Identification of transcriptional signals in Encephalitozoon cuniculi widespread among Microsporidia phylum: support for accurate structural genome annotation

<p>Abstract</p> <p>Background</p> <p>Microsporidia are obligate intracellular eukaryotic parasites with genomes ranging in size from 2.3 Mbp to more than 20 Mbp. The extremely small (2.9 Mbp) and highly compact (~1 gene/kb) genome of the human parasite <it>Encephalitozoon cuniculi </it>has been fully sequenced. The aim of this study was to characterize noncoding motifs that could be involved in regulation of gene expression in <it>E. cuniculi </it>and to show whether these motifs are conserved among the phylum Microsporidia.</p> <p>Results</p> <p>To identify such signals, 5' and 3'RACE-PCR experiments were performed on different E. cuniculi mRNAs. This analysis confirmed that transcription overrun occurs in E. cuniculi and may result from stochastic recognition of the AAUAAA polyadenylation signal. Such experiments also showed highly reduced 5'UTR's (<7 nts). Most of the <it>E. cuniculi </it>genes presented a CCC-like motif immediately upstream from the coding start. To characterize other signals involved in differential transcriptional regulation, we then focused our attention on the gene family coding for ribosomal proteins. An AAATTT-like signal was identified upstream from the CCC-like motif. In rare cases the cytosine triplet was shown to be substituted by a GGG-like motif. Comparative genomic studies confirmed that these different signals are also located upstream from genes encoding ribosomal proteins in other microsporidian species including <it>Antonospora locustae</it>, <it>Enterocytozoon bieneusi</it>, <it>Anncaliia algerae </it>(syn. <it>Brachiola algerae</it>) and <it>Nosema ceranae</it>. Based on these results a systematic analysis of the ~2000 E. cuniculi coding DNA sequences was then performed and brings to highlight that 364 translation initiation codons (18.29% of total CDSs) had been badly predicted.</p> <p>Conclusion</p> <p>We identified various signals involved in the maturation of E. cuniculi mRNAs. Presence of such signals, in phylogenetically distant microsporidian species, suggests that a common regulatory mechanism exists among the microsporidia. Furthermore, 5'UTRs being strongly reduced, these signals can be used to ensure the accurate prediction of translation initiation codons for microsporidian genes and to improve microsporidian genome annotation.</p

Unraveling biogeographical patterns and environmental drivers of soil fungal diversity at the French national scale

The fungal kingdom is among the most diversified kingdoms on Earth, with estimations of up to 12 million species. However, it remains poorly understood, with only 150 000 fungal species currently described. Given the major ecological role of fungi in ecosystem functioning, these numbers stress the importance of investigating fungal diversity description across different ecosystem types. Here, we explored the spatial distribution of the soil fungal diversity on a broad geographical scale, using the French Soil Quality Monitoring Network that covers the whole French territory (2171 soils sampled along a systematic grid). Fungal alpha diversity was assessed directly from soil DNA using a meta-barcoding approach by targeting the 18S rDNA gene. The total accumulated fungal diversity across France included 136 219 operational taxonomic units (OTUs), i.e., about 1 % of worldwide soil fungal diversity (based on a maximum diversity estimate of 12 million) for a territory representing only 0.3 % of the terrestrial surface on Earth. Based on this dataset, the first extensive map of fungal alpha diversity was drawn and showed a heterogeneous and spatially structured distribution in large biogeographical patterns of 231 km radius for richness (Hill diversity of order 0) and smaller patterns of 36 km radius for dominant fungi (Hill diversity of order 2). As related to other environmental parameters, the spatial distribution of fungal diversity (Hill numbers based on different orders of diversity) was mainly influenced by local filters such as soil characteristics and land management and also by global filters such as climate conditions with various relative influences. Interestingly, cropped soils exhibited the highest pool of fungal diversity relative to forest and vineyard soils. To complement this, soil fungal OTU network interactions were calculated for the different land uses across France. They varied hugely and showed a loss of 75 % of the complexity in crop systems and grasslands compared to forests and up to 83 % in vineyard systems. Overall, our study revealed that a nationwide survey with a high spatial-resolution approach is relevant for deeply investigating the spatial distribution and determinism of soil fungal diversity. Our findings provide novel insights for a better understanding of soil fungal ecology across the 18S rDNA gene and upgrade biodiversity conservation policies by supplying representative repositories dedicated to soil fungi.</p

Effect of different organic amendments on soil microbial communities

International audienc

Medicago truncatula genotype drives the plant nutritional strategy and its associated rhizosphere bacterial communities

