Soil biological, chemical and physical parameters and herbage yield in a field experiment with organic and inorganic fertilizers on peat grassland in the Netherlands

To evaluate the performance of organic and inorganic fertilizers for regeneration of ecosystem services in peat grasslands with biodiversity goals, we carried out a field experiment in the western peat district in the Netherlands. The fertilizers tested represent the current practice and potential alternatives for regenerative grassland management on drained peat.

Experimental setup

The field experiment (2013 – 2015) was conducted on a permanent grassland on peat soil (Terric Histosol; SOM 56 g 100 g⁻¹ and pH_KCl of 4.5 in 0-10 cm) at the experimental dairy farm at Zegveld (the Netherlands). In March 2013, a randomized block experiment (six blocks) was laid out with six fertilizer treatments and a control treatment (no fertilizer: “Contr”). The fertilizer used were: conventional dairy cattle slurry manure (“Slurry”), mature compost of kitchen and garden waste (“Comp”), dairy cattle farmyard manure (“FYM”), solid fraction of the cattle slurry manure (“SFrac”, obtained by pressurized filtration), inorganic N fertilizer (“IF”; calcium ammonium nitrate, 27% N) and a combination of inorganic N fertilizer and sawdust (“IF+SD”). Plot size was 4 × 10 m; for the Slurry treatment plots were 5.2 × 10 m. Slurry was applied by slit injection, the other fertilizers were applied by hand. Target application rate was 120 kg total N ha⁻¹ yr⁻¹, divided in two applications per year (February/March and May). This is relatively low for conventional grasslands but usual for grasslands with biodiversity goals (Kleijn et al., 2004). The amount of C_total applied in Comp was taken for the rate of sawdust to be applied. All plots were fertilized with 200 kg K₂O ha⁻¹ yr⁻¹ (applications in March and May) (Commissie Bemesting Grasland en Voedergewassen, 2019). Fertilizer application quantities and organic matter and nutrient inputs are provided in Fertilizer_intput.csv (dataset).

The grassland had an history of conventional management with mainly cutting, winter grazing with sheep and a normal fertilization regime with both slurry manure and inorganic fertilizer. The normal cutting and grazing regime was continued in the first two years of the experiment; during 2015, the monitoring year, the plots were not grazed and only cut for herbage measurements.

Measurements

From April to October 2015, soil and aboveground measurements were carried out. Most soil parameters were measured in October. Earthworms and insect larvae are an important food source for meadow birds during the pre-breeding period in spring (Galbraith, 1989) and were therefore sampled in April. Soil moisture and penetration resistance were measured both in April and October.

Soil biological parameters

Earthworms and insect larvae were sampled in the top soil layer in two soil cubes (20 × 20 × 20 cm) per plot. Earthworms were hand-sorted, counted, weighed and fixed in alcohol prior to identification. Both adults and juveniles were identified to species (Sims and Gerard, 1985; Stöp-Bowitz, 1969) and classified into functional groups (Bouché, 1977). Crane flies (Tipulidae; leatherjackets) or click beetles (Elateridae; wireworms) larvae were counted.

Phospholipid fatty acids (PLFA) were measured in October. PLFA were extracted from 4 g of fresh soil (Palojärvi, 2006), and analyzed by gas chromatography (Hewlett-Packard, USA). PLFA i15:0, a15:0, 15:0, i16:0, 16:1ω9, i17:0, a17:0, cy17:0, 18:1ω7 and cy19:0 were chosen to represent bacteria and PLFA 18:2ω6 was used as a marker of saprotrophic fungi (Hedlund, 2002). The neutral lipid fatty acid (NLFA) 16:1ω5 occurs in storage lipids of arbuscular mycorrhizal fungi (AMF) and was used as marker of AMF (Vestberg et al., 2012). PLFA i15:0, a15:0, i16:0, i17:0 and a17:0 were used as a measure of Gram-positive bacteria, and cy17:0 and cy19:0 for Gram-negative bacteria. PLFA 10Me16:0, 10Me17:0 and 10Me18:0 represented actinomycetes.

