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Metabolic pathway of anaerobic ammonium oxidation on the basis of 15N studies in a fluidized bed reactor
Summary: A novel metabolic pathway for anaerobic ammonium oxidation with nitrite as the electron acceptor has been elucidated using 15N-Iabelled nitrogen compounds. These experiments showed that ammonium was biologically oxidized with hydroxylamine as the most probable electron acceptor. The hydroxylamine itself is most likely derived from nitrite. Batch experiments in which ammonium was oxidized with hydroxylamine transiently accumulated hydrazine. The conversion of hydrazine to dinitrogen gas is postulated as the reaction generating electron equivalents for the reduction of nitrite to hydroxylamine. During the conversion of ammonium a small amount of nitrate was formed from some of the nitrite. The addition of NH2OH to an operating fluidized bed system caused a stoichiometric increase in the ammonium conversion rate (1 mmol I−1 h−1) and a decrease in the nitrate production rate (0.5 mmol I−1 h−1). Addition of hydrazine also caused a decrease in nitrate production. On the basis of these findings it is postulated that the oxidation of nitrite to nitrate could provide the anaerobic ammonium-oxidizing bacteria with the reducing equivalents necessary for CO2 fixation.
Autotrophic growth of anaerobic ammonium-oxidizing micro-organisms in a fluidized bed reactor
An autotrophic synthetic medium for the enrichment of anaerobic ammonium-oxidizing (Anammox) micro-organisms was developed. This medium contained ammonium and nitrite as the only electron donor and electron acceptor respectively while carbonate was the only carbon source provided. Preliminary studies showed that the presence of nitrite and the absence of organic electron donors were essential for Anammox activity. The conversion rate of the enrichment culture in a fluidized bed reactor was 3 kg NH4 + m−3 d−1 when fed with 30 mM NH4 +. This is equivalent to a specific anaerobic ammonium oxidation rate of 1000–1100 nmol NH4 +h−1 (mg volatile solids)−1. The maximum specific oxidation rate obtained was 1500 nmol NH4 +h−1 (mg volatile solids)−1. Per mol NH4 + oxidized 0.041mol CO2 were incorporated resulting in a estimated growth rate of 0.001 h−1. The main product of the Anammox reaction is N2 but about 10% of the N-feed is converted to NO3 −. The overall nitrogen balance gave a ratio of NH4 −-conversion to NO2 −-conversion and NO3 −-production of 1:1·31±0·06:2·02±0·02. During the conversion of NH4 + with NO2 − no other intermediates or end-products such as hydroxylamine NO and N2O could be detected. Acetylene phosphate and oxygen were shown to be strong inhibitors of the Anammox activity. The dominant type of micro-organism in the enrichment culture was an irregularly shaped cell with an unusual morphology. During the enrichment for Anammox micro-organisms on synthetic medium an increase in ether lipids was observed. The colour of the biomass changed from brownish to red which was accompanied by an increase in the cytochrome content. Cytochrome spectra showed a peak at 470 nm gradually increasing in intensity during enrichment.
Heterotrophic Nitrification in Thiosphaera pantotropha: Oxygen Uptake and Enzyme Studies
Thiosphaera pantotropha is a heterotrophic nitrifying bacterium which reduces nitrite produced from ammonia to nitrogen gas regardless of the ambient dissolved O2 concentration. Under certain growth conditions nitrous oxide may be produced. The ammonia oxygenase showed a number of similarities with that of autotrophic nitrifiers [e.g. light sensitivity Mg2+ requirement NAD(P)H utilization] as did the hydroxylamine oxidoreductase (cytochrome c oxidation hydrazine inhibition). However there were also differences (e.g. hydroxylamine inhibition of ammonia oxidation) and this apparent similarity may be superficial. Control experiments with a strain of Paracoccus denitrificans (which does not nitrify) did not show the presence of either enzyme.
Thiomicrospira crunogena sp. nov., a Colorless, Sulfur-Oxidizing Bacterium from a Deep-Sea Hydrothermal Vent †
A new species of the genus Thiomicrospira was isolated from the 21°N deep-sea (2600-m) hydrothermal vent area of the East Pacific Rise. This organism is an obligate chemolithoautotrophic sulfur oxidizer and differs from the two other species of this genus by its deoxyribonucleic acid base composition and by its growth rate and optimal pH in thiosulfate medium. The new species is named Thiomicrospira crunogena and has been deposited in the American Type Culture Collection as strain ATCC 35932 as well as in the Delft Culture Collection as strain LMD 84.00.
Thiosphaera pantotropha gen. nov. sp. nov., a Facultatively Anaerobic, Facultatively Autotrophic Sulphur Bacterium
During studies on a desulphurizing denitrifying effluent-treatment system an organism which is able to grow aerobically and anaerobically on reduced sulphur compounds and hydrogen while fixing carbon dioxide was isolated. The new isolate is also capable of mixotrophic and heterotrophic growth on a wide range of substrates and is therefore a facultatively aerobic facultative autotroph. Comparisons with two similar species Thiobacillus A2 and Paracoccus denitrificans showed that the new isolate is significantly different from the other two and merits separate classification. In view of its ability to oxidize reduced sulphur compounds and because it is a chain-forming coccus rather than a rod the new isolate has been given the generic name of Thiosphaera and the species name pantotropha in recognition of its wide range of possible substrates.