Nitric oxide reductase gene expression and nitrous oxide production in nitrate-grown Pseudomonas mandelii

Pure cultures of Pseudomonas mandelii were incubated with or without nitrate, which acts as a substrate and an electron acceptor for denitrification. Nitric oxide reductase (cnorB) gene expression was measured using a quantitative reverse transcription-PCR, and nitrous oxide emissions were measured by gas chromatography. P. mandelii cells in either the presence or absence of nitrate demonstrated an increase in cnorB gene expression during the first 3 h of growth. The level of expression of cnorB in nitrate-amended cells remained high (average, 2.06 x 10(8) transcripts/microg of RNA), while in untreated cells it decreased to an average of 3.63 x 10(6) transcripts/microg of RNA from 4 to 6 h. Nitrous oxide accumulation in the headspace was detected at 2 h, and cumulative emissions continued to increase over a 24-h period to 101 mumol in nitrate-amended cells. P. mandelii cnorB gene expression was not detected under aerobic conditions. These results demonstrate that P. mandelii cnorB gene expression was induced 203-fold at 4 h when nitrate was present in the medium. Accumulations of N(2)O indicated that the cNorB enzyme was synthesized and active.