Agriculture is responsible for over half of the input of reactive nitrogen (N) to terrestrial systems; however improving N availability remains the primary management technique to increase crop ...yields in most regions. In the majority of agricultural soils, ammonium is rapidly converted to nitrate by nitrification, which increases the mobility of N through the soil matrix, strongly influencing N retention in the system. Decreasing nitrification through management is desirable to decrease N losses and increase N fertilizer use efficiency. We review the controlling factors on the rate and extent of nitrification in agricultural soils from temperate regions including substrate supply, environmental conditions, abundance and diversity of nitrifiers and plant and microbial interactions with nitrifiers. Approaches to the management of nitrification include those that control ammonium substrate availability and those that inhibit nitrifiers directly. Strategies for controlling ammonium substrate availability include timing of fertilization to coincide with rapid plant update, formulation of fertilizers for slow release or with inhibitors, keeping plant growing continuously to assimilate N, and intensify internal N cycling (immobilization). Another effective strategy is to inhibit nitrifiers directly with either synthetic or biological nitrification inhibitors. Commercial nitrification inhibitors are effective but their use is complicated by a changing climate and by organic management requirements. The interactions of the nitrifying organisms with plants or microbes producing biological nitrification inhibitors is a promising approach but just beginning to be critically examined. Climate smart agriculture will need to carefully consider optimized seasonal timing for these strategies to remain effective management tools.
Soil extracellular enzymes play a significant role in the N mineralization process. However, few studies have documented the linkage between enzyme activity and the microbial community that performs ...the function. This study examined the effects of inorganic and organic N fertilization on soil microbial communities and their N mineralization functions over 4 years. Soils were collected from silage corn field plots with four contrasting N treatments: control (no additional N), ammonium sulfate (AS; 100 and 200 kg of N ha
), and compost (200 kg of N ha
). Illumina amplicon sequencing was used to comprehensively assess the overall bacterial community (16S rRNA genes), bacterial ureolytic community (
), and bacterial chitinolytic community (
). Selected genes involved in N mineralization were also examined using quantitative real-time PCR and metagenomics. Enzymes (and marker genes) included protease (
and
), chitinase (
), urease (
), and arginase (
). Compost significantly increased diversity of overall bacterial communities even after one application, while ammonium fertilizers had no influence on the overall bacterial communities over four seasons. Bacterial ureolytic and chitinolytic communities were significantly changed by N fertilization. Compost treatment strongly elevated soil enzyme activities after 4 years of repeated application. Functional gene abundances were not significantly affected by N treatments, and they were not correlated with corresponding enzyme activities. N mineralization genes were recovered from soil metagenomes based on a gene-targeted assembly. Understanding how the structure and function of soil microbial communities involved with N mineralization change in response to fertilization practices may indicate suitable agricultural management practices that improve ecosystem services while reducing negative environmental consequences.
Agricultural N management practices influence the enzymatic activities involved in N mineralization. However, specific enzyme activities do not identify the microbial species directly involved in the measured process, leaving the link between the composition of the microbial community and the production of key enzymes poorly understood. In this study, the application of high-throughput sequencing, real-time PCR, and metagenomics shed light on how the abundance and diversity of microorganisms involved in N mineralization respond to N management. We suggest that N fertilization has significantly changed bacterial ureolytic and chitinolytic communities.
Organic farming systems receive organic amendments to maintain soil fertility and supply nutrients for plant growth. This study investigated the effect of organic fertilizers (no amendment as ...control, compost, and manure), and their interaction with cover crops (millet, buckwheat, and black turtle bean) on soil enzyme activities, N transformation rates, and functional gene abundances under an organic production system. Organic N fertilizers had a stronger effect than cover crop type on soil function and functional gene abundances. Soil enzyme activities were increased by both compost and manure, but there were few differences between these treatments. Nitrification potential, nitrite oxidation potential, and denitrification potential were significantly higher in manure-treated than in control and compost-treated soils, indicating application of manure had a higher N loss potential than compost application in this organic farming system. Organic N fertilizers significantly increased the abundance of some genes involved in N mineralization, ammonification, and nitrification (
sub, ureC
, bacterial
amoA
and
nxrB
). The activity of ammonia-oxidizing bacteria and archaea were both increased by organic N fertilizers, and their activities were higher in manure-treated than in compost-treated soils. Overall, the abundance of functional genes was significantly correlated with their corresponding enzyme activity. However, functional gene abundance was less important than soil chemical and microbiological properties in explaining the variation in the corresponding enzyme activity.
