In vivo emission of dinitrogen by earthworms via denitrifying bacteria in the gut

Earthworms emit the greenhouse gas nitrous oxide (N2O), and ingested denitrifiers in the gut appear to be the main source of this N2O. The primary goal of this study was to determine if earthworms also emit dinitrogen (N-2), the end product of complete denitrification. When [N-15] nitrate was injected into the gut, the earthworms Aporrectodea caliginosa and Lumbricus terrestris emitted labeled N-2 (and also labeled N2O) under in vivo conditions; emission of N-2 by these two earthworms was relatively linear and approximated 1.2 and 6.6 nmol N-2 per h per g (fresh weight), respectively. Isolated gut contents also produced [N-15] nitrate-derived N-2 and N2O under anoxic conditions. N-2 is formed by N2O reductase, and acetylene, an inhibitor of this enzyme, inhibited the emission of [N-15]nitrate-derived N-2 by living earthworms. Standard gas chromatographic analysis demonstrated that the amount of N2O emitted was relatively linear during initial incubation periods and increased in response to acetylene. The calculated rates for the native emissions of N-2 (i.e., without added nitrate) by A. caliginosa and L. terrestris were 1.1 and 1.5 nmol N-2 per h per g (fresh weight), respectively; these emission bates approximated that of N2O. These collective observations indicate that (i) earthworms emit N-2 concomitant with the emission of N2O via the in situ activity of denitrifying bacteria in the gut and (ii) N2O is quantitatively an important denitrification-derived end product under in situ conditions.