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Nitrite affinities of Nitrospira (black), Nitrospina (orange), Nitrotoga (blue), Nitrobacter (red), Nitrococcus (dark green), and Nitrolancea (light green). Nitrospira lineages are indicated as I, II, IVa or unknown above the species name (modified after Nowka et al. 2015). Marine strains from this study are indicated with an asterisk (*). Filled symbols: pure cultures; open symbols: enrichment cultures. Further half-saturation constant (K m ) values are from references (a) Nowka et al. (2015), (b) Schramm et al. (1999), (c) Maixner et al. (2006), (d) Blackburne et al. (2007), (e) Vadivelu et al. (2006), (f) Both et al. (1992), (g) Laanbroek et al. (1994), and (h) Sorokin et al. (2012)
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Nitrification, the step-wise oxidation of ammonium to nitrite and nitrate, is important in the marine environment because it produces nitrate, the most abundant marine dissolved inorganic nitrogen (DIN) component and N-source for phytoplankton and microbes. This study focused on the second step of nitrification, which is carried out by a distinct g...
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In this study, ammonia (NH3) vapor was added to atmospheric pressure argon plasma jet generated by sinusoidal power supply (with an applied voltage of 4 kV and frequency of 35 kHz) and it was analyzed by electrical, optical, and chemical probe measurements. A total gas flow rate of 1200 sccm under 2% flow rate of either water or ammonia solution at...
Citations
... NOB is improving using metagenomic and other meta-omics methodologies, updated molecular techniques, and rectification cultivation methods, which offer a new approach to investigate NOB due to its importance in the nitrogen cycle (Albertsen et al., 2013;Shah, 2020Shah, , 2021. The majority of Nitrobacter species were extracted at a concentration of 29 mM nitrite, while marine strains needed a 1 mM nitrate (Table 1) (Jacob et al., 2017). However, it was later shown that different soil NOB populations have different nitrite preferences (Wang et al., 2015). ...
Nitrite-oxidizing bacteria (NOB) are primarily organo and/or chemoautotrophic Gram-negative bacteria that play a critical role in aerobic nitrification and gain energy by converting nitrite into nitrate. NOB is found in natural ecosystems and also in man-made ecosystems. Studies show that NOB lives in extreme environmental conditions such as concrete and natural stones, acidic soil, alkaline soil, and hot water springs. Some species can also survive in a long period of starvation and in dry conditions. They also need a specific type of enriched media composition and laboratory conditions for their growth. Chemotaxonomy helps to characterize NOB in mixed and pure cultures based on biochemical composition. In this chapter, different methods and techniques for the isolation and characterization of NOB from different environmental conditions will be discussed.KeywordsNitrificationNitrite-oxidizing bacteriaHeterotrophicNitriteIsolation
... Biological N transformations give rise to isotopic fractionation, leading to measurable shifts in the isotopic ratios of substrate and product pools (Kendall, 1998;Mariotti et al., 1982;Sebilo et al., 2006). These isotopic shifts can be compared with fractionation factors determined from pureculture studies (e.g., Buchwald et al., 2012;Casciotti et al., 2003;Granger et al., 2004;Jacob et al., 2018). For example, the 15 N fractionation factor (or isotope effect, 15 ε) for ammonia oxidation in bacterial and archaeal pure cultures ranges from −13 ‰ to −41 ‰ (Casciotti et al., 2003;Mariotti et al., 1981;Santoro and Casciotti, 2011), whereas nitrite oxidation has a fractionation factor from +10 ‰ to +12 ‰, reflecting a unique inverse isotope effect (Casciotti, 2009;Jacob et al., 2018). ...
... These isotopic shifts can be compared with fractionation factors determined from pureculture studies (e.g., Buchwald et al., 2012;Casciotti et al., 2003;Granger et al., 2004;Jacob et al., 2018). For example, the 15 N fractionation factor (or isotope effect, 15 ε) for ammonia oxidation in bacterial and archaeal pure cultures ranges from −13 ‰ to −41 ‰ (Casciotti et al., 2003;Mariotti et al., 1981;Santoro and Casciotti, 2011), whereas nitrite oxidation has a fractionation factor from +10 ‰ to +12 ‰, reflecting a unique inverse isotope effect (Casciotti, 2009;Jacob et al., 2018). The fractionation factor imparted during remineralization/ammonification of organic nitrogen (into ammonia) is generally small, with estimates ranging from ∼ 0 ‰ (Brandes and Devol, 2002;Kendall, 1998) to −2.5 ‰ (Möbius, 2013). ...
