Anammox Bacterial Abundance Research Articles

Anaerobic ammonium oxidation (anammox) bacterial diversity, abundance, and activity in marsh sediments of the Yangtze Estuary

AbstractAnaerobic ammonium oxidation (anammox) as an important process of nitrogen cycle has been studied in estuarine environments. However, knowledge about the dynamics of anammox bacteria and their interactions with associated activity remains scarce in these environments. Here we report the anammox bacterial diversity, abundance, and activity in the Yangtze Estuary, using molecular and isotope‐tracing techniques. The phylogenetic analysis of 16S rRNA indicated that high anammox bacterial diversity occurred in this estuary, including Scalindua, Brocadia, Kuenenia, and two novel clusters. The patterns of community composition and diversity of anammox bacteria differed across the estuary. Salinity was a key environmental factor defining the geographical distribution and diversity of the anammox bacterial community at the estuarine ecosystem. Temperature and organic carbon also had significant influences on anammox bacterial biodiversity. The abundance of anammox bacteria ranged from 2.63 × 106 and 1.56 × 107 gene copies g−1, and its spatiotemporal variations were related significantly to salinity, temperature, and nitrite content. The anammox activity was related to temperature, nitrite, and anammox bacterial abundance, with values of 0.94–6.61 nmol N g−1 h−1. The tight link between the anammox and denitrification processes implied that denitrifying bacteria may be a primary source of nitrite for the anammox bacteria in the estuarine marshes. On the basis of the 15N tracing experiments, the anammox process was estimated to contribute 6.6%–12.9% to the total nitrogen loss whereas the remainder was attributed to denitrification.

Broad Distribution of Diverse Anaerobic Ammonium-Oxidizing Bacteria in Chinese Agricultural Soils

Anaerobic ammonium-oxidizing (anammox) bacteria have been detected in many marine and freshwater ecosystems. However, little is known about the distribution, diversity, and abundance of anammox bacteria in terrestrial ecosystems. In this study, anammox bacteria were found to be present in various agricultural soils collected from 32 different locations in China. Phylogenetic analysis of the 16S rRNA genes showed "Candidatus Brocadia," "Candidatus Kuenenia," "Candidatus Anammoxoglobus," and "Candidatus Jettenia" in the collected soils, with "Candidatus Brocadia" being the dominant genus. Quantitative PCR showed that the abundance of anammox bacteria ranged from 6.38 × 10(4) ± 0.42 × 10(4) to 3.69 × 10(6) ± 0.25 × 10(6) copies per gram of dry weight. Different levels of diversity, composition, and abundance of the anammox bacterial communities were observed, and redundancy analysis indicated that the soil organic content and the distribution of anammox communities were correlated in the soils examined. Furthermore, Pearson correlation analysis showed that the diversity of the anammox bacteria was positively correlated with the soil ammonium content and the organic content, while the anammox bacterial abundance was positively correlated with the soil ammonium content. These results demonstrate the broad distribution of diverse anammox bacteria and its correlation with the soil environmental conditions within an extensive range of Chinese agricultural soils.

Anaerobic ammonium oxidation (anammox) bacteria and sulfide‐dependent denitrifiers coexist in the water column of a meromictic south‐alpine lake

In addition to organotrophic denitrification, alternative pathways, such as anaerobic ammonium oxidation (anammox) or sulfide‐dependent denitrification may be important modes for the removal of fixed nitrogen (N) from lakes. We used Lake Lugano as a model system with which to assess possible controls on the concurrence of multiple suboxic N2 production pathways in a lacustrine water column. In the northern basin of Lake Lugano, concentration gradients of dissolved inorganic N toward the hypolimnetic redox transition zone (RTZ) indicate ammonium oxidation and nitrate reduction occurring in close vicinity. Ammonium reaches undetectable levels 15 m below the depth of oxygen disappearance, indicating its anaerobic consumption. The presence of anammox bacteria was confirmed by 16S ribosomal ribonucleic acid gene sequencing. Quantitative polymerase chain reaction revealed a maximum in anammox bacterial abundance at the same water depth where ammonium becomes exhausted. 15N‐labeling experiments indicate that anammox activity within the Lake Lugano RTZ can contribute up to ∼ 30% of total N2 production. Incubation experiments with various potential electron donors—glucose, acetate, Mn(II), Fe(II), and H2S—revealed that N2 formation was sulfide‐dependent and that organotrophic denitrification is only of minor importance for the elimination of fixed N from the Lake Lugano north basin. Maximum potential rates of anammox and chemolithotrophic denitrification were comparatively low but consistent with nutrient fluxes calculated from concentration gradients. This study provides evidence for the coexistence of anammox bacteria and sulfide‐dependent denitrifiers in the stratified water column of a lacustrine environment.