Soil chemical parameters

A soil sample from the 0−10 cm layer (c. 50 randomly taken soil cores) per experimental plot was collected in October (auger diameter 2.3 cm; Eijkelkamp grass plot sampler, Giesbeek, the Netherlands), was sieved (1 cm mesh size) and homogenized. One sub-sample was taken for analysis of hot water extractable carbon (HWC) according to Ghani et al. (2003) and one for chemical analysis. Prior to analysis of soil acidity (pH_KCl), soil organic matter (SOM), total carbon (C_total), total nitrogen (N_total), total phosphorus (P_total) and ammonium-lactate extractable P (P_AL) by Eurofins Agro (Wageningen, the Netherlands), the sub sample was dried at 40°C. Soil pH_KCl was measured according to NEN-ISO 10390 2005. SOM was determined by loss-on-ignition (NEN 5754 2005). C_total was measured by incineration at 1150°C, and determination of the CO₂ produced by an infrared detector (LECO Corporation, St. Joseph, Mich., USA). For N_total, evolved gasses after incineration were reduced to N₂ and measured with a thermal-conductivity detector (LECO Corporation, St. Joseph, Mich., USA). P_total was analysed with Fleishmann acid (Houba et al., 1997). P_AL is used to assess the P supply capacity of grassland soils (Reijneveld et al., 2014) and was determined according to Egnér et al. (1960) (NEN 5793).

Soil physical parameters

Soil moisture was determined in April and October in a homogenized 0−10 cm soil sample after drying at 105°C for 24 hrs. Moisture content was expressed as percentage of fresh soil weight.

Penetration resistance was measured (April and October) with a penetrologger (Eijkelkamp, Giesbeek, the Netherlands; cone of 2.0 cm² penetration surface and 60° apex angle. Penetration resistance was expressed as an average of 7 penetrations per plot and per soil layer of 0−10, 10−20, and 20−30 cm.

Soil structure and rooting density were assessed in October in the 0−10 cm and 10−25 cm layers. The percentage of crumbs, sub-angular blocky elements and angular blocky elements was estimated by one experienced person as described by Peerlkamp (1959) and Shepherd (2000), Root density was estimated by scoring visible roots (score 1–10; 1 for no roots and 10 for above average).

Water infiltration rate was measured in October at three spots per experimental plot in 5 of the 6 blocks (35 plots). A PVC pipe (15 cm high, 15 cm diameter) was pushed into the soil to a depth of 10 cm. 500 ml water was poured into each pipe and the infiltration time was recorded. If the infiltration time exceeded 15 min, the remaining water volume was estimated to calculate the infiltration rate (mm min⁻¹).

Grass yield and botanical composition

Grass dry matter (DM) and N yield were determined during 2015 with a Haldrup plot harvester (J. Haldrup a/s, Løgstør, Denmark). The four harvest dates were May 15, June 29, August 19 and September 30. Fresh biomass, DM content (70°C for 24 hrs) and total N content (Kjeldahl) were determined for each harvest. Herbage DM yield (Mg DM ha⁻¹) and herbage N yield (kg N ha⁻¹) were calculated. Apparent N recovery (ANR; kg N.kg N⁻¹) was calculated as (N yield_(fertilized) – N yield_{(non-fertilized)})/(N fertilization rate) (Vellinga and André, 1999).

In June 2015, botanical composition was measured by visually estimating the relative soil cover of the sward and the proportion of each species therein (Sikkema, 1997).

Data files

Data_soil_grass.csv

Content:

Dataset with soil biological (earthworms, microbial PLFA), soil chemical, soil physical parameters, herbage dry matter and N yields, and botanical parameters.

Column names and units:

