NC1182: Management and Environmental Factors Affecting Nitrogen Cycling and Use Efficiency in Forage-Based Livestock Production Systems

(Multistate Research Project)

Status: Active

NC1182: Management and Environmental Factors Affecting Nitrogen Cycling and Use Efficiency in Forage-Based Livestock Production Systems

Duration: 10/01/2014 to 09/30/2019

Administrative Advisor(s):

NIFA Reps:

Statement of Issues and Justification

Nitrogen use efficiency and improvement thereof is a recurring subject of investigation regarding grazing systems. The amount of N applied annually to these systems exceeds plant uptake, and only little of this N is removed by animals and subsequently the ecosystem. Significant quantities of N fertilizers are applied to forage crop and grazing systems because N is usually the most limiting nutrient for plant growth.

Although nutrient cycles in grazing systems are less open than in systems that rely on concentrated feed such as poultry production, loss of N from pastures is an unsolved problem to which fertilizer N and supplemental feed contribute (Li et al. 2012). In the Midwest, loss of N from any agricultural systems may contribute in a large fashion to periodic hypoxia in the Gulf of Mexico. In addition, gaseous N emissions from pastures contribute to the greenhouse effect (Mosier et al., 1998). The magnitude of N loss and resulting negative impacts on ecosystems are influenced by the timing, frequency, and intensity of management practices within the ecosystems.

Increased demand for meat products by consumers during past decades has encouraged producers to respond with an increased intensification of forage-based livestock production. Hence, there is an urgent need for scientific information to help producers make decisions about how to best manage rural landscapes and to produce agricultural commodities while maintaining soil, water, and air quality. Our experiments will examine alternative strategies to enhance legume establishment and persistence, improve N harvest efficiency, and reduce greenhouse gas (GHG) footprints in pastures; assess secondary plant metabolites in forage legumes for increased N retention and altered N cycling in dung and urine excretions from grazing ruminant animals; and quantify the effects of intensive pasture management strategies on N harvest efficiency and spatiotemporal patterns of N cycling in grassland agro-ecosystems.

Expected outcomes and predictions will include advice on management strategies in terms of N use efficiency, particularly as it relates to the capture and excretion of N in the environment. The ultimate goal is to help producers adopt strategies/practices that ensure efficient use of N in order to positively influence environmental quality. In addition, this work will facilitate the identification forage systems that minimize N inputs and production costs. Minimizing expensive N inputs (e.g., fertilizers) in forage-based livestock production systems has tremendous potential to enhance their profitability. These impacts are most likely achieved through the development and implementation of a multiple-state project. The members of our proposed project represent a geographically diverse set of states from the Southeast through the Midwest and Great Plains and to the Intermountain West. Our objectives of analyzing N use efficiency of grassland production systems will be based on a wide range of environments (humid to semi-arid) and levels of management intensity (irrigated pasture to low-input pastures). The expertise, facilities and other resources required to design and conduct the proposed research are not found at a single institution. The synergy coming from a multiple-state effort in this area greatly enhances the likelihood of success in characterizing N use and developing appropriate management strategies for grassland agro-ecosystems. Furthermore, the technical feasibility of this type of research is questionable for a single university but becomes realistic when several institutions combine resources and expertise.

We propose to continue to conduct complementary experiments to help stakeholders make informed decisions about rural landscapes. Multifunctional farming systems provide multiple ecosystem services. These services can include provisioning (i.e., meat, milk, and fiber production) as well as supporting, regulating, and cultural services. Supporting services include soil building and nutrient retention whereas carbon sequestration and water storage are regulating services, and cultural services include spiritual, aesthetic, and educational factors. Perennial grasslands vary greatly in their ability to provide these types of ecosystem services because of differing environmental and management characteristics. We will assess tradeoffs amongst these services, which should allow more informed decision-making and long-range improvements in U.S. agriculture as a result.

Related, Current and Previous Work

The principle objectives of the NC-1182 Multi State Project have focused on factors affecting N use efficiency in forage-based livestock production systems. Members of the committee were able to generate competitive federal funding for experiments directly and indirectly related to the previous and current research objectives of NC-1182 and several manuscripts have been submitted to and published in scientific journals. During the past five years, combined funds of approximately $1 million were received to investigate topics related to N cycling in pastures affected by different grazing methods and analyze strategies for increasing stand persistence of annual and perennial legumes for enhanced N transfer. In addition, a multi-state proposal was funded in 2012 with $100,000 to plan a comprehensive proposal related to greenhouse gas (GHG) emissions and nutrient cycling in beef production. Currently, there are several proposals pending for USDA-funded studies regarding the effects of different types of legumes on rumen N-use efficiency and N-turnover in perennial grass pastures.

Research proposed for the new project will continue to identify management and environmental factors that will positively influence N cycling for improved ecosystem services from grasslands. Discussion of CRIS search results, current knowledge and our previous work on management and environmental factors that affect N cycling and N use efficiency in major U.S. grassland agro-ecosystems are described below.

