Nitrification and Denitrification Enzyme Activity: a successful tool in Arctic and Alpine soil ecology

• Nitrogen is consideration to be a limiting factor for plants and microbes in arctic and alpine ecosystems and the rates of mineralization, nitrification, and denitrifi¬cation are known to be generally low. However, Climate Change is expected to alter the nitrogen availability and dynamics and, as a consequence, affect plant community composition and production. The general consensus today is that increased temperature will lead to greater microbial activity and more plant-available nitrogen. Nevertheless, nitrification and denitrification are restricted by a number of environmental factors such as low tem¬perature and low pH. The C/N ratio and the water content of the soils also play an important role in determining the rates of nitrification and denitrification. Since 2002 microbial studies has been undertaken at Latnjajure, and comprise several microbial techniques, e.g. Nitrification Enzyme Activity (NEA), Denitrification Enzyme Activity (DEA), Phospholipid fatty acid analysis (PFLA), and Temperature Gradient Gel Electrophoresis (TGGE). These studies focuses on the interaction between plants and microbes along natural environmental gradients, both within plant communities and within the landscape, but also entails the OTCs used in the ITEX studies at Latnjajaure. Here we present the techniques NEA and DEA and give some brief results from how these have been successfully applied at Latnjajaure. In ecosystems with low nitrification activity, small amounts of NO3-/NO2- will be formed and it is thus difficult to measure low fluxes. However, NO3-/NO2- can be converted to N2O and then analysed by gas chromatography, whereby the detection limit is increased at least 1000 times compared to the spectroscopical technique. These techniques are referred to Nitrification (NEA) and Denitrification Enzyme Activity (DEA) and give a potential measure on the nitrification and denitrification rates, which goes back to the actual populations of nitrifiers and denitrifiers in the soil. For instance, NEA has been proved to better correlate with extractable NH4+ concentration than net nitrification does, and still after twelve weeks show a strong correlation with the initial extractable NH4+ concentration. Therefore, these variables, in particular NEA, have the advantage of being a much more stable variable than, for instance, extractable N concentrations and net nitrification, and NEA and DEA are therefore suitable when working in fringe environments with restricted logistics like the Latnjajaure catchment. NEA is measured using a two-step incubation technique; first by incubate the soil with a nutrient solution for 24 hours in darkness, at room temperature on a rotary shaker. Sub-samples are then withdrawn after a specified time schedule. The second step allows NO3- to be reduced to N2O by adding a modified denitrifying bacterium, Pseudomonas chlororaphis ATCC 43928, together with a carbon source. This strain of bacteria lacks the enzyme to reduce N2O to N2. The samples are then again incubated in darkness, at room temperature for 24 hours, and analysed by gas chromatography. This method was first used by Lensi et al. (1985, 1986), to study nitrification potentials in forest soils. Furthermore, the method has been developed for soils with low pH and small amounts of NO3- and the analysis makes the quantification without interference of organic matter, which makes it suitable for arctic and alpine ecosystems. To analyse DEA an anaerobic incubation technique is used, based on acetylene inhibition of the N2O-reductase resulting in N2O as the only end product. The soil sample is evacuated and flushed with N2. Thereafter acetylene is inserted to a final acetylene concentration of 10%, and the samples are shaken continuously and gas samples are withdrawn after a specified time schedule, which is then analysed by gas chromatography. This provides an estimate of the maximum concentration of functional denitrifying enzymes in the soil. Denitrifiers, in contrast to nitrifiers, are heterotrophs and can switch from using NO3- as an alternative electron acceptor to O2 under aerobic conditions. This makes other factors in the soil important determinants of DEA, e.g. availability of oxygen and C. Hence, the presence of denitrifiers is rarely a limitation for denitrification and they usually make up a reasonably large fraction of the soil bacteria. At Latnjajaure NEA shows a larger differentiation across plant communities than DEA. However, the spatial variability in the landscape, at the meso-scale, was in the same range in both variables and increased with altitude from 1000 to 1365 m a.s.l, particularly in heath plant communities. This result suggests that the decrease in mean annual temperature with altitude (0.6ºC with every one hundred meters) did not reduce nitrification and denitrification rates, as one might have expected. None of the other variable studied could explain the altitudinal increase in all cases, and the factors controlling the nitrification and denitrification rates seem to vary with the vegetation type. Furthermore, neither NEA nor DEA exhibited any changes between the ambient and warmed plots in the warming experiments. However, the warming experiment in the dry heath exhibited a change in root morphology via increased specific root length (SRL; m gDM-1) and specific root area (SRA; m2 kgDM -1). As both heterotrophic microbes and plants out-compete nitrifiers for NH4+, a change in root morphology, as seen in the warming experiment, may also explain the increased activity of nitrifying and denitrifying microbes with altitude.

Ämnesord

NATURVETENSKAP  -- Biologi -- Ekologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Ecology (hsv//eng)

Nyckelord

Arctic

Climate Change

Denitrification

Fine roots

ITEX

Nitrification

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