Source: COLORADO STATE UNIVERSITY submitted to
SOIL MICROBIAL STRUCTURAL AND FUNCTIONAL DIVERSITY FOR SOIL QUALITY AND ECOSYSTEM SERVICES
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
0203896
Grant No.
(N/A)
Project No.
COL00613
Proposal No.
(N/A)
Multistate No.
(N/A)
Program Code
(N/A)
Project Start Date
Jul 1, 2010
Project End Date
Jun 30, 2015
Grant Year
(N/A)
Project Director
Stromberger, M. E.
Recipient Organization
COLORADO STATE UNIVERSITY
(N/A)
FORT COLLINS,CO 80523
Performing Department
Soil and Crop Science
Non Technical Summary
Soil microorganisms are integral components of soil quality because of their crucial involvement in many ecosystem processes (e.g., organic matter decomposition and stabilization, nutrient cycling, and plant growth promotion) (Schutter et al. 2001). However, there are significant gaps in our knowledge of the microbes that live in the soil and how they function. This project will advance our ecological understanding of soil microbial communities so that microbial communities can be effectively utilized to monitor soil biological quality in response to disturbance and land management and managed to promote soil ecosystem services. In addition to microbial measurements, we will collect site history data and analyze soils for physical and chemical properties to identify abiotic and biotic factors that regulate microbial processes and ecosystem services across space and time. Our results will be used to generate simple models that growers and land use managers can use to predict how soil microbial communities and their ecosystem services will change in response to disturbances or land management practices.
Animal Health Component
(N/A)
Research Effort Categories
Basic
35%
Applied
60%
Developmental
5%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
1010110110050%
1010110110250%
Knowledge Area
101 - Appraisal of Soil Resources;

Subject Of Investigation
0110 - Soil;

Field Of Science
1100 - Bacteriology; 1102 - Mycology;
Goals / Objectives
The goal of the project is to increase our ecological understanding of soil microbial community structure and function in order to identify important ecosystem service providers and how they are regulated by abiotic and biotic factors that are spatially and temporally heterogenous. Specific objectives are to 1)combine structure and functional measurements of soil microbial communities to monitor soil biological quality in response to disturbance or management practice, 2) determine the relationship between microbial structural and functional diversity by identifying important microbial ecosystem service providers in soil, 3) identify important abiotic and biotic factors that predict soil microbial community structure and function, and 4)identify important spatial and temporal scales over which soil microbial communities and ecosystem service providers operate. The expected outcomes include new knowledge of soil microbial community ecology, including knowledge on how to apply microbial communities for soil quality monitoring and ecosystem services. Other outcomes will be management tools (models) to predict the presence and/or activity of ecosystem service providing microorganisms based on environmental conditions and management history across multiple spatial scales. Education-related outcomes include training for graduate and undergraduate students. Research approach and findings will be also incorporated into current undergraduate and graduate courses to maximize the longevity of the educational impact. Results will be disseminated to people through presentations at scientific meetings, field days, and publication in peer-reviewed journals.
Project Methods
The overall experimental approach will be to compare changes in microbial community biomass, structure and function among disturbed, managed and reference native soils to determine changes in soil biological quality and/or ecosystem services. Studies will be conducted at various spatial and temporal scales on native and working lands, including soils undergoing experimental manipulation. Diverse methods will be utilized to couple structural diversity with functional diversity (Objectives 1 and 2) by measuring abundance (e.g., culture-based methods, fumigation methods, fatty acid biomarkers, and quantitative PCR), composition (fatty acid profiling, metagenomics, pyrosequencing), and function (enzyme assays, activity measurements, molecular studies of functional genes). Site history data will be collected and soils will be analyzed for physical and chemical properties. These data will be co-analyzed with microbial data to identify important factors regulating microbial communities and develop models that predict microbial community structure or function (e.g., multiple linear regression or Classification and Regression Tree analysis) (Objective 3). The spatial and temporal distribution of microorganisms and their activities will be examined by determining the geographical location of each sampling point by Global Positioning System (GPS) and applying geostatistics to microbial data, and by measuring microbial variables across time (days, weeks, to months following disturbance or initiation of management practice) (Objective 4). Results of this work will include management tools (prediction models) that growers and land managers can use to predict the response of microbial communities and their ecosystem services to disturbance or land use practice. Our research approach and findings will be incorporated into current undergraduate and graduate courses to maximize the longevity of our educational impact.

Progress 07/01/10 to 06/30/15

Outputs
Target Audience:My target audience included undergraduate students, graduate students, scientists (domestic and international), growers in the Great Plains region, and Colorado agribusineses. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The research conducted during this period has supported the training of four international, sef-funded graduate studetns. How have the results been disseminated to communities of interest?My results have been disseminated to specific audiences the following ways: 1) undergraduate and graduate students by integrating research results into lectures and class discussions; 2) scientists through publications in peer-reviewed journals and presentations at national and international meetings (volunteered and invited talks); 3) growers in field day visits and workshops (1-3 per year, each year) 4) agribusinesses at annual Rocky Mountain Agribusiness and Independent Crop Consultantconferences (invited talks, 1-2 per year) What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Soil is home to the greatest diversity of life on the planet. This life primarily exists in the forms of bacteria and fungi, of which there are tens of millions of species in soil, and whose biomass in the top foot of soil is in the magnitude of tons per acre. My teaching, research and outreach goals have been to educate others on soil microbial diversity and function, increase our understanding of how land management practices affect soil microbial communities and therefore soil quality, and discover how microbial communities and species can be manipulated to promote ecosystem services, or specific functions that benefit humankind. Because of my efforts, students have been educated in soil biology, soil ecosystem services, and sustainable farming practices. Graduate students have been trained to conduct independent research in soil microbial ecology, and to communicate their findings through scientific writing or oral presentations. Four international students from Libya will return to their home institutions, and transfer the knowledge and research skills gained at CSU to a new generation of soil science students. Soil microorganisms and their activities are sensitive to environmental change, and are effective and responsive indicators of soil quality change in response to land management change. I've shown that soil microbial communities are sensitive to cultivation and crop rotations, and particularly by soil climate, including potential evapotranspiration, but may not be responsive to biochar amendments to soil.There is great interest right now in land application of biochar as a means to improve soil fertility and increase crop yields in temperate soils. Biochar has been promoted (almost feverishly) as a beneficial soil amendment, but without a lot of scientific evidence to support such claims.I and my collaborators conducted laboratory, greenhouse, and field studies to study the effects of biochar amendment on soil quality, microbial communities, and crop yield. We discovered thatwhen applied at practical rates (a few tons per acre, due to cost and limited availability), biochar has very little to no effect on soil microbial communities or nutrient cycling activities. In conclusion, biochar may be ineffective as a fertilizer for increasing crop yields. On the other hand, biochar has the potential to stabilize certain enzymes and protect enzymes from denaturing due to heat stress. Thus, there may be some unique applications of biochar in technologies or applications that require enzymes to be stabilized. My expertise in understanding how soil microbial communities respond to changes in their environment has evolved to an understanding of how to purposely manipulate soils to promote activities of beneficial species, or to inhibit activities of detrimental species. For this project, I focused on three groups of organisms and their ecosystem services: 1) atrazine degrading bacteria, 2) the endogeic earthworm Apporectodea caliginosa, and 3) ACC-deaminase positive bacteria. A phenomenon observed in Colorado is the rapid degradation of atrazine in some soils, which leads to loss of weed control and declines in yield. I and my collaboratorsconducted research to develop a model so that growers could predict whether their soils would degrade atrazine rapidly or not, based on the presence of atrazine-degrading bacteria, soil factors, and management practices. I discovered that atrazine degradation activity can be predicted based on soil pH, organic matter content, and history of atrazine use. Atrazine-degrading bacteria are nearly ubiquitous in Colorado soils, but their activity is high only when soil pH is above 6and the bacteria have been exposed to a previous atrazine application. In conclusion, growers should not apply atrazine repeatedly to soils with pH >6, and should alternate herbicides or grow crops in rotation to avoid atrazine being applied every year. Otherwise, there is a high probability that soils will develop the ability to degrade atrazine rapidly. Results of this work were disseminated in course instruction, scientific presentations at national meetings, a publication in a peer-reviewed journal article, at at multiple crop consultant and agribusiness conferences. Earthworms can have dramatic effects on soil structure and porosity, due to their burrowing habits, and therefore impact water infiltration and water holding capacity in soil. Apporectodea caliginosa is the most common endogeic earthworm in Colorado but is absent or rare in dryland, agricultural soils. I and my collaboratorsinvestigated the potential for A. caliginosa to survive in a Colorado agricultural soil under simulated cycles of drought. Wefound that A. caliginosa can live and reproduce in the soil with relatively low organic matter content (1%), and can survive a series of drought cycles (up to three weeks without precipitation). When active,A. caliginosa increased soil water content by 33 to 41% and altered soil physical properties so that water flow became more tortuous, solute dispersion increased under saturated and unsaturated conditions, and soil drained over a larger range of tensions. In conclusion, if introduced into dryland agricultural soils,A. caliginosa has the potential to reduce upward losses of water, increase water retention, and increase horizontal and vertical dispersion of agricultural chemicals applied to the soil surface.Results of this work were disseminated in course instruction, scientific presentations at national meetings, multiple publications in peer-reviewed journal articles, at at multiple crop consultant and agribusiness conferences. My third set of experiments was to study the potential of belowground interactions for improving drought tolerance in winter wheat.When exposed to drought or salt stress, plants produce elevated concentrations of ethylene ("stress" ethylene), whichreduces root and shoot growth and results in lowered productivity. Certain bacteria called ACC-deaminase positive (ACC+)bacteria produce an enzyme that degrades the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC), whichinhibits the production of stress ethylene and thus potentially can increase plant drought tolerance. My collaborators and I have conducted field and greenhouse studies to determine the effect of ACC+ bacteria on wheat growth under drought stress. We have discovered that interactions between wheat and ACC+ bacteria are cultivar specific, and that only certain cultivars, including RonL, have increased drought tolerance when inoculated with ACC+ bacteria. Other cultivars, including Byrd and Ripper, do not appear to rely on belowground interactions with ACC+ bacteria for drought tolerance. Wehypothesize that wheat-microbial interactions for drought tolerance require the production of specific root exudates that recruit and sustain ACC+ bacteria in the rhizosphere. In a greenhouse study, we demonstrated that Byrd yield under drought stresscould be improved upon if Byrd was grown in the presence of RonL and ACC+ bacteria.In conclusion,ACC+ bacteria can improve the drought tolerance of winter wheat, but require a cultivar-specific factor (presumably root exudates) for recruitment and to sustain their populations prior to onset of drought.Ultimately, our goal is to identify genetic markers of RonL associated with root exudates and ACC+ bacterial interactions, and incorporate these markers in a traditional plant breeding program to develop new cultivars of winter wheat with multiple drought tolerance traits.Results of greenhouse and field studies have been disseminated in course instruction, scientific presentations at national meetings, at at multiple crop consultant and agribusiness conferences.