CRIS search results:

A search with the terms: nitrogen cycling and pastures and nitrogen use efficiency and pastures found projects focused on cropland systems, confined animal feeding operations, and range and pastureland in different regions of the country. An example included: “Improving the efficiency and sustainability of diversified forage-based livestock production systems” that emphasized beef cattle genetics and utilization of native prairie and wheat pastures in the southern plains. Our project is focused on forage-based livestock production systems in pastures consisting of introduced perennial cool-season grasses. A project focused on “Environmentally friendly forage-livestock systems for the subtropical U.S.A” has a goal of improving understanding of N cycling in pastures, but the grass and legume species being evaluated are subtropical in origin compared to temperate species used in our project. A project entitled “Re-Coupling carbon and nitrogen cycling to increase nutrient use efficiency in annual cropping systems” is investigating the relationship between management practices that control the quantity and quality of C and N inputs and internal N and C cycling processes in agro-ecosystems absent of forage-based livestock. A project investigating “Feeding strategies for dairy cattle to reduce nutrient excretion to the environment” has a focus on commercial dairy herds and confined animal feedlot operations. Another project “Whole farm dairy and beef systems: gaseous emissions, P management, organic production, and pasture based production” is focused on use of models to impact several environmental challenges (GHG emissions, nutrient loading, pathogen transport, and pharmaceuticals) beyond our central focus on improving N use efficiency and cycling. The recent NC1181 project “Enhancing resiliency in beef production under shifting forage resources” is focused on improving grazing efficiency and utilization of crop residues, annual forage crops, and perennial pastureland without addressing the management and environmental challenges of improving N cycling and N use efficiency.

Legume establishment and persistence in pastures:

Legumes enhance forage production and quality in pastures, but competition from existing pasture grasses limits legume establishment and persistence. Under the previous NC1182 project, research in Arkansas found survival of crimson clover and white clover seedlings in bermudagrass pastures were improved with no-till drill seeding over broadcast seeding and stand establishment costs were lowest when stands were seeded at standard rates due to a greater likelihood of stand failure at lower seeding rates (Smith et al. 2012). Project states commonly recommend no-till drill seeding as the best method of interseeding legumes into established perennial grass sod, but the recent NC1182 research shows lower equipment costs accompany other seeding methods if successful stands can be established (Smith et al. 2012). From discussions among NC1182 project members, it has become apparent that more research is needed on lower cost establishment methods to facilitate greater use of legumes in forage-livestock operations. Light disking of grass sod prior to drilling legume seeds has been identified by committee members as a pre-drilling treatment with promise and thus will be evaluated in this project. In addition, we propose to set aside more desirable areas of each pasture and establish “legume banks”. These legume banks would be pure stands of legumes with grass competition removed and should make management of these stands less difficult to manage. The legume banks would provide a source of supplemental, high-quality forage and serve as a source of N for grasses in the remainder of the pasture area via animal excretions. Controlling grazing pressure in the legume banks would also enhance legume survival as a result of minimizing excessive and/or too frequent grazing.

Nitrogen fertilization is a factor affecting persistence of legumes in pastures but the N fertilization rate at which legumes decline due to excessive grass competition has not been evaluated in many pasture types, and project members have discovered that advances in slow-release fertilizer technology may have strategic value in improving legume persistence in N-fertilized pastures. While omission of N fertilizer use on pastures would be a simple solution to issues of legume persistence, it does not acknowledge that high soil N concentrations are needed to maintain the production and persistence of some grass species, and the amount of N required for adequate production of pasture grasses may exceed the amount provided by the interseeded legume. The use of slow-release fertilizers may provide a possible solution to the dilemma with N fertilizers and legume-grass pastures. In previous NC1182 research from Kentucky, it was observed these materials release N more consistently during the growing season, providing the supplemental N needed by pasture grasses without stimulating the excessive growth that is detrimental to maintaining the legume (Goff, unpublished data). In the new project, this research will be expanded to other states to evaluate forage yield, botanical composition, and N use efficiency of grass-legume mixtures fertilized at different N rates with conventional polymer-coated urea and uncoated urea fertilizers.

N cycling, GHG emissions, and N use efficiency in legume-interseeded and N-fertilized pastures:

Through symbiotic N fixation, forage legumes provide an economical source of protein to the diets of grazing livestock reducing or eliminating the need of forage-livestock enterprises to purchase N fertilizer. While economical sources of protein in diets are important to grazing livestock performance, high levels of available protein may lead to excessive proteolysis and deamination within the rumen. If ammonium released by these reactions is not reincorporated into microbial or animal tissues, it will be excreted in the urine of the animal. In recent NC1182 research, we found urine N contributes significantly to nitrous oxide (N2O) emissions from pastures, with cumulative emissions increasing exponentially with N fertilizer rates (Snell et al. 2014). In the new project, we plan to examine whether replacement of N fertilizer application with forage legumes as an N source in pastures reduces N2O emissions, as well as increases N harvest efficiency and soil organic C as GHG mitigation strategies in forage-based livestock production systems. Project states will evaluate N2O emissions and soil organic matter dynamics from legume-interseeded and N-fertilized pastures with the goal of identifying management practices that increase reproductive and growth efficiencies by grazing ruminants and thereby reduce GHG emissions per unit of meat produced. Recent NC1182 research demonstrated that the form and amount of N inputs to cool-season grass pasture impact cattle performance (Greenquist et al. 2009), N use efficiency (Greenquist et al. 2011), production economics (Watson et al. 2012), annual forage accumulation (Guretzky et al. 2013) and plant residue quality both for the grazing animal (Greenquist et al. 2009) and as a decomposer substrate in soil (Guretzky et al. 2014). With our proposed grass-legume research, we plan to investigate similar factors including their impact in altering relationships between residue decomposition, N2O emissions, and soil organic matter dynamics.

