Effect of Lake Trophic Status and Rooted Macrophytes on Community Composition and Abundance of Ammonia-Oxidizing Prokaryotes in Freshwater Sediments

Communities of ammonia-oxidizing archaea (AOA) and bacteria (AOB) in freshwater sediments and those in association with the root system of the macrophyte species Littorella uniflora, Juncus bulbosus, and Myriophyllum alterniflorum were compared for seven oligotrophic to mesotrophic softwater lakes and acidic heathland pools. Archaeal and bacterial ammonia monooxygenase alpha-subunit (amoA) gene diversity increased from oligotrophic to mesotrophic sites; the number of detected operational taxonomic units was positively correlated to ammonia availability and pH and negatively correlated to sediment C/N ratios. AOA communities could be grouped according to lake trophic status and pH; plant species-specific communities were not detected, and no grouping was apparent for AOB communities. Relative abundance, determined by quantitative PCR targeting amoA, was always low for AOB (<0.05% of all prokaryotes) and slightly higher for AOA in unvegetated sediment and AOA in association with M. alterniflorum (0.01 to 2%), while AOA accounted for up to 5% in the rhizospheres of L. uniflora and J. bulbosus. These results indicate that (i) AOA are at least as numerous as AOB in freshwater sediments, (ii) aquatic macrophytes with substantial release of oxygen and organic carbon into their rhizospheres, like L. uniflora and J. bulbosus, increase AOA abundance; and (iii) AOA community composition is generally determined by lake trophy, not by plant species-specific interactions.Oxygen release from the roots of macrophyte species such as Littorella uniflora (L.) Asch. (shore weed), Lobelia dortmanna L. (water lobelia), and Glyceria maxima (Hartm.) Holmb. (reed sweet grass) stimulates nitrification and coupled nitrification-denitrification in the rhizosphere compared to that in unvegetated sediment (2, 36, 40). These interactions are of high ecological relevance especially in oligotrophic systems, since enhanced nitrogen loss due to rhizosphere-associated denitrification can retard natural eutrophication and succession of plant communities (1). While the microbial communities involved in coupled nitrification-denitrification have been well studied in rice paddy soils (7, 11), less information is available for natural freshwater sediments, especially those from oligotrophic lakes (2, 26).The first key step of coupled nitrification-denitrification, the oxidation of ammonia to nitrite, is catalyzed by two groups of prokaryotes—the ammonia-oxidizing bacteria (AOB) (24) and the only recently recognized ammonia-oxidizing archaea (AOA) (22). For both groups, the gene encoding the alpha-subunit of ammonia monooxygenase (amoA) has been widely used as a functional marker to analyze their community compositions (15, 25); recent studies demonstrated the ubiquity of AOA and their predominance over AOB in a broad range of environments (32, 38). AOA, but not AOB, were also strongly enriched in the rhizosphere of the freshwater macrophyte Littorella uniflora in a mesotrophic Danish lake, suggesting that AOA were primarily responsible for increased rates of nitrification in the rhizosphere of this plant species (19). Moreover, ammonia oxidizer communities differed between rhizosphere and unvegetated sediment, indicating a plant-specific effect on AOA and AOB community composition. The objectives of this study were therefore to test whether (i) AOA generally predominate over AOB in freshwater sediments and especially in macrophyte rhizospheres and (ii) macrophytes have species-specific effects on abundance and community composition of AOA and AOB in rhizosphere sediments and on root surfaces.To address these questions, two shallow heathland pools and five lakes in Denmark and Germany, ranging from low-pH and dystrophic sites to neutral-pH and oligotrophic and mesotrophic sites, were chosen, and three macrophyte species—Littorella uniflora, Juncus bulbosus L. (bulbous rush), and Myriophyllum alterniflorum DC. (alternate water milfoil)—were selected as model systems. These plant species differ in nitrogen nutrition, extent of radial oxygen loss, and lifestyle, presumably resulting in differential, plant species-specific effects on rhizosphere- and root-associated AOA and AOB communities. L. uniflora prefers nitrate as the nitrogen source, while J. bulbosus prefers ammonium (41, 45); oxygen release is high to moderate from the roots of L. uniflora and J. bulbosus (9, 12) but is minor from the roots of M. alterniflorum (M. Herrmann, P. Stief, and A. Schramm, unpublished results); L. uniflora and J. bulbosus remain photosynthetically active throughout the year, while only the below-ground parts of M. alterniflorum are retained during winter.Rhizosphere sediments and roots from each plant species were sampled from three different sites per species, and unvegetated sediment was obtained from all seven sites. The comparison of samples from these different sites and compartments (rhizosphere, root surface, unvegetated sediment) allowed an evaluation of the importance of plant species relative to that of environmental conditions related to lake trophic status and pH on ammonia oxidizer communities.