Quantitative PCR of denitrification genes encoding the nitrate, nitrite, and nitrous oxide reductases was used to study denitrifiers across a glacier foreland. Environmental samples collected at different distances from a receding glacier contained amounts of 16S rRNA target molecules ranging from 4.9 ؋ 10 5 to 8.9 ؋ 10 5 copies per nanogram of DNA but smaller amounts of narG, nirK, and nosZ target molecules. Thus, numbers of narG, nirK, nirS, and nosZ copies per nanogram of DNA ranged from 2.1 ؋ 10 3 to 2.6 ؋ 10 4 , 7.4 ؋ 10 2 to 1.4 ؋ 10 3 , 2.5 ؋ 10 2 to 6.4 ؋ 10 3 , and 1.2 ؋ 10 3 to 5.5 ؋ 10 3 , respectively. The densities of 16S rRNA genes per gram of soil increased with progressing soil development. The densities as well as relative abundances of different denitrification genes provide evidence that different denitrifier communities develop under primary succession: higher percentages of narG and nirS versus 16S rRNA genes were observed in the early stage of primary succession, while the percentages of nirK and nosZ genes showed no significant increase or decrease with soil age. Statistical analyses revealed that the amount of organic substances was the most important factor in the abundance of eubacteria as well as of nirK and nosZ communities, and copy numbers of these two genes were the most important drivers changing the denitrifying community along the chronosequence. This study yields an initial insight into the ecology of bacteria carrying genes for the denitrification pathway in a newly developing alpine environment.Primary successional ecosystems, such as glacier forelands and volcanoes, present an ideal opportunity to study the biological colonization of substrates. Since the ice covers of many glaciers have receded over the past century, glacier forelands have released substrates for soil development. Autotrophic colonizers are expected to be important in the initial stages of primary community assembly. Organic substrates for microbial growth, however, might also come from allochthonous dead organic matter and living invertebrates in these environments. Hodkinson et al. (8) therefore recently proposed a previously unrecognized heterotrophic phase which should allow the initial establishment of functional communities. Accordingly, future studies in microbial ecology must account for both autotrophic and heterotrophic colonization along primary successional gradients such as glacier forelands, land lifts, floodplains, landslides, and volcanoes. In the past few years, studies have focused mainly on the composition and activities of the soil microbiota in primary succession of receding glaciers (19,21,24,25). Only a few studies have employed molecular tools to understand the diversity of archaeal and bacterial community structures along the forefields of receding glaciers (2,13,20). Analyses of activity and genetic structures of the nitrate reducer community at the Rotmoosferner glacier have shown that N cycling processes as well as microbial community composition depend on the successional age (...

Changes in community structure and activity of the dissimilatory nitrate-reducing community were investigated across a glacier foreland in the Central Alps to gain insight into the successional pattern of this functional group and the driving environmental factors. Bulk soil and rhizosphere soil of Poa alpina was sampled in five replicates in August during the flowering stage and in September after the first snowfalls along a gradient from 25 to 129 years after deglaciation and at a reference site outside the glacier foreland (>2000 years deglaciated). In a laboratory-based assay, nitrate reductase activity was determined colorimetrically after 24 h of anaerobic incubation. In selected rhizosphere soil samples, the community structure of nitrate-reducing microorganisms was analysed by restriction fragment length polymorphism (RFLP) analysis using degenerate primers for the narG gene encoding the active site of the membrane-bound nitrate reductase. Clone libraries of the early (25 years) and late (129 years) succession were constructed and representative clones sequenced. The activity of the nitrate-reducing community increased significantly with age mainly due to higher carbon and nitrate availability in the late succession. The community structure, however, only showed a small shift over the 100 years of soil formation with pH explaining a major part (19%) of the observed variance. Clone library analysis of the early and late succession pointed to a trend of declining diversity with progressing age. Presumably, the pressure of competition on the nitrate reducers was relatively low in the early successional stage due to minor densities of microorganisms compared with the late stage; hence, a higher diversity could persist in this sparse environment. These results suggest that the nitrate reductase activity is regulated by environmental factors other than those shaping the genetic structure of the nitrate-reducing community.

Rhizosphere soil was sampled in monocultures of Lolium perenne and Trifolium repens in June and October 2002, at two different nitrogen fertilisation levels (14 and 56 g N m(-2) year(-1)) and under two pCO(2) atmospheres (360 and 600 ppmv) at the Swiss FACE (Free Air Carbon dioxide Enrichment) site. Directly extracted soil DNA was analysed with restriction fragment length polymorphism (PCR-RFLP) by use of degenerated primers for the narG gene encoding the active site of the membrane-bound nitrate reductase. The corresponding enzyme activity of the nitrate reductase was determined colorimetrically after 24 h of anaerobic incubation. The narG PCR-RFLP fingerprints showed that the structure of the nitrate-reducing community was primarily affected by season and pH of the sampling site, whereas CO(2) enrichment, plant species or fertiliser treatment had no apparent effect. In contrast, the nitrate reductase activity responded to N fertilisation, CO(2) enrichment and plant species in October, whereas in June drought stress most likely kept the enzyme activity at a low level in all treatments. Apparently, the respiratory nitrate-reducing community adapted to different treatments primarily by altered enzyme activity.

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