Publications

  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Bertini, S.C.B., L.C.B Azevedo, M.E. Stromberger, and E.J.B.N. Cardosa. 2015. Soil properties discriminating Araucaria forests with different disturbance levels. Environmental Monitoring and Assessment 187:194 DOI 10.1007/s10661-015-4398-5.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: McDaniel, J.P., G. Butters, K.A. Barbarck, and M.E. Stromberger. 2015. Effects of Aporrectodea caliginosa on soil hydraulic properties and solute dispersivity. Soil Science Society of America Journal 79:838-847.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Ippolito, J.A., M.E. Stromberger, R.D. Lentz and R.S. Dungan. 2015. Hardwood biochar and manure co-application to a calcareous soil. Chemosphere doi.org/10.1016/j.chemosphere.2015.05.039
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Elzobair, K.A., M.E. Stromberger, and J.A. Ippolito. 2015. Stabilizing effect of biochar on soil extracellular enzymes after a denaturing stress. Chemosphere doi.org/10.1016/j.chemosphere.2015.03.018
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Elzobair, K.A., M.E. Stromberger, J.A. Ippolito, and R.D. Lentz. 2015. Contrasting effects of biochar versus manure on soil microbial communities and enzyme activities in an Aridisol. Chemosphere doi.org/10.1016/j.chemosphere.2015.06.044
  • Type: Other Status: Published Year Published: 2015 Citation: Soil Science Society of America. 2015. "Is it Bad to Have Bacteria in Soil? Diversity of Life Keeps 'Bad' Bacteria in Check". News release. July 15, 2015. Available at https://www.soils.org/newsroom/releases/2015/0715/680/
  • Type: Other Status: Published Year Published: 2015 Citation: Soil Science Society of America. 2015. "The Importance of Soil-Dwelling Animals". News release. July 1, 2015. Available at https://www.soils.org/newsroom/releases/2015/0701/674/
  • Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Stromberger, M., P. Byrne, D.K. Manter, T. Weir, and G. Saleh. 2015. Specificity of Root-Bacterial Interactions for Drought Stress Tolerance in Winter Wheat. Annual Meeting of ASA-CSSA-SSSA, November 15-19, 2015, Minneapolis, MN.


Progress 10/01/13 to 09/30/14

Outputs
Target Audience: The target audience for this rating period were wheat growers, soil scientists, crop consultants and other agribusiness professionals. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Training was provided to one graduate student. The student was trained in bacterial culture techniques, seed sterilization and inoculation, planting and maintenance of plants in the greenhouse, root exudate extraction, biomass harvesting, and measurements of root growth and yield traits. How have the results been disseminated to communities of interest? Dr. Stromberger spoke about this research project to the Independent Agriculture Consultants of Colorado meeting in Denver on January 13, 2014. Dr. Stromberger spoke about this research project to the Rocky Mountain Agribusiness Association Meeting in Denver on January 15, 2014. Drs. Byrne and Stromberger described this project in their presentation at the CSU Interdisciplinary Water Seminar, March 31, 2014. Dr. Byrne gave a presentation on this research at the CSU-USDA Central Great Plains Research Station Field Day in Akron, CO on June 11, 2014. Dr. Stromberger gave a presentation on this project to crop consultants and growers at the CSU Extension Crops Clinic in Sterling, CO on November 19, 2014. Dr. Stromberger presented this research to other wheat scientists at the Heat and Drought Wheat Improvement Consortium meeting in Frankfurt, Germany on December 2, 2014. Drs. Byrne, Stromberger, and Weir wrote an article about this project for the 2014 CSU Crops Testing Winter Wheat Report ‘Making Better Decisions’. The greenhouse experiments were featured in tours given to participants in the Biofuels Research Tour, April 17, 2014 and to students in the course Plant Breeding for Drought Tolerance, June 4, 2014. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? When exposed to drought or salt stress, plants produce elevated concentrations of ethylene (“stress” ethylene), which reduces root and shoot growth and results in lowered productivity. Certain bacteria called ACC-deaminase positive (ACC+) bacteria produce an enzyme that degrades the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC), which inhibits the production of stress ethylene and thus potentially can increase plant drought tolerance. A greenhouse study was conducted to determine the effect of ACC+ bacteria on wheat growth under drought stress. The study was conducted in PVC root tubes (1 m tall × 10 cm diameter) filled with a fritted clay growing medium that allows entire root masses to be isolated and characterized. Seven wheat varieties (Byrd, Hatcher, OK06318, Ripper, RonL, Tam112, and WB Cedar) were grown that differ in their drought response under well-watered and moisture stress conditions, with four replicates for each moisture treatment. Seeds were inoculated with a consortia of ACC+ bacteria, enriched from a drought-prone soil in Walsh, CO that mainly consisted of ~12 species of Pseudomonas, or with sterile physiological saline (as a control). Two seeds were planted per root tube and grown for 5 weeks, after which daily irrigation was stopped for half of the tubes. Four weeks later, plants were evaluated for leaf relative water content, biomass and root traits. Under continuous irrigation, roots were concentrated in the top section (0-33 cm) of the tube, whereas a greater proportion of roots grew in the middle (33-66 cm) and bottom (66-99 cm) sections of the tube under water stress (data not shown). Cultivars varied significantly in total root length and lengths of specific root diameter classes, with Hatcher having the greatest root lengths (data not shown). It was expected that inoculation effects on root traits would be dependent upon cultivar type, and indeed the interaction of inoculation x cultivar was significant for total biomass of roots that reached the bottom section of the root tubes. Specifically, RonL showed the largest increase (145%) in root biomass in response to inoculation. Lastly, leaf relative water content under drought stress was significantly greater when inoculated with ACC+ bacteria than without, and the effect was greatest for cultivars RonL and OK06318. The variation in root growth and leaf water content among cultivars suggests that inoculation effects of ACC+ bacteria are cultivar specific, with some cultivars being responsive to inoculation and demonstrating increased drought tolerance (eg., RonL and OK06318) whereas other cultivars are non-responsive (eg., Byrd and WB Cedar). A second greenhouse study was conducted to determine if drought tolerance of winter wheat genotypes that are non-responsive to ACC+ bacteria will be enhanced when grown in the presence of a wheat genotype that is capable of recruiting effective ACC+ bacteria (i.e., RonL). Paired plants were grown in root tubes in the following combinations: RonL-RonL, Byrd-Byrd, OK06318-OK06318, Byrd-RonL and Byrd-OK06318. It was predicted that the performance of Byrd would be enhanced by ACC+ bacteria if grown in the presence of RonL and OK06318. Vernalized seedlings were transplanted into root tubes filled with fritted clay after soaking in either ACC+ bacteria or in sterile physiological saline. Half the tubes were well-watered throughout the study and half were drought stressed by withholding water beginning at the booting stage (7-10 days before flowering). There were a total of six replicate tubes per treatment combination. Yield traits were measured for individual plants per tube at physiological maturity. Analysis of variance revealed that moisture treatment, inoculation, cultivar, and interactions of those factors were significant sources of variation, though not for every trait. Two primary indicators of plant productivity are above ground biomass and seed weight, and results for these traits in the water-stressed treatment are presented in Figure 2 below. RonL-RonL and the Byrd-RonL combinations showed significant increases for both traits in the inoculated tubes (Figure 2A, 2B). Data for individual Byrd plants in intra-specific and con-specific combinations were also examined. The presence of RonL and ACC+ bacteria resulted in highly significant increases in biomass and seed weight of the individual Byrd plant when grown under drought stress. Byrd with OK06318 also showed a significant increase in biomass, and a small (but not significant) increase in seed weight. Thus, this experiment provides additional preliminary evidence that ACC+ bacteria improve growth of RonL when grown under drought stress, and new evidence that aboveground biomass and seed weight of Byrd are better protected under drought stress when grown in the presence of RonL roots and ACC+ bacteria. Given the temporal separation between inoculation and induction of water stress, ACC+ bacteria would have to have been sustained in wheat rhizospheres, presumably through root exudates, to remain effective over the course of the growing season. Co-planting and inoculation had the greatest effect on Byrd when it was co-planted with RonL, which previously was shown to possess a unique root exudate profile and ability to sustain relatively large proportions of ACC+ bacteria in its rhizosphere community.

