Metabolism of nitric oxide by Pseudomonas stutzeri in culture and in soil

Abstract

Denitrification of nitrate with glucose as electron donor was measured in Pseudomonas stutzeri in batch culture and after inoculation into sterile soil. In closed culture, denitrification of 4 mM nitrate, NO and N₂O with maximum concentrations of 2.2 mM, 126 nM and 169 nM, respectively. Denitrification experiments in soil were done in a flow-through system in which the soil was continuously flushed with N₂. Denitrification of about 80 μg nitrate-N g⁻¹dw soil (= 67mM) resulted in the intermediate accumulation of about 3–6 μg nitrate-N g⁻¹dw soil (= 2.5–5.0 mM. The produced NO and N₂O whcih were flushed out of the system amounted to 37–61% and 2–6% of the reduced nitrate, respectively. Both glucose and nitrate were required for denitrificatiom and for release of NO and N₂O. NO consumption wasm studied in nitrate-exhausted soil using the flow-through system. NO consumption followed Michaelis-Menten kinetics. The apparent V_max was about 15–42 amol NO h⁻¹ cell⁻¹, and the apparent K_m was about 0.92 ppmv NO equivalent to 1.5 nM NO in the soil aqueous phase. At high soil water content (60–75% WHC), O₂ mixing ratios between 0 and 20% in the gas phase had no short-term effect on the V_max of NO consumption. At low soil water contents (10–30% WHC) however, the aerobic V_max reached only about 20% of the aneorobic control. At high soil water contents only prolonged incubation (up to 96 h) under 20% O₂ resulted also in decreased V_max being about 30% of the anaerobic control The K_m of NO consumption was unaffected by all these treatments. The NO reductase activity of P. stutzeri appeared to be relatively resistant towards O₂ when the bacteria had been inoculated into soil, and was sufficient to explain NO uptake in oxic soils.