Legume species, secondary plant metabolites and N retention in forage-livestock systems:

Degradation rate of protein in the rumen is a major factor affecting how ruminant animals utilize plant protein, and thus affects the mode of excretion of excess N and N-use efficiency. Increasing the amount of rumen undegradable protein may improve livestock performance while reducing N excretion. Protein in some legumes is degraded rapidly by ruminal microorganisms to ammonia and short-chain fatty acids (Cassida et al. 2000). While some of the ammonia is used by ruminal microorganisms to produce microbial protein, a large part of this ammonia is absorbed by the rumen wall, converted to urea, and excreted in urine. The reason this ammonia is absorbed is that the ammonia released from the legumes exceeds the capacity of the ruminal microorganisms to convert it into microbial protein. Some legumes, however, contain condensed tannins that suppress degradation of plant protein by rumen microorganisms and allow it to be digested posterior to the rumen in the small intestine with reduced urinary N excretion and altered fecal N chemistry. Other studies, however, have found higher rumen undegradable protein concentrations in red clover than alfalfa and birdsfoot trefoil (Broderick et al., 2004; Grabber and Coblentz, 2009). The higher concentrations of rumen undegradable protein in red clover compared to other forage legume species is believed to be due to the presence of the enzyme polyphenol oxidase, which oxidizes soluble phenolic compounds into forms that covalently bind to proteins and protect them from proteolysis in the rumen (Peirpoint, 1969; Macheix et al., 1991). This line of research has not been addressed in previous NC1182 research, but project members in Michigan have evaluated how legume forages affect protein digestion in ruminant animals (Cassida et al. 2000) and improved understanding of these relationships has potential to alter N cycling and losses in pastures. In the proposed project, we plan to compare rumen undegradable protein levels in legumes such as red clover, alfalfa, and birdsfoot trefoil for their impact on animal production, GHG emissions, and N contribution to the pasture system; examine the effects of polyphenol oxidase activity and soluble phenolic concentrations on red clover protein fractions and availability across multiple varieties to determine if there is sufficient variability that may be exploited in breeding programs; and evaluate if alfalfa and white clover have a similar polyphenol oxidase mechanism to increase rumen protein utilization.

Tall fescue management and N use efficiency:

Cool-season grasses are a major component of pastures, and management strategies that affect these grasses are likely to influence N cycling and N harvest efficiency in forage-based livestock systems. Tall fescue, the predominant cool-season grass used for cow-calf production in much of the Southeast and lower Midwest, is infected with an endophytic fungus that limits N harvest efficiency by beef cattle. This “wild-type” endophytic fungus (E+) imparts desirable properties to the plant, providing a persistent forage, but it is toxic to cattle resulting in reduced animal gains, immune function, and reproductive performance. This project will build upon previous NC1182 research and investigate use of non-toxic “novel” endophyte-infected tall fescue associations (NE+) as a means by which cow-calf operations can improve cattle and forage management with a resulting improvement in production efficiency and a reduction in off-farm inputs. Current efforts to counter the toxicosis associated with E+ have centered on dilution of E+ pastures and establishment of endophyte-free (E-) pastures. Dilution of E+ pastures with legumes has been beneficial for beef cattle performance, but in past NC1182 research, legumes provided inconsistent production and animals displayed many of the adverse symptoms of fescue toxicity (Coffey et al. 2005). Dilution of E+ with bermudagrass and other forages also did not eliminate the effects of grazing E+ (Coblentz et al. 2004), and E- is less tolerant and adaptable to diverse environments, leading to poor persistence and reduced production. In contrast, research on NE+ tall fescue suggests it has greater persistence than E- and does not produce symptoms of tall fescue toxicosis (Parish et al. 2003; Watson et al. 2004). In previous NC1182 research we found moving spring-calving cows from E+ to NE+ pastures one month prior to breeding and weaning improved calving rate and partial returns ($/ha) compared with those grazing E+ for the entire year (Smith et al. 2012; Caldwell et al. 2013). Steers weaned from E+ also maintained a lower weight differential throughout a subsequent wheat-pasture grazing period and feedlot period compared with steers weaned from NE+, and heifers weaned from E+ had lower subsequent calving rates compared with heifers weaned from NE+ (Coffey et al. 2007). Our NC1182 research also has evaluated performance of fall-calving cows grazing stockpiled tall fescue pastures (Caldwell et al. 2009).

Patch burning in warm-season grass pastures, N cycling, and N use efficiency:

In the Midwest, the cool-season grasses smooth bromegrass and Kentucky bluegrass provide important forage resources but similar to tall fescue, their production is limited to spring and dependent on N fertilizer. A means to improve forage production during summer and N use efficiency by beef cattle is to increase use of perennial warm-season grasses. Warm-season grass stands, however, face invasion from cool-season pasture grasses without burning in spring. Patch burning, a strategy to enhance heterogeneous vegetation cover (Fuhlendorf and Engle, 2004), may also enhance N harvest efficiency of beef cattle grazing warm-season grass pastures relative to N-fertilized pastures. In contrast to traditional management where homogeneous vegetation cover is emphasized through uniform grazing and fire distribution, patch burning increases heterogeneity through livestock grazing preferences for recently burned patches. As fire and grazing distribution is shifted to different landscapes, succession creates a patchwork landscape of plant communities, and soil N availability is increased as shifting cattle grazing patterns distribute dung and urine to different patches. A research gap that we aim to address with the new NC1182 project is how patch burning and interactions with N fertilization affects pasture N cycling and N use efficiency of beef cattle.

