Peaks of in situ N2O emissions are influenced by N2O‐producing and reducing microbial communities across arable soils

Agriculture is the main source of terrestrial N₂O emissions, a potent greenhouse gas and the main cause of ozone depletion. The reduction of N₂O into N₂ by microorganisms carrying the nitrous oxide reductase gene (nosZ) is the only known biological process eliminating this greenhouse gas. Recent studies showed that a previously unknown clade of N₂O‐reducers (nosZII) was related to the potential capacity of the soil to act as a N₂O sink. However, little is known about how this group responds to different agricultural practices. Here, we investigated how N₂O‐producers and N₂O‐reducers were affected by agricultural practices across a range of cropping systems in order to evaluate the consequences for N₂O emissions. The abundance of both ammonia‐oxidizers and denitrifiers was quantified by real‐time qPCR, and the diversity of nosZ clades was determined by 454 pyrosequencing. Denitrification and nitrification potential activities as well as in situ N₂O emissions were also assessed. Overall, greatest differences in microbial activity, diversity, and abundance were observed between sites rather than between agricultural practices at each site. To better understand the contribution of abiotic and biotic factors to the in situ N₂O emissions, we subdivided more than 59,000 field measurements into fractions from low to high rates. We found that the low N₂O emission rates were mainly explained by variation in soil properties (up to 59%), while the high rates were explained by variation in abundance and diversity of microbial communities (up to 68%). Notably, the diversity of the nosZII clade but not of the nosZI clade was important to explain the variation of in situ N₂O emissions. Altogether, these results lay the foundation for a better understanding of the response of N₂O‐reducing bacteria to agricultural practices and how it may ultimately affect N₂O emissions.     

Role of respiratory nitrate reductase in ability of Pseudomonas fluorescens YT101 to colonize the rhizosphere of maize

Selection of the denitrifying community by plant roots (i.e., increase in the denitrifier/total heterotroph ratio in the rhizosphere) has been reported by several authors. However, very few studies to evaluate the role of the denitrifying function itself in the selection of microorganisms in the rhizosphere have been performed. In the present study, we compared the rhizosphere survival of the denitrifying Pseudomonas fluorescens YT101 strain with that of its isogenic mutant deficient in the ability to synthesize the respiratory nitrate reductase, coinoculated in nonplanted or planted soil. We demonstrated that under nonlimiting nitrate conditions, the denitrifying wild-type strain had an advantage in the ability to colonize the rhizosphere of maize. Investigations of the effect of the inoculum characteristics (density of the total inoculum and relative proportions of mutant and wild-type strains) on the outcome of the selection demonstrated that the selective effect of the plant was expressed only during the phase of bacterial multiplication and that the intensity of selection was dependent on the magnitude of this phase. Moreover, application of the de Wit replacement series technique to our results suggests that the advantage of the wild-type strain was maximal when the ratio between the two strains in the inoculum was close to 1:1. This work constitutes the first direct demonstration that the presence of a functional structural gene encoding the respiratory nitrate reductase confers higher rhizosphere competence to a microorganism.     

Disruption of narG, the Gene Encoding the Catalytic Subunit of Respiratory Nitrate Reductase, Also Affects Nitrite Respiration in Pseudomonas fluorescens YT101

The Pseudomonas fluorescens YT101 gene narG, which encodes the catalytic alpha subunit of the respiratory nitrate reductase, was disrupted by insertion of a gentamicin resistance cassette. In the Nar(-) mutants, nitrate reductase activity was not detectable under all the conditions tested, suggesting that P. fluorescens YT101 contains only one membrane-bound nitrate reductase and no periplasmic nitrate reductase. Whereas N(2)O respiration was not affected, anaerobic growth with NO(2) as the sole electron acceptor was delayed for all of the Nar(-) mutants following a transfer from oxic to anoxic conditions. These results provide the first demonstration of a regulatory link between nitrate and nitrite respiration in the denitrifying pathway.     

Involvement of Nitrate Reductase and Pyoverdine in Competitiveness of Pseudomonas fluorescens Strain C7R12 in Soil

Involvement of nitrate reductase and pyoverdine in the competitiveness of the biocontrol strain Pseudomonas fluorescens C7R12 was determined, under gnotobiotic conditions, in two soil compartments (bulk and rhizosphere soil), with the soil being kept at two different values of matric potential (−1 and −10 kPa). Three mutants affected in the synthesis of either the nitrate reductase (Nar⁻), the pyoverdine (Pvd⁻), or both (Nar⁻ Pvd⁻) were used. The Nar⁻ and Nar⁻ Pvd⁻ mutants were obtained by site-directed mutagenesis of the wild-type strain and of the Pvd⁻ mutant, respectively. The selective advantage given by nitrate reductase and pyoverdine to the wild-type strain was assessed by measuring the dynamic of each mutant-to-total-inoculant (wild-type strain plus mutant) ratio. All three mutants showed a lower competitiveness than the wild-type strain, indicating that both nitrate reductase and pyoverdine are involved in the fitness of P. fluorescens C7R12. The double mutant presented the lowest competitiveness. Overall, the competitive advantages given to C7R12 by nitrate reductase and pyoverdine were similar. However, the selective advantage given by nitrate reductase was more strongly expressed under conditions of lower aeration (−1 kPa). In contrast, the selective advantage given by nitrate reductase and pyoverdine did not differ in bulk and rhizosphere soil, indicating that these bacterial traits are not specifically involved in the rhizosphere competence but rather in the saprophytic ability of C7R12 in soil environments.     

