STABLE-ISOTOPE ANALYSIS OF A COMBINED NITRIFICATION-DENITRIFICATION SUSTAINED BY THERMOPHILIC METHANOTROPHS UNDER LOW-OXYGEN CONDITIONS

To simulate growth conditions experienced by microbiota at O-2-limited interfaces of organic matter in compost, an experimental system capab le of maintaining dual limitations of oxygen and carbon for extended p eriods, i.e., a pO(2)-auxostat, has been used. N-15 tracer studies on thermophilic (53 degrees C) decomposition processes occurring in manur e-straw aggregates showed the emission of dinitrogen gas from the reac tor as a result of simultaneous nitrification and denitrification at l ow pO(2) values (0.1 to 2.0%, vol/vol). The N loss was confirmed by ni trogen budget studies of the system, Depending on the imposed pO(2), 0 .6 to 1.4 mmol of N/day (i.e., 20 to 40% of input N) was emitted as N- 2. When the pO(2) was raised, the rates of both nitrification and deni trification increased instantaneously, indicating that ammonia oxidati on was limited by oxygen. In auxostats permanently running at pO(2) gr eater than or equal to 2% (vol/vol), the free ammonium pool was almost completely oxidized and was converted to nitrite plus nitrate and N-2 gas. Labelling of the auxostat with [C-13]carbonate was conducted to reveal whether nitrification was of autotrophic or heterotrophic origi n, Incorporation of (CO2)-C-13 into population-specific cellular compo unds was evaluated by profiling the saponifiable phospholipid fairy ac ids (FAs) by using capillary gas chromatography and subsequently analy zing the C-13/C-12 ratios of the individual FAs, after their combustio n to CO2, by isotope ratio mass spectrometry. Apart from the observed label incorporation into FAs originating from a microflora belonging t o the genus Methylococcus (type X group), supporting nitrification of a methylotrophic nature, this analysis also corroborated the absence o f truly autotrophic nitrifying populations, Nevertheless, the extent t o which ammonia oxidation continued to exist in this thermophilic comm unity suggested that a major energy gain could be associated with it.