Abstract
Nitrogen is essential for life and its transformations are an important part of the global biogeochemical cycle. Being an essential nutrient, nitrogen exists in a range of oxidation states ...from +5 (nitrate) to −3 (ammonium and amino-nitrogen), and its oxidation and reduction reactions catalyzed by microbial enzymes determine its environmental fate. The functional annotation of the genes encoding the core nitrogen network enzymes has a broad range of applications in metagenomics, agriculture, wastewater treatment and industrial biotechnology. This study developed an alignment-free computational approach to determine the predicted nitrogen biochemical network-related enzymes from the sequence itself. We propose deepNEC, a novel end-to-end feature selection and classification model training approach for nitrogen biochemical network-related enzyme prediction. The algorithm was developed using Deep Learning, a class of machine learning algorithms that uses multiple layers to extract higher-level features from the raw input data. The derived protein sequence is used as an input, extracting sequential and convolutional features from raw encoded protein sequences based on classification rather than traditional alignment-based methods for enzyme prediction. Two large datasets of protein sequences, enzymes and non-enzymes were used to train the models with protein sequence features like amino acid composition, dipeptide composition (DPC), conformation transition and distribution, normalized Moreau–Broto (NMBroto), conjoint and quasi order, etc. The k-fold cross-validation and independent testing were performed to validate our model training. deepNEC uses a four-tier approach for prediction; in the first phase, it will predict a query sequence as enzyme or non-enzyme; in the second phase, it will further predict and classify enzymes into nitrogen biochemical network-related enzymes or non-nitrogen metabolism enzymes; in the third phase, it classifies predicted enzymes into nine nitrogen metabolism classes; and in the fourth phase, it predicts the enzyme commission number out of 20 classes for nitrogen metabolism. Among all, the DPC + NMBroto hybrid feature gave the best prediction performance (accuracy of 96.15% in k-fold training and 93.43% in independent testing) with an Matthews correlation coefficient (0.92 training and 0.87 independent testing) in phase I; phase II (accuracy of 99.71% in k-fold training and 98.30% in independent testing); phase III (overall accuracy of 99.03% in k-fold training and 98.98% in independent testing); phase IV (overall accuracy of 99.05% in k-fold training and 98.18% in independent testing), the DPC feature gave the best prediction performance. We have also implemented a homology-based method to remove false negatives. All the models have been implemented on a web server (prediction tool), which is freely available at http://bioinfo.usu.edu/deepNEC/.
Soil ammonia-oxidizing bacteria and archaea (AOB and AOA) convert ammonium/ammonia to nitrite in the process of nitrification. However, the potentially differential responses of these AO to substrate ...and temperature and the effects of conventional and organic nitrogen management on these responses remains poorly understood. We determined the response of nitrification to ammonium substrate concentration and temperature using an AOB specific inhibitor to distinguish the contribution of AOB and AOA to nitrification. Soils were sampled from cornfield plots that had been treated for four years with contrasting nitrogen sources: control (no additional N), ammonium sulfate at two rates and compost. Nitrification potential and net rates were stimulated for one month after fertilization with ammonium sulfate compared to relatively lower and stable rates in control and compost treated soils. For soils that had been fertilized with ammonium sulfate, the proportion of nitrification mediated by AOB in slurry assays was over 90% at 1.0 mM but less than 50% at 0.01 mM. Kinetic analysis showed maximum nitrification activity (Vmax) for AOB ranged from 0.32 to 4.8 mmol N kg−1d−1 with a half saturation constant (Km) of 14–160 μM ammonium; parameters were higher for soils from ammonium sulfate treated plots. Vmax and Km for AOA averaged 0.24 mmol N kg−1d−1 and 4.28 μM ammonium with no effect of field treatment. The proportion of nitrification due to AOA was lowest at 5 °C, increased with temperature, and was near to 100% at 50 °C; optimum temperature was 41 °C for AOA versus 31 °C for AOB. Understanding the kinetic and temperature response of microbes responsible for nitrification may allow ecosystem models to include these populations as dynamic components driving nitrogen flux.