Publications

  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Ippolito, J.A., M.E. Stromberger, R.D. Lentz, and R.S. Dungan. 2014. Hardwood biochar influences calcareous soil physicochemical and microbiological status. Journal of Environmental Quality 43:681-689.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Bell, C., M. Stromberger, and M. Wallenstein. 2014. New insights into enzymes in the environment. Biogeochemistry 117:1-4.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Arnosti, C., C. Bell, L. McCallister, D. Moorhead, B. Sinsabaugh, M. Stromberger, M. Wallenstein, and M. Weintraub. 2014. Extracellular enzymes in terrestrial, freshwater, and marine environments: New perspectives on system variability and common research needs. Biogeochemistry 117:5-21.
  • Type: Theses/Dissertations Status: Published Year Published: 2013 Citation: Elzobair, K. 2013. Biochar effects on soil microbial communities and resistance of enzymes to stress. Thesis, Colorado State University.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2014 Citation: Stromberger, M., P. Byrne, and T. Weir. 2014. Root-microbial interactions to enhance wheat productivity under drought stress. Heat and Drought Wheat Improvement Consortium, Frankfurt, Germany, December 2-4, 2014
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2014 Citation: Ippolito, J., K. Spokas, J. Novak, R. Lentz, M. Stromberger, and T. Ducey. 2014. Biochar Research: Land Application Advances to Reap its Multifunctional Abilities. Annual meeting of the American Geophysical Union, San Francisco, CA, December 15-19, 2014.


Progress 01/01/13 to 09/30/13

Outputs
Target Audience: The target audience for this rating period were soil scientists and graduate students who conduct research on extracellular enyzmes, biochar, and soil nutrient cycling. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? This project provided training opportunities and professional development for one international graduate student. How have the results been disseminated to communities of interest? Results of this study were presented to professional scientists, academics and graduate students at the 2013 international meeting of the Soil Science Society of America. What do you plan to do during the next reporting period to accomplish the goals? A greenhouse study is underway to investigate the role of plant-growth promoting rhizobacteria (PGPR's) in enhancing drought tolerance in winter wheat.

Impacts
What was accomplished under these goals? Soil microbial extracellular enzymes are involved in numerous ecosystem services, such residue and waste decomposition, C sequestration, and water purification via contaminant degradation. Enzyme stabilization may maintain enzymatic activity and protect enzymes from proteolysis and other denaturing events such as soil heating. Yet, we are still at the beginning of practical applications to manipulate stabilized enzymes for beneficial ecosystem services such as bioremediation, C sequestration, and plant growth promotion. Currently, there is interest in biochar creation and land application for the purposes of biogas production, C sequestration, and increasing soil. Because of its porous nature, researchers have speculated that biochar can provide habitats for bacteria and fungi. If biochar can attract soil microbes and sorb extracellular enzymes, it is possible that biochar could stabilize enzymes and protect enzymes from degradation or denaturation during environmental stress. Biochar could thus be a useful material in cases where enzyme stabilization is desired. Therefore, my laboratory group conducted a study to determine whether a fast pyrolysis biochar (CQuest) derived from oak and hickory hardwood would stabilize extracellular enzymes in soil and prohibit the loss of potential enzyme activity following a denaturing stress such as microwaving. Soil was incubated in the presence of biochar (0, 1, 2, 5, or 10% by weight) and exposed to increasing levels of microwave energy (0, 400, 800, 1600, and 3200 J/g soil). Soil enzymes (β-glucosidase, β-D-cellobiosidase, N-acetyl-β-glucosaminidase, phosphatase, leucine aminopeptidase, and β-xylosidase) were analyzed using fluorescence-based microassays. Results showed that extracellular enzymes responded differently to biochar rate, stress level and their interactions. The main effect of stress level was highly significant on the potential activities of β-glucosidase, β-D-cellobiosidase, N-acetyl-β-glucosaminidase, and phosphatase enzymes, whereby enzyme activity decreased with increasing microwave energy. Potential activity of leucine aminopeptidase was significantly affected by biochar rate, stress level, and their interaction. In addition, potential activity of β-xylosidase was marginally affected by biochar’s interaction with microwave stress. The potential activity of the two enzymes were reduced after a 36-day incubation in the presence of biochar, indicating that biochar absorbs these particular enzymes and/or their substrates and inhibits enzyme function. Although the potential activity of β-xylosidase was reduced in the presence of biochar, intermediate rates (1 and 5 %) of biochar prevented a complete loss of this enzyme’s potential activity after soil was exposed to 400 (1% biochar treatment) or 1600 (5% biochar treatment) J microwave energy / g soil. We conclude that biochar has the potential to stabilize certain extracellular enzymes and prohibit the loss of their activity in soil when exposed to a denaturing stress. While most of the enzymes were not stabilized by biochar, β-xylosidase and leucine aminopeptidase were stabilized and protected to some degree from microwave stress by intermediate rates of biochar application. More research is needed to understand the mechanism(s) by which biochar stabilizes some extracellular enzymes but not others, and how stabilization is affected by different biochars and biochar concentrations in soil.

