The Effect of Oxygen Supply on Nitrite Reduction by Tallgrass Prairie Soil Bacteria

Nitrite reduction, catalyzed by nitrite reductase, is a key step in the denitrification pathway because it catalyzes the reduction of soluble nitrite (NO2-) into nitric oxide gas (NO). The production of nitric oxide gas therefore decreases the amount of nitrogen in soils. Other studies have investigated the effect of oxygen on denitrification in a few specific microorganisms (e.g. Aquaspirillum magnetotacticum, Thiosphaera pantotropha, and Paracoccus denitrificans). In this study, we tested the effect of oxygen on nitrite reduction in seven strains representing 5 different genera obtained from Tallgrass Prairie soil. The strains were chosen based on positive detection of at least one functional gene in the denitrification pathway along with positive results for nitrate (NO3-) and/or nitrite reduction after growth in nitrate broth in a microtiter plate assay. Under these conditions which did not totally exclude oxygen, three strains were able to reduce nitrite while four strains did not reduce nitrite. All seven strains were retested for the production/consumption of nitrite under strictly anaerobic conditions using nitrate as the electron acceptor. In addition, the strains were screened for genes encoding the copper nitrite reductase (nirK) and the cytochrome cd1-nitrite reductase (nirS) and other functional markers of denitrification pathway, namely nitric oxide reductase gene (norB) and nitrous oxide (N2O) reductase gene (nosZ) by PCR amplification using specific primers. Our results show that under strict anaerobic conditions, two additional strains demonstrated nitrite reduction. Although none of the molecular markers showed perfect correlation with the ability to reduce nitrite, nirS/nirK, commonly used to screen environmental samples for denitrifying bacteria, was detected in only two of the five strains shown in this study to reduce nitrite. More nitrite reducing strains were correctly identified when both the cnorB and nosZ primer sets were used. ©2015 Oklahoma Academy of Science