Quantification of anammox populations enriched in an immobilized microbial consortium with low levels of ammonium nitrogen and at low temperature

Anaerobic ammonium oxidizing (anammox) bacteria present in microbial communities in two laboratory-scale upflow anoxic reactors supplied with small amounts of ammonium (< 3 mg/l) at low temperature were detected and quantified. The reactors, operated at 20 degrees C, were seeded with an immobilized microbial consortium (IMC) and anaerobic granules (AG) from an upflow anaerobic sludge blanket (UASB) treating brewery wastewater. Our results showed that complete ammonium and nitrite removal with greater than 92% total nitrogen removal efficiency was achieved in the reactor inoculated with both the IMC and AG, while that of the reactor inoculated with only the IMC was lower than 40%; enrichment was successful after the addition of AG. Quantitative fluorescence in situ hybridization (FISH) analysis confirmed that anammox bacteria were present only in the reactor inoculated with an IMC and AG. The copy number of the 16S-rRNA gene of the anammox bacteria calculated by most probable number-polymerase chain reaction (MPN-PCR) from the total DNA extracted from both reactors (2.5 x 10(4) copies/mu g of DNA) was two orders lower than that of the domain bacteria (2.5 x 10(6) copies/mu g of DNA). The results revealed that immobilized multiple seed sludges were optimal for anammox enrichment at low temperature and ammonium concentrations.