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This paper originates from an address at the 8th International Symposium on Nitrogen Fixation with Non-Legumes, Sydney, NSW, December 2000 Different Azospirillumstrains and some other plant growth-promoting rhizobacteria (PGPR) were screened for the occurrence of genes coding for denitrification and nitrogenase reductase (nifH) using polymerase chain reaction (PCR)-based techniques. All PGPR examined were nitrogenase-positive. Azospirillum strains were remarkably dissimilar with respect to denitrification capabilities, in particular with respect to genes of the dissimilatory nitrite reductase. A. brasilense, A. lipoferum and A. halopraeferens strains possess a cytochrome cd1-containing nitrite reductase with low sequence similarities among them. A. irakense and A. doebereinerae have a Cu-containing nitrite reductase and A. amazonense is unable to denitrify. The molecular data were corroborated by activity measurements. The current results indicate that the inability to perform denitrification is unlikely a selective advantage for Azospirillum spp. and other associative bacteria for forming an association with plant roots.

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The heterotrophic nitrifier Pseudomonas putida aerobically oxidized ammonia to hydroxylamine, nitrite, and nitrate. Product formation was accompanied by a small but significant release of NO, whereas N2O evolution could not be detected under the assay conditions employed. The isolate reduced nitrate to nitrite and partially further to NO under anaerobic conditions. Aerobically grown cells utilized gamma-aminobutyrate as a carbon source and as a N-source by ammonification. The physiological experiments, in particular the inhibition pattern by C2H2, indicated that P. putida expressed an ammonia monooxigenase. DNA-hybridization with an amoA gene probe coding for the smaller subunit of the ammonia monooxigenase of Nitrosomonas europaea allowed us to identify, to clone, and to sequence a region with an open reading frame showing distinct sequence similarities to the amoA gene of autotrophic ammonia oxidizers.

Bacteria isolated from different layers of four soils of the Cologne area were analyzed for denitrifying, nitrifying and N2-fixing isolates by colony hybridization using gene probes. In the soils tested, the percentage of denitrifying bacteria among the total population isolated was 3 -8 % (in one case exceptionally 15% ) and thus small. Denitrifying bacteria were partic ularly enriched in the upper layer (depth ~ 5 cm) and were present only in low amounts at 25 cm depth in two gleysol soils. Nitrate apparently did not determine the distribution of denitrifying bacteria in these soils. The potential denitrification activity of different soil layers coincided with the distribution pattern of isolates assessed by DNA-probing. The total number of bacteria and of denitrifying isolates was considerably higher in or at the roots of plants than in the bulk, root-free soil adjacent to the plants. The percentage of the isolated aerobic N2-fixing bacteria varied between 0 -3 % , and these bacteria could be isolated mainly from the upper 5 cm layer. A small portion of the isolates hybridized with the probe coding for part of one subunit of ammonia monooxygenase from Nitrosomonas europaea. The inves tigation showed that DNA-probing can provide useful information about the relative distri bution of denitrifying and N2-fixing bacteria in different soils and their layers.