Co-Occurring Anammox, Denitrification, and Codenitrification in Agricultural Soils

Anammox and denitrification mediated by bacteria are known to be the major microbial processes converting fixed N to N(2) gas in various ecosystems. Codenitrification and denitrification by fungi are additional pathways producing N(2) in soils. However, fungal codenitrification and denitrification have not been well investigated in agricultural soils. To evaluate bacterial and fungal processes contributing to N(2) production, molecular and (15)N isotope analyses were conducted with soil samples collected at six different agricultural fields in the United States. Denitrifying and anammox bacterial abundances were measured based on quantitative PCR (qPCR) of nitrous oxide reductase (nosZ) and hydrazine oxidase (hzo) genes, respectively, while the internal transcribed spacer (ITS) of Fusarium oxysporum was quantified to estimate the abundance of codenitrifying and denitrifying fungi. (15)N tracer incubation experiments with (15)NO(3)(-) or (15)NH(4)(+) addition were conducted to measure the N(2) production rates from anammox, denitrification, and codenitrification. Soil incubation experiments with antibiotic treatments were also used to differentiate between fungal and bacterial N(2) production rates in soil samples. Denitrifying bacteria were found to be the most abundant, followed by F. oxysporum based on the qPCR assays. The potential denitrification rates by bacteria and fungi ranged from 4.118 to 42.121 nmol N(2)-N g(-1) day(-1), while the combined potential rates of anammox and codenitrification ranged from 2.796 to 147.711 nmol N(2)-N g(-1) day(-1). Soil incubation experiments with antibiotics indicated that fungal codenitrification was the primary process contributing to N(2) production in the North Carolina soil. This study clearly demonstrates the importance of fungal processes in the agricultural N cycle.

Co-occurrence and distribution of nitrite-dependent anaerobic ammonium and methane-oxidizing bacteria in a paddy soil

The anaerobic ammonium-oxidizing (anammox) and nitrite-dependent anaerobic methane-oxidizing (n-damo) bacteria in a paddy soil core (0-100 cm) were investigated with newly designed primers targeting the hydrazine synthase β-subunit (hzsB) of anammox bacteria and the recently published primers targeting the pmoA and 16S rRNA genes of n-damo bacteria. The hzsB gene was identified as a proper biomarker to explore the anammox bacterial biodiversity and abundance in soil. The anammox bacteria were present throughout the soil core with the highest abundance of 2.7 × 10(6) hzsB copies g(-1) dry soil at 40-50 cm and were not detectable below 70 cm. Sequences related to at least three species of known anammox bacteria, 'Brocadia anammoxidans', 'Brocadia fulgida', and 'Jettenia asiatica' were detected. By combining the analysis of pmoA and 16S rRNA genes, the n-damo bacteria were observed to be present in 30-70 cm with abundance from 6.5 × 10(3) (60-70 cm) to 7.5 × 10(4) (30-40 cm) copies g(-1) dry soil. The pmoA sequences retrieved from different depths closely related to each other and formed a unique clade. Our results showed that anammox and n-damo bacteria co-occurred in the paddy soil. Both of them were abundant in deep layers (30-60 cm) and the community structures changed along depths in the soil core.

Anammox Bacterial Abundance, Activity, and Contribution in Riparian Sediments of the Pearl River Estuary