Estuaries and rivers are important biogeochemical reactors that act to modify the loads and composition of nutrients in the coastal zone. In a case study during July 2013, we sampled an 80 km transect along the Elbe Estuary under low-oxygen conditions. To better elucidate specific mechanisms of estuarine nitrogen processing, we tracked the evolution of the stable isotopic composition of nitrate, nitrite, particulate matter, and ammonium through the water column. This approach allowed assessment of the in situ isotope effects of ammonium and nitrite oxidation and of remineralization at the reach scale. The isotope effects of nitrite oxidation and ammonium oxidation were consistent with pure-culture assessments. We found that the nitrogen budget of the Elbe Estuary is governed by settling, resuspension, and reminer-alization of particulate matter, and we further used our stable isotope data to evaluate sources and sinks of nitrogen in the Elbe Estuary via an isotope mass-balance approach. We find that the reactivity of particulate matter, through its reminer-alization in the estuary, is the main control on the isotope dynamics of inorganic nitrogen species. Moreover, while underscoring this role of particulate matter delivery and reactivity , the isotope mass balance also indicated additional sinks of reactive nitrogen, such as possible denitrification of water column nitrate in the intensively dredged and deep Hamburg Harbor region.
... This scarcity of measurements is unsurprising given that in situ NO − 2 oxidation kinetics studies are generally limited; indeed, to our knowledge, there are only two studies from the coastal ocean (Olson, 1981a;Zhang et al., 2020) and two from the eastern tropical North Pacific oxygen deficient zone (ETNP ODZ; with these experiments conducted across a range of ambient oxygen concentrations; Sun et al., 2017Sun et al., , 2021. By contrast, there exist numerous estimates of NO − 2 oxidation kinetic parameters determined using cultured marine NOB (e.g., Jacob et al., 2017;Kits et al., 2017;Nowka et al., 2015;Sorokin et al., 2012;Zhang et al., 2020). In general, culture experiments suggest far higher kinetic constants compared to the limited in situ observations from the ocean, particularly for K m (i.e., culture-based K m estimates of 9-544 µM; Blackburne et al., 2007;Nowka et al., 2015;Ushiki et al., 2017). ...
Across the Southern Ocean in winter, nitrification is the dominant
mixed-layer nitrogen cycle process, with some of the nitrate produced
therefrom persisting to fuel productivity during the subsequent growing
season. Because this nitrate constitutes a regenerated rather than a new
nutrient source to phytoplankton, it will not support the net removal of
atmospheric CO2. To better understand the controls on Southern Ocean
nitrification, we conducted nitrite oxidation kinetics experiments in
surface waters across the western Indian sector in winter. While all
experiments (seven in total) yielded a Michaelis–Menten relationship with
substrate concentration, the nitrite oxidation rates only increased
substantially once the nitrite concentration exceeded 115±2.3 to
245±18 nM, suggesting that nitrite-oxidizing bacteria (NOB) require a
minimum (i.e., “threshold”) nitrite concentration to produce nitrate. The
half-saturation constant for nitrite oxidation ranged from 134±8 to
403±24 nM, indicating a relatively high affinity of Southern Ocean
NOB for nitrite, in contrast to results from culture experiments. Despite
the high affinity of NOB for nitrite, its concentration rarely declines
below 150 nM in the Southern Ocean's mixed layer, regardless of season. In
the upper mixed layer, we measured ammonium oxidation rates that were two-
to seven-fold higher than the coincident rates of nitrite oxidation,
indicating that nitrite oxidation is the rate-limiting step for
nitrification in the winter Southern Ocean. The decoupling of ammonium and
nitrite oxidation, combined with a possible nitrite concentration threshold
for NOB, may explain the non-zero nitrite that persists throughout the
Southern Ocean's mixed layer year-round. Additionally, nitrite oxidation may
be limited by dissolved iron, the availability of which is low across the
upper Southern Ocean. Our findings have implications for understanding the
controls on nitrification and ammonium and nitrite distributions, both in
the Southern Ocean and elsewhere.
... Further downstream, ammonium and nitrite concentrations increased, so we assume that remineralization no longer determines the overall isotope effect of nitrification. Instead, there was a combined influence of ammonium oxidation with an isotope effect 15 ε = −14 ‰ to −41 ‰ (Mariotti et al., 1981;Casciotti et al., 2003;Santoro and Casciotti, 2011) and nitrite oxidation with 15 ε = +9 ‰ to +20 ‰ Buchwald and Casciotti, 2010;Jacob et al., 2017). As we measured elevated ammonium and nitrite concentrations, both processes influenced the fractionation caused by nitrification. ...
