In this study, microcosms were used to investigate the influence of temperature (4 and 28 degrees C) and water content (45% and 90% WHC) on microbial communities and activities in carbon-rich fen soil. Bacterial, archaeal and denitrifier community composition was assessed during incubation of microcosms for 12 weeks using terminal restriction fragment length polymorphism (T-RFLP) profiling of 16S rRNA and nitrous oxide reductase (nosZ) genes. In addition, microbial and denitrifier abundance, potential denitrification activity and production of greenhouse gases were measured. No detectable changes were observed in prokaryote or denitrifier abundance. In general, cumulatively after 12 weeks more carbon was respired at the higher temperature (3.7 mg CO(2) g(-1) soil), irrespective of the water content, whereas nitrous oxide production was greater under wet conditions (98-336 microg N(2)O g(-1) soil). After an initial lag phase, methane emissions (963 microg CH(4) g(-1) soil) were observed only under warm and wet conditions. T-RFLP analyses of bacterial 16S rRNA and nosZ genes revealed small or undetectable community changes in response to temperature and water content, suggesting that bacterial and denitrifying microbial communities are stable and do not respond significantly to seasonal changes in soil conditions. In contrast, archaeal microbial community structure was more dynamic and was strongly influenced by temperature.

The effect of standard agricultural management on the genetic heterogeneity of nitrous oxide reductase (nosZ) fragments from denitrifying prokaryotes in native and cultivated soil was explored. Thirty-six soil cores were composited from each of the two soil management conditions. nosZ gene fragments were amplified from triplicate samples, and PCR products were cloned and screened by restriction fragment length polymorphism (RFLP). The total nosZ RFLP profiles increased in similarity with soil sample size until triplicate 3-g samples produced visually identical RFLP profiles for each treatment. Large differences in total nosZ profiles were observed between the native and cultivated soils. The fragments representing major groups of clones encountered at least twice and four randomly selected clones with unique RFLP patterns were sequenced to verify nosZ identity. The sequence diversity of nosZ clones from the cultivated field was higher, and only eight patterns were found in clone libraries from both soils among the 182 distinct nosZ RFLP patterns identified from the two soils. A group of clones that comprised 32% of all clones dominated the gene library of native soil, whereas many minor groups were observed in the gene library of cultivated soil. The 95% confidence intervals of the Chao1 nonparametric richness estimator for nosZ RFLP data did not overlap, indicating that the levels of species richness are significantly different in the two soils, the cultivated soil having higher diversity. Phylogenetic analysis of deduced amino acid sequences grouped the majority of nosZ clones into an interleaved Michigan soil cluster whose cultured members are ␣-Proteobacteria. Only four nosZ sequences from cultivated soil and one from the native soil were related to sequences found in ␥-Proteobacteria. Sequences from the native field formed a distinct, closely related cluster (D mean ‫؍‬ 0.16) containing 91.6% of the native clones. Clones from the cultivated field were more distantly related to each other (D mean ‫؍‬ 0.26), and 65% were found outside of the cluster from the native soil, further indicating a difference in the two communities. Overall, there appears to be a relationship between use and richness, diversity, and the phylogenetic position of nosZ sequences, indicating that agricultural use of soil caused a shift to a more diverse denitrifying community.

Respiratory denitrification is not always adequately established when bacteria are characterized. We have tested a simple method that allows one to evaluate whether the two necessary criteria to claim denitrification have been met, namely, that N(inf2) or N(inf2)O is produced from nitrate or nitrite and that this reduction is coupled to a growth yield increase. Microorganisms were cultured in sealed tubes under a helium headspace and in the presence of 0, 2, 4, 7, and 10 mM nitrate or nitrite. After growth had ceased, N(inf2) and N(inf2)O were quantified by gas chromatography and the final protein concentration was measured. Net protein production was linearly related to nitrate concentration for all denitrifiers tested and ranged from 2 to 6 g of protein per mol of electron equivalent reduced. Nitrogen recovery as N(inf2) plus N(inf2)O from nitrate and nitrite transformed exceeded 80% for all denitrifiers. We also suggest that a rate of N gas production of >10 (mu)mol/min/g of protein can be used as an additional characteristic definitive of denitrification since this process produces gas more rapidly than other processes. These characteristics were established after evaluation of a variety of well-characterized respiratory denitrifiers and other N(inf2)O-producing nitrate reducers. Several poorly characterized denitrifiers were also tested and confirmed as respiratory denitrifiers, including Aquaspirillum itersonii, Aquaspirillum fasciculus, Bacillus azotoformans, and Corynebacterium nephridii. These criteria distinguished respiratory denitrifiers from other groups that reduce nitrate or produce N(inf2)O. Furthermore, they correctly identified respiratory denitrification in weak denitrifiers, a group in which the existence of this process may be overlooked.

