Project description:The abundance of bacterial (AOB) and archaeal (AOA) ammonia oxidisers, assessed using quantitative PCR measurements of their respective a-subunit of the ammonia monooxygenase (amoA) genes, and ammonia oxidation rates were measured in four contrasting coastal sediments in the Western English Channel. Sediment was sampled bimonthly from July 2008 to May 2011, and measurements of ammonia oxidiser abundance and activity compared to a range of environmental variables including salinity, temperature, water column nutrients and sediment carbon and nitrogen content. Despite a higher abundance of AOA amoA genes within all sediments, and at all time-points, rates of ammonia oxidation correlated with AOB and not AOA amoA gene abundance. Other than ammonia oxidation rate, sediment particle size was the only variable that correlated with the spatial and temporal patterns of AOB amoA gene abundance, implying a preference of the AOB for larger sediment particles. This is possibly due to deeper oxygen penetration into the sandier sediments, increasing the area available for ammonia oxidation to occur, higher concentrations of inhibitory sulphide with pore waters of muddier sediments or a combination of both oxygen and sulphide concentrations. Similar to many other temporal studies of nitrification within estuarine and coastal sediments, decreases in AOB amoA gene abundance were evident during summer and autumn, with maximum abundance and ammonia oxidation rates occurring in winter and early spring. The lack of correlation between AOA amoA gene abundance and ammonium oxidation rate suggests an alternative role for amoA-carrying AOA within these sediments.
Project description:The abundance of bacterial (AOB) and archaeal (AOA) ammonia oxidisers, assessed using quantitative PCR measurements of their respective a-subunit of the ammonia monooxygenase (amoA) genes, and ammonia oxidation rates were measured in four contrasting coastal sediments in the Western English Channel. Sediment was sampled bimonthly from July 2008 to May 2011, and measurements of ammonia oxidiser abundance and activity compared to a range of environmental variables including salinity, temperature, water column nutrients and sediment carbon and nitrogen content. Despite a higher abundance of AOA amoA genes within all sediments, and at all time-points, rates of ammonia oxidation correlated with AOB and not AOA amoA gene abundance. Other than ammonia oxidation rate, sediment particle size was the only variable that correlated with the spatial and temporal patterns of AOB amoA gene abundance, implying a preference of the AOB for larger sediment particles. This is possibly due to deeper oxygen penetration into the sandier sediments, increasing the area available for ammonia oxidation to occur, higher concentrations of inhibitory sulphide with pore waters of muddier sediments or a combination of both oxygen and sulphide concentrations. Similar to many other temporal studies of nitrification within estuarine and coastal sediments, decreases in AOB amoA gene abundance were evident during summer and autumn, with maximum abundance and ammonia oxidation rates occurring in winter and early spring. The lack of correlation between AOA amoA gene abundance and ammonium oxidation rate suggests an alternative role for amoAÂ-carrying AOA within these sediments. Two color array (Cy3 and Cy5): the universal standard 20-mer oligo is printed to the slide with a 70-mer oligo (an archetype). Environmental DNA sequences (fluoresced with Cy3) within 15% of the 70-mer conjugated to a 20-mer oligo (fluoresced with Cy5) complementary to the universal standard will bind to the oligo probes on the array. Signal is the ratio of Cy3 to Cy5. Three replicate probes were printed for each archetype. Two replicate arrays were run on duplicate targets.
