Evidence for Clostridia as Agents of Dissimilatory Reduction of Nitrate to Ammonium in Soils

Abstract

Abstract Reduction of NO 3 ‐ to NH 4 + was studied in fresh and air‐dried soils which were amended with 15 NO 3 ‐ and glucose, acetate, or water and incubated anaerobically. Soils were either untreated or heat‐shocked at 68°C for 1 hour prior to amendment. Ammonium was produced rapidly after the onset of anaerobiosis, and thereafter was incorporated into organic matter. 15 NH 4 + plus 15 N‐organic matter production was observed in glucose‐amended fresh soil in quantities up to 43% of added 15 NO 3 ‐ in untreated samples and 55% in heat‐shocked samples after 5 days incubation. In soils unamended with organic carbon, NO 3 ‐ reduction to NH 4 + was minor. Pretreatment of the soils either by air‐drying or heat‐shocking increased to a similar extent the amount of NO 3 ‐ reduced to NH 4 + . The activity of clostridia during the NO 3 ‐ reduction was indicated by the absence of any effect exerted by heat‐treatment, the production of H 2 and CO 2 , and the presence of higher numbers of anaerobic sporeforming bacteria relative to denitrifying bacteria in the air‐dried soil. Also, the most common isolate capable of reducing NO 3 ‐ to NH 4 + was a Clostridium spp. The addition of washed spores of a NO 3 ‐ ‐reducing Clostridium isolated from the soil increased the formation of 15 NH 4 + ‐N plus 15 N‐organic N fourfold, an accumulation equivalent to 83% of added 15 NO 3 ‐ ‐N. The reduction of NO 3 ‐ to NH 4 + in soils was not inhibited by NH 4 + or glutamine, indicating that the mechanism of reduction was dissimilatory. This conclusion is supported by studies with several Clostrdium spp. isolated from the soils. The isolates were capable of reducing NO 3 ‐ to NH 4 + and exhibited increased cell yields when NO 3 ‐ was included in the growth medium. These studies suggest that the potential for significant dissimilatory NO 3 ‐ reduction to NH 4 + exists in most soils, principally in the sporeforming genera of Clostridium and Bacillus , but that this potential is likely expressed only when soils become anoxic and are rich in C.