International audienceWith the growing concern of developing a more sustainable agriculture, decreasing the use of inputs, and promoting biological diversity, harnessing plant microbiome through plant genetics is gaining of interest to improve plant growth, nutrition, and health. While genome-wide association studies have been conducted to identify plant genes driving the plant microbiome, more multidisciplinary studies are required to assess the relationships among plant the genetic effects, the plant microbiome and plant fitness. Using a metabarcoding approach, we characterized the rhizosphere bacterial communities of a core collection of 155 Medicago truncatula genotypes together with the plant phenotype, using an ecophysiological framework, and investigate the plant genetic effects through genome-wide association studies. The different genotypes within the M. truncatula core collection showed contrasted growth and carbon and nitrogen nutritional strategies but few loci were associated to these ecophysiological traits. To go further, we described its associated rhizosphere bacterial communities, dominated by Proteobacteria, Actinobacteria and Bacteroidetes, and defined acore rhizosphere bacterial community. Next, occurrence of bacterial candidates predicting plant ecophysiological traits of interest were identified using random forest analyzes. Some of them were heritable and plant loci were identified, pinpointing genes related to response to hormone stimulus, systemic acquired resistance, response to stress, nutrient starvation ortransport, and root development.Together, these results suggest that plant genetic can affect the plant growth and nutritional strategies harnessing keystones bacteria in a well-connected network community

Medicago truncatula genotype drives the plant nutritional strategy and its associated rhizosphere bacterial communities

International audienceWith the growing concern of developing a more sustainable agriculture, decreasing the use of inputs, and promoting biological diversity, harnessing plant microbiome through plant genetics is gaining of interest to improve plant growth, nutrition, and health. While genome-wide association studies have been conducted to identify plant genes driving the plant microbiome, more multidisciplinary studies are required to assess the relationships among plant the genetic effects, the plant microbiome and plant fitness. Using a metabarcoding approach, we characterized the rhizosphere bacterial communities of a core collection of 155 Medicago truncatula genotypes together with the plant phenotype, using an ecophysiological framework, and investigate the plant genetic effects through genome-wide association studies. The different genotypes within the M. truncatula core collection showed contrasted growth and carbon and nitrogen nutritional strategies but few loci were associated to these ecophysiological traits. To go further, we described its associated rhizosphere bacterial communities, dominated by Proteobacteria, Actinobacteria and Bacteroidetes, and defined acore rhizosphere bacterial community. Next, occurrence of bacterial candidates predicting plant ecophysiological traits of interest were identified using random forest analyzes. Some of them were heritable and plant loci were identified, pinpointing genes related to response to hormone stimulus, systemic acquired resistance, response to stress, nutrient starvation ortransport, and root development.Together, these results suggest that plant genetic can affect the plant growth and nutritional strategies harnessing keystones bacteria in a well-connected network community

Optimized and standardized 192-plex solution for 16S rDNA gene sequencing on Illumina Miseq platform to assess soil biodiversity

The growing need to survey the tremendous microbial diversity in a culture independent manner, has led to the development of molecular methods through sequence profiling of conserved genes such as 16S rDNA. Next-generation sequencing technologies are now used routinely to assess bacterial communities composition in complex environmental samples. Recently, the improvement of the Illumina MiSeq platform to a 2×300 bases paired-end version made it much more attractive for 16S rDNA amplicons sequencing enabling the analysis of a longer region thus facilitating taxonomic assignation. Additionally, the Illumina V3 chemistry enables high-throughput metagenomics analysis at the greatest coverage yet possible with a lower cost per sequence. Nevertheless, known limitations associated with the sequencing of low sequence diversity samples have hampered harnessing its true potential to sequence 16S rDNA gene amplicons. Moreover, validated Illumina 16S rDNA amplicon sequencing protocols do not support more than 96 samples per run, which underutilized the overall capacity of a sequencing run. We therefore worked on the development of a standardized and optimized high-multiplexed metagenomic solution for the exploration of complex environments. We developed an integrated solution combining several technical and bioinformatical strategies that are (i) a custom and standardized amplification protocol for 16S rDNA library preparation to minimize as full as possible technical biases; (ii) a custom 192-index strategy within a single run; (iii) the inclusion of a nucleotide sequence variability at the first sequencing cycles; (iv) an accurate paired-end reads assembly process for full length amplicon analysis and (v) a dedicated pipeline for taxonomic affiliation of 16S rDNA sequences and microbial communities analysis. Our solution validated within soil samples provides high sequencing accuracy and technical reproducibility with no index biases based on taxonomic distribution analysis. The association of technical and bioinformatical improvements yields substantial cost reductions and provides greater target flexibility to assess soil biodiversity as well as functions of microbial communities through carbon, phosphorus or nitrogen cycles in future developments

Etude de la diversité bactérienne et caractérisation des voies métaboliques du méthane par une approche biopuces ADN

National audienc

Next-generation sequencing propels environmental genomics to the front line of research

International audienc