Nitrous oxide (N2O) production pathway in a signal-stage nitritation-anammox sequencing batch reactor (SBR) was investigated based on a multilateral approach including real-time N2O monitoring, N2O ...isotopic composition analysis, and in-situ analyses of spatial distribution of N2O production rate and microbial populations in granular biomass. N2O emission rate was high in the initial phase of the operation cycle and gradually decreased with decreasing NH4+ concentration. The average emission of N2O was 0.98 ± 0.42% and 1.35 ± 0.72% of the incoming nitrogen load and removed nitrogen, respectively. The N2O isotopic composition analysis revealed that N2O was produced via NH2OH oxidation and NO2− reduction pathways equally, although there is an unknown influence from N2O reduction and/or anammox N2O production. However, the N2O isotopomer analysis could not discriminate the relative contribution of nitrifier denitrification and heterotrophic denitrification in the NO2− reduction pathway. Various in-situ techniques (e.g. microsensor measurements and FISH (fluorescent in-situ hybridization) analysis) were therefore applied to further identify N2O producers. Microsensor measurements revealed that approximately 70% of N2O was produced in the oxic surface zone, where nitrifiers were predominantly localized. Thus, NH2OH oxidation and NO2 reduction by nitrifiers (nitrifier-denitrification) could be responsible for the N2O production in the oxic zone. The rest of N2O (ca. 30%) was produced in the anammox bacteria-dominated anoxic zone, probably suggesting that NO2− reduction by coexisting putative heterotrophic denitrifiers and some other unknown pathway(s) including the possibility of anammox process account for the anaerobic N2O production. Further study is required to identify the anaerobic N2O production pathways. Our multilateral approach can be useful to quantitatively examine the relative contributions of N2O production pathways. Good understanding of the key N2O production pathways is essential to establish a strategy to mitigate N2O emission from biological nitrogen removal processes.
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•N2O production pathways were studied by N2O isotopic, FISH, and microsensor analyses.•Ca. 70% of N2O was produced in nitrifiers-dominated oxic surface zone of granules.•The rest of N2O (30%) was produced in the anammox bacteria-dominated anoxic zone.•Overall, N2O was produced via NH2OH oxidation and NO2− reduction pathways equally.
ABSTRACT
Cryoconite granules are naturally occurring microbial structures on glacier surfaces worldwide. They play a key role in carbon and nitrogen cycling in glacier ecosystems and can accelerate ...the melting of snow and ice. However, detailed mechanism of nitrogen cycling in cryoconite granules remains unclear. Here, we demonstrate that redox stratification affects the spatial distribution of N cycling processes in cryoconite granules. Based on microsensor measurements for O2, NH4+, NO2– and NO3–, we identified the presence of fine-scale redox stratification within cryoconite granules. Cyanobacteria at the surface layer of the granules created oxic conditions, whereas the inner core of the granules was anoxic. Metatranscriptomic analyses indicated the active occurrences of nitrification in the inner core, whereas denitrification actively occurred both in the inner core and the surface layer of the granules. Cyanobacteria in the inner core of the granules were inactive, and likely dead and being degraded, providing carbon and nitrogen to support nitrifiers and denitrifiers. Quantities of nitrification genes/transcripts were greater in large cryoconite granules than small ones, most likely because nitrogen substrates were more abundantly present in the inner core of large granules due to distinct redox stratification. Our results suggest that the development of a granular structure of cryoconite granules can largely affect carbon and nitrogen cycling on glaciers.
Development of layered microbial structure and resulting redox stratification influence the nitrogen cycling within cryoconite granules, naturally occurring microbial structures found in glacier environments.
An autotrophic partial nitrification granule. Display omitted
•The detection frequency of clones related to Nitrosomonas AOB was 62%.•Hydroxylamine oxidation was a key N2O production pathway in PN ...granules.•AOB were dominant and N2O was produced in the upper 200μm of PN granules.•N2O emission increased with increasing DO concentration.•In situ analyses explained N2O emission mechanisms in the PN process.
The effects of dissolved oxygen (DO) and pH on nitrous oxide (N2O) production rates and pathways in autotrophic partial nitrification (PN) granules were investigated at the granular level. N2O was primarily produced by betaproteobacterial ammonia-oxidizing bacteria, mainly Nitrosomonas europaea, in the oxic surface layer (<200μm) of the autotrophic PN granules. N2O production increased with increasing bulk DO concentration owing to activation of the ammonia (i.e., hydroxylamine) oxidation in this layer. The highest N2O emissions were observed at pH 7.5, although the ammonia oxidation rate was unchanged between pH 6.5 and 8.5. Overall, the results of this study suggest that in situ analyses of PN granules are essential to gaining insight into N2O emission mechanisms in a granule.
