Potential Anammox Rates Research Articles

Anaerobic ammonium oxidation and its contribution to nitrogen removal in China's coastal wetlands.

Over the past several decades, human activities have caused substantial enrichment of reactive nitrogen in China’s coastal wetlands. Although anaerobic ammonium oxidation (anammox), the process of oxidizing ammonium into dinitrogen gas through the reduction of nitrite, is identified as an important process for removing reactive nitrogen, little is known about the dynamics of anammox and its contribution to nitrogen removal in nitrogen-enriched environments. Here, we examine potential rates of anammox and associate them with bacterial diversity and abundance across the coastal wetlands of China using molecular and isotope tracing techniques. High anammox bacterial diversity was detected in China’s coastal wetlands and included Candidatus Scalindua, Kuenenia, Brocadia, and Jettenia. Potential anammox rates were more closely associated with the abundance of anammox bacteria than to their diversity. Among all measured environmental variables, temperature was a key environmental factor, causing a latitudinal distribution of the anammox bacterial community composition, biodiversity and activity along the coastal wetlands of China. Based on nitrogen isotope tracing experiments, anammox was estimated to account for approximately 3.8–10.7% of the total reactive nitrogen removal in the study area. Combined with denitrification, anammox can remove 20.7% of the total external terrigenous inorganic nitrogen annually transported into China’s coastal wetland ecosystems.

Nitrogen loss through anaerobic ammonium oxidation in agricultural drainage ditches

Agricultural drainage ditches are a key conduit of nitrogen from agricultural fields to surface water. Denitrification has been proven to be an important N removal pathway in agricultural drainage ditches; however, it is currently unknown about the role of anaerobic ammonium oxidation (anammox). Here, we investigated the N loss through anammox in six different agricultural drainage ditches, located in Nanjing City, Jiangsu Province (China). Stable isotope-labeling experiments confirmed the anammox activity in every examined ditch, and the potential anammox rates ranged from 4.4 to 23.8 nmol N2 g−1 sediment day−1. The anammox process contributed 2.1–18.8 % to sediment dinitrogen gas production, while the remainder would be due to denitrification. It is estimated that a total loss of 6.1–32.9 g N m-2 per year could be attributed to the anammox process in the examined ditches. Illumina-based 16S rRNA gene sequencing confirmed the presence of diverse anammox bacteria in the examined sediments, including Candidatus Brocadia, Candidatus Kuenenia, and Candidatus Anammoxoglobus. Quantitative PCR further confirmed the presence of anammox bacteria, varying between 1.9 × 104 and 1.2 × 105 copies g−1 sediment. The results of our study showed the evidence of anammox process in agricultural drainage ditches and demonstrated that the contribution of anammox to N removal from agricultural drainage ditches may not be negligible.

Evidence for anaerobic ammonium oxidation process in freshwater sediments of aquaculture ponds

The anaerobic ammonium oxidation (anammox) process, which can simultaneously remove ammonium and nitrite, both toxic to aquatic animals, can be very important to the aquaculture industry. Here, the presence and activity of anammox bacteria in the sediments of four different freshwater aquaculture ponds were investigated by using Illumina-based 16S rRNA gene sequencing, quantitative PCR assays and (15)N stable isotope measurements. Different genera of anammox bacteria were detected in the examined pond sediments, including Candidatus Brocadia, Candidatus Kuenenia and Candidatus Anammoxoglobus, with Candidatus Brocadia being the dominant anammox genus. Quantitative PCR of hydrazine synthase genes showed that the abundance of anammox bacteria ranged from 5.6 × 10(4) to 2.1 × 10(5) copies g(-1) sediment in the examined ponds. The potential anammox rates ranged between 3.7 and 19.4 nmol N2 g(-1) sediment day(-1), and the potential denitrification rates varied from 107.1 to 300.3 nmol N2 g(-1) sediment day(-1). The anammox process contributed 1.2-15.3% to sediment dinitrogen gas production, while the remainder would be due to denitrification. It is estimated that a total loss of 2.1-10.9 g N m(-2) per year could be attributed to the anammox process in the examined ponds, suggesting that this process could contribute to nitrogen removal in freshwater aquaculture ponds.