Estuaries are nutrient filters and change riverine nutrient loads before
they reach coastal oceans. Their morphology have been extensively changed by
anthropogenic activities like draining, deepening and dredging to meet
economic and social demand, causing significant regime changes like tidal
amplifications and in some cases to hyper-turbid conditions. Furthermore,
increased nutrient loads, especially nitrogen, mainly by agriculture cause
coastal eutrophication. Estuaries can either act as a sink or as a source of
nitrate, depending on environmental and geomorphological conditions. These
factors vary along an estuary, and change nitrogen turnover in the system.
Here, we investigate the factors controlling nitrogen turnover in the
hyper-turbid Ems estuary (Northern Germany), which has been strongly
impacted by human activities. During two research cruises in August 2014 and
June 2020, we measured water column properties, dissolved inorganic
nitrogen, dual stable isotopes of nitrate and dissolved nitrous oxide
concentration along the estuary. We found that three distinct biogeochemical
zones exist along the estuary. A strong fractionation (∼26 ‰) of nitrate stable isotopes points towards nitrate
removal via water column denitrification in the hyper-turbid tidal river,
driven by anoxic conditions in deeper water layers. In the middle reaches of
the estuary nitrification gains importance, turning this section into a net
nitrate source. The outer reaches are dominated by mixing, with nitrate
uptake in 2020.
We find that the overarching control on biogeochemical nitrogen cycling,
zonation and nitrous oxide production in the Ems estuary is exerted by
suspended particulate matter concentrations and the linked oxygen deficits.
... A commamox, for example, Nitrospira inopinata, has a low ammonia half-saturation constant (K m , NH 3 ) of 0.063 μM and therefore is highly abundant in biofilm with a low range of ammonia concentrations (Kits et al., 2017). At low nitrite concentrations, nitrite oxidation is driven by Nitrospira, while Nitrobacter perform nitrite oxidation in environments high in nitrite (Jacob et al., 2017;Ushiki et al., 2017). ...
... For NOMs, Nitrospira as the NOMs with high affinity to nitrite (K m = 6-54 μM-NO 2 − , Jacob et al., 2017;Ushiki et al., 2017) dominated in A8-S2 and A40-S2, where low nitrite concentrations of 0.02 ± 0.04 mgN/L and 0.04 ± 0.10-0.53 ± 0.54 mgN/L, respectively, were observed, (Fig. 4a and b). ...
Bioaugmentation of nitrifying cultures can accelerate nitrification during startup and transition periods of recirculating aquaculture systems (RASs) operations. To formulate nitrifying cultures for RASs, impacts of ammonia and salinity (NaCl) on culturing nitrifying microorganisms were comprehensively investigated by including currently known groups of nitrifying microorganisms (ammonia oxidizing bacteria (AOB), ammonia-oxidizing archaea (AOA), comammox, Nitrospira, and Nitrobacter). By varying ammonia loading rate (ALRs of 1.6, 8, 20, 40, 60 and 150 mgN/L/d) of continuous-flow bioreactors fed with inorganic medium experimented for culture preparation, cultures containing distinct patterns of nitrifying communities were produced. Operating the reactors at the ALRs of ≤40 mgN/L/d, resulting in the effluent total ammonia nitrogen (TAN) and nitrite concentrations of ≤2.64 and ≤0.53 mgN/L, respectively, delivered the consortia consisting of a broad spectrum of substrate affinity nitrifying microorganisms. At the lower ranges of these ALRs (≤8 mgN/L/d), the most desirable consortia comprising comparable numbers of AOB, AOA, and comammox could be produced (the effluent TAN concentrations of ≤0.20 mgN/L), which would be resilient for applying in various RAS types. Enriching the cultures at the ALRs of ≥60 mgN/L/d allowed only the nitrifying microorganisms with low substrate affinity to dominate, incongruent with the consortia found in actual RASs. AOB were adaptable at all salinity studied (2, 15, and 30 g/L), while AOA and comammox were sensitive to elevated salinity (15 and 30 g/L, respectively). The ammonia removal rate of a culture prepared at 2 g/L salinity decreased largely when applied at 15 and 30 g/L. In contrast, those prepared at 15 and 30 g/L were more robust to different salinity. Separately preparing the cultures at different salinity for uses in freshwater and brackish-marine RASs is recommended. The findings of this work enhance our understanding on how to formulate nitrifying culture augmentation for used in different RAS types.