•Field studies show ammonium fertilizer rapidly increased AOB activity but not AOA.•AOB have 20× higher Vmax and 40× higher Km compared to AOA kinetic parameters.•AOB mediated 90% of nitrification at 1.0 mM ammonium versus <50% at 0.01 mM.•Optimum temperature for nitrification was 31 °C for AOB versus 41 °C for AOA.•Target ammonia oxidizing bacteria for inhibition immediately after fertilization.
Autotrophic nitrification is mediated by ammonia oxidizing bacteria (AOB) or ammonia oxidizing archaea (AOA) and nitrite oxidizing bacteria (NOB). Mounting studies have examined the impact of ...nitrogen (N) fertilization on the dynamic and diversity of AOA and AOB, while we have limited information on the response of the activity, abundance, and diversity of NOB to N fertilization. We investigated the influence of organic and inorganic N fertilizers on soil NOB in silage corn field plots that received contrasting nitrogen (N) treatments: control (no additional N), ammonium sulfate (AS 100 and 200 kg N ha
), and compost (200 kg N ha
). Nitrifying community was examined using a universal marker (16S rRNA gene), functional gene markers (AOB
and
), and metagenomics. The overall nitrifying community was not altered after the first fertilization but was significantly shifted by 4-year repeated application of ammonium fertilizers.
were the dominant NOB (>99.7%) in our agricultural soil. Both community compositions of AOB and
were significantly changed by ammonium fertilizers but not by compost after 4 years of repeated applications. All nitrifiers, including comammox, were recovered in soil metagenomes based on a gene-targeted assembly, but their sequence counts were very low. Although N treatment did not affect the abundance of
determined by real-time quantitative PCR, ammonium fertilizers significantly promoted rates of potential nitrite oxidation determined at 0.15 mM nitrite in soil slurries. Understanding the response of both ammonia oxidizers and nitrite oxidizers to N fertilization may initiate or improve strategies for mitigating potential environmental impacts of nitrate production in agricultural ecosystems.
Understanding nitrification rates and their regulation continues as a key area of research for assessing human's increasing impact on the terrestrial N cycle. We review the organisms and processes ...responsible for nitrification in terrestrial systems. The control of nitrification by substrate availability is discussed with particular attention to the factors affecting ammonia/ammonium availability. The effects on nitrification rates of environmental controls including oxygen, water potential, temperature and pH are described. With this general understanding of the factors affecting nitrification rates as a basis, we present an in depth analysis of methods used to measure nitrification in terrestrial systems. Net, gross and potential nitrification rate measurements are explained including the use of isotopes and inhibitors to measure rates in soils. Methods for the estimation of nitrification kinetics and modeling are briefly described. Future challenges will require understanding the factors controlling nitrification across spatial scales from ecosystems to soil microsites if we are to sustainably manage reactive nitrogen in terrestrial environments.
