Diffusion Chamber for Nitrogen-15 Determination of Coupled Nitrification-Denitrification around Soil-Manure Interfaces

Denitrifying activity in soil is often associated with anoxic microsites rich in organic material, but previously described procedures for determining denitrification rates are not satisfactory when high rates of both nitrification and denitrification are associated with such hot spots. Coupled nitrification-denitrification associated with hot spots of cattle manure was analyzed by use of a recently developed isotope pairing method in combination with a newly designed diffusion chamber. The diffusion chamber contained three separate phases: an anaerobic manure layer on top, an aerobic soil layer below that placed on top of a highly permeable aluminum oxide filter, and below the filter sterile water containing ¹⁵NO⁻₃. The soil could be kept at a water potential below field capacity by suction of a hanging water column connected to the water phase. The isotopically labeled NO⁻₃ entered the soil through the filter and diffused through the soil to the manure where it was denitrified. While diffusing through the soil, ¹⁵NO⁻₃ was mixed with ¹⁴NO⁻₃ formed by nitrification. The isotopic composition of the N₂ formed by denitrification of the mixed NO⁻₃ could then be used to determine the rate of denitrification based on ¹⁵NO⁻₃ from the water as well as the rate of coupled nitrification-denitrification within the soil. The rate of coupled nitrification-denitrification increased exponentially during the first days after application of manure, indicating exponential growth of the nitrifiers with a doubling time of 1.7 d. The rate reached a maximum of 116 nmol N₂ cm⁻² h⁻¹ after 10 d and subsequently decreased to 20 nmol N₂ cm⁻² h⁻¹ after 3 wk. After 3 wk, about 40% of the NH⁺₄ initially present had been removed through coupled nitrification-denitrification. Supported by the Center for Microbial Ecology under the Danish Biotechnology Program.