Molecular and geochemical constraints on anaerobic ammonium oxidation (anammox) in a riparian zone of the Seine Estuary (France)

To expand the limited knowledge about the ecological significance of anaerobic ammonium oxidation (anammox) in continental aquatic and terrestrial ecosystems, we studied community structure, abundance, and activity of anammox bacteria in soils and sediments in the wetland of Trou Deshayes, a riparian zone in the Seine Estuary, France. Combining (i) molecular analyses of the genes coding for anammox bacterial 16S rRNA and the enzyme hydrazine oxidoreductase (hzo), (ii) quantification of unique anammox bacterial membrane lipids (i.e. ladderanes), and, (iii) 15N-isotope label incubation experiments with intertidal sediments and irregularly flooded soils nearby, we demonstrated that anammox bacteria were ubiquitous in the studied wetland ecosystem. In both soils and sediments, detected anammox bacteria were related to Candidatus ‘Brocadia’. 16S rRNA genes were generally lower in the more oxygenated soils, but on the same order of magnitude (107–108 copies g−1 d.w.) as found for other river estuaries, riparian zones and agricultural soils. While the C20-ladderane fatty acid with five cyclobutane moieties (C20-[5]-FA) was found in both sediments and soils, other ladderane species were detected only in the wetland sediments. The observed differential ladderane distribution suggests intra-genus differences in the community composition of anammox bacteria between the sediments and the floodplain soils. While the abundance of anammox bacteria was significantly lower in the soils versus the sediments, the potential anammox rates were similar (≤15 and ≤22 nmol N2 d−1 g−1 w.w. sediment and soil, respectively), suggesting lower cell-specific anammox rates in the sediments. The observed potential rates of anammox were rather low, leaving canonical denitrification as the main fixed N removal pathway in this riparian zone. The relative contribution of anammox to the total N2 production (between 3 and 8 %) was similar at all sites, highlighting the dependence of the anammox process on nitrite supply from denitrification across environmental boundaries. Due to this coupling, the dependence of organotrophic denitrification on the quality and stoichiometry of OM also seems to affect the anammox bacterial community. Our results suggest that N removal and mitigation of N supply from agriculture in wetlands by anammox is limited, and much less important than denitrification.

Evidence for the cooccurrence of nitrite-dependent anaerobic ammonium and methane oxidation processes in a flooded paddy field.

Anaerobic ammonium oxidation (anammox) and nitrite-dependent anaerobic methane oxidation (n-damo) are two of the most recent discoveries in the microbial nitrogen cycle. In the present study, we provide direct evidence for the cooccurrence of the anammox and n-damo processes in a flooded paddy field in southeastern China. Stable isotope experiments showed that the potential anammox rates ranged from 5.6 to 22.7 nmol N2 g(-1) (dry weight) day(-1) and the potential n-damo rates varied from 0.2 to 2.1 nmol CO2 g(-1) (dry weight) day(-1) in different layers of soil cores. Quantitative PCR showed that the abundance of anammox bacteria ranged from 1.0 × 10(5) to 2.0 × 10(6) copies g(-1) (dry weight) in different layers of soil cores and the abundance of n-damo bacteria varied from 3.8 × 10(5) to 6.1 × 10(6) copies g(-1) (dry weight). Phylogenetic analyses of the recovered 16S rRNA gene sequences showed that anammox bacteria affiliated with "Candidatus Brocadia" and "Candidatus Kuenenia" and n-damo bacteria related to "Candidatus Methylomirabilis oxyfera" were present in the soil cores. It is estimated that a total loss of 50.7 g N m(-2) per year could be linked to the anammox process, which is at intermediate levels for the nitrogen flux ranges of aerobic ammonium oxidation and denitrification reported in wetland soils. In addition, it is estimated that a total of 0.14 g CH4 m(-2) per year could be oxidized via the n-damo process, while this rate is at the lower end of the aerobic methane oxidation rates reported in wetland soils.