Intensive grazing strategies, N cycling and N use efficiency:

As a whole, grassland agro-ecosystems tend to be low input systems where management of nutrient cycles becomes critical to system productivity, sustainability, and resilience. Intensive grazing strategies (i.e., rotational grazing) designed to target the spatial and temporal utilization of vegetation may regulate nutrient cycling through defoliation frequency and intensity. Grazing induces pulses of energy and nutrients through trampling of vegetation and litter, deposition of dung and urine, and root exudation. A strong body of knowledge of how grazing strategies impact plant productivity, botanical composition, animal performance and water quality has been recently summarized (Briske, 2011). However, effects of grazing strategies on nutrient cycling, carbon sequestration, and mitigation of GHG emissions are knowledge gaps. Knowledge-gap questions related to N cycling include: (1) what are the effects of grazing strategies on the size and composition of nutrient return pools? (2) How do grazing strategies impact the temporal and spatial distribution of nutrient return pools? (3) What are the influences of the spatial and temporal patterns of nutrient return on physical and biological processes for nutrient transport, cycling and losses? (4) What are the levels of association of grazing strategies to patterns of N return and soil nutrient availability? The new NC1182 project will address these questions with a rotational grazing experiment that evaluates ultrahigh stocking densities in cool-season grass pastures.

Supplementation strategies and N use efficiency of beef cattle grazing annual forages:

Annual forages such as winter wheat in the southern Great Plains (Horn et al. 2005) and cover crops (Sulc and Tracy, 2007) in the northern Great Plains may be used as forage resources for beef cattle. These forages are often high in nitrogen, but a majority of the plant protein is degraded in the rumen, so animal growth may be limited by amino acids. Therefore, the efficiency of N use by the animal is low. Research by NC1182 scientists (Buckner et al. 2013) has shown that supplementation strategies which provide amino acids to the small intestine, fermentable carbohydrate to the rumen, or both may improve growth and the efficiency of N use. Additionally, there is essentially no information available to evaluate the potential to reduce N from fertilizer if fields are grazed and N is being recycled to the plant.


  1. Evaluate legume cultural and management strategies emphasizing legume establishment, N cycling and use efficiency, and GHG emissions. (AR, KY, NE, UT). Specific objectives: (i) Identify practices that optimize legume establishment and persistence. (ii) Compare N cycling and use efficiency of ruminants grazing pastures with and without forage legumes. (iii) Determine the impact of legumes on the GHG footprint of livestock production systems.
  2. Assess the efficacy of secondary plant metabolites in legume species for increasing N retention and improving N cycling in forage-livestock systems. (AR, KY, MI, UT) Specific objectives: (i) Evaluate effects of birdsfoot trefoil, a tannin-containing legumes, on N partitioning in dung and urine excretions. (ii) Determine soluble phenolic and genotypic effects on forage legume protein fractionation and nitrogen availability. (iii) Evaluate effects of genetic variability in tannin concentration on soil N availability in mixed birdsfoot trefoil/tall fescue swards.
  3. Quantify effects of pasture management strategies on N use efficiency by ruminant animals and N cycling in herbage and soils of grassland agro-ecosystems. (AR, NE, MI, OK) Specific objectives: (i) Investigate effects of management strategies that alter spatiotemporal distribution of grazing and nutritive value of forage on ruminant performance and N harvest efficiency. (ii) Evaluate effects of management strategies on herbage mass and accumulation, nutritive value, botanical composition, and N use efficiency across growing seasons and pasture landscapes. (iii) Determine N pool and cycling responses to management strategies across variable soil environments and climatic conditions. (iv) Evaluate byproduct supplementation as a source of N for annual forages in integrated cropping livestock systems.
  4. Disseminate research results through coordinated extension/education activities, including extension publications, university course material, and regional and state conferences on nitrogen cycling and use efficiency and management of grass-legume mixtures. (AR, KY, MI, NE, OK, UT)


