Nitrification inhibitor chlorate and nitrogen substrates differentially affect comammox Nitrospira in a grassland soil.

Introduction: Through the combined use of two nitrification inhibitors, Dicyandiamide (DCD) and chlorate with nitrogen amendment, this study aimed to investigate the contribution of comammox Nitrospira clade B, ammonia oxidizing bacteria (AOB) and archaea (AOA) to nitrification in a high fertility grassland soil, in a 90-day incubation study. Methods: The soil was treated with nitrogen (N) at three levels: 0 mg-N kg-1 soil, 50 mg-N kg-1 soil, and 700 mg-N kg-1 soil, with or without the two nitrification inhibitors. The abundance of comammox Nitrospira, AOA, AOB, and nitrite oxidising bacteria (NOB) was measured using qPCR. The comammox Nitrospira community structure was assessed using Illumina sequencing. Results and Discussion: The results showed that the application of chlorate inhibited the oxidation of both NH4+ and NO2- in all three nitrogen treatments. The application of chlorate significantly reduced the abundance of comammox Nitrospira amoA and nxrB genes across the 90-day experimental period. Chlorate also had a significant effect on the beta diversity (Bray-Curtis dissimilarity) of the comammox Nitrospira clade B community. Whilst AOB grew in response to the N substrate additions and were inhibited by both inhibitors, AOA showed litle or no response to either the N substrate or inhibitor treatments. In contrast, comammox Nitrospira clade B were inhibited by the high ammonium concentrations released from the urine substrates. These results demonstrate the differential and niche responses of the three ammonia oxidising communities to N substrate additions and nitrification inhibitor treatments. Further research is needed to investigate the specificity of the two inhibitors on the different ammonia oxidising communities.

Soil moisture is a primary driver of comammox Nitrospira abundance in New Zealand soils.

The objectives of this study were to investigate the abundance and community composition of comammox Nitrospira under: (i) pasture-based dairy farms from different regions, and (ii) different land uses from the same region and soil type. The results clearly showed that comammox Nitrospira were most abundant (3.0 × 106 copies) under the west coast dairy farm conditions, where they were also significantly more abundant than canonical ammonia oxidisers. This was also true in the Canterbury dairy farm. The six land uses investigated were pine monoculture, a long term no input ecological trial, sheep + beef and Dairy, both irrigated and non-irrigated. It was concluded that comammox Nitrospira was most abundant under the irrigated dairy farm (2.7 × 106 copies). Contrary to the current industry opinion, the relatively high abundance of comammox Nitrospira under fertile irrigated dairy land suggests that comammox Nitrospira found in terrestrial ecosystems may be copiotrophic. it was also determined that comammox Nitrospira was more abundant under irrigated land use than their non-irrigated counterparts, suggesting that soil moisture is a key environmental parameter influencing comammox abundance. Comammox abundance was also positively correlated with annual rainfall, further supporting this theory. Phylogenetic analysis of the comammox Nitrospira detected determined that 17 % of the comammox community belonged to a newly distinguished subclade, clade B.2. The remaining 83 % belonged to clade B.1. No sequences from clade A were found.

Comammox Nitrospira Clade B is the most abundant complete ammonia oxidizer in a dairy pasture soil and inhibited by dicyandiamide and high ammonium concentrations.

The recent discovery of comammox Nitrospira, a complete ammonia oxidizer, capable of completing the nitrification on their own has presented tremendous challenges to our understanding of the nitrification process. There are two divergent clades of comammox Nitrospira, Clade A and B. However, their population abundance, community structure and role in ammonia and nitrite oxidation are poorly understood. We conducted a 94-day microcosm study using a grazed dairy pasture soil amended with urea fertilizers, synthetic cow urine, and the nitrification inhibitor, dicyandiamide (DCD), to investigate the growth and community structure of comammox Nitrospira spp. We discovered that comammox Nitrospira Clade B was two orders of magnitude more abundant than Clade A in this fertile dairy pasture soil and the most abundant subcluster was a distinctive phylogenetic uncultured subcluster Clade B2. We found that comammox Nitrospira Clade B might not play a major role in nitrite oxidation compared to the role of canonical Nitrospira nitrite-oxidizers, however, comammox Nitrospira Clade B is active in nitrification and the growth of comammox Nitrospira Clade B was inhibited by a high ammonium concentration (700 kg synthetic urine-N ha-1) and the nitrification inhibitor DCD. We concluded that comammox Nitrospira Clade B: (1) was the most abundant comammox in the dairy pasture soil; (2) had a low tolerance to ammonium and can be inhibited by DCD; and (3) was not the dominant nitrite-oxidizer in the soil. This is the first study discovering a new subcluster of comammox Nitrospira Clade B2 from an agricultural soil.

The response of ammonia-oxidizing microorganisms to trace metals and urine in two grassland soils in New Zealand.

An incubation experiment was conducted to investigate the response of ammonia-oxidizing bacteria (AOB), ammonia-oxidizing archaea (AOA), and the nitrification rate to the contamination of Cu, Zn, and Cd in two New Zealand grassland soils. The soils spiked with different concentrations of Cu (20 and 50 mg kg-1), Zn (20 and 50 mg kg-1), and Cd (2 and 10 mg kg-1) were incubated for 14 days and then treated with 500 mg kg-1 urine-N before continuing incubation for a total of 115 days. Soils were sampled at intervals throughout the incubation. The nitrification rate in soils at each sampling period was determined, and the abundance of AOB and AOA was measured by real-time quantification polymerase chain reaction (qPCR) assay of the amoA gene copy numbers. The results revealed that moderate trace metal stress did not significantly affect the abundance of AOB and AOA in the two soils, probably due to the high organic matter content of the soils which would have reduced the toxic effect of the metals. Nitrification rates were much greater and the observable nitrification period was much shorter in the dairy farm (DF) soil, in which the AOB and AOA abundances were greater than those of the mixed cropping farm (MF) soil. AOB were shown to grow under high nitrogen conditions, whereas AOA were shown to grow under low N environments, with different metal concentrations. Therefore, nitrogen status rather than metal applications was the main determining factor for AOB and AOA growth in the two soils studied.