The hypothesis of an anammox hotspot in river riparian zones was put forward based on our investigation on freshwater ecotones for over 25 years and previous anammox research. Here we used a complementary array of methods including isotope-pairing technique, quantitative PCR assays, and 16S rRNA and hydrazine synthase gene (hzsB) clone libraries to document the spatiotemporal evidence for a high abundance zone of anammox bacteria in river riparian sediment with observed abundance of 1.3-12 × 10(6) (summer) and 1.4-20 × 10(8) (winter) hydrazine synthase gene copies g(-1), which is the highest abundance in natural environments recorded so far. Meanwhile high anammox bacterial biodiversity were detected with 'Brocadia' and 'Kuenenia' dominating. However, the high anammox bacterial abundances were not related with high activities and contributions for nitrogen gas generation. The anammox activities ranged from 0.07 to 0.15 nmol N cm(-3) h(-1) (summer) to 1.0-2.6 nmol N cm(-3) h(-1) (winter) with high temporal heterogeneity. The retrieval of archaeal and bacterial amoA sequences indicated that nitrifying microbes might be the major source of nitrite for anammox bacteria in winter, while in summer the anaerobic nitrate reduction is more likely the main source. On the basis of (15)N tracing technology, it was estimated that a total loss of 0.67-9.62 g N m(-2) yr(-1) is linked to anammox in the riparian zone while denitrification contributed 96.2-170.3 g N m(-2) yr(-1) in Pearl River riparian sediments.

Distribution and diversity of anaerobic ammonium‐oxidizing bacteria in the sediments of the Qiantang River

Anaerobic ammonium oxidation (anammox) is an important process in the marine nitrogen cycle. However, little is known about the distribution, diversity and abundance of anammox bacteria in inland river ecosystems. Here, we found the presence of diverse anammox bacteria in a freshwater river - the Qiantang River, Zhejiang Province (China). The phylogenetic analysis of 16S rRNA genes showed that Brocadia genus, Kuenenia genus, Scalindua genus and three new anammox bacterial clusters could be detected together in Qiantang River sediments, suggesting a higher anammox bacterial diversity in the Qiantang River ecosystem than in open ocean environments where only Scalindua genus was detected. Brocadia and Kuenenia appeared to be the most common anammox bacterial genera in the Qiantang River. Redundancy analysis showed that the sediment organic carbon (OrgC) content had significant influence on the distribution of anammox bacteria in Qiantang River sediments. Pearson correlation analyses showed that OrgC content significantly influenced the anammox bacterial diversity. The results of real-time quantitative PCR showed spatial variations of anammox bacterial abundances which were highly correlated with the sediment total inorganic nitrogen content. These results demonstrated the distribution of diverse anammox bacteria and the influences of environmental factors on anammox bacterial communities in Qiantang River sediments.

Microbial community structure and productivity in the oxygen minimum zone of the eastern tropical North Pacific

Microbial abundances and activities (heterotrophic and chemoautotrophic) were profiled in the fall of 2007 along a transect of four stations in the upwelling region between the Tehuantepec Bowl, off the coast of southern Mexico, and the Costa Rica Dome. Heterotrophic potential varied with depth exhibiting relatively low maximum values of 4–8nM leucine d−1, but varied among stations by a factor of 2. Generally, heterotrophic potential varied inversely with nitrate implying NO3− draw down by nitrate-reducing chemoorganotrophs. Dark dissolved inorganic carbon assimilation (≈chemoautotrophy) was also highly variable with depth and reached a maximum value of 6.4μMC d−1 at one station, but only attained maxima of 0.02 and 0.2μMCd−1 at other stations. Elevated values of chemoautotrophy corresponded with enrichments in nitrite concentrations suggesting nitrification as well as with depletions in nitrite and ammonium concentrations suggesting anammox processes. Additions of 30μM NH4+ at selected depths stimulated chemoautotrophic activity up to 33-fold. Fluorescence in situ hybridization (FISH) analyses revealed elevated abundances of Crenarchaeota and β-proteobacteria (up to 1.0×107 and 10.0×107cellsL−1, respectively) associated with NH4+ minima and NO3- maxima. Nitrifying β-proteobacteria were most abundant in the upper oxycline (1.1–1.8×107cellsL−1) at three stations and exhibited a secondary peak of 0.3×107cellsL−1 in the lower oxycline at one station. Anammox bacterial (Planctomycetes) abundances were as high as 2.1×106cellsL−1, but accounted for less than 1% of total DAPI counts. The concentration of diploptene (a hopanoid biomarker found in bacteria) varied from 2 to 15pgL−1 in the upper oxycline and from 51 to 160pgL−1 in the lower oxycline. Delta 13C values of particulate organic carbon (POC) varied from −30 to −21‰ in the water column (avg. −25‰). Distributions of isotopically light POC coincided with enrichments in diploptene, Planctomycetes cells, nitrite, and chemoautotrophy as well as with nitrate and ammonium depletions, suggesting that anammox and nitrification are important for secondary biological production in this system.

