The anaerobic oxidation of ammonium.

From recent research it has become clear that at least two different possibilities for anaerobic ammonium oxidation exist in nature. 'Aerobic' ammonium oxidizers like Nitrosomonas eutropha were observed to reduce nitrite or nitrogen dioxide with hydroxylamine or ammonium as electron donor under anoxic conditions. The maximum rate for anaerobic ammonium oxidation was about 2 nmol NH4+ min-1 (mg protein)-1 using nitrogen dioxide as electron acceptor. This reaction, which may involve NO as an intermediate, is thought to generate energy sufficient for survival under anoxic conditions, but not for growth. A novel obligately anaerobic ammonium oxidation (Anammox) process was recently discovered in a denitrifying pilot plant reactor. From this system, a highly enriched microbial community with one dominating peculiar autotrophic organism was obtained. With nitrite as electron acceptor a maximum specific oxidation rate of 55 nmol NH4+ min-1 (mg protein)-1 was determined. Although this reaction is 25-fold faster than in Nitrosomonas, it allowed growth at a rate of only 0.003 h-1 (doubling time 11 days). 15N labeling studies showed that hydroxylamine and hydrazine were important intermediates in this new process. A novel type of hydroxylamine oxidoreductase containing an unusual P468 cytochrome has been purified from the Anammox culture. Microsensor studies have shown that at the oxic/anoxic interface of many ecosystems nitrite and ammonia occur in the absence of oxygen. In addition, the number of reports on unaccounted high nitrogen losses in wastewater treatment is gradually increasing, indicating that anaerobic ammonium oxidation may be more widespread than previously assumed. The recently developed nitrification systems in which oxidation of nitrite to nitrate is prevented form an ideal partner for the Anammox process. The combination of these partial nitrification and Anammox processes remains a challenge for future application in the removal of ammonium from wastewater with high ammonium concentrations.

Anaerobic oxidation of ammonium is a biologically mediated process.

A newly discovered process by which ammonium is converted to dinitrogen gas under anaerobic conditions (the Anammox process) has now been examined in detail. In order to confirm the biological nature of this process, anaerobic batch culture experiments were used. All of the ammonium provided in the medium was oxidized within 9 days. In control experiments with autoclaved or raw wastewater, without added sludge or with added sterilized (either autoclaved or gamma irradiated) sludge, no changes in the ammonium and nitrate concentrations were observed. Chemical reactions could therefore not be responsible for the ammonium conversion. The addition of chloramphenicol, ampicillin, 2,4-dinitrophenol, carbonyl cyanide m-chlorophenyl-hydrazone (CCCP), and mercuric chloride (HgIICl2) completely inhibited the activity of the ammonium-oxidizing sludge. Furthermore, the rate of ammonium oxidation was proportional to the initial amount of sludge used. It was therefore concluded that anaerobic ammonium oxidation was a microbiological process. As the experiments were carried out in an oxygen-free atmosphere, the conversion of ammonium to dinitrogen gas did not even require a trace of O2. That the end product of the reaction was nitrogen gas has been confirmed by using 15NH4+ and 14NO3-. The dominant product was 14-15N2. Only 1.7% of the total labelled nitrogen gas produced was 15-15N2. It is therefore proposed that the N2 produced by the Anammox process is formed from equimolar amounts of NH4+ and NO3-.