Aerobic ammonium oxidizing bacteria were first isolated more than 100 years ago and hydroxylamine is known to be an intermediate. The enzymatic steps involving hydroxylamine conversion to nitrite are still under discussion. For a long time it was assumed that hydroxylamine was directly converted to nitrite by a hydroxylamine oxidoreductase. Recent enzymatic evidences suggest that the actual product of hydroxylamine conversion is NO and a third, yet unknown, enzyme further converts NO to nitrite. More recently, ammonium oxidizing archaea and complete ammonium oxidizing bacteria were isolated and identified. Still the central nitrogen metabolism of these microorganisms presents to researchers the same puzzle: how hydroxylamine is transformed to nitrite. Nitrogen losses in the form of NO and N2O have been identified in all three types of aerobic ammonium oxidizing microorganisms and hydroxylamine is known to play a significant role in the formation. Yet, the pathways and the factors promoting the greenhouse gas emissions are to be fully characterized. Hydroxylamine also plays a yet poorly understood role on anaerobic ammonium oxidizing bacteria and is known to inhibit nitrite oxidizing bacteria. In this review, the role of this elusive intermediate in the metabolism of different key players of the nitrogen cycle is discussed, as well as the putative importance of hydroxylamine as a key nitrogen metabolite for microbial interactions within microbial communities and engineered systems. Overall, for the first time putting together the acquired knowledge about hydroxylamine and the nitrogen cycle over the years in a review, setting potential hypothesis and highlighting possible next steps for research.@en

Hydroxylamine is a nitrogen intermediate of ammonium oxidizing bacteria (AOB) that can transiently accumulate during nitrification. The impact of hydroxylamine on aerobic ammonium oxidations is still obscure. In the present study the short and long term impact of hydroxylamine on partial nitritation granular sludge was investigated. Dissolved oxygen was the governing factor determining the hydroxylamine impact in short term studies with continuous hydroxylamine feeding. Continuous short term hydroxylamine feeding together with low dissolved oxygen resulted in higher hydroxylamine accumulation, higher N2O production and decreased or maintained ammonium consumption. Instead, high dissolved oxygen reduced hydroxylamine accumulation and N2O production and increased ammonium consumption. Long term continuous hydroxylamine feeding reduced ammonium consumption rate while the constant nitrite production rate indicated that dosed hydroxylamine was mainly transformed to nitrite. This indicates that hydroxylamine was preferred over ammonium as substrate. Nitrosomonas sp. was shown to be predominant during continuous hydroxylamine feeding while the side community shifted.@en

Hydroxylamine is a key intermediate in several biological reactions of the global nitrogen cycle. However, the role of hydroxylamine in anammox is still not fully understood. In this work, the impact of hydroxylamine (also in combination with other substrates) on the metabolism of a planktonic enrichment culture of the anammox species Ca. Kuenenia stuttgartiensis was studied. Anammox bacteria were observed to produce ammonium both from hydroxylamine and hydrazine, and hydroxylamine was consumed simultaneously with nitrite. Hydrazine accumulation - signature for the presence of anammox bacteria - strongly depended on the available substrates, being higher with ammonium and lower with nitrite. Furthermore, the results presented here indicate that hydrazine accumulation is not the result of the inhibition of hydrazine dehydrogenase, as commonly assumed, but the product of hydroxylamine disproportionation. All kinetic parameters for the identified reactions were estimated by mathematical modelling. Moreover, the simultaneous consumption and growth on ammonium, nitrite and hydroxylamine of anammox bacteria was demonstrated, this was accompanied by a reduction in the nitrate production. Ultimately, this study advances the fundamental understanding of the metabolic versatility of anammox bacteria, and highlights the potential role played by metabolic intermediates (i.e. hydroxylamine, hydrazine) in shaping natural and engineered microbial communities.@en

A lab-scale partial nitritation granular sludge air-lift reactor was operated in continuous mode treating low strength synthetic medium (influent ca. 50 mg-N-NH4 +/L). Granules were initially stratified with AOB in the external shell and NOB in the inner core at 20 °C. Once temperature was decreased progressively from 20 °C to 15 °C, nitrate production was initially observed during several weeks. However, by maintaining relatively high ammonium concentrations in the liquid (ca. 28 mg-N-NH4 +/L), effluent nitrate concentrations in the reactor decreased in time and process performance was recovered. Batch tests were performed in the reactor at different conditions. To understand the experimental results an existing one-dimensional biofilm model was used to simulate batch tests and theoretically assess the impact of stratification, dissolved oxygen (DO) and short-term effects of temperature on time course concentrations of ammonium, nitrite and nitrate. This theoretical assessment served to develop an experimental methodology for the evaluation of in-situ batch tests in the partial nitritation reactor. These batch tests proved to be a powerful tool to easily monitor the extent of stratification of nitrifier guilds in granular sludge and to determine the required bulk ammonium concentration to minimize nitrite oxidation. When nitrifier guilds were stratified in the granular sludge, a higher bulk ammonium concentration was required to efficiently repress NOB at lower temperature (ca. 19 versus 7 mg-N-NH4 +/L at 15 and 20 °C, respectively).@en

One of the least studied nitrogen compounds is hydroxylamine. Hydroxylamine is a highly reactive and toxic inorganic compound that some microorganisms use as intermediate during nitrogen conversions. The aim of this thesis was to further understand the role of hydroxylamine in the nitrogen cycle.@en