Occurrence and activity of anammox bacteria in surface sediments of the southern North Sea

We investigated the occurrence and activity of anaerobic ammonia oxidation (anammox) bacteria in sandy and muddy sand sediments of the southern North Sea. The presence of anammox bacteria was established through the detection of specific phosphocholine-monoether ladderane lipids, 16S rRNA gene, and hydrazine synthase (hzsA) genes. Anammox activity was measured in intact sediment cores (in situ rate) and in sediment slurries (potential rate) as the rate of N2 evolution from (15) N-labeled substrates and compared to the transcriptional activity of genes of anammox bacteria. The contribution of anammox to N2 production ranged between 0% and 29%. The potential rate of anammox agreed well with the abundance of anammox bacteria 16S rRNA and hzsA gene copies and the transcriptional activity of the anammox bacteria 16S rRNA gene. We found a higher abundance and activity of anammox bacteria in sediments with higher organic carbon content and also higher activity in summer than in winter. The abundance of anammox bacteria and their potential anammox rates were similar to those reported for other marine coastal sediments, suggesting that potentially they are important contributors to the nitrogen cycle in sandy sediments of shallow continental shelf areas.

Environmental controls of anammox and denitrification in southern New England estuarine and shelf sediments

We measured denitrification and anammox potential rates in homogenized sediments over two annual cycles from two estuarine sites (Providence River estuary and Narragansett Bay) and two continental shelf sites (Block Island and Rhode Island sounds). Denitrification varied both spatially and seasonally, but anammox only varied spatially. Denitrification ranged from 2.5 to 112 nmol N h−1 mL−1 sediment, and anammox from 0 to 8.7 nmol N h−1 mL−1 sediment, with the contribution of anammox to N2 fluxes (ra) ranging from undetectable to 4% in estuarine sites and 8–42% on the shelf. Oxygen microprofiles were analyzed in intact cores and concentrations of nitrate, nitrite, and ammonium were measured in the sediment slice corresponding to the depth layer in which potential rates were measured. Denitrification rates correlated positively with diffusive O2 consumption and temperature and potential anammox rates correlated positively with pore‐water concentrations. Anammox was positively correlated with denitrification in shelf sediments where anammox was regularly detected, but not in estuarine sediments. ra was negatively correlated with diffusive O2 consumption and positively correlated with pore‐water nitrate. Both organic matter and nitrate were important factors regulating the contribution of anammox to N2 production. As a whole, our results demonstrate that incorporating processes that control these two factors into models of N cycling, rather than focusing on organic matter availability alone, may improve predictions of the relative importance of anammox to N2 production in marine sediments.

Impacts of freshwater flushing on anammox community structure and activities in the New River Estuary, USA

Anaerobic ammonium oxidation (anammox) and denitrification are 2 microbial nitro- gen removal processes that may play an important role in controlling the intensity and duration of estuarine and coastal eutrophication. Sediment communities in the New River Estuary, North Car- olina were investigated to determine the dynamics of anammox activity and community structure in conjunction with environmental conditions. 15 N tracer incubation experiments with sediment slurries were used to measure anammox and denitrification rates and estimate anammox contri- bution to total N2 production. Molecular analyses targeting the hydrazine oxidoreductase (hzo) gene were conducted to examine the structure of anammox communities and quantify the abun- dance of anammox bacteria in sediments. Potential anammox rates ranged from 0.02 to 1.4 nmol N2 g �1 h �1 , with the highest potential activities observed during winter and spring when the estu- ary received large doses of nitrogen from the watershed. Anammox contributed up to 14.1% of total N2 production in upstream estuarine sediments and abundance of anammox communities ranged from 1.55 × 10 2 to 2.59 × 10 5 hzo gene copies g �1 sediment. Both activities and abundance of anammox communities were correlated with percent sediment organics (%organics) and the porewater concentrations of hydrogen sulfide. Based on hzo sequence analysis, anammox bacte- ria related to 'Candidatus Jettenia spp.' were widespread in estuarine sediments, which may be attributed to freshwater flushing and associated changes in environmental parameters as well as the geomorphology of the estuary. This is the first study to describe a dominance of 'Candidatus Jettenia spp.' in relation to %organics and hydrogen sulfide in an estuarine ecosystem driven by meteorological forcing.