Objective 1 Specific objective i: Three studies will be conducted to identify practices that optimize legume establishment and persistence. Influence of no-till drilling vs. light disking soil cultivation on establishment of perennial legumes will be evaluated in bermudagrass at the University of Arkansas. The objective is to evaluate establishment success for various white and subterranean clover entries under two establishment scenarios consisting of no-till drilling and light soil cultivation (disking at a 8-cm depth) to break up the existing pasture sod. Number of clover seedlings in each treatment will be evaluated after planting in autumn and again the following spring. Whole plots will be grazed in early summer. Regrowth of legumes will be measured after each grazing cycle. Forage mass will be sampled from each plot to determine legume/bermudagrass forage mass ratios. A second study at the University of Arkansas will evaluate the hypothesis that use of multiple types of legumes planted in monoculture legume banks associated with grass pastures will enhance legume persistence and reduce risk to producers for adopting these practices. Spring- and fall-born calves used in this project will be 7-8 months of age when allocated randomly to one of three post-weaning forage treatments: 1) N-fertilized grass pasture (control), 2) fertilized grass pasture with red clover/ladino clover bank, or 3) fertilized grass pasture with birdsfoot trefoil/sericea lespedeza bank. The legume banks will be seeded in separate areas within each pasture. The forage base of these pastures will be NE+ fescue and bermudagrass, which coupled with rotational stocking and stockpiling management, can provide forage throughout the entire year. Calf performance through the grazing season and the feedlot finishing will be quantified. Cost and GHG footprint per pound of grass-fed beef produced will be tracked and compared to current literature estimates of conventionally produced beef. Existing spreadsheet based software, the Forage & Cattle Planner, developed at the University of Arkansas (Smith, 2013; Keeton, 2013) to model returns and GHG emissions at the cow-calf level, will also be used in conjunction with grazing and feedlot GHG estimates to determine cost and GHG footprint from birth to slaughter. Separate samples of the base pasture forage and the particular legume treatment will be taken on a monthly basis for determination of crude protein (CP) and in vitro dry matter digestibility (IVDMD). Forage species composition will be assessed twice annually in late May and late October of each year to assess changes in forage species associated with N fertilization by pasture treatment. Legume persistence under varying N fertilizer rates and formulations will be evaluated at the University of Kentucky. ‘Durana’ white clover was drilled into existing stands of ‘Wrangler’ and ‘Quickstand’ bermudagrass during fall 2013. Five formulations of urea will be used as treatments and will include: commercially available urea, urea mixed with volatilization and nitrification inhibitors (Agrotain® and dicyandiamide, respectively), polymer-coated urea, branched-chain methylene urea, and blend of commercially available urea and the polymer-coated form (25 and 75%, respectively). Each fertilizer will be applied at total seasonal rates of 100, 200, and 400 lbs N acre-1 that will be distributed in two equal applications at the start and approximate mid-point during each growing season. Two additional plots will be incorporated into the experimental design and will serve as controls to evaluate the effect of no fertility addition (i.e., no N fertilizer or clover) or only white clover on stand productivity and species dynamics. Data will be collected on approximately 28-day cycles to simulate the effects of a rotational grazing system. Species composition (bermudagrass, white clover, and weeds), forage availability, and forage quality (CP and IVDMD) of the standing crop in each of the treatments will be determined. Specific objective ii: Nitrogen cycling and use efficiency of pastures with and without legumes will be evaluated in studies conducted in Arkansas, Utah, and Nebraska. This research will be conducted in annual forage systems as well as pastures with perennial forage species. At the University of Arkansas, teff, an annual summer grass, will be seeded into plots that had annual legumes (crimson clover, arrowleaf clover, and hairy vetch) in them the year before. Forage production and quality (including N content) of teff in the legume plots will be compared to plots of teff fertilized at different rates (0 to 150 kg N ha-1). These data will be used to estimate the amount of N made available for teff production by the legumes. Research also will be conducted at Utah State University and the University of Nebraska-Lincoln to compare forage and animal production between fertilized monocultures of cool-season grasses (smooth bromegrass and tall fescue) and grass-legume pasture over a 5-year period. The experimental paddocks will be grazed rotationally by beef cattle yearlings annually from late April to late September. Daily gains of the cattle will be measured annually and N removal from the pastures will be estimated with NRC (1996) equations. Diet samples will be collected monthly with ruminally-fistulated cattle. Intakes will be estimated with the use of NRC (1996) energy equations. Leachate samples will be collected weekly using suction cup lysimeters. N losses to volatilization will be estimated using a mass balance approach, and nitrogen use efficiency will be determined. Economics and environmental aspects of grass-legume mixtures versus grass monocultures will be compared. Specific objective iii: Impact of legumes on GHG footprint of livestock production systems will be measured seasonally to coincide with seasonal changes in temperature and moisture, and with consideration to cattle rotation on and off legume banks in pastures at the University of Arkansas. Sampling events will be approximately 48 hours after cattle are moved from a legume bank to a grass pasture to allow for fecal deposits to contain material from the legume bank. Gas emission samples will be taken at 1, 2, and 5-7 days after manure deposition events. Sample locations will be “stratified” by manure deposition, such that areas with and without evident manure deposits will be sampled at each sampling event. Sample collection will occur in as short a time as possible during the same time period (10:00am to noon) on each sampling day. Sampling for GHG will cover five replications of locations with and without manure deposits during each time series sampling event within each pasture. Independent cells within a pasture will be the experimental unit. A known volume of soil headspace will be enclosed for a timeframe determined empirically during preliminary experiments. A gas sample will be removed using an air-tight syringe piercing a septum. Analysis of N2O, CO2, and CH4 will be conducted by gas chromatograph (Agilent Technologies, Santa Clara, CA). Fluxes of gases will be measured throughout 2 years of the experiment. Gases will be measured from the “system,” so if grass is present, GHG fluxes will be measured with aboveground plant biomass in place. Temperature (thermometer) and soil moisture (theta probe) will be measured at the 10-cm depth in each plot as each GHG sample is collected. Objective 2 Specific objective i: Effects of birdsfoot trefoil, a tannin-containing legume, on N partitioning in dung and urine excretions will be evaluated at Utah State University. Treatments will include: 1) tall fescue and birdsfoot trefoil paddocks (TFBFT); 2) tall fescue and alfalfa paddocks (TFALF); and 3) fertilized tall fescue (TFWF) paddocks. Ammonium sulfate fertilizer (35 kg ha-1) will be applied to the TFWF paddocks approximately every 30 days during the growing season. Fifteen heifers with similar weights, age, and body condition will be outfitted with urine and fecal collection systems. The heifers will be randomly assigned to one of the treatments and placed in the appropriate paddock. The heifers will be allowed to adapt to the diet for a minimum of 48 hours prior to data collection. Fecal samples and urine samples will be collected every 12 hours for 72 hours (3 days). The feces will be weighed and the volume of urine determined. Manure and urine samples will be frozen for further analysis. Manure samples will be analyzed for CP, total N, and NH4. Urine samples will be analyzed for total N and NH4. Specific objective ii: Soluble phenolic and genotypic effects on forage legume protein fractionation and N availability will be determined in forage samples collected from the University of Kentucky forage variety trials. Fifty varieties of alfalfa, 16 varieties of red clover, and 18 varieties of white clover will be included in the experiment. During collection of the samples, half of the harvested forage will be frozen immediately and stored at -20°C, while the other half will remain in the field to air-dry to allow for activation of the enzyme polyphenol oxidase. All samples will be lyophilized and ground to pass through a 1-mm screen with a Udy mill. A micro-Kjeldahl procedure utilizing a salicylic acid modification will used to determine sample N concentrations. The methods summarized in Licitra et al. (1996) will be used to determine how the total CP is allocated among the different fractions. The amount of rumen degradable protein and rumen undegradable protein will be estimated using the Cornell net carbohydrate and protein system. Near infra-red spectroscopy (NIRS) will be used to determine the concentrations of total soluble phenolics (TSP) and the various fractions of forage protein. Reflectance spectrum (400-2500 nm) will be obtained from each sample using a Foss NIRSystems 6500 spectrophotometer. The total number of collected spectra will be subdivided to allocate samples for the development and validation of calibration curves for each of the estimated parameters. A significant reduction in TSP concentrations following air-drying will represent the presence of polyphenol oxidase, and depending upon the relationships between this decrease and the various protein fractions, polyphenol oxidase activity and concentrations of individual phenolics may also be determined. Specific objective iii: Effects of genetic variability in tannin concentration on soil N availability in mixed birdsfoot trefoil/tall fescue swards will be evaluated at Michigan State University using plot s that contain two tall fescue varieties (‘Kentucky 31’ with wild-type endophyte and ‘Duramax’ with low-alkaloid endophyte); two fescue endophyte infection levels (E+ and E-) and eight commercially available birdsfoot trefoil entries that differ in condensed tannin content (Grabber, Cassida, MacAdam et al., unpublished data). Birdsfoot trefoil will be planted in spring 2014 into a tilled seedbed. In spring 2015, plots will be overseeded with tall fescue using a no-till drill. During the second and third growing seasons, plots will be harvested whenever mean forage dry biomass reaches an estimated 1.5 to 2.0 tons/acre. At each harvest, forage mass will be measured, corrected to dry matter basis, and ground through 1-mm screens in preparation for analysis of nutritive value, including condensed tannins, with NIRS. An NIRS calibration equation will be developed to estimate the proportions of birdsfoot trefoil and tall fescue in harvested forages. Birdsfoot trefoil shoots and roots, collected by digging three plants per plot immediately before and after each harvest and at weekly intervals between harvests, also will be analyzed for condensed tannin and water soluble carbohydrate concentrations. Soil samples will be analyzed for total soil organic C and N and mineral N (NO3-N and NH4-N) concentrations. Objective 3 Specific objective i: Effects of management strategies that alter spatiotemporal distribution of grazing and nutritive value of forage on ruminant performance and N use efficiency will be evaluated in three independent grazing experiments at the Univ. of Arkansas, Univ. of Nebraska-Lincoln, and Michigan State Univ. Pastures will be based on cool-season pasture grasses that provide appropriate forage resources for each region (i.e., tall fescue in AR; smooth bromegrass, Kentucky bluegrass, and quackgrass in NE; and orchardgrass and tall fescue in MI). Each of the experiments will contain multiple grazing management strategies that differ in intensity and are replicated within sites. High-intensity management strategies aim to increase ruminant livestock production and N use efficiency through use of rotational stocking, stockpiling of vegetation, ultra-high stocking densities, strategic overseeding of grassland resources with legumes and annual forages, and patch burning. These practices shift grazing to different pasture or forage resources in time and are designed to increase harvest efficiency, provide rest and recovery periods to intensively utilized forage resources, and enhance forage nutritive value. In contrast, livestock in medium-intensity management strategies will be placed in larger pastures at lower stocking densities, and will be rotated at less frequent intervals or graze without restriction in the allocated pasture area. Hay will be offered as needed and N fertilization and general overseeding of pastures without strategic site selection will be employed as techniques to enhance forage mass and nutritive value in medium-intensity management strategies. Body weights and body condition scores will be taken before and after grazing of treatment pastures. Nitrogen in the livestock pool will be computed with NRC (1996) equations based on initial and final body weights and protein contents. Nitrogen removal by cattle will be computed as the difference between the initial and final body weight N contents. Nitrogen harvest efficiency will be computed by dividing annual body weight N gains by N fertilizer inputs on per animal and land area bases. Specific objective ii: Evaluate effects of management strategies on herbage mass and accumulation, nutritive value, botanical composition, and N use efficiency across growing seasons and pasture landscapes. Herbage mass will be measured on biweekly to monthly bases with a disk meter and calibrated with harvested samples in each of the grazing experiments described in specific objective i. Upon harvesting of the samples, the herbage will be separated into live (current year’s growth), standing dead, and litter pools. Nutritive value (CP, neutral detergent fiber, acid detergent fiber, and in vitro organic matter digestibility) will be determined on the live forage pool with NIRS calibrated with wet chemistry techniques. Botanical composition will be determined by separating grasses from forbs (legumes) and other (weedy) species in the harvested forage samples, as well as with visual canopy and ground cover assessment procedures. Cattle exclosures will be established in the grazing experiments for measurement of annual herbage accumulation and N use efficiency. Nitrogen use efficiency will be computed by dividing annual herbage accumulation by N fertilizer inputs. Specific objective iii: Determine N pool and cycling responses to management strategies across variable soil environments and climatic conditions. Pools and pulses of N in herbage, soil, and animal resources will be measured in each of the grazing experiments. The herbage N pool will be determined from samples harvested biweekly to monthly across the grazing season and annual herbage N uptake will be determined from samples harvested from grazing exclosures. The litter N pool will be measured through collection of senescent and detached plant material from the soil surface when herbage is harvested. Carbon and N in root and rhizome pools will be measured by sieving and extracting roots and rhizomes from soil cores. Soil cores will be analyzed for total soil organic C and N and mineral (NO3-N, NH4-N) concentrations. Carbon and N pulses will be quantified by determining the size of nutrient return in dung, urine, trampled green tissue, and litter deposition. Carbon and N pulses from litter deposition on the soil surface will be measured during vegetative growth (spring), reproductive growth (June-July), and post reproductive tiller senescence (August-Sept) to coincide with developmental stages. Litter deposition will be measured during time periods when cattle are not on the pasture or within grazing exclosures to avoid mixing trampled and senescent plant material. Effects of grazing management strategies on N pulses from trampled plant material and its entry into the litter pool will be determined through measurement of differences between trampled material before, during, and after grazing. Nitrogen balance under each management strategy and grassland agro-ecosystem will be determined after accounting for total N inputs from fertilizer and supplemental feeds. In light of difficulties in obtaining precise measurements of intake rates on pasture, herbage N consumption will be computed with NRC (1996) equations for intake rates in the pastures multiplied by the herbage N concentration (Greenquist et al. 2009; Greenquist et al. 2011). Supplemental feed N consumption will be computed through multiplication of daily intake of the feed by its N concentration. Total N consumption will be computed through addition of herbage and supplemental feed N intake. Nitrogen retention by cattle will be determined from actual cattle gains and NRC (1996) equations which estimate the proportion of gain that is protein (N). Nitrogen excretion in dung and urine will be computed through subtraction of N retention from N consumption. Surplus N (balance) with respect to the high and medium-intensity grazing management strategies will be determined through subtraction of N retention from total N inputs. Specific objective iv: Effects of byproduct supplementation as a source of N for annual forages in integrated cropping livestock systems will be evaluated at the Univ. of Nebraska-Lincoln. Growing cattle grazing annual forages (wheat pasture, cover crops, forage crops, etc.) will be supplemented with co-product feeds to evaluate management and supplementation strategies to improve N use efficiency in an integrated cropping livestock system. Mass balance approaches will be used to evaluate N inputs through fertilizer and supplemental feed, N outputs in animal products and crops. Where appropriate, animal digestion models will be used to elucidate the animal effects on the total system. Cattle with duodenal and/or rumen cannulas will be used to evaluate site of N digestion as well as N retention for different supplements utilized in the system. Objective 4: The objectives cover research projects in six different states across climatic gradients with varying seasonal temperatures and amounts of precipitation. Our projects will address unique challenges and problems related to nitrogen cycling and use efficiency in each of participating locations. While specific methodologies are necessarily site-specific, implications from experiments will be transferable across states. Two examples: (a) Outcomes from the proposed fertilizer study in KY (Obj. 1, Specific Obj. i) will be directly transferable to neighboring states including AR, but may not be of high importance to MI, where the prevailing climate does not allow the production of bermudagrass forage. (b) There are also direct links between the projects of Obj. 1/Specific Obj. iii and Obj. 2/Specific Obj. i. With the former, we seek to investigate the GHG emission impact of ruminant production from grass-legume forage, while the latter focuses on N partitioning in urine and feces. The studies complement each other because the quality of chemical N-compounds in ruminant excrements has direct impact on the quality of GHG emission coming from legume-containing diets. The members of the committee will meet annually to report on project-related results and to identify research questions and outreach activities that are project-related to develop research proposals that the group may submit jointly to granting agencies. Committee members also are committed to extend communications beyond the annual meetings and to share ideas and data through frequent communications, including email messaging and teleconferences. As studies are completed, coordinated extension programming and efforts will be developed by project personnel to facilitate effective dissemination of project results throughout the region. This regional effort will be realized through extension publications (e.g., extension circulars and fact sheets), web sites [examples are UNL's Beef site ( and UA extension forage/livestock site (], and regional and local conferences and tours. These methods of information dissemination will ensure exposure of project results to extension educators, producers, federal and state agency personnel, and consultants/advisors. Results also will be published in scientific journals. As made apparent in this proposal, there will be numerous opportunities for project scientists to jointly publish research articles that have regional significance. This regional effort is important because it brings together expertise from different states in discovering and disseminating solutions to the research questions. Several faculty members have joint research and cooperative extension appointments and will take the lead in dissemination of project results. Extension activities are discussed at the annual meetings and coordinated via email discussions.