Anammox Bacterial Abundance, Biodiversity and Activity in a Constructed Wetland

An integrated approach to document high anammox activity and biodiversity in a constructed wetland (CW) was performed and showed that substantial anammox activity could mitigate undesirable N(2)O emission. The enhanced anammox bacterial abundance, biodiversity and activity were achieved by supplementing activated sludge to the CW. Up to 3.38 × 10(7) gene copies g(-1) dry soil of anammox bacteria were enriched in the CW. The activity measured by isotope pairing technique increased from 1.6 nmol N g(-1) sludge h(-1) in the original activated sludge to 18 nmol N g(-1) soil h(-1) in the CW, with the specific cellular activity increased from 5.1 to 12.8 fmol cell(-1) d(-1). Up to 33% of produced N(2) could be attributed to anammox process in the CW, with the remainder being due to denitrification. Phylogenetic analysis of anammox bacterial 16S rRNA genes indicated a shift of community from single Candidatus "Brocadia fulgida" in sludge to multiple "Jettenia", "Brocadia", and "Anammoxoglobus" species in the CW. With static chambers and control experiments, the CW with supplemented sludge had a 30% reduced N(2)O emission flux compared with the tests without adding biomass during an 8 month testing period.

Water column anammox and denitrification in a temperate permanently stratified lake (Lake Rassnitzer, Germany)

We studied microbial N 2 production via anammox and denitrification in the anoxic water column of a restored mining pit lake in Germany over an annual cycle. We obtained high-resolution hydrochemical profiles using a continuous pumping sampler. Lake Rassnitzer is permanently stratified at ca. 29 m depth, entraining anoxic water below a saline density gradient. Mixed-layer nitrate concentrations averaged ca. 200 μmol L −1, but decreased to zero in the anoxic bottom waters. In contrast, ammonium was <5 μmol L −1 in the mixed layer but increased in the anoxic waters to ca. 600 μmol L −1 near the sediments. In January and October, 15N tracer measurements detected anammox activity (maximum 504 nmol N 2 L −1 d −1 in 15NH 4 +-amended incubations), but no denitrification. In contrast, in May, N 2 production was dominated by denitrification (maximum 74 nmol N 2 L −1 d −1). Anammox activity in May was significantly lower than in October, as characterized by anammox rates (maximum 6 vs. 16 nmol N 2 L −1 d −1 in incubations with 15NO 3 −), as well as relative and absolute anammox bacterial cell abundances (0.56% vs. 0.98% of all bacteria, and 2.7×10 4 vs. 5.2×10 4 anammox cells mL −1, respectively) (quantified by catalyzed reporter deposition-fluorescence in situ hybridization (CARD-FISH) with anammox bacteria-specific probes). Anammox bacterial diversity was investigated with anammox bacteria-specific 16S rRNA gene clone libraries. The majority of anammox bacterial sequences were related to the widespread Candidatus Scalindua sorokinii/brodae cluster. However, we also found sequences related to Candidatus S. wagneri and Candidatus Brocadia fulgida, which suggests a high anammox bacterial diversity in this lake comparable with estuarine sediments.

Biogeographical distribution of diverse anaerobic ammonium oxidizing (anammox) bacteria in Cape Fear River Estuary

Anaerobic ammonium oxidation (anammox) specific PCR method was developed to examine diversity and distribution of anammox bacteria in sediments collected from three different sites at Cape Fear River Estuary, North Carolina, where environmental parameters vary greatly over the year. Abundance and activities of anammox bacteria in these sediments were measured using the quantitative PCR (Q-PCR) method and (15)N isotope tracer incubations. Different anammox bacterial communities composed with Brocadia, Kuenenia, Jettenia or Scalindua were found among sites along the estuarine gradient. Seasonal variations of anammox community structures were observed along the estuary based on terminal restriction fragment length polymorphism (T-RFLP) analysis of 16S rRNA genes. Correlation analysis suggested that salinity variation influenced the diversity and distribution of different anammox bacteria in the estuary. Q-PCR assays of anammox bacteria showed temporal and spatial variations of their abundances, which were highly correlated to salinity variation. (15)N isotope tracer incubations measured different anammox rates and its per cent contribution to total N(2) production among sites. The highest anammox rate was found at the site where Scalindua organisms dominated with the highest anammox bacterial abundance. Thus, we demonstrated a biogeographical distribution of diverse anammox bacteria influenced by salinity, and provide evidence to link anammox abundance and activities in estuarine sediments.