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Harvard Forest Data Archive


Extracellular Enzyme Activity in Rhizosphere Soil at Harvard Forest and Pisgah State Forest 2010

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  • Lead: Edward Brzostek, Adrien Finzi
  • Investigators: John Drake, Allison Greco
  • Contact: Information Manager
  • Start date: 2008
  • End date: 2010
  • Status: completed
  • Location: Prospect Hill Tract (Harvard Forest), Pisgah State Forest (NH)
  • Latitude: +42.50 to +42.87
  • Longitude: -72.45 to -72.18
  • Elevation:
  • Taxa: Acer saccharum (sugar maple), Fagus grandifolia (american beech), Fraxinus americana (white ash), Quercus rubra (red oak), Tsuga canadensis (eastern hemlock)
  • Release date: 2015
  • Revisions:
  • EML file: knb-lter-hfr.251.3
  • DOI: digital object identifier
  • EDI: data package
  • DataONE: data package
  • Related links:
  • Study type: short-term measurement
  • Research topic: soil carbon and nitrogen dynamics
  • LTER core area: organic matter
  • Keywords: amino acids, microbes, mycorrhizae, nitrogen, rhizosphere, soil water content
  • Abstract:

    The exudation of carbon (C) by tree roots stimulates microbial activity and the production of extracellular enzymes in the rhizosphere. Here, we investigated whether the strength of rhizosphere processes differed between temperate forest trees that vary in soil organic matter (SOM) chemistry and associate with either ectomycorrhizal (ECM) or arbuscular mycorrhizal (AM) fungi. We measured rates of microbial and extracellular enzyme activity, and nitrogen (N) availability in samples of rhizosphere and bulk soil influenced by four temperate forest tree species (i.e., to estimate a rhizosphere effect). The magnitude of the rhizosphere effects could not be easily characterized by mycorrhizal associations or SOM chemistry. Ash had the lowest rhizosphere effects and beech had the highest rhizosphere effects, representing one AM and one ECM species, respectively. Hemlock and sugar maple had equivalent rhizosphere effects on enzyme activity. However, the form of N produced in the rhizosphere varied with mycorrhizal association. Enhanced enzyme activity primarily increased amino acid availability in ECM rhizospheres and increased inorganic N availability in AM rhizospheres. These results show that the exudation of C by roots can enhance extracellular enzyme activity and soil-N cycling. This work suggests that global changes that alter belowground C allocation have the potential to impact the form and amount of N to support primary production in ECM and AM stands.

  • Methods:

    Site Description

    This research was conducted at two sites, the Harvard Forest and the Pisgah State Forest. The sites have similar land use history and stand age. Soils at both sites are inceptisols classified as Typic Dystrochrepts derived from glacial till overlying granite-schist-gneiss bedrock. Study plots dominated by one of five target tree species were identified at each site. Plots of sugar maple (Acer saccharum) and American beech (Fagus grandifolia) were located in Pisgah State Forest. Plots of eastern hemlock (Tsuga canadensis), red oak (Quercus rubra), and white ash (Fraxinus americana) were located in the Harvard Forest. These species differed in mycorrhizal association, with ash and sugar maple supporting AM fungi and hemlock and beech supporting ECM fungi. Further, hemlock and beech have recalcitrant leaf litter and SOM that is characterized by higher ratios of C-to-N and lower pH than sugar maple and ash. At each site we located six replicate, 8-m radius, monodominant plots within larger mixed hardwood/conifer stands where the target tree species constituted 100 % of standing basal area in the inner 5-m core and 80 % in the entire 8-m radius plot.

    Soil Sampling Protocol

    Soil samples were collected in May, June, and August of 2010 from each of the six replicate plots for each species. We chose the May, June and August time-points because they span the majority of the growing season and are coincident with major phenological events, including leaf out, peak leaf area index, and the start of the seasonal decline in C uptake at the Harvard Forest, respectively. At each sampling point, we collected the top 15 cm of mineral soil using a 5-cm diameter soil bulk-density sampler from each plot. The samples were processed in the laboratory within 24 h of collection. We separated the mineral soil horizons into rhizosphere and bulk soil fractions. Fine roots were removed from each sample. Soil adhering to fine roots was operationally classified as rhizosphere soil.The remaining soil was defined as the bulk soil fraction. After separation, the bulk soil fraction was then sieved through a 2-mm mesh.

