Effect of Nitrate and Acetylene on nirS, cnorB, and nosZ Expression and Denitrification Activity in Pseudomonas mandelii

Nitrate acts as an electron acceptor in the denitrification process. The effect of nitrate in the range of 0 to 1,000 mg/liter on Pseudomonas mandelii nirS, cnorB, and nosZ gene expression was studied, using quantitative reverse transcription-quantitative PCR. Denitrification activity was measured by using the acetylene blockage method and gas chromatography. The effect of acetylene on gene expression was assessed by comparing denitrification gene expression in P. mandelii culture grown in the presence or absence of acetylene. The higher the amount of NO₃⁻ present, the greater the induction and the longer the denitrification genes remained expressed. nirS gene expression reached a maximum at 2, 4, 4, and 6 h in cultures grown in the presence of 0, 10, 100, and 1,000 mg of KNO₃/liter, respectively, while induction of nirS gene ranged from 12- to 225-fold compared to time zero. cnorB gene expression also followed a similar trend. nosZ gene expression did not respond to NO₃⁻ treatment under the conditions tested. Acetylene decreased nosZ gene expression but did not affect nirS or cnorB gene expression. These results showed that nirS and cnorB responded to nitrate concentrations; however, significant denitrification activity was only observed in culture with 1,000 mg of KNO₃/liter, indicating that there was no relationship between gene expression and denitrification activity under the conditions tested.