Publications

  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Nelson, S.D., H.A. Ajwa, T. Trout, M. Stromberger, S.R. Yates, and S. Sharma. 2013. Water and methyl isothiocyanate distribution in soil after drip fumigation. Journal of Environmental Quality 42:1555-1564.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: McDaniel, J.P., K.A. Barbarick, M.E. Stromberger, and W. Cranshaw. 2013. Survivability of Aporrectodea caliginosa in response to drought stress in a Colorado soil. Soil Science Society of America Journal 77:1667-1672.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: McDaniel, J.P., M.E. Stromberger, K.A. Barbarick, and W. Cranshaw. 2013. Survival of Aporrectodea caliginosa and its effects on nutrient availability in biosolids amended soil. Applied Soil Ecology 71:1-6.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Mellor, N., E. Paul, M. Stromberger, and S. Morris. 2013. Changes in ecosystem carbon following afforestation of native sand prairie. Soil Science Society of America Journal 77:1613-1624.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Comerford, N., A. Franzleubbers, M.E. Stromberger, L. Morris, D. Markewitz, and R. Moore. 2013. Assessment and evaluation of soil ecosystem services. Soil Horizons 54 doi: 10.2136/sh12-10-0028.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Burns, R.G., J.L. DeForest, J.C. Marxen, R.L. Sinsabaugh, M.E. Stromberger, M.D. Wallenstein, M.N. Weintraub, and A. Zoppini. 2013. Soil enzymes in a changing environment: Current knowledge and future directions. Soil Biology & Biochemistry 58:216-234.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Hurisso, T., J.G. Davis, J.E. Brummer, M.E. Stromberger, M.M. Mikha, M.L. Haddix, M.R. Booher, and E.A. Paul. 2013. Rapid changes in microbial biomass and aggregate size distribution in response to changes in organic matter management in grass pasture. Geoderma 193-194:68-75.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Elzobair, K., M.E. Stromberger, and J.A. Ippolito. 2013. Biochar effects on soil microbial communities and resistance of enzymes to stress. Annual meeting of the Soil Science Society of America, Tampa, FL, November 3-6, 2013.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: McDaniel, J.P., G. Butters, K.A. Barbarick, and M.E. Stromberger. 2013. Earthworm effect on chemical leaching. Annual meeting of the Soil Science Society of America, Tampa, FL, November 3-6, 2013.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Sukor, A., F. Stonaker, M.E. Stromberger, H. Storteboom, and J.G. Davis. 2013. Fertilizer recovery of cyanobacterial and commonly-used organic fertilizers and lettuce growth response on different soil textures. Annual meeting of the Soil Science Society of America, Tampa, FL, November 3-6, 2013.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Maskus, C., M.E. Stromberger, T. Weir, and P. Byrne. 2013. Metabolomic profiling of root exudates for wheat-microbial interactions. Annual meeting of the Soil Science Society of America, Tampa, FL, November 3-6, 2013.


Progress 01/01/12 to 12/31/12

Outputs
OUTPUTS: My research focuses on soil microorganisms that provide valuable ecosystem services, including microbial activities that promote crop productivity under abiotic stress. When exposed to drought or salt stress, plants produce elevated concentrations of the phytohormone ethylene. "Stress" ethylene then induces defense responses, including reduced root and shoot growth and reduced productivity. Certain plant growth-promoting rhizobacteria produce an enzyme that degrades the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC), which inhibits the production of stress ethylene and results in root elongation and greater tolerance to water stress. These bacteria are referred to as ACC deaminase positive (ACC+) bacteria. In order to maximize the potential of ACC+ bacteria to increase drought tolerance of winter wheat, I am studying how these bacteria are affected by plant genotype, plant growth stage, and agricultural management practice in eastern Colorado. In one of the first studies to assess the natural abundance of these bacteria, my research group found that the abundance of ACC+ bacteria is relatively high in Colorado soil (16,900,000 to 3,280,000,000 colony forming units per g of soil). Moreover, abundance varies under different wheat cultivars when water-stressed, with two- to three-times greater relative abundances of ACC+ bacteria under RonL than under Baca, Hatcher and Ripper when grown under limited irrigation practices. Moreover, the diversity index of ACC+ bacteria was greater in RonL root-associated soil (2.28) than in soil from Ripper (1.19) under dryland conditions. In a collaborative study with Dr. Tiffany Weir (Department of Horticulture and Landscape Architecture), root exudates of RonL and Ripper were collected under hydroponic growth conditions and fractionated in polar, non-polar, and aqueous fractions. These fractions were added to soil to determine their effect on ACC+ bacteria. We discovered that the polar (ethyl acetate) fraction of RonL had a different chemical profile than that of Ripper and when added to soil, this exudate fraction of RonL but not Ripper doubled the relative abundance of ACC+ bacteria. And finally, in a greenhouse study, RonL benefited from inoculation with ACC+ bacteria, where the number of fertile heads was increased by 16-28% compared to uninoculated control, biomass by ~50%, and seed number by 95-130%. In contrast, Ripper productivity was not enhanced by inoculation with ACC+ bacteria. The results of this research have been dessiminated as a M.S. thesis and multiple presentations at the 2012 American Society of Agronomy/Crop Science Society of America/Soil Science Society of America meeting. One manuscript is in preparation, and data have been used as preliminary data in multiple proposals to USDA-NIFA. Ongoing research includes a metabolomics study with Dr. Tiffany Weir to isolate the root exudate chemical(s) that recruit ACC+ bacteria, and a genetic screening of a Doubled Haploid (DH) population of RonL and Ripper to identify plant genetic markers associated with production and secretion of root exudates that select for ACC+ bacteria (with Dr. Patrick Byrne). PARTICIPANTS: Collaborators on this project are former M.S. student Ibrahem Abduelafez, Ph.D. student Asma Elamari, and Dr. Tiffany Weir. The two graduate students were mentored by me, and received training from me. Mr. Abduelafez completed his thesis and defended in Summer 2012. Dr. Tiffany Weir is an Assistant Professor with a joint appointment in the Department of Food Science and Human Nutrition, and the Department of Horticulture and Landscape Architecture. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
This research demonstrates that the abundance of ACC+ bacteria and the potential effects of these bacteria on winter wheat productivity are dependent on winter wheat genotype and irrigation practice/level of water stress. Certain wheat varieties including RonL are able to accumulate more numbers of ACC+ bacteria in their rhizospheres under water stress, and I hypothesize that this is due to the ability of such cultivars to actively select for ACC+ bacteria through specific root exudate chemical(s). Two graduate students have gained research training directly related to this project. One student has subsequently defended his M.S. degree (Summer 2012). This research will result in a new fundamental understanding of a strategy by which winter wheat utilizes interactions with root exudates and soil bacteria for enhanced drought tolerance. Identification of root exudates that recruit ACC+ bacteria will support future research to identify genetic markers associated with exudates, and therefore potential for ACC+ bacterial interaction and improved drought tolerance.

Publications

  • Comerford, N., Stromberger, M., Morris, L., Franzleubbers, A.J., Markewitz, D., and Moore, R. 2012. Soil ecosystem services: a task force report for a SSSA position paper. Annual meeting of the Soil Science Society of America, Cincinnati, OH, October 21-25, 2012.
  • Neghaban-Azar, M., Stromberger, M.E., and Sharvelle, S. 2012. Fate of graywater constituents after long-term application for landscape irrigation. Water Air and Soil Pollution 223:4733-4749.
  • Busby, R.R., Paschke, M.W., Stromberger, M.E., and Gebhart, D.L. 2012. Seasonal variation in arbuscular mycorrhizal fungi root colonization of downy brome (Bromus tectorum), an invasive winter annual. Journal of Ecosystem and Ecography S8:001 doi: 10.4172/2157-7625.S8-001.
  • Busby, R.R., Paschke, M.W., Stromberger, M.E., and Gebhart, D.L. 2012. Arbuscular mycorrhizal fungal community differs between a coexisting native shrub and introduced annual grass. Mycorrhiza doi: 10.1007/s00572-012-0455-x.
  • Stromberger, M.E., Keith, A.M., and Schmidt, O. 2012. Distinct microbial and microeukaryotic communities and translocated carbon in Lumbricus terrestris drilospheres. Soil Biology and Biochemistry 46:155-162.
  • Wallenstein, M., Stromberger, M., and Bell, C. 2012. Bridging the gap between modelers and experimentalists. Eos 93:312.
  • Sharvelle, S., Roesner, L.A., Qiang, Y., Stromberger, M., and Azar, M.N. 2012. Long-term study on landscape irrigation using household graywater experimental study. Final Report. Water Environment Research Foundation WERF) document no. 06-CTS-1CO.
  • Sukor, A., Storteboom, H., Stromberger, M., and Davis, J. 2012. Nitrogen mineralization potential of cyanobacterial fertilizers compared to traditional organic fertilizers applied to clayey and sandy soils. Annual meeting of the Soil Science Society of America, Cincinnati, OH, October 21-25, 2012.
  • Barminski, R., Davis, J., Storteboom, H., Yang, Y.-Y., Dominick, S., Athey, K., Wang, H., and Stromberger, M. Development of an organically certified growth-medium for cyanobacteria. Annual meeting of the Soil Science Society of America, Cincinnati, OH, October 21-25, 2012.
  • Elamari, A., Wier, T., Abduelafez, I., Moragues, M., and Stromberger, M. 2012. Diversity and abundance of ACC deaminase-positive bacteria from winter wheat rhizospheres in Colorado. Annual meeting of the Soil Science Society of America, Cincinnati, OH, October 21-25, 2012.
  • McDaniel, J., Stromberger, M., and Barbarick, K. 2012. Drought stress affects earthworms in a biosolids amended Colorado soil. Annual meeting of the Soil Science Society of America, Cincinnati, OH, October 21-25, 2012.