Measurement of Progress and Results


  • Refereed publications in the Agronomy Journal, Crop Science, Forage and Grazinglands, Journal of Animal Science, and Nutrient Cycling in Agroecosystems.
  • Outreach publications and popular-press articles for audiences of forage/livestock producers, crop and nutrition consultants, advisors with federal and state agencies, managers of public and private agricultural lands, and policy makers in the state and federal governments. The publications will present strategies/methods of efficiently using N in forage-based livestock production systems, identify factors affecting N uptake and cycling in forage-based livestock systems, and describe the pertinent N dynamics in common forage plants.
  • Research summaries in university progress reports, in producer field day proceedings, and on web sites.
  • Regional conferences, workshops, and field days.
  • Presentations at extension meetings and workshops and at annual meetings of professional societies (e.g., Crop Science Society of America and Society for Range Management).

Outcomes or Projected Impacts

  • Fertilizer recommendations for optimizing forage production and N harvest efficiency.
  • Adoption by producers of strategies/practices that ensure efficient use of N, which will have a positive influence on environmental quality in the NCR.
  • Identification of management strategies and forage systems that minimize N inputs and production costs. Optimizing types and amounts of fertilizers, legume management, and forage/grazing management in forage-based livestock production systems will enhance their N use efficiency and profitability.
  • Multi-functional farming systems provide multiple ecosystems services. In this new project, we will continue to conduct complementary experiments and help stakeholders make informed decisions about the future use of rural landscapes. We will assess the tradeoffs among these services which should contribute to a long-term improvement in agricultural sustainability.


