Impact of copper on the diversity, abundance and transcription of nitrite and nitrous oxide reductase genes in an urban European estuary

In this study, the effects of increasing copper (Cu) concentrations on the denitrification pathway and on the diversity of a denitrifier community and gene expression were evaluated in intertidal sandy sediments of the Douro River estuary (north-west Portugal). The results demonstrated that sediment denitrification rates were highly affected by Cu; almost complete inhibition (85%) of the process was observed in sediments amended with 60 μg Cu g(-1) wet wt sediment. Moreover, the addition of Cu stimulated the accumulation of both N(2)O and NO(2)(-) and inhibited the rate of NO(3)(-) uptake. Further, the amendment with even the lowest Cu concentration (4 μg Cu g(-1) wet wt sediment) yielded a drastic decrease in the abundance of nirK, nirS and nosZ (between 79% and 81%) assessed by means of real-time PCR. In agreement, reverse transcription-PCR-denaturing gradient gel electrophoresis analysis of nirK, nirS and nosZ transcripts showed a progressive decrease in the diversity of the transcription products of these genes with an increase of the Cu concentration.