• plot: Experimental plot number (1-42)
• treatment: Treatment code (see text)
• block: Block number (1-6)
• EW_species_number: Earthworm - number of species
• EW_totalnumber: Earthworm - total number per m2
• EW_epigeic: Earthworm - number of epigeic adults and juveniles per m2
• EW_endogeic: Earthworm - number of endogeic adults and juveniles per m2
• EW_adults: Earthworm - number of adults per m2
• EW_juveniles: Earthworm - number of juveniles per m2
• EW_adult_epigeic: Earthworm - number of epigeic adults per m2
• EW_adult_endogeic: Earthworm - number of endogeic adults per m2
• EW_juven_epigeic: Earthworm - number of epigeic juveniles per m2
• EW_juven_endogeic: Earthworm - number of endogeic juveniles per m2
• EW_L_rubellus: Earthworm - number of L. rubellus adults and juveniles per m2
• EW_A_chlorotica: Earthworm - number of A. chlorotica adults and juveniles per m2
• EW_A_caliginosa: Earthworm - number of A. caliginosa adults and juveniles per m2
• EW_O_lacteum: Earthworm - number of O. lacteum adults and juveniles per m2
• EW_A_rosea: Earthworm - number of A. rosea adults and juveniles per m2
• EW_O_cyaenum: Earthworm - number of O. cyaneum adults and juveniles per m2
• EW_L_castaneus: Earthworm - number of L. castaneus adults and juveniles per m2
• EW_D_rubida: Earthworm - number of D. rubida adults and juveniles per m2
• EW_adult_L_rubellus: Earthworm - number of L. rubellus adults per m2
• EW_adult_A_chlorotica: Earthworm - number of A. chlorotica adults per m2
• EW_adult_A_caliginosa: Earthworm - number of A. caliginosa adults per m2
• EW_adult_O_lacteum: Earthworm - number of O. lacteum adults per m2
• EW_adult_A_rosea: Earthworm - number of A. rosea adults per m2
• EW_adult_O_cyaenum: Earthworm - number of O. cyaneum adults per m2
• EW_adult_L_castaneus: Earthworm - number of L. castaneus adults per m2
• EW_adult_D_rubida: Earthworm - number of D. rubida adults per m2
• EW_juven_L_rubellus: Earthworm - number of L. rubellus juveniles per m2
• EW_juven_A_chlorotica: Earthworm - number of A. chlorotica juveniles per m2
• EW_juven_A_caliginosa: Earthworm - number of A. caliginosa juveniles per m2
• EW_non_determined: Earthworm - number of non determined individuals per m2
• EW_total_biomass: Earthworm - total fresh biomass per m2
• Leatherjackets: number of leatherjackets per m2
• Wireworms: number of wireworms per m2
• TOTmicrPLFA: total microbial PLFA in nmol.g-1 dry soil
• bactPLFA: bacterial PLFA in nmol.g-1 dry soil
• saprofungPLFA: saprotrophic fungal PLFA in nmol.g-1 dry soil
• Fung_bactPLAF_ratio: ratio of fungal to bacterial PLFA
• GramPLUSplfa: gram positive PLFA in nmol.g-1 dry soil
• GramMINplfa: gram negative PLFA in nmol.g-1 dry soil
• ratioGram_PLUS_MIN: ratio of gram positive to gram negative PLFA
• AMFsporNLFA: AMF spores NLFA in nmol.g-1 dry soil
• ActinomPLFA: Actinomycetes PLFA in nmol.g-1 dry soil
• ShannonPLFA: PLFA shannon diversity index
• SOM: soil organic matter in g.100 g-1 dry soil
• Ctotal: total C in g.100 g-1 dry soil
• HWC: hot water extractable C in μg.100 g-1 dry soil
• Ntotal: total N in g.100 g-1 dry soil
• Ptotal: total P2O5 in mg.100 g-1 dry soil
• P_AL: total P-AL in mg.100 g-1 dry soil
• pH_KCl: pH-KCl
• CN_ratio: C:N ratio
• C_SOM: C:SOM ratio
• Soilmoisture_April: soil moisture content in April in g.100g-1 fresh soil
• Penetrationresistance_April_cm010: penetration resistance in April in 10-20 cm in Newton
• Penetrationresistance_April_cm1020: penetration resistance in April in 20-30 cm in Newton
• Penetrationresistance_April_cm2030: penetration resistance in April in 0-10 cm in Newton
• Soilmoisture_October: soil moisture content in October in g.100g-1 fresh soil
• Penetrationresistance_October_cm010: penetration resistance in October in 10-20 cm in Newton
• Penetrationresistance_October_cm1020: penetration resistance in October in 20-30 cm in Newton
• Penetrationresistance_October_cm2030: penetration resistance in October in 0-10 cm in Newton