Project description:The ecophysiology of complete ammonia oxidizing Nitrospira (CMX) and their widespread occurrence in groundwater suggests that CMX bacteria have a competitive advantage over ammonia-oxidizing bacteria (AOB) and archaea (AOA) in these environments. However, the relevance of their activity from the ecosystem-level process perspective has remained unclear. We investigated oligotrophic carbonate rock aquifers as a model system to assess the contribution of CMX, AOA and AOB to nitrification and to identify the environmental drivers of their niche differentiation at different levels of ammonium and oxygen. CMX accounted for up to 95% of the ammonia oxidizer communities. Nitrification rates were positively correlated to CMX clade A-associated phylotypes and AOB affiliated with Nitrosomonas ureae. Surprisingly, short-term incubations amended with the nitrification inhibitors allylthiourea and chlorate suggested that AOB contributed more than 90% to overall ammonia oxidation, while metaproteomics analysis confirmed an active role of CMX in both ammonia and nitrite oxidation. Ecophysiological niche differentiation of CMX clades A and B, AOA and AOB was linked to their requirements for ammonium, oxygen tolerance, and metabolic versatility. Our results demonstrate that despite numerical predominance of CMX, the first step of nitrification in oligotrophic groundwater is primarily governed by AOB. Higher growth yields at lower NH4+ turnover rates and energy derived from nitrite oxidation most likely enable CMX to maintain consistently high populations. Activity measurements combined with differential inhibition allowed a refined understanding of ammonia oxidizer coexistence, competition and cooperation beyond the insights from molecular data alone.
Project description:Salinity strongly influences the physiology and distribution of nitrifying microorganisms, yet the effects of low salinity on these key players in nitrogen cycling remain understudied. This study investigates the impact of hypoosmolarity on different groups of ammonia oxidizers in soil and lake environments, as well as in pure culture isolates. In soil microcosms amended with ammonium, at low salinity levels (~120 µS/cm), which are comparable to values commonly found in pristine terrestrial and aquatic environments, the abundance of ammonia-oxidizing bacteria (AOB), dominated by Nitrosomonas oligotropha, significantly increased. In contrast, the growth of ammonia-oxidizing archaea (AOA), dominated by “Ca. Nitrosotenuis” of the Nitrosopumilaceae family, was stimulated by high salinity (~760 µS/cm). In ammonium-fed lake microcosms, the abundance of AOB, dominated by N. oligotropha, significantly increased under both low (~170 µS/cm) and high salinity (~850 µS/cm) conditions. In the presence of allylthiourea, a bacterial nitrification inhibitor, AOA were found to be sensitive to low salinity in both soil and lake microcosms. Consistently, pure culture studies revealed marked growth inhibition of AOA, especially of members of the Nitrosopumilaceae, under hypoosmolarity, unlike AOB and complete ammonia oxidizers (comammox) strains. Comparative genomic analyses with AOB and comammox, along with transcriptomic studies, suggested that the sensitivity of AOA to hypoosmolarity stress is attributed to a lack of sophisticated osmoregulatory transport systems and their S-layer cell wall structure. Overall, this study highlights the importance of hypoosmolarity as a key factor shaping the ecological niches and distribution of ammonia oxidizers as well as nitrification activities in terrestrial and aquatic environments increasingly affected in their salinities by intensified water cycles due to climate change.
Project description:Contaminated aquifer (Dusseldorf-Flinger, Germany) templates extracted from 5 sediment depths ranging between 6.4 and 8.4 m below ground and over 3 years of sampling were amplified for amplicon pyrosequencing using the primers Ba27f (5’-aga gtt tga tcm tgg ctc ag-3’) and Ba519r (5’- tat tac cgc ggc kgc tg-3’), extended as amplicon fusion primers with respective primer A or B adapters, key sequence and multiplex identifiers (MID) as recommended by 454/Roche. Amplicons were purified and pooled as specified by the manufacturer. Emulsion PCR (emPCR), purification of DNA-enriched beads and sequencing run were performed following protocols and using a 2nd generation pyrosequencer (454 GS FLX Titanium, Roche) as recommended by the developer. Quality filtering of the pyrosequencing reads was performed using the automatic amplicon pipeline of the GS Run Processor (Roche), with a slight modification concerning the valley filter (vfScanAllFlows false instead of TiOnly) to extract the sequences. Demultiplexed raw reads were furhter trimmed for quality and lenght (>250 bp).