Progress 01/01/11 to 12/31/11

Outputs
OUTPUTS: The need to identify microbial community parameters that predict microbial activity is becoming more urgent, due to the desire to manage microbial communities for ecosystem services as well as the desire to incorporate microbial community parameters within ecosystem models. Microbial biomass C (MBC) in soil can be increased by adopting alternative management strategies that increase crop residue retention, nutrient reserves, improve soil structure and result in greater water retention. Changes in MBC could subsequently affect microbial activities related to decomposition, C stabilization and sequestration. In dryland agroecosystems, however, changes in MBC and their subsequent activities in response to management may be constrained by water availability. We hypothesized that MBC and potential microbial activities that broadly relate to decomposition (basal and substrate-induced respiration, N mineralization, and β-glucosidase and arylsulfatase enzyme activities) would be differentially affected by no-till, dryland winter wheat rotations distributed along a potential evapotranspiration (PET) gradient in eastern Colorado. Specifically, MBC and activities broadly related to organic matter decomposition (basal and substrate-induced respiration, net N mineralization, and beta-glucosidase and arylsulfatase enzyme activities) would decline along a potential evapotranspiration (PET) gradient. The experiment was conducted across three locations in eastern Colorado that have approximately the same long-term annual precipitation (420 mm yr-1) but different levels of potential evapotranspiration (PET) as measured by an open pan evaporation: Sterling (low PET, 1016 mm y-1), Stratton (medium PET, 1270 mm y-1), and Walsh (high PET, 1900 mm y-1). Results of my CRIS project have been disseminated as four published, peer-reviewed papers in leading soil science journals. My research has also been shared with fellow scientists as seven different presentations at two national and two international meetings. I have also incorporated my research findings into at least four lectures of my SOCR 455 Soil Microbiology course in Fall 2011. PARTICIPANTS: Not relevant to this project. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
I found that MBC was consistently smaller in soils from the high PET soil (191 μg g-1) than in the medium and low PET soils (379 and 398 μg g-1, respectively). Among treatments, MBC was largest under perennial grass (398 μg g-1). In contrast to my hypothesis, potential microbial activities did not consistently follow the same trends as MBC, and the only activities significantly correlated with MBC were beta-glucosidase (r = 0.61) and substrate induced respiration (r = 0.27). In contrast to MBC, specific microbial activities (expressed on a per MBC basis) were greatest in the high PET soils. Specific but not total activities were correlated with microbial community structure, which was determined in a previous study. High specific activity in low biomass, high PET soils may be due to higher microbial maintenance requirements, as well as to the unique microbial community structure (lower bacterial-to-fungal fatty acid ratio and lower 17:0 cy-to-16:1ω7c stress ratio) associated with the high PET soil. In conclusion, microbial biomass should not be utilized as the sole predictor of microbial activity when comparing soils with different community structures and levels of physiological stress, due to the influence of these factors on specific activity.

Publications

  • Busby, R.R., Gebhart, D.L., Stromberger, M.E., Meiman, P.J., and Paschke, M.W. 2011. Early seral plant species' interactions with an arbuscular mycorrhizal community are highly variable. Applied Soil Ecology 48:257-262.
  • Stromberger, M. 2011. Managing microbial communities for ecosystem services: Do we know enough Annual meeting of the Soil Science Society of America, San Antonio, TX, October 16-19, 2011.
  • Abduelafez, I., Moragues, M., Elamari, A.A., Buchleiter, G., and Stromberger, M. 2011. Growth promotion of winter wheat under drought stress by ACC deaminase-positive bacteria. Annual meeting of the Soil Science Society of America, San Antonio, TX, October 16-19, 2011.
  • Busby, R., Paschke, M., Gebhart, D., and Stromberger, M. 2011. Cheatgrass and AMF: Impacts of invasion and mitigation for restoration. Annual meeting of the Soil Science Society of America, San Antonio, TX, October 16-19, 2011.
  • McDaniels, J., Butter, G., Stromberger, M., and Barbarick, K. 2011. Earthworm effects on solute leaching in a biosolids amended soil. Annual meeting of the Soil Science Society of America, San Antonio, TX, October 16-19, 2011.
  • Busby, R.R., Paschke, M.W., Stromberger, M.E., Gebhart, D.L., and Meiman, P.J. 2011. Cheatgrass and AMF: Understanding interactions for improved restoration of invaded lands. Annual meeting of the Ecology Society of America, Austin, TX, August 7-12, 2011.
  • Stromberger, M. 2011. Genes, enzymes, and environment: What do we need to know to manage bioremediation activity in soil International Enzymes in the Environment Conference, Bad Nauheim, Germany, July 17-21, 2011.
  • Stromberger, M. 2011. Strategies to assess and manage microbial communities for ecosystem services. INDO-US Workshop on Precision Agricultural Techniques and Technologies. Ludhiana, India, February 28-March 3, 2011.
  • Shaner, D., Stromberger, M., Khosla, R., Helm, A., Bosley, B., and Hansen, N. 2011. Spatial distribution of enhanced atrazine degradation across northeastern Colorado cropping systems. Journal of Environmental Quality 40:46-56.
  • Hurisso, T.T., Davis, J.G., Brummer, J.E., Stromberger, M.E., Stonaker, F.H., Kondratieff, B.C., Booher, M.R., and Goldhamer, D.A. 2011. Earthworm abundance and species composition in organic forage production systems of northern Colorado receiving different soil amendments. Applied Soil Ecology 48:219-226.
  • Stromberger, M.E., Shah, Z., and Westfall, D.G. 2011. High specific activity in low microbial biomass soils across a no-till evapotranspiration gradient in Colorado. Soil Biology & Biochemistry 43:97-105.