Comparison of the internalization efficiency of LDL and transferrin receptors on L2C guinea pig lymphocytes

We demonstrate that L2C lymphocytes have about 10-times more receptors for transferrin (Tf) than healthy lymphocytes, as has been shown in the case of LDL receptors. The dissociation constant is the same in the two cell types (about 4 X 10(-7) M). In contrast to LDL, Tf enters L2C lymphocytes with very rapid kinetics. It is shown by cross-reaction that each receptor is internalized independently of the other.     

Activity and Composition of the Denitrifying Bacterial Community Respond Differently to Long-Term Fertilization

The objective of this study was to explore the long-term effects of different organic and inorganic fertilizers on activity and composition of the denitrifying and total bacterial communities in arable soil. Soil from the following six treatments was analyzed in an experimental field site established in 1956: cattle manure, sewage sludge, Ca(NO₃)₂, (NH₄)₂SO₄, and unfertilized and unfertilized bare fallow. All plots but the fallow were planted with corn. The activity was measured in terms of potential denitrification rate and basal soil respiration. The nosZ and narG genes were used as functional markers of the denitrifying community, and the composition was analyzed using denaturing gradient gel electrophoresis of nosZ and restriction fragment length polymorphism of narG, together with cloning and sequencing. A fingerprint of the total bacterial community was assessed by ribosomal intergenic spacer region analysis (RISA). The potential denitrification rates were higher in plots treated with organic fertilizer than in those with only mineral fertilizer. The basal soil respiration rates were positively correlated to soil carbon content, and the highest rates were found in the plots with the addition of sewage sludge. Fingerprints of the nosZ and narG genes, as well as the RISA, showed significant differences in the corresponding communities in the plots treated with (NH₄)₂SO₄ and sewage sludge, which exhibited the lowest pH. In contrast, similar patterns were observed among the other four treatments, unfertilized plots with and without crops and the plots treated with Ca(NO₃)₂ or with manure. This study shows that the addition of different fertilizers affects both the activity and the composition of the denitrifying communities in arable soil on a long-term basis. However, the treatments in which the denitrifying and bacterial community composition differed the most did not correspond to treatments with the most different activities, showing that potential activity was uncoupled to community composition.     

Kinetics of phospholipid transfer between liposomes (neutral or negatively charged) and high-density lipoproteins: a spin-label study of early events

The kinetics of spin-labeled phosphatidylcholine transfer between vesicles and HDL particles exhibited a two-phase process, as seen by ESR spectroscopy. The results were analyzed by considering several possible steps in the overall transfer, whose aspects were also studied: (i) micellar complex formation after HDL apolipoprotein-vesicle mixture, (ii) the rate of PC transfer from the micellar complex to HDL, (iii) the rate of the reverse reaction between overloaded HDL particles and other particles such as HDLs, LDLs, and lipid vesicles. The results agree most convincingly with a mechanism in which the diffusion of phospholipids into the HDL-endogenous lipids is the limiting step, occurring as a two-step process. In addition, we observed a negative charge effect on the lipid transfer rates and yields.     

Effect of soil type and plant species on the fluorescent pseudomonads nitrate dissimilating community

The distribution of nitrogen dissimilative abilities among 618 isolates of fluorescent pseudomonads was studied. These strains were isolated from two uncultivated soils (C and D; collected at Chateaurenard and Dijon, France, respectively) and from rhizosphere, rhizoplane and root tissue of two plant species (flax and tomato) cultivated on these two soils. According to their ability to dissimilate nitrogen, the isolates have been distributed into three metabolic types: non-dissimilators, NO₂ ^- accumulators and denitrifiers. While the three metabolic types were recovered in all the compartments of soil D experiments, only two (non-dissimilators and denitrifiers) were recovered in all the compartments of soil C experiments. Even under the contrasting conditions of the two soil types, both plants were able to select the nitrate dissimilating community among the total community of fluorescent Pseudomonas, but the mode of this selection seems to be dependent on both plant and soil type. The soil type appears to be unable to significantly modulate the strong selective effect of tomato. Indeed, similar dissimilator to non-dissimilator ratios were found in the root tissue of this plant species cultivated in both soils. In contrast, the different dissimilator to non-dissimilator ratios observed in flax roots between soils C and D suggest that the selective effect of flax was modulated by the soil type. Taxonomic identifications showed that the 618 isolates were distributed among three species (P. chlororaphis, P. fluorescens, P. putida) plus an intermediate type between P. fluorescens and P. putida. However, no clear relationship between the distribution of the metabolic types (functional diversity) and the distribution of bacterial species has been found. This revised version was published online in June 2006 with corrections to the Cover Date.     

Unraveling negative biotic interactions determining soil microbial community assembly and functioning

Microbial communities play important roles in all ecosystems and yet a comprehensive understanding of the ecological processes governing the assembly of these communities is missing. To address the role of biotic interactions between microorganisms in assembly and for functioning of the soil microbiota, we used a top-down manipulation approach based on the removal of various populations in a natural soil microbial community. We hypothesized that removal of certain microbial groups will strongly affect the relative fitness of many others, therefore unraveling the contribution of biotic interactions in shaping the soil microbiome. Here we show that 39% of the dominant bacterial taxa across treatments were subjected to competitive interactions during soil recolonization, highlighting the importance of biotic interactions in the assembly of microbial communities in soil. Moreover, our approach allowed the identification of microbial community assembly rule as exemplified by the competitive exclusion between members of Bacillales and Proteobacteriales. Modified biotic interactions resulted in greater changes in activities related to N- than to C-cycling. Our approach can provide a new and promising avenue to study microbial interactions in complex ecosystems as well as the links between microbial community composition and ecosystem function.Subject terms: Soil microbiology, Ecology