Thaumarchaeal ammonia oxidation in an acidic forest peat soil is not influenced by ammonium amendment

Abstract

Both bacteria and thaumarchaea contribute to ammonia oxidation, the first step in nitrification. The abundance of putative ammonia oxidizers is estimated by quantification of the functional gene amoA, which encodes ammonia monooxygenase subunit A. In soil, thaumarchaeal amoA genes often outnumber the equivalent bacterial genes. Ecophysiological studies indicate that thaumarchaeal ammonia oxidizers may have a selective advantage at low ammonia concentrations, with potential adaptation to soils in which mineralization is the major source of ammonia. To test this hypothesis, thaumarchaeal and bacterial ammonia oxidizers were investigated during nitrification in microcosms containing an organic, acidic forest peat soil (pH 4.1) with a low ammonium concentration but high potential for ammonia release during mineralization. Net nitrification rates were high but were not influenced by addition of ammonium. Bacterial amoA genes could not be detected, presumably because of low abundance of bacterial ammonia oxidizers. Phylogenetic analysis of thaumarchaeal 16S rRNA gene sequences indicated that dominant populations belonged to group 1.1c, 1.3, and "deep peat" lineages, while known amo-containing lineages (groups 1.1a and 1.1b) comprised only a small proportion of the total community. Growth of thaumarchaeal ammonia oxidizers was indicated by increased abundance of amoA genes during nitrification but was unaffected by addition of ammonium. Similarly, denaturing gradient gel electrophoresis analysis of amoA gene transcripts demonstrated small temporal changes in thaumarchaeal ammonia oxidizer communities but no effect of ammonium amendment. Thaumarchaea therefore appeared to dominate ammonia oxidation in this soil and oxidized ammonia arising from mineralization of organic matter rather than added inorganic nitrogen.

FIG. 1.

The influence of acetylene and ammonium amendment on nitrification in peat soil microcosms. (a and b) Changes in ammonium (a) and nitrate (b) concentrations in soil microcosms incubated at 28°C for 30 days containing 0, 10, or 100 Pa acetylene headspace concentration. The y axes for these two graphs show the ammonium concentration (NH₄⁺-N g dry soil⁻¹) and nitrate concentration (NO₃⁻-N g dry soil⁻¹ day⁻¹), respectively. (c and d) Changes in ammonium (c) and nitrate (d) in soil microcosms amended with 0, 10, or 100 μg NH₄⁺-N g dry soil⁻¹ and incubated at 22°C for 20 days. The y axes for these two graphs show the ammonium concentration (NH₄⁺-N g dry soil⁻¹) and nitrate concentration (NO₃⁻-N g dry soil⁻¹ day⁻¹), respectively. For both experiments, triplicate microcosms were destructively sampled at each time point and mean values are plotted. Error bars represent standard errors but were usually smaller than the plotted symbols. Symbols overlap in panels b and c where values are close to zero.

FIG. 2.

DGGE analysis of archaeal amoA gene transcripts from soil incubated with 0, 10, or 100 μg NH₄⁺-N g dry soil⁻¹. (a) DGGE profiles of mRNA transcripts. Each lane represents a profile derived from an individual microcosm. The numbers of bands highlighted by arrows were derived using Phoretix 1-D (Phoretix International, Newcastle-Upon-Tyne, United Kingdom), with highlighted bands showing a decrease (band 4) or increase (bands 13, 14, and 19) in relative intensity during incubation. (b) PCA of the different archaeal amoA gene transcript communities based on the relative intensities of DGGE bands. Each symbol represents one community derived from an individual microcosm.

FIG. 3.

Archaeal amoA gene abundance (a) and transcript abundance (b) in soil microcosms amended with 0, 10, or 100 μg NH₄⁺-N g dry soil⁻¹ after incubation for 0 or 20 days. Values plotted are means and standard errors from triplicate microcosms. Values on the y axes are the gene or transcript abundance per g of dry soil.

FIG. 4.

Maximum likelihood phylogenetic analysis of thaumarchaeal 16S rRNA genes from three clone libraries (A, B, and C) from bog soil (a) and translated amoA gene sequences from soil microcosms (b) sampled after incubation for 0 and 20 days, with reference sequences of environmental clones and cultivated organisms described as follows: clone name (environmental source, accession number). For panel a, names of lineages are given according to the methods described by Prosser and Nicol (38), except for the deep peat (DP) group (40). The majority of sequences fell within three clades, highlighted by gray blocks (group 1.1c, deep peat, and group 1.3) and are presented fully expanded in Fig. S1 of the supplemental material. Multifurcation indicates where the relative branching order of major lineages could not be determined in the majority of bootstrap replicates. For panel b, 86 of 87 sequences formed one specific clade within the soil (assumed group 1.1b) clade. In both trees, the length of each branch represents the maximum branch length obtained. Bars, an estimated 0.1 change per nucleotide (a) or amino acid (b) position.