(2015): Initiate study on no-till drilling vs. light disking on establishment of perennial legumes (Objective 1). Initiate study on using legume banks as a means of increasing legume stand persistence (Obj. 1). Establish the N fertilizer rates and formulations study on the experimental site where white clover was drilled into bermudagrass in fall 2013 (Obj. 1). Initiate studies on N availability and use efficiency of pastures with and without legumes (Obj. 1). Measure GHG emissions on the grass pastures associated with the legume banks (Obj. 1). Initiate the study testing the effect of a tannin-containing legume on N partitioning in dung and urine excretions (Obj. 2). Collect the tissue samples of the varieties of alfalfa, red clover, and white clover for analysis of soluble phenolic and genotypic effects on legume protein fractionation and N availability (Obj. 2). Drill tall fescue into birdsfoot trefoil plots (Obj. 2). Initiate grazing studies relating spatiotemporal distribution of grazing and nutritive value of forage on herbage production, cattle performance, and N cycling and harvest efficiency (Obj. 3). Initiate studies dealing with byproduct supplementation as a source of N (Obj. 3).

(2016): Conduct the second year of the following Obj. 1 studies: no-till drilling vs. light disking, legume banks, fertilizing white clover/bermudagrass for legume persistence, grazing grass pastures with and without legumes, and GHG emissions on grass pastures. Conduct the second year of the study on the effect of a tannin-containing legume on N partitioning in dung and urine excretions (Obj. 2). Analyze tissue samples of alfalfa, red clover, and white clover for protein fractionation and N availability (Obj. 2). The tall fescue/birdsfoot trefoil plots will be harvested to determine production and nutritive value of the forages; soil samples will be analyzed for C and N content (Obj. 2). Continue the Obj. 3 grazing management studies and the evaluation of byproduct supplementation.

(2017): Complete and evaluate the results of the following Obj. 1 studies: no-till drilling vs. light disking and GHG emissions on grass pastures. Conduct the third year of Obj. 1 studies: legume banks, fertilizing white clover/bermudagrass for legume persistence, and grazing grass pastures with and without legumes. Complete and evaluate the results of the two Obj. 2 studies: the effect of a tannin-containing legume on N partitioning in dung and urine excretions and protein fractionation and N availability of three legumes. Harvest the tall fescue/birdsfoot trefoil plots again (Obj. 2). Continue the Obj. 3 grazing management studies and the evaluation of byproduct supplementation. Report/disseminate findings of completed studies through coordinated extension programming (e.g., conferences and web sites) (Obj. 4). Prepare manuscripts for scientific journals (Obj. 4).

(2018): Conduct the fourth year of Obj. 1 studies: legume banks, fertilizing white clover/bermudagrass for legume persistence, and grazing grass pastures with and without legumes. Complete and evaluate the results of the Obj. 2 tall fescue/birdsfoot trefoil study. Continue the Obj. 3 grazing management studies and the evaluation of byproduct supplementation. Report/disseminate findings of completed studies through coordinated extension programming (e.g., conferences and web sites) (Obj. 4). Prepare manuscripts for scientific journals (Obj. 4).

(2019): Summarize and evaluate the results of all studies that were still being conducted in 2018. Report/disseminate findings of completed studies through coordinated extension programming (e.g., conferences and web sites) (Obj. 4). Prepare manuscripts for scientific journals (Obj. 4).

Projected Participation

View Appendix E: Participation

Outreach Plan

The timing and purpose of the proposed project is extremely relevant because N use efficiency and the dynamics of associated soil nutrients are a major focus of production agriculture systems in both the North Central Region and the entire country. Furthermore, there is a large audience needing research-based information for making decisions.Results of the project will be made available to intended users, listed in the Outputs section, through a number of avenues. Several of the participants have extension appointments and most participating research scientists work closely with extension specialists in the region.

Results of this project will be incorporated in extension programming of each participating state. Methods used will include extension publications available to producers and extension educators, publication of results in such journals as the Forage and Grazing Land Journal, Crop Management Journal, and Journal of Animal Science which are used by crop and livestock consultants, county/area training sessions, grazing workshops, field days, and individual consultations. Establishment of on-farm demonstrations also will be used in Arkansas and other states after data become available from the research effort. The Cooperative Extension Service in member states has extensive listings of fact sheets for livestock and forage production. Information from this work will be used to develop new fact sheets and to update existing fact sheets on forage and livestock systems. These fact sheets are distributed to producers region wide. Summaries of the research also will be included on the Extension Service websites and the projects website for use by producers and other clientele throughout the region and nation. The websites also will provide access to project updates and publications and other relevant resources. Finally, project results will be available through refereed publications. These publications should be particularly important to scientists and policy makers seeking the most current information on N dynamics and N use efficiency in livestock based agricultural production systems.


The technical committee will organize and function in accordance with the procedures described in "Manual for Cooperative Regional Research." The voting members will elect two officers (Chair and Secretary). These officers plus the immediate past Chair (after the first year) will constitute the executive committee. The executive committee will conduct any necessary business in close coordination with the administrative advisor between annual meetings of the technical committee. The Chair will be responsible for preparing the annual meeting agenda, presiding over the annual meeting, appointing necessary subcommittees, and helping the Secretary coordinate any other reports or proposals as required. The Secretary will take meeting minutes, prepare the approved minutes and the annual report, and perform other duties as assigned by the Chair. Specific task subcommittees and coordinators will be appointed as necessary to help coordinate activities among states. At the end of each annual meeting, the Secretary will become Chair and a new Secretary will be elected.

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