Use of Tn5 Mutants To Assess the Role of the Dissimilatory Nitrite Reductase in the Competitive Abilities of Two Pseudomonas Strains in Soil

We examined the influence of soil aeration state and plant root presence on the comparative survival of wild-type bacteria and isogenic Tn5 (Nir(sup-)) mutants lacking the ability to synthesize nitrite reductase. Two denitrifying Pseudomonas strains with different nitrite reductase types were used. Enumeration of bacteria in sterile and nonsterile soils was based on differential antibiotic resistance. The validity of the bacterial models studied (i.e., equal growth of wild-type and mutant bacteria under aerobic conditions and significantly better growth of wild-type bacteria under denitrifying conditions) was verified in pure-culture studies. In sterile soil, both strains survived better under aerobic than under anaerobic conditions. The lower efficiency of denitrification than O(inf2) respiration in supporting bacterial growth explained this result, and the physical heterogeneity of soil did not strongly modify the results obtained in pure-culture studies. In nonsterile soil, one of the Pseudomonas strains survived better under anaerobic conditions while the other competed equally with the indigenous soil microflora under aerobic and anaerobic conditions. However, when the Nir(sup-)-to-total inoculant ratios (wild type plus Nir(sup-) mutant) were analyzed, it appeared that the presence of nitrite reductase conferred on both Pseudomonas strains a competitive advantage for anaerobic environment or rhizosphere colonization. This is the first attempt to demonstrate with isogenic nondenitrifying mutants that denitrification can contribute to the persistence and distribution of bacteria in fluctuating soil environments

Influence of Two Plant Species (Flax and Tomato) on the Distribution of Nitrogen Dissimilative Abilities within Fluorescent Pseudomonas spp

The distribution of nitrogen-dissimilative abilities among 317 isolates of fluorescent pseudomonads was studied. These strains were isolated from an uncultivated soil and from the rhizosphere, rhizoplane, and root tissue of two plant species (flax and tomato) cultivated on this same soil. The isolates were distributed into two species, Pseudomonas fluorescens (45.1%) and Pseudomonas putida (40.4%), plus an intermediate type (14.5%). P. fluorescens was the species with the greatest proportion of isolates in the root compartments and the greatest proportion of dissimilatory and denitrifying strains. According to their ability to dissimilate nitrogen, the isolates have been distributed into nondissimilatory and dissimilatory strains, nitrate reducers and true denitrifiers with or without N(inf2)O reductase. The proportion of dissimilatory isolates was significantly enhanced in the compartments affected by flax and tomato roots (55% in uncultivated soil and 90 and 82% in the root tissue of flax and tomato, respectively). Among these strains, the proportion of denitrifiers gradually and significantly increased in the root vicinity of tomato (44, 68, 75, and 94% in uncultivated soil, rhizosphere, rhizoplane, and root tissue, respectively) and was higher in the flax rhizoplane (66%) than in the uncultivated soil. A higher proportion of N(inf2)O reducers was also found in the root compartments. This result was particularly clear for tomato. It is hypothesized that denitrification could be a selective advantage for the denitrifiers in the root environment and that this process could contribute to modify the specific composition of the bacterial communities in the rhizosphere

Early-stage bacterial colonization between a sterilized remoulded soil clod and natural soil aggregates of the same soil

The early stages of bacterial colonization of a sterilized soil put in contact with the same non-sterile soil were investigated. Soil mesocosms of 269 cm3 composed of sterilized spherical remoulded soil clods of 27mm diameter surrounded by non-sterilized 2–4mm soil aggregates of the same soil (considered as a source of bacterial colonizers) were designed. Bacterial colonization was monitored from 0 to 14 days in three concentric portions of the sterilized clods (outer, intermediate and inner) and in the surrounding non-sterile soil, by measuring cell numbers and substrate-induced respiration (SIR). In addition, modifications of genetic diversity of the soil bacterial community associated to colonization were monitored with the ribosomal intergenic spacer analysis (RISA) technique in the intermediate portion of the sterilized soil clod and in the surrounding non-sterile soil aggregates. Assessments of bacterial cell numbers and SIR rates showed that the sterilized soil clod was colonized rapidly during incubation time from its outer to its inner portion. In addition, the bacterial genetic structure of the clod varied during incubation time, suggesting succession of bacteria during recolonization. Comparison of cell numbers, SIR rates and bacterial genetic structure between the soil clod and the surrounding soil aggregates showed that the colonization process in the clod led to the establishment of a bacterial community different from the surrounding soil aggregates

Effects of steam disinfestation on community structure, abundance and activity of heterotrophic, denitrifying and nitrifying bacteria in an organic farming soil

Steam disinfestation is an ecologically less harmful alternative to synthetic chemical fumigants such as methyl bromide, which is being phased out of use due to its ozone-depleting properties. Although previous studies have characterized the effects of steaming on targeted pests, soil microorganisms, including beneficial ones, may be strongly influenced by this agricultural practice, since: (1) high temperature disturbs most soil microorganisms; and (2) disinfestation-induced changes in the soil environment can indirectly affect soil microbiota. The impact of soil disinfestation on functional bacterial communities was evaluated particularly in view of their role in nitrogen cycling. The short-term effects of steam disinfestation on heterotrophic bacteria, denitrifying and nitrifying bacteria, and their ability to recover after this disturbance were examined by surveying the enzyme activity, size and genetic structure of each community. Our results show that: (1) steaming immediately induced significant decrease in community activity and size, and changes in community composition, nitrifying bacteria being mostly affected; (2) abundances of each community reached values equal or higher than those observed in control soil within 15–60 days after steaming, but community structures remained very different as compared to those in control soil; and (3) for each activity, no complete recovery was observed after the disturbance: substrate induced respiration and denitrification increased but remained lower in steamed soil, whereas nitrification was not detectable after 62 days. Our results show that these effects of steaming on key soil functional communities can have important, long-lasting implications for nitrogen cycle that should be taken into account when evaluating the influence of such an agricultural practice

Appendix C. A photograph of DGGE analysis of 16S rRNA gene sequences of ammonia oxidizers from the intensive grazing (IG) and light grazing (LG) grassland sites, at two topographical locations (downslope and upslope).

A photograph of DGGE analysis of 16S rRNA gene sequences of ammonia oxidizers from the intensive grazing (IG) and light grazing (LG) grassland sites, at two topographical locations (downslope and upslope)

Appendix E. A table of size (in bp) of bands specific of either IG or LG treatments (i.e., bands discriminating IG vs. LG samples in PCA) for the nitrate reducing and free N2-fixing communities.

A table of size (in bp) of bands specific of either IG or LG treatments (i.e., bands discriminating IG vs. LG samples in PCA) for the nitrate reducing and free N2-fixing communities

Appendix A. A photograph showing a view of the grassland sites upslope.

A photograph showing a view of the grassland sites upslope