• crumb_struct_cm010: percentage of crumb elements in 0-10 cm
• round_struct_cm011: percentage of sub-angular elements in 0-10 cm
• rootdensity_cm010: score (1-10) of root density in 0-10 cm
• crumb_struct_cm1025: percentage of crumb elements in 10-25 cm
• round_struct_cm1025: percentage of sub-angular elements in 10-25 cm
• sharp_struct_cm1025: percentage of angular elements in 10-25 cm
• rootdensity_cm1025: score (1-10) of root density in 10-25 cm
• water_infiltration: water infiltration rate in mm per minute
• DM_yield_year: total herbage dry matter yield in kg.ha-1 per year
• DM_yield_H1: herbage dry matter yield of harvest 1 in kg.ha-1
• DM_yield_H2: herbage dry matter yield of harvest 2 in kg.ha-1
• DM_yield_H3: herbage dry matter yield of harvest 3 in kg.ha-1
• DM_yield_H4: herbage dry matter yield of harvest 4 in kg.ha-1
• N_yield_year: total herbage N yield in kg.ha-1 per year
• N_yield_H1: herbage N yield of harvest 1 in kg.ha-1
• N_yield_H2: herbage N yield of harvest 2 in kg.ha-1
• N_yield_H3: herbage N yield of harvest 3 in kg.ha-1
• N_yield_H4: herbage N yield of harvest 4 in kg.ha-1
• DMperc_yield_year: herbage dry matter content (per year; weighed average over the 4 harvests) in g.100g-1 fresh weight
• DMperc_yield_H1: herbage dry matter content of harvest 1 in g.100g-1 fresh weight
• DMperc_yield_H2: herbage dry matter content of harvest 2 in g.100g-1 fresh weight
• DMperc_yield_H3: herbage dry matter content of harvest 3 in g.100g-1 fresh weight
• DMperc_yield_H4: herbage dry matter content of harvest 4 in g.100g-1 fresh weight
• Ncontent_yield_year: herbage N content (per year; weighed average over the 4 harvests) in g.kg-1 dry matter
• Ncontent_yield_H1: herbage N content of harvest 1 in g.kg-1 dry matter
• Ncontent_yield_H2: herbage N content of harvest 2 in g.kg-1 dry matter
• Ncontent_yield_H3: herbage N content of harvest 3 in g.kg-1 dry matter
• Ncontent_yield_H4: herbage N content of harvest 4 in g.kg-1 dry matter
• fresh_yield_H1: herbvage fresh yield of harvest 1 in Mg.ha-1
• ANR: apparent N recovery in kg N.kg N-1
• productive_grasses: cover percentage of L. perenne and P trivialis
• monocotyledons: cover percentage of monocotyledons
• dicotyledons: cover percentage of dicotyledons
• plant_species: number of plant species
• monocot_species: number of monocotyledon species
• dicot_species: number of dicotyledon species
• Lolium_perenne: plant cover %
• Poa_trivialis: plant cover %
• Phleum_pratense: plant cover %
• Elytrigia_repens: plant cover %
• Poa_annua: plant cover %
• Agrostis_stolonifera: plant cover %
• Holcus_lanatus: plant cover %
• Alopecurus_pratensis: plant cover %
• Alopecurus_geniculatus: plant cover %
• Trifolium_repens: plant cover %
• Taraxacum_officinale: plant cover %
• Ranunculus_arvensis: plant cover %
• Rumex_obtusifolius: plant cover %
• Rumex_crispus: plant cover %
• Ranunculus_acris: plant cover %
• Stellaria_media: plant cover %
• Cardamine_pratensis: plant cover %
• Bellis_perennis: plant cover %
• Rumex_acetosa: plant cover %
• Ranunculus_sceleratus: plant cover %
• Polygonum_aviculare: plant cover %
• Capsella_bursa-pastoris: plant cover %
• Glechoma_hederacea: plant cover %
• Geranium_molle: plant cover %

Fertilizer_input.csv

Content:

Application quantities of fertilizers and ash, organic matter, C and mineral inputs, and fertilizer C:N ratio. Total N input is the sum of mineral N (Nmin) and organic N (Norg). Average values per hectare and per year over the years 2013−2015.

Column names and units:

• Treatment: Treatment code (see text)
• Fertilizer_fresh: Applied fertilizer in Mg.ha^-1 per year (fresh weight)
• Fertilizer_DM: Applied fertilizer in Mg.ha^-1 per year (dry matter weight); for IF+SD this is the sum of 2.72 Mg sawdust + 0.45 Mg N fertilizer
• Ash: Mineral fraction in kg.ha^-1 per year
• OM: Organic matter in kg.ha^-1 per year
• C: Total C in kg.ha^-1 per year
• Nmin: Mineral N in kg.ha^-1 per year
• Norg: Organic N in kg.ha^-1 per year
• P2O5: kg.ha^-1 per year
• C_N_ratio: C:N ratio