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Shifts in nitrifying community structure and function in response to different ammonium concentrations (50, 500, 1,000, and 3,000 mg of N liter−1), pH values (pH 6.0, 7.0, and 8.2), and oxygen concentrations (1, 7, and 21%) were studied in experimental reactors inoculated with nitrifying bacteria from a wastewater treatment plant. The abilities of the communities selected for these conditions to regain their original structures after conditions were returned to the original conditions were also determined. Changes in nitrifying community structure were determined by performing an amplified ribosomal DNA (rDNA) restriction analysis of PCR products obtained with ammonia oxidizer-specific rDNA primers, by phylogenetic probing, by small-subunit (SSU) rDNA sequencing, and by performing a cellular fatty acid analysis. Digestion of ammonia-oxidizer SSU rDNA with five restriction enzymes showed that a high ammonium level resulted in a great community structure change that was reversible once the ammonium concentration was returned to its original level. The smaller changes in community structure brought about by the two pH extremes, however, were irreversible. Sequence analysis revealed that the highest ammonium environment stimulated growth of a nitrifier strain that exhibited 92.6% similarity in a partial SSU rRNA sequence to its nearest relative, Nitrosomonas eutropha C-91, although the PCR product did not hybridize with a general phylogenetic probe for ammonia oxidizers belonging to the β subgroup of the classProteobacteria. A principal-component analysis of fatty acid methyl ester data detected changes from the starter culture in all communities under the new selective conditions, but after the standard conditions were restored, all communities produced the original fatty acid profiles.

The objective of the present study was to asses the effect of watertable level on N mineralization in a Histosol and a Humic Gleysol profile under natural meadows in Ljubljana marsh, Slovenia. The two soils differ significantly in organic matter content (27—40 % in Histosol and 14—20 % in Humic Gleysol) but not in C : N ratio (13—20) and pH (6.5—7.0). For each soil, the watertable was maintained at two levels (above or below 50 cm from the soil surface) for approximately one year. The four main plots, according to soil carbon content and watertable level were divided into 4 subplots, according to 4 fertilization treatments (unfertilized control, PK, PK + 50 kg N ha—1, PK + 3 × 50 kg N ha—1). Net N mineralization in unfertilized subplots was estimated from indices of N mineralization obtained by incubation of soil samples in the laboratory and by seasonal dynamics of mineral N content in the field. Annual uptake of N in herbage under the 4 fertilization treatments was also measured. Total mineral N content in topsoil was 20—80 % higher in Histosol than in Humic Gleysol. Similarly, aerobic N mineralization potentials along the entire soil profile (0—90 cm) were 20—130 % higher in Histosol than in Humic Gleysol. By contrast, anaerobic N mineralization potentials in subsoil were 10—60 % lower in Histosol than in Humic Gleysol. Both, aerobic and anaerobic N mineralization potentials strongly depended on watertable levels at sampling time. Seasonal dynamics of soil mineral N content as well as N mineralization potentials indicated that the N mineralization in the Histosol could be 10—40 % higher at low than at high watertable level. In the Humic Gleysol the N mineralization could be 10—100 % higher at high watertable level. Higher N availability in Histosol at low watertable and in Humic Gleysol at high watertable was also reflected in higher N uptake in herbage. These results indicate that N mineralization in Histosol and Humic Gleysol, was proportional to soil organic matter content, whereas in both soils, higher N mineralization rates can be expected at watertable levels between 40 and 60 cm below the soil surface, than at higher/lower watertable levels.

Bacteriophage morphotype diversity and latent period duration upon induction were correlated with the host population growth. The prophages of the lysogenic Vibrio sp. (DSM14379) were induced with mitomycin C in a batch culture with different salinity, substrate concentration or composition, and at different temperatures. Under all experimental conditions, phages were induced and a population of different complete and incomplete phage-like particles was observed in the lysate. Under favorable growth conditions, the phage-like particle community in the lysate was overpopulated with phage tail-like rigid rods. The number of rods was reduced in samples with low organic carbon concentration, samples with 8% and 10% NaCl, and samples induced at 40 and 43 degrees C. Although all lysates contained all phage-like particle-size fractions, their relative abundances varied. Up to a fivefold difference in phage-like particle size was observed in lysates. Size distribution of phage-like particles changed along temperature, salinity and organic carbon gradients. Results also indicated that the latent period of the induced phage-like particle population converged to approximately 90 min above a growth rate of 1.0 h(-1). At lower host growth rates, the latent period generally increased. However, at 40 and 43 degrees C and at low peptone-yeast extract concentration in the growth medium, the latent period remained short. We propose that different host physiological conditions influence organic matter composition upon prophage induction and may thus affect virus-controlled flow of the energy and carbon in the ecosystem.

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