An ionophore-doped sensing membrane phosphate (PO4) microsensor based on bis(dibromophenylstannyl)methane (Bis microsensor) is described. The Bis microsensor showed a Nernstian response. The response ...of the Bis microsensor was log-linear down to a monohydrogen phosphate ion (HPO42−) concentration of 0.5 μM (corresponding to 1.0 μM of orthophosphate at pH 7.2), whereas the detection limit of PO4-microsensors based on trialkyl/aryltin chloride was 50 μM of HPO42−. The Bis microsensor showed excellent selectivity for HPO42− against nitrite, nitrate, chloride, bicarbonate and sulfate, as compared with PO4 microsensors based on trialkyl/aryltin chloride. Dissolved oxygen, which is known to interfere with the response of a previously developed cobalt-based potentiometric solid-state PO4 microsensor, had no effect on the response of the ionophore-doped sensing membrane-type microsensors described herein. Only OH− (i.e., pH) interfered with the ionophore-doped sensing membrane-type microsensors.
Anaerobic ammonium oxidation (anammox) is a promising process for NH4+-rich wastewaters such as anaerobic digester liquids. In the present study, we investigated various properties of an up-flow ...column reactor containing anammox granules and fed with a real digester liquid at four different concentrations (Phases 1 to 4). The efficiencies of NH4+ and NO2− removal decreased by up to 32% and 42%, respectively, in the digester-liquid-fed reactor (reactor-DL). When the performance of reactor-DL deteriorated, the community structure, spatial distribution, and in situ anammox activity in the two reactors were further investigated using 16S rRNA gene-based phylogenetic analysis, fluorescence in situ hybridization (FISH), and microelectrode measurements. The phylogenetic analysis and FISH results showed that non-anammox bacteria were predominant in the granule outer layers in reactor-DL, whereas anammox bacteria still dominated the granule interiors. Microelectrode measurements showed clear evidence of NH4+ oxidation activity in the interiors of granules from reactor-DL. Batch experiments using anammox granules at different acetate concentrations indicated that concentrations up to 50 mM had no effects on the anammox activity, whereas inorganic carbon uptake decreased in the presence of acetate. The present study clearly shows that the anammox activity and anammox bacterial density in the granules were maintained after feeding the digester liquid to the reactor for 140 days.
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•Digester liquids were fed to an anammox reactor.•Anammox activity and bacterial density were lower in the outer layer of granules.•Number of coexisting non-anammox bacteria increased in the outer layer of granules.•Anammox activity in the inner part of the granules was maintained.
Hematological studies of any animal species comprise an important diagnostic method in veterinary medicine and an essential tool for the conservation of species. In Sri Lanka, this essential ...technique has been ignored in studies of many species including reptiles. The aim of the present work was to establish a reference range of hematological values and morphological characterization of wild spectacled cobras (
) in Sri Lanka in order to provide a diagnostic tool in the assessment of health condition in reptiles and to diagnose diseases in wild populations.
Blood samples were collected from the ventral caudal vein of 30 wild-caught
(18 males and 12 females). Hematological analyses were performed using manual standard methods.
Several hematological parameters were examined and their mean values were: red blood cell count 0.581 ± 0.035 × 10
/μL in males; 0.4950 ± 0.0408 × 10
/μL in females; white blood cell count 12.45 ± 1.32 × 10
/μL in males; 11.98 ± 1.62 × 10
/μL in females; PCV (%) in males was 30.11 ± 1.93 and in females was 23.41 ± 1.67; hemoglobin (g/dL) was 7.6 ± 0.89 in males and 6.62 ± 1.49 in females; plasma protein (g/dL) was 5.11 ± 0.75 in males and 3.25 ± 0.74 in females; whereas cholesterol (mg/mL) was 4.09 ± 0.12 in males and 3.78 ± 0.42 in females. There were no significant differences in hematological parameters between the two genders except for erythrocyte count, thrombocyte count, hematocrit, hemoglobin, plasma protein, percentage of azurophil and heterophil. Intracellular parasites were not found in any of the studied specimens.
Hematological and plasma biochemical parameters indicated a difference between geographically isolated populations and some values were significantly different between the two genders. These hematological results provide a reference range for Sri Lankan population of adult
.
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