Plant presence and species combination, but not diversity, influence denitrifier activity and the composition of nirK-type denitrifier communities in grassland soil

To explore potential links between plant communities, soil denitrifiers and denitrifier function, the impact of presence, diversity (i.e. species richness) and plant combination on nirK -type denitrifier community composition and on denitrifier activity was studied in artificial grassland plant assemblages over two consecutive years. Mesocosms containing zero, four and eight species and different combinations of two species were set up. Differences in denitrifier community composition were analysed by canonical correspondence analyses following terminal restriction fragment length polymorphism analysis of PCR-amplified nirK gene fragments coding for the copper-containing nitrite reductase. As a measure of denitrifier function, denitrifier enzyme activity (DEA) was determined in the soil samples. The presence as well as the combination of plants and sampling time, but not plant diversity, affected the composition of the nirK -type denitrifier community and DEA. Denitrifier activity significantly increased in the presence of plants, especially when they were growing during summer and autumn. Overall, we found a strong and direct linkage of denitrifier community composition and functioning, but also that plants had additional effects on denitrifier function that could not be solely explained by their effects on nirK -type denitrifier community composition.     

Perspectives on measurement of denitrification in the field including recommended protocols for acetylene based methods

Of the biogeochemical processes, denitrification has perhaps been the most difficult to study in the field because of the inability to measure the product of the process. The last decade of research, however, has provided both acetylene and¹⁵N based methods as well as undisturbed soil core andin situ soil cover sampling approaches to implementing these methods. All of these methods, if used appropriately, give comparable results. Thus, we now have several methods, each with advantages for particular sites or objectives, that accurately measure denitrification in nature. Because of the general usefulness of the acetylene methods, updated protocols for the following three methods are given: gas-phase recirculation soil cores; static soil cores; and the denitrifying enzyme assay also known as the phase 1 assay. Despite the availability of these and other methods, denitrification budgets remain difficult to accurately establish in most environments because of the high spatial and temporal variability inherent in denitrification. Appropriate analysis of those data includes a distribution analysis of the data, and if highly skewed as is typically the case, the most accurate method to estimate the mean and the population variance is the UMVUE method (uniformly minimum variance unbiased estimator). Geostatistical methods have also been employed to improve spatial and temporal estimates of denitrification. These have occasionally been successful for spatial analysis but in the attempt described here for temporal analysis the approach was not useful.Discussions of the importance of denitrification have always focused on quantifying the process and whether particular measured quantities are judged to be a significant amount of nitrogen. A second line of evidence discussed here is the extant genetic record that results from natural selection. These analysis lead to the conclusion that strong selection for denitrification must currently be occurring, which implies that the process is of general significance in soils.     

Ecophysiology of abundant denitrifying bacteria in activated sludge

The abundance of potential denitrifiers in full-scale wastewater treatment plants with biological nitrogen and phosphorus removal was investigated by FISH and various oligonucleotide probes. The potential denitrifiers were characterized as probe-defined populations that were able to consume radiolabelled substrate with oxygen, nitrate and nitrite as electron acceptor as determined by microautoradiography. The most abundant potential denitrifiers were related to the genera Aquaspirillum, Azoarcus, Thauera and Rhodocyclus, all within the Betaproteobacteria. They made up 20-49% of all bacteria in most of the 17 nitrogen removal plants investigated and were hardly present in four plants without denitrification. The ecophysiology of Aquaspirillum, Azoarcus and Thauera-related bacteria was consistent within each probe-defined group in the plants investigated. These three groups showed distinct physiological differences, with the Aquaspirillum-related bacteria appearing as the most specialized one, consuming only amino acids among the substrates tested, and Thauera as the most versatile consuming some volatile fatty acids, ethanol and amino acids. The coexistence of Aquaspirillum, Azoarcus and Thauera-related bacteria in a range of treatment plants with differences in wastewater, design and operation suggest that the populations ensure a functional stability of the plants by occupying different ecological niches related to the carbon transformation.     

Storage and growth of denitrifiers in aerobic granules: part I. model development

A mathematical model, based on the Activated Sludge Model No.3 (ASM3), is developed to describe the storage and growth activities of denitrifiers in aerobic granules under anoxic conditions. In this model, mass transfer, hydrolysis, simultaneous anoxic storage and growth, anoxic maintenance, and endogenous decay are all taken into account. The model established is implemented in the well-established AQUASIM simulation software. A combination of completely mixed reactor and biofilm reactor compartments provided by AQUASIM is used to simulate the mass transport and conversion processes occurring in both bulk liquid and granules. The modeling results explicitly show that the external substrate is immediately utilized for storage and growth at feast phase. More external substrates are diverted to storage process than the primary biomass production process. The model simulation indicates that the nitrate utilization rate (NUR) of granules-based denitrification process includes four linear phases of nitrate reduction. Furthermore, the methodology for determining the most important parameter in this model, that is, anoxic reduction factor, is established.     

