Abundance and diversity of ammonia-oxidizing archaea and bacteria in sediments of trophic end members of the Laurentian Great Lakes, Erie and Superior

Abstract

Ammonia oxidation is the first step of nitrification carried out by ammonia-oxidizing Archaea (AOA) and Bacteria (AOB). Lake Superior and Erie are part of the Great Lakes system differing in trophic status with Lake Superior being oligotrophic and Lake Erie meso- to eutrophic. Sediment samples were collected from both lakes and used to characterize abundance and diversity of AOA and AOB based on the ammonia monooxygenase (amoA) gene. Diversity was accessed by a pyro-sequencing approach and the obtained sequences were used to determine the phylogeny and alpha and beta diversity of the AOA and AOB populations. In Lake Erie copy numbers of bacterial amoA genes were in the same order of magnitude or even higher than the copy numbers of the archaeal amoA genes, while in Lake Superior up to 4 orders of magnitude more archaeal than bacterial amoA copies were detected. The AOB detected in the samples from Lake Erie belonged to AOB that are frequently detected in freshwater. Differences were detected between the phylogenetic affiliations of the AOA from the two lakes. Most sequences detected in Lake Erie clustered in the Nitrososphaera cluster (Thaumarchaeal soil group I.1b) where as most of the sequences in Lake Superior were found in the Nitrosopumilus cluster (Thaumarchaeal marine group I.1a) and the Nitrosotalea cluster. Pearson correlations and canonical correspondence analysis (CCA) showed that the differences in abundance and diversity of AOA are very likely related to the sampling location and thereby to the different trophic states of the lakes.

Figure 1. Sampling sites for Lake Superior (A) and Lake Erie (B) sediments.

Figure 2. Abundance (amoA gene copy number) of AOA and AOB in the sediment of Lake Erie and Superior (mean ± SD, n = 3; different letters above the columns indicate significant differences between samples determined by one-way ANOVA followed by Tukey test; p<0.05).

Figure 3. Alpha-diversity at 98% identity and singletons removed of the AOA and AOB amoA sequence libraries.

(mean ± SD, n = 100 rarefactions; different letters in figure A above the columns indicate significant differences between samples determined by one-way ANOVA followed by Tukey test; p<0.05; data for AOA and AOB were tested separately).

Figure 4. Relative frequency [%] of the AOA in the different phylogenetic groups.

Representative sequences were picked based on 98% identity, aligned in ARB to the AOA tree published by Pester et al. (2012) and assigned to different phylogenetic groups. Cultivated members of the AOA can be found in Nitrosopumilus cluster 1.1: Nitrosopumilus maritimus, Candidatus Nitrosoarchaeum limnia, Candidatus Nitrosoarchaeum korensis MY1 and Enrichment AOA-AC2; Nitrosopumilus cluster 5: Enrichments AOA-AC1, AOA-AC5 and AOA-DW. Most cultivated strains are integrated in Figure S3.

Figure 5. Relative frequency [%] of the AOB in the different phylogenetic groups.

Representative sequences were picked based on 98% identity, aligned in ARB and assigned to phylogenetic groups.

Figure 6. UPGMA clustering of Weighted Unifrac distances (A) and Jaccard abundances (B) of AOA in the sediment of Lake Erie and Superior at 98% similarity.

Numbers at the nodes represent statistical analysis of 100 rarefactions.

Figure 7. Canonical correspondence analysis (CCA) triplot (arrows: environmental variables; circles: samples; triangles: species) for quantitative data as presented in Fig. 3 of the AOA amoA sequence libraries in Lake Erie and Superior (Eigenvalues: Axis 1: 0.5865, Axis 2: 0.0421, Axis 3: 0.0073).

Explanatory value of the environmental factors was determined using forward selection: AOA_log explained 47%; ammonium concentration 18.3% and nitrate concentration 0.8% of the variation. (Abbreviations: Npum11: Nitrosopumilus subcluster 1.1; Npum51: Nitrosopumilus subcluster 5.1; Npum15: Nitrosopumilus subcluster 15; Nspae21: Nitrososphaera subcluster 2.1; Nspae32: Nitrososphaera subcluster 3.2; Nspae41: Nitrososphaera subcluster 4.1; Nspae51: Nitrososphaera subcluster 5.1; Nspae7: Nitrososphaera subcluster 7; Nspae81: Nitrososphaera subcluster 8.1; Nspae9: Nitrososphaera subcluster 9; Nspae11: Nitrososphaera subcluster 11; NspaeX: Nitrososphaera subcluster diverse; Ntalea: Nitrosotalea subcluster 2)