Progress 01/01/10 to 12/31/10

Outputs
OUTPUTS: My research activities this year have included conducting experiments and analyzing data to develop a grower-friendly classification model based on soil and management variables to predict soil atrazine degradation activity; to characterize the effects of soil-feeding earthworms (Aporrectodea caliginosa) on soil hydraulic properties; the response of soil microbial communities to biochar amendments under field and greenhouse conditions; the abundance, diversity and activity of ACC deaminase positive bacteria under different winter wheat varieties and irrigation regimes; and the effect graywater irrigation on urban soil quality. These activities have provided training and research experience to one undergraduate student, three graduate students that I advise, plus one additional graduate student in Civil Engineering (graywater project). I have incorporated my research activities as case studies in SOCR 755 Advanced Soil Microbiology, SOCR 455 Soil Microbiology and SOCR 456 Soil Microbiology Laboratory. My research findings have been disseminated through eight presentations at four national and international meetings, one technical report to the Water and Environment Research Foundation, and two publications in peer-reviewed journals. PARTICIPANTS: Graywater Project: Drs. Sybil Sharvelle and Larry Roesner, Department of Civil Engineering, Colorado State University. Dr. Yaling Qian, Department of Horticulture, Colorado State University. Graduate students Masoud Azar and Chris Olson, Department of Civil Engineering, Colorado State University. Atrazine Degradation Project: Dr. Raj Khosla, Department of Soil and Crop Sciences, Colorado State University. Dr. Dale Shaner, Water Management Research Unit, USDA-ARS, Fort Collins. Earthworm Project: Dr. Ken Barbarick, Department of Soil and Crop Sciences, Colorado State University. Graduate student Jacob McDaniel, Department of Soil and Crop Sciences, Colorado State University. Biochar Project: Dr. Jim Ippolito, USDA-ARS, Kimberly, Idaho. Collaborator Mr. Morgan Williams, Flux Farm, Carbondale, Colorado. ACC Deaminase Bacteria Project: Drs. Marc Moragues and Scott Haley, Department of Soil and Crop Sciences, Colorado State University. Dr. Tiffany Weir, Department of Horticulture, Colorado State University. TARGET AUDIENCES: Graywater Project: Water use managers, Proctor and Gamble soap company, household owners, and scientists and engineers at professional meetings. Atrazine Degradation Project: Colorado growers attending outreach meetings with Dr. Shaner. Scientists at professional meetings and readers of scientific journals and USDA ARS publications. Undergraduate and graduate students enrolled in SOCR 755, 455 and 456. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
The graywater study provides one example of the impact of my research. Soil quality data were collected from three homes located in Escondido, CA, Fort Collins, CO, and Dallas, TX, where graywater had been applied for five or more years. For each site, a sample area was selected where graywater was applied for irrigation and a control area was also sampled with similar vegetation that had been irrigated with fresh water. Analysis of soil samples included measurement of several microbial and chemical properties, including sodium adsorption ratio (SAR), counts of fecal indicator organisms (total coliforms, E. coli, enterococci, and Clostridium perfringens), and concentration of the two most widely used surfactants in personal care products surfactants: alkyl ethoxy sulfate (AES) and linear alkyl benzene sulphonates (LAS). All soil samples collected for this study had a SAR below 5. The highest SAR values were observed at the California household (3.3), where soils received manure as an amendment. In general, SAR values were slightly elevated in areas irrigated with graywater compared to fresh water among the households (except Colorado), although levels were not found to be high enough to cause concern for soil quality. Across all households, the average number (and standard deviation) of E. coli was 9 cells/g soil (sd=19) for fresh water-irrigated soil and 11 cells/g soil (sd=16) for graywater-irrigated soil, with no statistically significant effects of graywater irrigation (P=0.80; paired t-test). The average number (and standard deviation) of enterococci was 2,630 cells/g soil (sd=20) for fresh water-irrigated soil and 480 cells/g soil (sd=80) for graywater-irrigated soil, with no statistically significant effects of graywater irrigation (P=0.40; paired t-test). Clostridium perfringens were below the limits of detection in all soils (<10 colony forming units/g soil), except for the fresh water-irrigated soil in Texas, where C. perfringens was quantified as 300 colony forming units/g soil. Surfactant concentrations varied among the four households and ranged between 5-to-65 ppb AES and ~8-30 ppb LAS. Concentrations of AES were 2-to-6 times greater in graywater irrigated areas compared to areas irrigated with fresh water. LAS was detected only in graywater-irrigated soils, except at Texas, where the concentration was 3 times greater in the soil receiving fresh water compared to graywater. This research indicates that graywater has minimal impact to soil SAR as compared to reclaimed wastewater, and LAS and AES do not appear to accumulate to alarming levels in soil irrigated with graywater. While graywater may contribute fecal bacteria to soil, fresh-water irrigated soils were also contaminated with fecal organisms due to the presence of pets and wildlife. These findings will be critical to support the legalization or broader application of graywater re-use in states where its use is illegal or restricted (such as Colorado).

Publications

  • Sharvelle, S., Roesner, L.A., Qiang, Y., Stromberger, M. 2010. Long-term study on landscape irrigation using household graywater: experimental study. Water Environment Research Foundation (WERF) document no. 06-CTS-1CO.
  • Shaner, D., Stromberger, M., Khosla, R., Helm, A., Bosley, B., Hansen, N. 2011. Spatial distribution of enhanced atrazine degradation across northeastern Colorado cropping systems. J. Environ. Qual. doi:10.2134/jeq2010.0193
  • Stromberger, M.E., Shah, Z., Westfall, D.G. 2011. High specific activity in low microbial biomass soils across a no-till evapotranspiration gradient in Colorado. Soil Biol. Biochem. 43:97-105.
  • Sharvelle, S., Azar, M.N., Stromberger, M., Roesner, L., Olson, C. 2010. Effect of graywater application for landscape irrigation on soil quality. In: Proceedings of the 2010 WEFTEC Conference, New Orleans, LA, October 2-6, 2010, 6 p.
  • Stromberger, M.E., Khosla, R., Shaner, D. 2010. Spatio-temporal analysis of atrazine degradation and associated attributes in eastern Colorado soils. In: Proceedings of the 10th International Conference on Precision Agriculture, Denver, CO, July 18-21, 2010, 15 p.
  • Shaner, D.L., Khosla, R., Stromberger, M. 2010.Spatial and temporal changes in atrazine degradation rates in soil. In: Proceedings of the 10th International Conference on Precision Agriculture, Denver, CO, July 18-21, 2010, 11 p.


Progress 01/01/09 to 12/31/09

Outputs
OUTPUTS: Study Title: Spatial Analysis of Atrazine Degradation and Associated Attributes in Eastern Colorado Soils Background: Atrazine [2-chloro-4-(ethylamino)-6-(isopropylamino)-s triazine] is a widely used, soil-applied herbicide for controlling many broadleaf weeds, and growers depend on atrazine to provide season-long residual control. However, in the last 20 years, atrazine catabolism has rapidly evolved within soil microorganisms, and many soils have developed enhanced capacities to degrade atrazine. As a result, the half-life of atrazine in some soils has been reduced from 60 to a few days or less. While the presence of atrazine-degrading bacteria determine a soil's potential to degrade atrazine, the actual manifestation of enhanced atrazine degradation is likely regulated by soil physicochemical properties and management strategies that are spatially and temporally heterogeneous and control bacterial expression of atrazine degradation enzymes. Goal: To conduct a comprehensive field study to identify the relative influence of atrazine-degrading microorganisms/genes, management, and environmental factors on enhanced atrazine degradation in eastern Colorado soils. Methods: Soils were collected from eastern Colorado at multiple spatial scales and analyzed for soil physical and chemical properties, the presence of an atrazine degradation gene (atzC), and atrazine dissipation time (DT50) under laboratory conditions. Management practices, including atrazine use history, were also recorded. Data were subjected Classification and Regression Tree (CART) analysis to develop a prediction model for atrazine degradation based on management practice and soil predictor variables. PARTICIPANTS: 1) Dr. Mary Stromberger, Soil Microbiologist, Department of Soil and Crop Sciences, 1170 Campus Delivery, Colorado State University, Fort Collins, CO 80523; 2) Dr. Raj Khosla,Precision Agriculturalist, Department of Soil and Crop Sciences, 1170 Campus Delivery, Colorado State University, Fort Collins, CO 80523; and 3) Dr. Dale Shaner, Plant Physiologist, USDA-ARS, Water Management Research Unit, 2150 Centre Avenue, Building D, Suite 320, Fort Collins, CO 80526. Field sites were selected and soil samples were collected by Drs. Khosla and Shaner. Dr. Shaner conducted the atrazine degradation half-life assays in his laboratory. Dr. Stromberger analyzed soils for the presence of the atzC atrazine degradation gene and performed the CART analysis of the data. Dr. Khosla analyzed the spatial distribution of data generated in the study. All participated in the manuscript preparation. TARGET AUDIENCES: Target audiences are mainly weed scientists seeking a greater understanding of the mechanism(s) of enhanced atrazine degradation, as well as growers who routinely use atrazine to control broadleaf weeds. Other target audiences will be microbiologists interested in spatial distribution of microbial genes for catabolism, or in rapid evolution of microbial catabolic activities. Study results will be communicated to via scientific presentations, publications, and extension and outreach activities. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
Study Results: There was good agreement between atrazine DT50 and the presence of atzC gene for Larimer, Yuma, Kit Carson and Morgan county soils. Discrepancies were observed, however, for soils from Phillips or Logan counties. For example, all Phillips soils were positive for the atzC gene, despite having a wide range in degradation half-lives that ranged from 14.9 to 4.4 d. Also, one Logan field soil had a slow atrazine degradation half-life of Logan soils (13.6 d) but was positive for atzC. In an earlier study conducted in 2007, this soil had a much faster atrazine degradation half-life of 3.5 d. From 2007 to 2009, it appears that this particular soil has lost its enhanced atrazine degradation capability while maintaining the atzC gene. Soils that rapidly degraded atrazine (DT50 < 3 days) were correctly classified by the CART prediction model 80% of the time based on atrazine use history, soil pH and organic matter content. Specifically, atrazine degraded rapidly in soils where atrazine had also been applied the previous year, and if the soil pH was > 6.3. In addition, soils that no recent history of atrazine application could also degrade atrazine at moderate to rapid rates if the organic matter content was > 1.75%. Changes in Knowledge: This study demonstrated that while atrazine biodegradation is ultimately dependent on the presence and activity of atrazine-degrading microorganisms, these bacteria and their atrazine degradation genes were widespread and therefore their presence did not predict actual soil degradation activity. Instead, recent atrazine use and soil pH were the main model predictors of atrazine degradation activity in Colorado soils. These factors likely regulate the population size, diversity and activity of atrazine degraders and thus the expression of atrazine degradation activity. The impact of these findings are that 1) growers should adopt cropping rotations and/or limit atrazine applications to reduce the risk of a soil developing enhanced atrazine degradation activity and 2) as an environmental contaminant, atrazine should be relatively easy to mitigate provided organic matter content and pH are adequate. Impacts: 1) Greater understanding of management and environmental conditions that support enhanced degradation of atrazine in eastern Colorado soils, 2) one abstract submitted so far for presentation at a scientific meeting, 3) one manuscript in final draft stage for submission to a peer-reviewed journal, 4) supporting data for a proposal to USDA Soil Processes program to continue research on the spatial and temporal dynamics of atrazine degradation, but with expansion to regional and global scales.