    Amino Acid Concentrations

    The 2 M KCl extractable pool size of amino acids was determined for every soil sample across the three sampling dates. Eight grams of each mineral soil fraction were extracted in 30 mL of 2 M KCl. The concentration of amino acids for all extracts was quantified using the o-phthaldialdehyde and b-mercaptoethanol (OPAME) method. Concentrations of amino acid N were determined by comparing the fluorescence of the samples relative to a standard curve composed of glycine.

    Extracellular Enzyme Activity

    We assayed proteolysis following a method modified from Watanabe and Hayano (1995) and Lipson et al. (1999) for every soil fraction and sample date. Initial and incubated subsamples (2–3 g) received 10 ml of a 0.5 mM sodium acetate buffer (pH 5.0) with a small volume of toluene (400 μL) added to inhibit microbial uptake. After the reagent addition, the initial samples were immediately terminated and the incubated subsamples were incubated at 23°C for 4 h. Enzyme activity in all the initial and incubated subsamples was terminated through the addition of 3 mL of a trichloroacetic acid solution. Proteolytic rates for each soil were calculated from the difference between amino acid concentrations in the incubated and initial subsamples of each soil assayed using the OPAME method. For every soil fraction at each sample date, we also assayed the potential activities of the chitinolytic enzyme, n-acetyl-glucosaminidase (NAG) and acid phosphatase and the lignolytic enzymes, phenol oxidase and peroxidase. All the assays were run using a 1 g subsample of each soil homogenized in a pH 5.0 sodium acetate buffer at 23°C. NAG and acid phosphatase activities were determined using a fluorometric microplate assay, while phenol oxidase and peroxidase activities were determined using a colorimetric microplate assay.

    Soil Moisture

    Subsamples of fresh soils (5-10g) were weighed and then dried in an oven at 60°C for 48 hours to determine soil moisture content.

  • Use:

    This dataset is released to the public under Creative Commons CC0 1.0 (No Rights Reserved). Please keep the dataset creators informed of any plans to use the dataset. Consultation with the original investigators is strongly encouraged. Publications and data products that make use of the dataset should include proper acknowledgement.

  • Citation:

    Brzostek E, Finzi A. 2015. Extracellular Enzyme Activity in Rhizosphere Soil at Harvard Forest and Pisgah State Forest 2010. Harvard Forest Data Archive: HF251 (v.3). Environmental Data Initiative:

Detailed Metadata

hf251-01: soil moisture

  1. species: tree species
    • ACSA: sugar maple
    • FAGR: American beech
    • FRAM: white ash
    • TSCA: eastern hemlock
  2. horizon: soil horizon
    • B: mineral soil (bulk)
    • O: organic horizon
  3. site: site
  4. year: year of measurement
  5. may.avg: soil moisture content measured in May, average of 6 plots (%) (unit: dimensionless / missing value: NA)
  6. standard error on soil moisture content average of 6 plots, measured in May (%) (unit: dimensionless / missing value: NA)
  7. june.avg: soil moisture content measured in June, average of 6 plots (%) (unit: dimensionless / missing value: NA)
  8. standard error on soil moisture content average of 6 plots, measured in June (%) (unit: dimensionless / missing value: NA)
  9. august.avg: soil moisture content measured in August, average of 6 plots (%) (unit: dimensionless / missing value: NA)
  10. standard error on soil moisture content average of 6 plots, measured in August (%) (unit: dimensionless / missing value: NA)

hf251-02: enzyme activity

  1. year: year of sampling
  2. month: month of sampling
  3. site: site
  4. species: tree species
    • ASCA: sugar maple
    • FAGR: American beech
    • FRAM: white ash
    • TSCA: eastern hemlock
    • QURU: red oak
  5. plot: plot number
  6. soil: soil
    • B: mineral soil (bulk)
    • O: organic horizon
    • R: mineral soil (rhizosphere)
  7. proteolysis: proteolysis, the rate of protein depolymerization into free amino acids (converted to μg amino-acid-N/g dry soil/per hour) (unit: microgramsPerGramPerHour / missing value: NA)
  8. aa: extractable amino acid concentration (unit: microgramsPerGram / missing value: NA)
  9. nag: b-N-acetylglucosaminidase activity (unit: micromolesPerGramPerHour / missing value: NA)
  10. ap: acid phosphatase activity (unit: micromolesPerGramPerHour / missing value: NA)
  11. phenox: phenol oxidase activity (unit: micromolesPerGramPerHour / missing value: NA)
  12. perox: peroxidase activity (unit: micromolesPerGramPerHour / missing value: NA)