Ammonia-oxidizing bacteria (AOB) within the genus Nitrosomonas perform the first step in nitrification, ammonia oxidation, and are found in diverse aquatic and terrestrial environments. Nitrosomonas ...AOB were grouped into six defined clusters, which correlate with physiological characteristics that contribute to adaptations to a variety of abiotic environmental factors. A fundamental physiological trait differentiating Nitrosomonas AOB is the adaptation to either low (cluster 6a) or high (cluster 7) ammonium concentrations. Here, we present physiological growth studies and genome analysis of Nitrosomonas cluster 6a and 7 AOB. Cluster 6a AOB displayed maximum growth rates at ≤ 1 mM ammonium, while cluster 7 AOB had maximum growth rates at ≥ 5 mM ammonium. In addition, cluster 7 AOB were more tolerant of high initial ammonium and nitrite concentrations than cluster 6a AOB. Cluster 6a AOB were completely inhibited by an initial nitrite concentration of 5 mM. Genomic comparisons were used to link genomic traits to observed physiological adaptations. Cluster 7 AOB encode a suite of genes related to nitrogen oxide detoxification and multiple terminal oxidases, which are absent in cluster 6a AOB. Cluster 6a AOB possess two distinct forms of ribulose-1, 5-bisphosphate carboxylase/oxygenase (RuBisCO) and select species encode genes for hydrogen or urea utilization. Several, but not all, cluster 6a AOB can utilize urea as a source of ammonium. Hence, although Nitrosomonas cluster 6a and 7 AOB have the capacity to fulfill the same functional role in microbial communities, i.e., ammonia oxidation, differentiating species-specific and cluster-conserved adaptations is crucial in understanding how AOB community succession can affect overall ecosystem function.
Previous studies comparing invaded and non-invaded sites suggest that cheatgrass (Bromus tectorum L.) causes soil N cycling to increase. Unfortunately, these correlative studies fail to distinguish ...whether cheatgrass caused the differences in N cycling, or if cheatgrass simply invaded sites where N availability was greater. We measured soil C and N concentrations and net and gross N-cycling rates on 24-year-old replicated field plots in a sagebrush–steppe ecosystem that had been plowed, fumigated, and seeded to different plant communities in 1984. Laboratory assays of soil collected throughout the soil profiles (0–60 cm) showed that soil NO₃⁻, organic C and N, and net N mineralization, net nitrification, and soil respiration rates were all greater beneath cheatgrass than in sagebrush–perennial grass plots. In surface soils (0–10 cm), field and lab assays on five sampling dates during 2 years showed gross N mineralization, net N mineralization, and net nitrification rates were all faster beneath cheatgrass than in sagebrush–perennial grass plots. Modeling analyses based on soil respiration and gross N-cycling rates suggest that cheatgrass provides soil microbes with lower C:N substrates and that this could explain the faster N-cycling rates beneath cheatgrass. This is the first long-term replicated field study to conclusively show that cheatgrass created greater soil organic N pool sizes and stimulated N-cycling rates compared to similar-aged stands of sagebrush and native perennial grasses. Increased N-cycling rates may represent a positive plant–soil feedback that promotes continued dominance by cheatgrass, even in the absence of soil disturbance or fire.
Cadmium (Cd) and benzo apyrene (BaP) often co-occur in the environment, and the critical body residue of organisms is used as an indicator of the toxic effects of contaminants. However, little is ...known about their distributions and toxicities when pollution of Cd and BaP are combined. Semi-static solution culture experiment was used to study the impacts of BaP on the subcellular distribution of the toxic metal Cd in the earthworm Eisenia fetida. We explored the mechanisms by which this organism responds to combined exposure to these pollutants by measuring the protein content of each of three subcellular fractions, as well as acetylcholinesterase (AChE) and glutathione S-transferase (GST) activities. The subcellular partitioning of Cd was heterogeneous and Cd mainly accumulated in the cytosolic fraction (Fraction C), which was previously reported to be involved in metal immobilization. In Fraction C, Cd accumulation was correlated with the external concentration to which the earthworm had been exposed; however, in the presence of BaP, Cd accumulation was inhibited and plateaued at high external Cd concentrations. A principal component analysis revealed that this decreased Cd accumulation might be caused by increases in GST activity, which likely increased the excretion of Cd. BaP was also found to stimulate protein biosynthesis and upregulate AChE and GST activities in the debris fraction (Fraction E), indicating other potential detoxification mechanisms in this fraction. Granule fraction (Fraction D) had a lower protein content, AChE and GST activities than the other subcellular fractions, supporting previous findings that Fraction D is largely inert.
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•Cadmium (Cd) subcellular partitioning is heterogeneous.•Benzo apyrene (BaP) inhibits Cd accumulation.•BaP promoted the production of total protein in the cellular debris fraction.•Glutathione S-transferase (GST) is the main influencer of Cd accumulation.
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