Publications

  • Shaner, D.L., Khosla, R., Stromberger, M. 2010. How Rapidly Does Enhanced Atrazine Degradation Develop. Weed Science Society of America Meeting Abstracts. Denver,Colorado Feb 17-11, 2010.


Progress 01/01/08 to 12/31/08

Outputs
OUTPUTS: Current restoration of the ponderosa pine forests in the Colorado Rocky Mountains involves thinning and reintroduction of fire through prescribed burns to bring back the historical forest structure and function. Thinning for restoration or timber harvesting produces large amounts of slash material, which commonly is disposed of by burning in large piles. However, soil heating as a result of pile burning is detrimental to soil microorganisms, results in unattractive scars, and may assist in the establishment and spread of non-native plants. Alternatively, thinned forest fuel can be masticated into chips or chunks which are then spread on the forest floor and left to decompose naturally. A demonstration study was initiated in 2004 in a ponderosa pine-Douglas fir forest in Colorado, USA to determine changes in soil organic C (SOC; 0-15 cm depth) after forest thinning, with either removal of thinned material or surface addition of masticated wood. I expected that wood chips added to the forest floor would increase soil C levels over time based on studies of previously published studies on wood chips and slash residue management in loblolly pine clearcuts and a mixed conifer and hardwood forest. In my study, mean SOC concentration was highest in soil under wood chips in 2004, but SOC levels declined significantly under wood chips during 2005-2006. In 2006 and 2007, SOC in thinned-and-chipped soil was 30-to-33% lower than in untreated and thinned-and-removed soils. I also found in 2007 that fungal biomass C in the thinned-and-chipped soil was similar to that of the untreated soil but greater than in the thinned-and-removed soil, which indicated a stimulatory effect of wood chips on fungal biomass following tree thinning. Peroxidase activity was highly variable, but activity was highest in the thinned-and-chipped soil, compared to activities in the thinned-and-removed and untreated soils, respectively. These results have been disseminated nationally as an invited seminar at Purdue University, and internationally as a poster presentation at the European Geosciences Union meeting and as a submitted publication to the journal Soil Biology and Biochemistry. PARTICIPANTS: Dr. Mary Stromberger (CSU) collected soil samples in 2007 and analyzed samples for total carbon, total inorganic C, fungal biomass and peroxidase enzyme activity. She prepared the manuscript and presented the findings as an invited talk and a volunteered poster. Dr. Peter Marchand (Catamount Center for Geography of the Southern Rockies, Colorado Springs, CO) collected soil samples in 2004-2006 for carbon analysis. Dr. Howard Drossman (Colorado College, Colorado Springs, CO) analyzed 2004-2006 soil samples for total carbon content. Dr. William Massman (USDA Forest Service) provided access to the research site. TARGET AUDIENCES: Target audience is USDA Forest Service personnel and forest land managers. Efforts include laboratory instruction and a research experience for one undergraduate student. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
On federal, state and private forestland in the Rocky Mountain West, mechanical fuel reduction treatments are widely prescribed, but very little information exists on mastication treatment effects for the Rocky Mountains and the Colorado plateau. My study found that soil organic carbon levels declined over time under wood chips. I concluded that this was due to a priming effect, whereby dissolved organic matter that presumably leached from wood chips into soil stimulated soil fungi and resulted in the additional decomposition of native soil organic matter. This is important information for forest managers, who promote carbon sequestration as an important forest ecosystem service and therefore should consider the potential negative impact of thinning and mastication on soil carbon levels.

Publications

  • Stromberger, M., and Massman, W. (2008) Response of soil microbial community structure and C cycling functions to forest fuel mastication treatments. Annual meeting of the European Geosciences Union, Vienna, Austria, April 13-18, 2008.
  • Stromberger, M.E., Marchand, P., Drossman, H., and Massman, W.J. 2008. A demonstration of reduced soil carbon under wood chips in a Colorado ponderosa pine forest. Soil Biol. Biochem. (pending)


Progress 01/01/07 to 12/31/07

Outputs
OUTPUTS: Growing urban population, declining groundwater levels, and drought are factors leading to reduced water quantities for irrigated land in the semiarid western U.S. Developing sustainable limited irrigation systems necessitates an understanding of how reduced water availability affects soil microbial processes and ecological interactions critical to crop productivity and soil conservation. My laboratory, along with collaborator Dr. Neil Hansen, conducted a two year (2006-2007) study to determine the effects of deficit-irrigated maize cropping systems on rhizosphere microbial communities and activities. Maize rhizosphere soil samples were collected from furrow or sprinkler irrigated field plots in August 2006 and 2007, shortly after tasseling. 2006 was an abnormally dry year for the study location, whereas there were no drought conditions in 2007. Maize roots were excavated with a shovel from four replicate plots of the following cropping systems x furrow or sprinkler irrigated treatment: maize-alfalfa rotation, 740 mm anticipated annual consumptive maize water use (ET) (fully irrigated); maize-alfalfa rotation, 410 mm ET; and maize-sunflower-winter wheat rotation, 310 mm ET. Soil adhering to roots was considered to be rhizosphere soil, and this was analyzed for microbial biomass carbon (MBC), community fatty acid methyl ester (EL-FAME) structure, and C mineralization and beta-glucosidase activities. For both years, MBC and C mineralization activity were not significantly affected by type or amount of irrigation, indicating that soil C pools are not yet affected by deficit irrigation practices. In 2006 but not 2007, beta-glucosidase activity was significantly lower in rhizosphere soil from the deficit-irrigated maize-sunflower-wheat rotation compared to other treatments. Beta-glucosidase is an enzyme involved in cellulose decomposition, and its activity in this study was sensitive to soil water stress. Principle components analysis of EL-FAMEs separated rhizosphere communities according to deficit irrigation-rotation systems in 2006, and deficit irrigation significantly reduced relative amounts of fungal EL-FAME but increased relative amounts of Gram-positive bacterial EL-FAMEs. Bacterial stress ratio 17:0cy:16:1&#969;7c was not affected by deficit irrigation but was greater in furrow than in sprinkler-irrigated soil. This study found that fungal biomass and beta-glucosidase activity responded negatively to deficit irrigation in drought years. However, these properties recovered in deficit-irrigated soils the following year, indicating the capacity of the microbial community to tolerate deficit irrigation in non-drought years. Interestingly, furrow irrigation, which floods soils, posed a greater stress to soil bacteria than did reduced water availability because of deficit irrigation. Considering the increased frequency of drought in the Great Plains, growers should consider management practices that conserve and protect soil fungal populations, such as no-tillage and organic amendments, due to the sensitivity of fungi to water-stress and the critical roles fungi have in residue decomposition, soil aggregation, and soil stabilization. PARTICIPANTS: 1. Dr. Mary Stromberger, Department of Soil and Crop Sciences, CSU, project PI. 2. Kiera Coffin, B.S. Soil Science, December 2007. Kiera was employed in Dr. Stromberger's laboratory from February 2005 to December 2007. Dr. Stromberger mentored and trained Kiera in laboratory techniques and scientific communication. 3. Dr. Neil Hansen, Department of Soil and Crop Sciences, CSU, project collaborator. Dr. Hansen provided access to the research plots and field sampling crews for Stromberger's study. Dr. Stromberger and Kiera Coffin conducted the research on this project. Dr. Stromberger analyzed the data, and Kiera presented some of the findings at the 2007 Soil Science Society of America meeting. TARGET AUDIENCES: Irrigated agriculture growers of the Great Plains

Impacts
We know very little about how the soil environment will respond to diminishing supplies of water in irrigated agriculture. This short-term study found that maize rhizosphere soil microbial biomass and bacterial community structure were resilient to reduced water availability due to deficit irrigation. In a drought year, however, deficit irrigation resulted in reduced fungal biomass and activity of beta-glucosidase, an enzyme involved in cellulose degradation. Deficit-irrigated agroecosystems should incorporate management practices, such as no-tillage and organic matter amendments, to protect fungal populations due to their sensitivity to water stress and their residue decomposition and soil stabilization activities.