Temperature response of denitrification and anammox reveals the adaptation of microbial communities to in situ temperatures in permeable marine sediments that span 50° in latitude

Abstract. Despite decades of research on the physiology and biochemistry of nitrate/nitrite-respiring microorganisms, little is known regarding their metabolic response to temperature, especially under in situ conditions. The temperature regulation of microbial communities that mediate anammox and denitrification was investigated in near shore permeable sediments at polar, temperate, and subtropical sites with annual mean temperatures ranging from −5 to 23 °C. Total N2 production rates were determined using the isotope pairing technique in intact core incubations under diffusive and simulated advection conditions and ranged from 2 to 359 μmol N m−2 d−1. For the majority of sites studied, N2 removal was 2–7 times more rapid under simulated advective flow conditions. Anammox comprised 6–14% of total N2 production at temperate and polar sites and was not detected at the subtropical site. Potential rates of denitrification and anammox were determined in anaerobic slurries in a temperature gradient block incubator across a temperature range of −1 °C to 42 °C. The highest optimum temperature (Topt) for denitrification was 36 °C and was observed in subtropical sediments, while the lowest Topt of 21 °C was observed at the polar site. Seasonal variation in the Topt was observed at the temperate site with values of 26 and 34 °C in winter and summer, respectively. The Topt values for anammox were 9 and 26 °C at the polar and temperate sites, respectively. The results demonstrate adaptation of denitrifying communities to in situ temperatures in permeable marine sediments across a wide range of temperatures, whereas marine anammox bacteria may be predominately psychrophilic to psychrotolerant. The adaptation of microbial communities to in situ temperatures suggests that the relationship between temperature and rates of N removal is highly dependent on community structure.

Seasonal variation and controlling factors of anaerobic ammonium oxidation in freshwater river sediments in the Taihu Lake region of China

Anaerobic ammonium oxidation (anammox) has been recently recognized as an important pathway for the removal of fixed nitrogen (N) from aquatic systems. However, the functions of anammox in freshwater river systems remain uncertain. In this study, we evaluated the occurrence of anammox activity in two rivers in the Taihu Lake region in China during a seasonal survey. Homogenized sediments were incubated with 15N-labeled NO3- and NH4+ amendments to determine the potential importance of the anammox process relative to canonical denitrification. Production of 29N2 and 30N2 in slurries was determined using membrane inlet mass spectrometry. Potential anammox rates in the two river sediments ranged from 0.11±0.07 to 6.79±1.28μmolNm−2h−1 and the remove of N by anammox accounted for 0.8±0.00% to 10.7±0.03% of total N2 production. Potential anammox rates varied spatially and temporally in the two rivers, with the highest and lowest mean anammox rates appearing during summer and early autumn and during winter, respectively. The variation of the percentage of anammox to total N2 production displayed the same trend with potential anammox rates. Water temperature and NO3- content in sediments were the main factors affecting anammox activity. Anammox bacteria were detected in sediment samples using barcode pyrosequencing. The 16S rRNA anammox gene sequences in the river sediments were affiliated with Candidatus Kuenenia, Candidatus Jettenia, and Candidatus Scalindua, among which C. Kuenenia dominated the anammox bacterial communities. Our results confirmed the presence of anammox bacteria but their role is relatively small in removing fixed N from freshwater river systems.

