Molecular analysis of ammonia-oxidizing bacterial populations in aerated-anoxic Orbal processes

Aerated-anoxic processes operate under the principle that small additions of oxygen to an anoxic reactor induce simultaneous nitrification and denitrification. In these systems, ammonia oxidation in the anoxic zone can easily account for 30–50% of the total nitrification in the reactor, even though the dissolved oxygen concentration is usually below detection limit. To investigate whether the nitrification efficiency in aerated-anoxic processes was due to the presence of specialized ammonia-oxidizing bacteria (AOB), an analysis of the AOB population in an aerated-anoxic Orbal process and a conventional nitrogen removal process was carried out using phylogenetic analyses based on the ammonia monooxygenase A (amoA) gene. Terminal restriction fragment length polymorphism (TRFLP) analyses revealed that Nitrosospira-like organisms were one of the major contributors to ammonia oxidation in a full-scale aerated-anoxic Orbal reactor. However, the relative populations of Nitrosospira-like and Nitrosomonas-like AOB were not constant and appeared to have seasonal variability. Cloning and sequence comparison of amoA gene fragments demonstrated that most of the AOB in the aerated-anoxic Orbal process belonged to the Nitrosospira sp. and Nitrosomonas oligotropha lineages. The abundance of Nitrosospira-like organisms in aerated-anoxic reactors is significant, since this group of AOB has not been usually associated with nitrification in wastewater treatment plants.