Publications

  • Stromberger, M., Shah, Z.. and Westfall, D. 2007. Soil microbial communities of no-till dryland agroecosystems across an evapotranspiration gradient. Appl. Soil Ecol. 35:94-106.
  • Coffin, K., Stromberger, M., and Hansen, N. Rhizosphere microbial communities under deficit irrigation. Annual meeting of the Soil Science Society of America, New Orleans, LA, November 4-8, 2007.


Progress 01/01/06 to 12/31/06

Outputs
The spread of antibiotic resistance has been identified as one of the world's most pressing public health concerns. My laboratory has already determined that biosolids land application affects microbial structural and functional diversity in soil (see Sullivan et al. publications below). We have also determined that biosolids contain significant quantities of antibiotics within the tetracyclines class (average of 2,000 ppb total tetracyclines, n=6) as well as significant numbers of oxytetracycline-resistant bacteria (at least 10,000 to 100,000 cells/g biosolids). Thus, biosolids land application introduces antibiotics and antibiotic-resistance genes into the environment, which might influence the spread of antibiotic resistance. In 2006, a field study was conducted to determine the effects of biosolids land application on antibiotic resistance patterns of soil bacteria. There were two field locations with a long-term (+10 y) history of winter wheat-fallow cropping rotation and biosolids land application, with 4 plots per treatment. Biosolids are surface applied without incorporation at agronomic rates as needed at Byers, CO (4-7 tons/ha). At Bennett, CO, biosolids are biannually applied at 11 tons/ha and incorporated to a depth of 20 cm. Control plots receive inorganic fertilizer at equivalent N-based rates. Soil samples were collected in May (0-5 cm depth at Byers; 0-20 cm depth at Bennett) and analyzed for percent OTC-resistant bacteria by plating diluted soil suspensions onto 10% TSB agar plates with or without OTC. Representative OTC-resistant isolates were identified by their fatty acid profiles and tested for their sensitivities to other antibiotics. At Byers, the percentage of OTC-resistant bacterial isolates was similar between control plots and plots receiving biosolids. However, when biosolids were applied at relatively high rates and incorporated into soil, as at Bennett, the percent of OTC-resistant bacteria were 3-4x greater compared to background levels. Specifically, 60% of culturable soil bacteria were resistant to 1 ppm OTC compared to 27% in control plots, and 4% of bacteria were resistant to 10 ppm OTC in biosolids-amended plots compared to <1% in control plots. Many of the OTC-resistant bacteria cultured from control and biosolids-amended plots at Byers and Bennett were actinomycetes, which naturally produce tetracyclines and have an intrinsic resistance to this class of antibiotics. Some OTC-resistant isolates were unique to the biosolids-amended plots only, and isolation of Yersinia from biosolids-amended plots suggests some biosolids bacteria may persist in soil for relatively long periods of time. Many isolates resistant to OTC were also resistant to at least one other antibiotic, and there was evidence for multidrug resistance development in biosolids-amended plots but not in control plots. Specifically, Stenotrophomonas maltophilia from Bennett control plots showed only an intermediate resistance against tetracycline, whereas S. maltophilia isolates from Bennett biosolids-amended plots were also resistant to streptomycin, penicillin, erythromycin, kanamycin, and novobiocin.

Impacts
The spread of antibiotic resistance has been identified as one of the world's most pressing public health concerns. My findings show that land application of biosolids is a means of introducing tetracyclines and oxytetracyline-resistance genes to soil. When applied at relatively high rates, biosolids can increase antibiotic resistance among soil bacteria, thus contributing to the spread of antibiotic resistance in the environment.

Publications

  • Massman, W.J., Frank, J.M., Jimenez Esquilin, A.E., Stromberger, M.E., and Shepperd, W.D. 2006. Long term consequences of a controlled slash burn and slash mastication to soil moisture and CO2 at a southern Colorado site. Proceedings of the 27th Conference on Agricultural and Forest Meteorology, American Meteorological Society, San Diego, CA, May 22-26, 2006, 6 p.
  • Stromberger, M., and Coffin, K. 2006. Impacts of biosolids-borne antibiotics on soil microbial communities. Annual meeting of the Soil Science Society of America, Indianapolis, IN, November 12-16, 2006.
  • Coffin, K., and Stromberger, M. 2006. Emerging contaminants in the environment: biosolids-borne antibiotics. Annual meeting of the Soil Science Society of America, Indianapolis, IN, November 12-16, 2006.
  • Jimenez, A. 2006. Fire effects on soil microbial community structure and function in a ponderosa pine ecosystem. PhD Dissertation. Colorado State University, Fort Collins, CO.
  • Sullivan, T.S., Stromberger, M.E, and Paschke, M.W. 2006. Parallel shifts in plant and soil microbial communities in response to biosolids in a semi-arid grassland. Soil Biol. Biochem. 38:449-459.
  • Sullivan, T.S., Stromberger, M.E., Paschke, M.W., and Ippolito, J.A. 2006. Long-term impacts of infrequent biosolids applications on chemical and microbial properties of a semi-arid rangeland soil. Biol. Fertil. Soils 42:258-266.


Progress 01/01/05 to 12/31/05

Outputs
Biosolids contain therapeutic agents whose activities can persist in the environment. Risks posed by these agents following biosolids land application are unknown, but there is the potential for soil microorganisms and their nutrient cycling funcions to be affected. One concern is the development of antibiotic resistance in indigenous microbes exposed to biosolids containing antibiotics, particularly if antibiotic-resistant microorganisms come into human contact. My goals are 1) to screen for the occurrence of antibiotics in biosolids collected from several wastewater treatment facilities, 2) to determine the persistence of biosolids-associated antibiotics in soils under aerobic incubation, and 3) determine effects of antibiotics on microbial activity and antibiotic resistance patterns. A survey of biosolids obtained from six treatment facilities (three aerobic and three anaerobic digestor systems) revealed the presence of macrolides (erythromycin, roxithromycin, and tylosin), sulfonamides (including sulfathiazole, sulfachlorpyridazine, and sulfadimethoxine), and tetracycline antibiotics (tetracycline, oxytetracycline, chlortetracycline, demeclocycline, meclocycline, and doxycycline). Of the three antibiotic classes, tetracyclines were present in the highest concentrations. Concentrations of tetracyline, oxytetracycline and meclocycline ranged from 100 to over 10,000 ppb, while macrolide and sulfonamide antibiotics were generally less than 10 ppb. There was no consistent pattern of a particular antibiotic being in greater concentration in aerobically versus anaerobically digested biosolids. I also isolated numerous oxytetracycline-resistant bacteria from the biosolids samples. The number of oxytetracycline bacteria was less than 1% when bacteria were isolated on agar plates supplemented with 5 ppm oxytetracycline; however, this still represents a significant number of antibiotic-resistant organisms, considering that the total number of culturable bacteria in the biosolids samples was greater than 100,000,000 per g biosolids (dry weight basis). Thus, land application of biosolids, particularly if applied multiple times at high rates, will not only add antibiotics to soil, but antibiotic-resistance bacteria as well. There is the potential for these bacteria to persist and even multiply in soil, and if the resistance genes are carried on plasmids, the genes may be spread to indigenous soil bacteria via horizontal gene transfer, or conjugation events. Future studies will address the persistance of antibiotics, specifically oxytetracycline, in aerobically-incubated soil and the potential for indigenous soil microbes to acquire resistance to oxytetracycline either by direct exposure to the antibiotic or by acquisition of the resistance genes from bacteria originating from the biosolids material.

Impacts
Historically, the environmental risks associated with land-application of biosolids were largely considered to be over-application of phosphorous and heavy metals. However, we now know that antibiotics are not destroyed by waste water treatment practices and can be detected in biosolids materials. The application of biosolids to land introduces not only antibiotics, but antibiotic-resistant bacteria as well, to soil. A social impact would arise if humans come into contact with soil-borne bacteria carrying antibiotic resistance genes (either from direct contact with soil or from crops contaminated with soil and/or biosolids), or if antibiotic-resistant bacteria move from soil into water (surface runoff events or downward migration into groundwater) which is then consumed by humans. I am conducting research to determine whether such risk concerns are warranted.

Publications

  • No publications reported this period