Anammox bacterial populations in deep marine hypersaline gradient systems

To extend the knowledge of anaerobic ammonium oxidation (anammox) habitats, bacterial communities were examined in two hypersaline sulphidic basins in Eastern Mediterranean Sea. The 2m thick seawater-brine haloclines of the deep anoxic hypersaline basins Bannock and L'Atalante were sampled in intervals of 10cm with increasing salinity. (15)N isotope pairing incubation experiments showed the production of (29)N2 and (30)N2 gases in the chemoclines, ranging from 6.0 to 9.2% salinity of the L'Atalante basin. Potential anammox rates ranged from 2.52 to 49.65nmolN2 L(-1) day(-1) while denitrification was a major N2 production pathway, accounting for more than 85.5% of total N2 production. Anammox-related 16S rRNA genes were detected along the L'Atalante and Bannock haloclines up to 24% salinity, and the amplification of the hydrazine synthase genes (hzsA) further confirmed the presence of anammox bacteria in Bannock. Fluorescence in situ hybridisation and sequence analysis of 16S rRNA genes identified representatives of the marine anammox genus 'Candidatus Scalindua' and putatively new operational taxonomic units closely affiliated to sequences retrieved in marine environments that have documented anammox activity. 'Scalindua brodae' like sequences constituted up to 84.4% of the sequences retrieved from Bannock. The anammox community in L'Atalante was different than in Bannock and was stratified according to salinity increase. This study putatively extends anammox bacterial habitats to extremely saline sulphidic ecosystems.

The Contribution of Anammox and Denitrification to Sediment N2 Production in a Surface Flow Constructed Wetland

This study used anaerobic slurry assays and intact core incubations to quantify potential rates of anammox (anaerobic ammonia oxidation) in sediments along the flow path of a surface flow constructed wetland receiving secondary treated sewage effluent. Anammox occurred at two of the four sites assayed with a maximum rate of 199.4 +/- 18.7 micromol N x m(-2) x hr(-1) (24% of total N2 production) at the discharge end of the wetland. Denitrification was the major producer of N2, with a maximum rate of 965.3 +/- 122.8 micromol N x m(-2) x hr(-1) at site 2. Oxygen was probably the key regulator of anammox activity within the studied CW. In addition to anammox, we found evidence that nitrifier-denitrification was potentially responsible for the production of N2O. Total production of N2O was 15.1% of the total gaseous N produced. Limitations to the methodology for quantifying anammox in CW's are outlined. This study demonstrated that denitrification is not the only pathway for gaseous production in constructed wetlands and that wetlands may be significant sources of greenhouse gases such as N2O.

Correlation between Anammox Activity and Microscale Distribution of Nitrite in a Subtropical Mangrove Sediment

The distribution of anaerobic ammonium oxidation (anammox) in nature has been addressed by only a few environmental studies, and our understanding of how anammox bacteria compete for substrates in natural environments is therefore limited. In this study, we measure the potential anammox rates in sediment from four locations in a subtropical tidal river system. Porewater profiles of NO(x)(-) (NO2- plus NO3-) and NO2- were measured with microscale biosensors, and the availability of NO2- was compared with the potential for anammox activity. The potential rate of anammox increased with increasing distance from the mouth of the river and correlated strongly with the production of nitrite in the sediment and with the average concentration or total pool of nitrite in the suboxic sediment layer. Nitrite accumulated both from nitrification and from NO(x)(-) reduction, though NO(x)(-) reduction was shown to have the greatest impact on the availability of nitrite in the suboxic sediment layer. This finding suggests that denitrification, though using NO2- as a substrate, also provides a substrate for the anammox process, which has been suggested in previous studies where microscale NO2- profiles were not measured.