... The inverse isotope effect of NXR most likely originates at the enzyme level, where larger force constants in the transition state explain the inverse kinetic isotope effect when stretching vibrational contributions dominate the kinetic isotope effect (13). Follow-up studies demonstrated inverse isotope effects of the NOB Nitrococcus and Nitrobacter to be similar and around 20.5%, whereas the inverse isotope effects of the NOB Nitrospira and Nitrospina were less pronounced and close to 9.5% (11,14), and those of anammox bacteria were larger with 30.1 to 45.3% (15). The cause for the quantitatively different isotope effects of 15 « NO2-!NO3-in the diverse NOB remained unclear. ...
... The 15 « NH41!NO2-was 227.1% 6 0.8% based on the residual substrate (equation 2, Fig. 1C, and Table 1) and 232.2% 6 1.4% based on the cumulative product (Table 1), which was in agreement with the 15 « NH41!NO2-(238 to 214%) of canonical AOB and AOA (7,8). The 15 « NO2-!NO3-was 7.6% 6 0.2% based on the residual substrate from the Solver model (Table 1), which was close to the canonical Nitrospira NOB (9%) (11,14). Experiment 2: nitrite oxidation with an initial concentration of 1 mM NO 2 . ...
... Hence, the kinetic isotope effects of ammonia oxidizers may be modulated by environmental factors, some of which have been investigated in our study (see below). Like canonical NOB (10,11,14), N. inopinata displayed an inverse 15 « NO2-!NO3-, meaning that 15 NO 2 2 was preferentially oxidized to NO 3 2 during NO 2 2 oxidation. The measured value of 15 « NO2-!NO3-(9.2% 6 0.5% at pH 8.2 and 1 mM NO 2 2 ) was in line with previously determined 15 « NO2-!NO3-values (9.1 to 10.2%) of canonical Nitrospira NOB (11,14) ( Table 2 and Fig. S3). ...
Nitrification is an important nitrogen cycle process in terrestrial and aquatic environments. The discovery of comammox has changed the view that canonical AOA, AOB, and NOB are the only chemolithoautotrophic organisms catalyzing nitrification.
... Further, nitrite oxidation in ODZs has been shown to have a half-saturation constant (K m ) of <1 μM for O 2 (Bristow et al., 2016), below most of the in situ concentrations observed here. (Jacob et al., 2017;Zhang et al., 2020). ...
Marine oxygen deficient zones are dynamic areas of microbial nitrogen cycling. Nitrification, the microbial oxidation of ammonia to nitrate, plays multiple roles in the biogeochemistry of these regions, including production of the greenhouse gas nitrous oxide (N2O). We present here the results of two oceanographic cruises investigating nitrification, nitrifying microorganisms, and N2O production and distribution from the offshore waters of the Eastern Tropical South Pacific. On each cruise, high‐resolution measurements of ammonium ([NH4⁺]), nitrite ([NO2⁻]), and N2O were combined with ¹⁵N tracer‐based determination of ammonia oxidation, nitrite oxidation, nitrate reduction, and N2O production rates. Depth‐integrated inventories of NH4⁺ and NO2⁻ were positively correlated with one another and with depth‐integrated primary production. Depth‐integrated ammonia oxidation rates were correlated with sinking particulate organic nitrogen flux but not with primary production; ammonia oxidation rates were undetectable in trap‐collected sinking particulate material. Nitrite oxidation rates exceeded ammonia oxidation rates at most mesopelagic depths. We found positive correlations between archaeal amoA genes and ammonia oxidation rates and between Nitrospina‐like 16S rRNA genes and nitrite oxidation rates. N2O concentrations in the upper oxycline reached values of >140 nM, even at the western extent of the cruise track, supporting air‐sea fluxes of up to 1.71 μmol m⁻² day⁻¹. Our results suggest that a source of NO2⁻ other than ammonia oxidation may fuel high rates of nitrite oxidation in the offshore Eastern Tropical South Pacific and that air‐sea fluxes of N2O from this region may be higher than previously estimated.
... In the modern oxic ocean, ammonium released from organic matter remineralization is oxidized to nitrite by AOA, which is in turn oxidized to nitrate by NOB. Efficient cycling of ammonia and nitrite results in consistently low concentrations of these substrates and the proliferation of energyefficient NOB (Nitrospina) and AOA (Thaumarchaeota) with high substrate affinity (K strategists) (82)(83)(84)(85)(86)(87). Similarly, in the redox transition zones of modern oxygen minimum zones and anoxic basins, nitrite is maintained at low concentrations not only by NOB but also, by competing anaerobic ammoniaoxidizing (anammox) bacteria that use nitrite as an electron acceptor. ...
