Effect of free ammonia on the respiration and growth processes of an enriched Nitrobacter culture

The inhibitory effect of free ammonia (FA;NH3) on the metabolism of Nitrobacter is investigated using a method that allows decoupling energy generation from growth processes. A lab-scale sequencing batch reactor (SBR) was operated for the enrichment of Nitrobacter. Fluorescent in situ hybridization (FISH) analysis showed that 73% of the bacterial population in the reactor was Nitrobacter, while no Nitrospira was detected. Batch tests were carried out to measure the oxygen uptake rate (OUR) by the culture at various FA levels, in the presence (OURwithCO2) or absence (OURwithoutCO2) of inorganic carbon (CO2, HCO3− and CO32−). The FA inhibition on the respiration initiated at below 1 mgNH3–N L−1 in both cases. OURwithoutCO2 gradually decreased by 12% when the FA concentration increased from 0 to approximately 4 mgNH3–N L−1 and remained at the same level till an FA level of 9 mgNH3–N L−1 (the highest FA concentration applied in this study). This indicates that FA has a limited inhibitory effect on the respiratory capability of Nitrobacter. Starting from a level that is 15% higher than OURwithoutCO2 when no FA was present, OURwithCO2 decreased more rapidly than OURwithoutCO2 reaching the same level as OURwithoutCO2 when FA was between 6–9 mgNH3–N L−1. This implies that in this range of FA the presence of inorganic carbon did not cause any increase in the respiration activity of Nitrobacter. The results suggest that, while still oxidizing nitrite at approximately 75% of the non-inhibited rate, Nitrobacter likely ceased to grow at an FA level of above 6 mgNH3–N L−1. While the real mechanisms remain to be identified, this study indicates that the FA inhibition on Nitrobacter is likely much more serious than suggested by previous studies where OURwithCO2 (or the equivalent nitrite oxidation rate) was used as the sole measure of the inhibitory effects.