Accumulation of nitrogen oxides in copper-limited cultures of denitrifying bacteria

Three strains of heterotrophic denitrifying bacteria were cultured in artificial seawater medium under trace metal clean conditions to investigate their physiological response to changes in copper concentration. Decreasing the copper concentration in cultures of Pseudomonas stutzeri and Paracoccus denitrificans resulted in accumulation of nitrous oxide (N₂O) gas compared to copper-replete cultures and cessation of growth before complete denitrification to dinitrogen. Correspondingly, the activity of the copper enzyme nitrous oxide reductase, measured for P. denitrificans cultures, was high in copper-replete cultures and was below detection in copper-deficient cultures. Addition of copper to copper-deficient cultures following the accumulation of N₂O resulted in resumption of growth and complete consumption of N₂O in solution. Growth of the third strain, WLB20, in copper-deficient medium caused a marked accumulation of nitrite, suggesting that WLB20 has the copper form of nitrite reductase. These observations suggest a role for trace metals in regulating redox cycling of nitrogen and trace gas production in the ocean.