A BIOLUMINESCENCE ASSAY USING NITROSOMONAS-EUROPAEA FOR RAPID AND SENSITIVE DETECTION OF NITRIFICATION INHIBITORS

An expression vector for the luxAB genes, derived from Vibrio harveyi, was introduced into Nitrosomonas europaea, Although the recombinant s train produced bioluminescence due to the expression of the luxAB gene s under normal growing conditions, the intensity of the light emission decreased immediately, in a time-and dose-dependent manner, with the addition of ammonia monooxygenase inhibitors, such as allylthiourea, p henol, and nitrapyrin. When whole cells were challenged with several n itrification inhibitors and toxic compounds, a close relationship was found between the change in the intensity of the light emission and th e level of ammonia-oxidizing activity. The response of bioluminescence to the addition of allylthiourea,vas considerably faster than the cha nge in the ammonia-oxidizing rate, measured as both the O-2 uptake and NO2- production rates. The bioluminescence of cells inactivated by am monia monooxygenase inhibitor was recovered rapidly by the addition of certain substrates for hydroxylamine oxidoreductase. These results su ggested that the inhibition of bioluminescence was caused by the immed iate decrease of reducing power in the cell due to the inactivation of ammonia monooxygenase, as well as by the destruction of other cellula r metabolic pathways. We conclude that the assay system using luminous Nitrosomonas can be applied as a rapid and sensitive detection test f or nitrification inhibitors, and it will be used to monitor the nitrif ication process in wastewater treatment plants.