and (c) encourage maximal conservation of nutrients. Examples of microenvironmental recycling of carbon, phosphorus, and oxygen among epiphytic, epipelic, and epilithic communities are explained. Recalcitrant dissolved organic compounds from decomposition can serve both as carbon and energy substrates as well as be selectively inhibitory to microbial metabolism and nutrient recycling. Rapid recycling of nutrient and organic carbon within micro-environments operates at all levels, planktonic as well as attached, and is mandatory for high sustained productivity.     

Kinetic flux profiling for quantitation of cellular metabolic fluxes

This protocol enables quantitation of metabolic fluxes in cultured cells. Measurements are based on the kinetics of cellular incorporation of stable isotope from nutrient into downstream metabolites. At multiple time points, after cells are rapidly switched from unlabeled to isotope-labeled nutrient, metabolism is quenched, metabolites are extracted and the extract is analyzed by chromatography-mass spectrometry. Resulting plots of unlabeled compound versus time follow variants of exponential decay, with the flux equal to the decay rate multiplied by the intracellular metabolite concentration. Because labeling is typically fast (t(1/2)相似文献   

Sinorhizobium meliloti metabolism in the root nodule: a proteomic perspective

The proteome of the model symbiotic bacterium, Sinorhizobium meliloti was examined to determine the enzymatic reactions and cell processes that occur when S. meliloti occupies the root nodules of Medicago truncatula and Melilotus alba. The proteomes of the nodule bacteria were compared to that of S. meliloti grown under laboratory cultured conditions as an additional control. All the detectable protein spots on the two-dimensional (2-D) gels between pH 4-7 were analyzed. In total, the identity of proteins in 1545 spots from 2-D gels was determined using peptide mass fingerprinting. There were clear differences in the proteome of nodule bacteria and cultured bacteria and putative nodule-specific and nodule suppressed proteins were identified. The data were analyzed using metabolic pathway prediction programs and used to review the biochemical and genetic studies that had been done previously on S. meliloti over several decades. There was a broad congruency between the proteomic and biochemical data when the overall pathways of central carbon and nitrogen metabolism were considered. A selective suite of ABC-type transporters was present in nodule bacteria that were biased towards the transport of amino acids and inorganic ions (P and Fe) suggesting that a highly specialized nutrient exchange was occurring between the nodule bacteria and the host. Proteins prominent in nodule bacteria were those involved in the pathways for vitamin synthesis and stress-related processes (chaperoning, heat shock, detoxification of reactive oxygen species, regulation of stress and osmo-regulation). Some of these proteins were found only in nodule bacteria. These results show the extent of the shift in metabolism that occurs when S. meliloti invades legume plants and establishes a nitrogen fixing symbiosis.     

人工遮光和营养添加对河流反硝化活性和反硝化细菌群落结构的影响

河流生态系统受到人类活动例如河岸带森林植被毁损和农业活动施肥等的干扰日益加剧,而这些活动使河流接收的光照增多、河流的氮磷营养盐浓度增加。微生物的反硝化作用是河流去除氮的有效途径。在汉江的一级支流金水河上游核心保护区内选取6条溪流开展野外控制实验,利用营养添加模拟河流中营养的增加,遮盖河面来模拟源头溪流的隐蔽状态,来研究河流沉积物中微生物的反硝化作用对光照和营养改变的响应,并利用高通量测序(Mi Seq)技术研究在两种处理下河流沉积物中nir S型反硝化细菌的群落结构变化。结果显示:营养元素添加促进了沉积物的反硝化活性,河面遮盖抑制了沉积物的反硝化活性。营养添加和遮盖两种处理均降低了控制实验区域内脱氯单胞菌属(优势菌属)的相对丰度,同时也降低了该区域nir S型反硝化菌群落的Chao多样性。本研究初步证实了光照增加和河流的营养增加提高了河流沉积物反硝化活性,并为提高河流的脱氮能力提供科学依据。     

Autotrophic and mixotrophic metabolism of an anammox bacterium revealed by in vivo 13C and 2H metabolic network mapping

Anaerobic ammonium-oxidizing (anammox) bacteria mediate a key step in the biogeochemical nitrogen cycle and have been applied worldwide for the energy-efficient removal of nitrogen from wastewater. However, outside their core energy metabolism, little is known about the metabolic networks driving anammox bacterial anabolism and use of different carbon and energy substrates beyond genome-based predictions. Here, we experimentally resolved the central carbon metabolism of the anammox bacterium Candidatus ‘Kuenenia stuttgartiensis’ using time-series ¹³C and ²H isotope tracing, metabolomics, and isotopically nonstationary metabolic flux analysis. Our findings confirm predicted metabolic pathways used for CO₂ fixation, central metabolism, and amino acid biosynthesis in K. stuttgartiensis, and reveal several instances where genomic predictions are not supported by in vivo metabolic fluxes. This includes the use of the oxidative branch of an incomplete tricarboxylic acid cycle for alpha-ketoglutarate biosynthesis, despite the genome not having an annotated citrate synthase. We also demonstrate that K. stuttgartiensis is able to directly assimilate extracellular formate via the Wood–Ljungdahl pathway instead of oxidizing it completely to CO₂ followed by reassimilation. In contrast, our data suggest that K. stuttgartiensis is not capable of using acetate as a carbon or energy source in situ and that acetate oxidation occurred via the metabolic activity of a low-abundance microorganism in the bioreactor’s side population. Together, these findings provide a foundation for understanding the carbon metabolism of anammox bacteria at a systems-level and will inform future studies aimed at elucidating factors governing their function and niche differentiation in natural and engineered ecosystems.Subject terms: Environmental microbiology, Metabolism     

Can ectomycorrhizal weathering activity respond to host nutrient demands?

Ectomycorrhizal fungi may make a significant contribution to mineral weathering in temperate and boreal forests. It is important to know how this weathering activity will be affected by the changing nutrient demands of forests impacted by global change and nitrogen deposition. This review examines what is known about how plants sense and respond to nutrient demand and discusses the existing literature on ectomycorrhizal weathering in relation to plant nutrient demand. Plant physiology literature indicates that plants can respond to P limitation by allocating more carbon belowground and increasing root branching in areas of high P availability. Increased expression and upregulation of phosphorus and potassium uptake transporters has been observed under P- and K-limitation, respectively. There is evidence for a negative feedback between Mg- and K-deficiency and belowground carbon allocation. There are very few ectomycorrhizal weathering experiments that explicitly test how weathering activity responds to nutrient demand. Field studies suggest that hyphal colonization of readily available P sources does increase with increased P demand of the host. In microcosm studies there is indirect evidence that weathering activity may increase in response to P, K, or Mg demand. Recommendations are made for how future ectomycorrhizal research can better address this question. More research on how plants sense and respond to nutrient limitation, as well as genomic data from gymnosperms would also aid our understanding of this important aspect of forest ecology.     

Analyzing the significance of defoliation in growth, photosynthetic compensation and source-sink relations

Leaf canopy plays a determining role influencing source-sink relations as any change in source activity (photosynthesis) affects sink metabolism. Defoliation (removal of leaves) influences growth and photosynthetic capacity of plants, remobilizes carbon and nitrogen reserves and accelerates sink metabolism, leading to improved source-sink relations. The response of plants to defoliation could be used to manipulate source-sink relations by removing lower and senescing leaves to obtain greatest photosynthetic capacity and efficient carbon and nitrogen metabolism under optimal and stressful environments. The present work enhances our current understanding on the physiological responses of plants to defoliation and elaborates how defoliation influences growth, photosynthetic capacity and source-sink relations under optimal and changing environmental conditions.