Determinants of the distribution of nitrogen-cycling microbial communities at the landscape scale

Little information is available regarding the landscape-scale distribution of microbial communities and its environmental determinants. However, a landscape perspective is needed to understand the relative importance of local and regional factors and land management for the microbial communities and the ecosystem services they provide. In the most comprehensive analysis of spatial patterns of microbial communities to date, we investigated the distribution of functional microbial communities involved in N-cycling and of the total bacterial and crenarchaeal communities over 107 sites in Burgundy, a 31 500 km² region of France, using a 16 × 16 km² sampling grid. At each sampling site, the abundance of total bacteria, crenarchaea, nitrate reducers, denitrifiers- and ammonia oxidizers were estimated by quantitative PCR and 42 soil physico-chemical properties were measured. The relative contributions of land use, spatial distance, climatic conditions, time, and soil physico-chemical properties to the spatial distribution of the different communities were analyzed by canonical variation partitioning. Our results indicate that 43–85% of the spatial variation in community abundances could be explained by the measured environmental parameters, with soil chemical properties (mostly pH) being the main driver. We found spatial autocorrelation up to 739 km and used geostatistical modelling to generate predictive maps of the distribution of microbial communities at the landscape scale. The present study highlights the potential of a spatially explicit approach for microbial ecology to identify the overarching factors driving the spatial heterogeneity of microbial communities even at the landscape scale.     

电极生物膜反应器中反硝化菌的初步研究

以电极生物膜反应器中分离的三株反硝化菌(12E、22A、25C)为实验材料,为菌株的形态及生理生化特征进行了初步研究,并进一步研究了单一菌株去除硝酸盐氮的能力,结果表明:三株菌株去除(NO3-)-N(硝酸盐氮)的能力存在差异,25C菌株去除率最高.C/N(碳氮比)对菌株的(NO3-)-N去除率有影响,C/N越大,去除率越高.     

Storage and growth of denitrifiers in aerobic granules: part II. model calibration and verification

A mathematical model to describe the simultaneous storage and growth activities of denitrifiers in aerobic granules under anoxic conditions has been developed in an accompanying article. The sensitivity of the nitrate uptake rate (NUR) toward the stoichiometric and kinetic coefficients is analyzed in this article. The model parameter values are estimated by minimizing the sum of squares of the deviations between the measured and model-predicted values. The model is successfully calibrated and a set of stoichiometric and kinetic parameters for the anoxic storage and growth of the denitrifiers are obtained. Thereafter, the model established is verified with three set of experimental data. The comparison between the model established with the ASM1 model and ASM3 shows that the present model is appropriate to simulate and predict the performance of a granule-based denitrification system.     

Identification of the bacterial community involved in methane-dependent denitrification in activated sludge using DNA stable-isotope probing

Methane is used as an alternative carbon source in the denitrification of wastewater lacking organic carbon sources because it is nontoxic and may be efficiently produced by anaerobic biological processes. Methane-dependent denitrification (MDD) in the presence of oxygen requires the co-occurrence of methanotrophy and denitrification. Activated sludge was incubated with 13C-labeled methane in either a nitrate-containing medium or a nitrate-free medium. Then, bacterial and methanotrophic populations were analyzed by cloning analysis and terminal restriction fragment length polymorphism analysis targeting 16S rRNA gene and cloning analysis targeting pmoA genes. DNA-based stable-isotope probing (DNA-SIP) analysis of the 16S rRNA gene revealed an association of the Methylococcaceae and the Hyphomicrobiaceae in a MDD ecosystem. Furthermore, supplementation of nitrate stimulated methane consumption and the activity of methanotrophic populations (i.e. the stimulation of uncultivated relatives of distinct groups of the Methylococcaceae). In particular, uncultured type-X methanotrophs of Gammaproteobacteria were dominant when nitrate was added, i.e. in the MDD incubations. On the other hand, most methanotrophs (types I, II, and X methanotrophs) were found to have been labeled with 13C under nitrate-free conditions. This DNA-SIP study identifies key bacterial populations involved in a MDD ecosystem.     

文峪河上游河岸带不同植被类型土壤nirS反硝化菌群结构及功能

以文峪河上游河岸带不同演替阶段的8种植被类型五花草甸(WH)、沙棘林(HR)、柳树林(SS)、山杨林(PC)、山杨白桦林(PQ)、山杨白桦落叶松林(PQL)、落叶松云杉林(LP)和云杉林(PM)土壤为研究对象,采用高通量测序技术测定nirS反硝化细菌群落组成及相对丰度,乙炔抑制法测定反硝化酶活性(DEA)。对其土壤理化性质及反硝化细菌群落组成及相对丰度进行方差分析,采用冗余分析(RDA)和Spearman相关性分析不同植被类型及土层反硝化细菌群落结构及功能及土壤理化因子的关联性。结果表明:1)不同植被类型及土层土壤理化因子存在显著差异,柳树林(SS)0—15 cm土层硝态氮(NO~+_3-N)含量显著高于其他植被类型各土层;2)土壤反硝化菌群多样性指数在五花草甸(WH)、山杨白桦林(PQ)和云杉林(PM)中较其他植被类型高;3)沙棘林(HR)及柳树林(SS)反硝化酶活性(DEA)显著高于其他植被类型;4)不同植被类型反硝化优势菌群分布存在显著差异及特异性,如浮霉菌门(Planctomycetes)仅在落叶松云杉混交林(LP)和云杉林(PM)植被类型15—30 cm土层中分布;5)土壤pH、土壤有机碳(SOC)、土壤铵态氮(NH~+_4-N)和硝态氮(NO~+_3-N)等是影响土壤反硝化细菌群落结构及组成的重要因子,其中土壤铵态氮和硝态氮含量变化是导致土壤反硝化菌群多样性和反硝化酶活性差异的关键因子。本研究揭示了文峪河上游河岸带不同植被类型土壤反硝化细菌群落结构及功能的变化和分布特征,为进一步研究该区域河岸带氮素循环及水体污染防治提供重要参考依据。