Contribution of single-cell omics to microbial ecology.

Micro-organisms play key roles in various ecosystems, but many of their functions and interactions remain undefined. To investigate the ecological relevance of microbial communities, new molecular tools are being developed. Among them, single-cell omics assessing genetic diversity at the population and community levels and linking each individual cell to its functions is gaining interest in microbial ecology. By giving access to a wider range of ecological scales (from individual to community) than culture-based approaches and meta-omics, single-cell omics can contribute not only to micro-organisms' genomic and functional identification but also to the testing of concepts in ecology. Here, we discuss the contribution of single-cell omics to possible breakthroughs in concepts and knowledge on microbial ecosystems and ecoevolutionary processes.

atz gene expressions during atrazine degradation in the soil drilosphere.

One of the various ecosystemic services sustained by soil is pollutant degradation mediated by adapted soil bacteria. The pathways of atrazine biodegradation have been elucidated but in situ expression of the genes involved in atrazine degradation has yet to be demonstrated in soil. Expression of the atzA and atzD genes involved in atrazine dechlorination and s-triazine ring cleavage, respectively, was investigated during in situ degradation of atrazine in the soil drilosphere and bulked samples from two agricultural soils that differed in their ability to mineralize atrazine. Interestingly, expression of the atzA gene, although present in both soils, was not detected. Atrazine mineralization was greatest in Epoisses soil, where a larger pool of atzD mRNA was consistently measured 7 days after atrazine treatment, compared with Vezin soil (146 vs. 49 mRNA per 10(6)16S rRNA, respectively). Expression of the atzD gene varied along the degradation time course and was profoundly modified in soil bioturbated by earthworms. The atzD mRNA pool was the highest in the soil drilosphere (casts and burrow-linings) and it was significantly different in burrow-linings compared with bulk soil (e.g. 363 vs. 146 mRNA per 10(6)16S rRNA, 7 days after atrazine treatment in Epoisses soil). Thus, consistent differences in atrazine mineralization were demonstrated between the soil drilosphere and bulk soil. However, the impact of bioturbation on atrazine mineralization depended on soil type. Mineralization was enhanced in casts, compared with bulk soil, from Epoisses soil but in burrow-linings from Vezin soil. This study is the first to report the effects of soil bioturbation by earthworms on s-triazine ring cleavage and its spatial variability in soil.

Short-term response of soil bacteria to carbon enrichment in different soil microsites.

The response of bacteria in bulk soil and earthworm casts to carbon enrichment was studied by an RNA stable-isotope probing/terminal restriction fragment length polymorphism strategy with (13)C-labeled glucose and acetate. Both the soil microsite status and the carbon enrichment selected rapidly for different active bacterial communities, which resulted in different degradation kinetics. Our study clearly illustrates the biases that are generated by adding C substrates to detect metabolically active bacteria in soil.

Co-existing grass species have distinctive arbuscular mycorrhizal communities.

Arbuscular mycorrhizal (AM) fungi are biotrophic symbionts colonizing the majority of land plants, and are of major importance in plant nutrient supply. Their diversity is suggested to be an important determinant of plant community structure, but the influence of host-plant and environmental factors on AM fungal community in plant roots is poorly documented. Using the terminal restriction fragment length polymorphism (T-RFLP) strategy, the diversity of AM fungi was assessed in 89 roots of three grass species (Agrostis capillaris, Festuca rubra, Poa pratensis) that co-occurred in the same plots of a field experiment. The impact of different soil amendments (nitrogen, lime, nitrogen and lime) and insecticide application on AM fungal community was also studied. The level of diversity found in AM fungal communities using the T-RFLP strategy was consistent with previous studies based on clone libraries. Our results clearly confirm that an AM fungal host-plant preference exists, even between different grass species. AM communities colonizing A. capillaris were statistically different from the others (P < 0.05). Although grass species evenness changed in amended soils, AM fungal community composition in roots of a given grass species remained stable. Conversely, in plots where insecticide was applied, we found higher AM fungal diversity and, in F. rubra roots, a statistically different AM fungal community.

Arbuscular mycorrhizal community composition associated with two plant species in a grassland ecosystem.

Arbuscular mycorrhizal (AM) fungi are biotrophic symbionts colonizing about two-thirds of land plant species and found in all ecosystems. They are of major importance in plant nutrient supply and their diversity is suggested to be an important determinant of plant community composition. The diversity of the AM fungal community composition in the roots of two plant species (Agrostis capillaris and Trifolium repens) that co-occurred in the same grassland ecosystem was characterized using molecular techniques. We analysed the small subunit (SSU) ribosomal RNA gene amplified from a total root DNA extract using AM fungal-specific primers. A total of 2001 cloned fragments from 47 root samples obtained on four dates were analysed by restriction fragment length polymorphism, and 121 of them were sequenced. The diversity found was high: a total of 24 different phylotypes (groups of phylogenetically related sequences) colonized the roots of the two host species. Phylogenetic analyses demonstrate that 19 of these phylotypes belonged to the Glomaceae, three to the Acaulosporaceae and two to the Gigasporaceae. Our study reveals clearly that the AM fungal community colonizing T. repens differed from that colonizing A. capillaris, providing evidence for AM fungal host preference. In addition, our results reveal dynamic changes in the AM fungal community through time.

High genetic diversity in arbuscular mycorrhizal fungi: evidence for recombination events.

The genetic diversity of spores of two indigenous species of Glomus isolated from three soils of a long-term field experiment amended by different quantities of sewage sludges has been evaluated. Three populations of spores of Glomus claroideum (W2537) and three populations of spores of Glomus DAOM 225952 (W2538) were analysed using a microsatellite primer and aliquots of genomic DNA were obtained from single spores (Inter Simple Sequence Repeat (ISSR) fingerprints). 39 polymorphic bands were found for G. claroideum, and 43 in Glomus DAOM 225952. The intraspecific diversity was high, ranging from 22 to 33 different electrophoretic types for G. claroideum, and 15-27 for Glomus DAOM 225952 depending on the population. Resampling experiments showed that the number of polymorphic bands was sufficient to score all multilocus profiles in the populations and to describe the clonality structure within populations. On average, one multilocus profile was represented by about four spores whatever the population and the species. Partitioning of the within-species phenotypic variance showed that more than 92% of the variation was found within populations, while the among-population variance component accounted for less than 8%, even though it was statistically different from 0. This result is confirmed by the fact that only few multilocus profiles were shared by two populations of G. claroideum, and none by populations of Glomus DAOM 225952. In addition to the high level of diversity observed within populations, linkage disequilibria analyses and association indices calculated across loci indicates that reproduction cannot be solely clonal. Recombination or recombination-like events are likely to occur in these arbuscular mycorrhizal fungi. An 'epidemic' population structure was found for both fungal species in the soil that had received high amounts of sewage sludge.