Significance
Hopanoids are bacterial membrane lipids that are abundant in the geologic record. Hopanoids methylated at the C-2 position (2-methylhopanoids) are particularly abundant during Mesozoic episodes of ocean anoxia, yet the sources and factors that contribute to accumulation of these biomarkers remain poorly understood. Here, we report 2-methylhopanoid production in Nitrobacter , a ubiquitous group of nitrite-oxidizing bacteria that are favored under nutrient-rich growth environments, and we demonstrate that 2-methylhopanoid biosynthesis depends on cobalamin (vitamin B 12 ). We propose that Nitrobacter spp. acquire their cobalamin from ammonia-oxidizing archaea in a mutualistic relationship that promotes nitrification. Therefore, accumulation of 2-methylhopanoids during the Mesozoic could have resulted from intensification of the oxidative steps of the marine nitrogen cycle in a nutrient-rich ocean.
... Nitronauta litoralis" EB; n = 8 for N. gracilis). The other K m(app) values were retrieved from previous studies [54,55,81]. ...
Chemolithoautotrophic nitrite-oxidizing bacteria (NOB) are key players in global nitrogen and carbon cycling. Members of the phylum Nitrospinae are the most abundant, known NOB in the oceans. To date, only two closely affiliated Nitrospinae species have been isolated, which are only distantly related to the environmentally abundant uncultured Nitrospinae clades. Here, we applied live cell sorting, activity screening, and subcultivation on marine nitrite-oxidizing enrichments to obtain novel marine Nitrospinae. Two binary cultures were obtained, each containing one Nitrospinae strain and one alphaproteobacterial heterotroph. The Nitrospinae strains represent two new genera, and one strain is more closely related to environmentally abundant Nitrospinae than previously cultured NOB. With an apparent half-saturation constant of 8.7 ± 2.5 µM, this strain has the highest affinity for nitrite among characterized marine NOB, while the other strain (16.2 ± 1.6 µM) and Nitrospina gracilis (20.1 ± 2.1 µM) displayed slightly lower nitrite affinities. The new strains and N. gracilis share core metabolic pathways for nitrite oxidation and CO 2 fixation but differ remarkably in their genomic repertoires of terminal oxidases, use of organic N sources, alternative energy metabolisms, osmotic stress and phage defense. The new strains, tentatively named “ Candidatus Nitrohelix vancouverensis” and “ Candidatus Nitronauta litoralis”, shed light on the niche differentiation and potential ecological roles of Nitrospinae.
... This phylum was followed by Chloroflexi, Acidobacteria, Bacteroidetes, Nitrospirae, Actinobacteria, Verrucomicrobia, Nitrospinae, Ignavibacteriae, and Firmicutes, which collectively accounted for 90.99% of total bacterial sequences ( Figure S1). Nitrospirae and Nitrospinae are both commonly observed in freshwater environments and are essential participants in the nitration process, 45 while Firmicutes is related to the ammonization of organic nitrogen, which illustrated that nitrogen cycle processes could be active in this area. 46 However, no significant difference was found in the microbial community at the phylum level among different hydraulic regions. ...
Understanding the characteristics of biogeochemical processes in urban channel confluences is essential for the evaluation and improvement of water environmental capacity. However, influences of biogeochemical processes in confluence were always overlooked or simply parameterized since the transformation processes controlled by microbial community assembly were hard to be quantified. To address this knowledge gap, the present study proposed a novel mathematical modelling system, based on microbial community assembly theory and fluid kinetics, to decouple nitrogen dynamics into flow-induced transport and microorganism-induced transformation processes, and quantified their contributions to nitrogen concentrations. Results revealed that variable selection processes (including hydrodynamic conditions), contributed to significant difference in microbial communities among different hydraulic regions. Variation in microbial communities further shifted transformation processes. Rhodobacterales and Sphingomonadales, which were reported to be vital participants in denitrification process, were enriched in flow separation region, and promoted it as a hotspot of nitrogen removal. In flow separation region, microorganism-induced transformation processes accounted for 56% of total nitrogen removal, which was significantly higher than that in other regions (12% on average; p<0.01). Results and findings could provide useful information for the improvement of water environmental capacity.
