SNiPlay: a web-based tool for detection, management and analysis of SNP (C900)

The rapidly increasing amount of re-sequencing and genotyping data generated by large-scale genetic diversity projects requires the development of integrated bioinformatics tools able to efficiently manage, analyze, and combine genetic data with the genome's structure and with external data available. In this context, we developed SNiPlay, a flexible, user-friendly and integrative web-based tool dedicated to polymorphism discovery and analysis. It integrates: 1) a pipeline, freely accessible through the internet, combining existing software with new tools to detect SNPs and to compute different kinds of statistical indices and graphical layouts for SNP data. From Sanger sequencing traces, multiple sequence alignments or genotyping data given as input, SNiPlay detects SNPs and insertion/deletion events. In a second time, it sends sequences and genotyping data into a series of modules in charge of various post-processes: physical mapping to a reference genome, annotation (genomic position, intron/exon location, synonymous/non-synonymous substitutions), SNP frequency determination in user-defined groups, haplotype reconstruction and networking, linkage disequilibrium evaluation, and diversity analysis (Pi, Watterson's Theta, Tajima's D). 2) a database storing polymorphisms, genotyping data and sequences of grapevine produced by nationally-funded public projects. It allows one to retrieve SNPs using various filters (such as genomic position, missing data, polymorphism type, allele frequency), to compare SNP patterns between populations, and to export genotyping data or sequences in various formats. SNiPlay is available at: http://www.sniplay.cirad.fr/. (Texte intégral

Do we need to consider grape phyllosphere microbiome in breeding schemes?

The aerial surface of the plant (phyllosphere) is the habitat of complex microbial communities. These communities may have profound effects on host plant health and its performance traits. When breeding new cultivars, i.e. the aerial component of a grape plant, one can simply ignore the phyllosphere in breeding schemes if its composition is mainly dependent on the environment. It is considered an important component if the genotype is the main driver of the phyllosphere composition. In order to answer this question, we have analysed several factors influencing the structure of the phyllosphere microbial community. Using amplicon sequencing of the 16S rRNA gene and of the internal transcribed spacer (ITS), we explored the microbial diversity at genus level for both bacteria and fungi present in the phyllosphere of leaves and grape berries. We analysed it on different grape taxonomic level (between five Vitis species or a set of Vitis vinifera cultivars chosen to represent the three genetic pools of the species), for different years and on five commercially important varieties of Vitis vinifera that were sampled from three different French terroirs. Our results indicated the presence of complex microbial diversity and assemblages in the phyllosphere. A significant effect of several factors (organ, grape species, growing year and terroir) on taxa abundance was observed with varying degrees of effect. At a given location, genotypes have an impact on microbial assemblage in the phyllosphere of leaf and berries, most pronounced on fruits but the effect of terroir was much stronger than the cultivar identity when the leaf phyllosphere of five grapevine varieties grown in different agro-climatic zones was compared. Limitations of the study as well as implied consequences of this work will be discussed

Role of Galaxy in a bioinformatic plant breeding platform

With NGS development, bioinformatics has become central in plant breeding laboratories, and researchers are in need of some autonomy in its use. The Southgreen platform (CIRAD, IRD, INRA) performs bioinformatics analyses for many plant breeding research teams in Montpellier (France), and offers many systems to users: for example GNPAnnot (automatic genomic sequence annotation), Greenphyl (phylogenetic orthology prediction), ESTtik (EST annotation) or the Bacchus analysis pipeline. Most of these systems have been translated into Galaxy workflows. As for the Bacchus pipeline, it has been created at INRA Montpellier (France) to investigate clonal diversity in grapevine genomes. For this task, many softwares have been wrapped in the Galaxy framework. Bacchus can be decomposed in three steps: i) Genome reconstruction, ii) test of reconstruction results, and iii) diversity analysis. This last step is done using SNP's and structural variations. To detect SNP's, the latest Freebayes version was used, while the IDfixe software was developped for structural variation detection. Some of the softwares developped for this pipeline are now used in the international project Grapereseq. Today, the Galaxy framework is widely used by Southgreen plateform users as an alternative to the command line system. In this context, dozens of users have already been trained in Galaxy-using bioinformatics. During weekly collective pair-programming sessions, platform engineers and interested scientists integrate new tools and functionalities.Thus, Galaxy is now a core component of the plant breeding community around the Southgreen platform, and the main access portal for non- bioinformatics specialists to our computing clusters. (Résumé d'auteur

Phaeoacremonium species associated with Eutypa dieback and esca of grapevines in Algeria

Algerian grapevines showing symptoms of Eutypa dieback and esca were examined for the presence of Phaeoacremonium species. Species were identified on the basis of morphological and cultural characteristics as well as DNA sequence data (β-tubulin and actin). From a total of 200 vines sampled, 61 Phaeoacremonium isolates were obtained corresponding to four different species. Pm. aleophilum was the most frequently isolated (n=42) followed by Pm. parasiticum (n=10) and Pm. venezuelense (n=8). Pm. hispanicum was also found but only once. Phaeoacremonium species were more frequently associated with Eutypa dieback than esca symptoms. This correlates with their frequent association with sectorial brown necrosis (V-shaped necrosis).Algerian grapevines showing symptoms of Eutypa dieback and esca were examined for the presence of Phaeoacremonium species. Species were identified on the basis of morphological and cultural characteristics as well as DNA sequence data (β-tubulin and actin). From a total of 200 vines sampled, 61 Phaeoacremonium isolates were obtained corresponding to four different species. Pm. aleophilum was the most frequently isolated (n=42) followed by Pm. parasiticum (n=10) and Pm. venezuelense (n=8). Pm. hispanicum was also found but only once. Phaeoacremonium species were more frequently associated with Eutypa dieback than esca symptoms. This correlates with their frequent association with sectorial brown necrosis (V-shaped necrosis)

Genetic diversity, linkage disequilibrium and power of a large grapevine (Vitis vinifera L) diversity panel newly designed for association studies

UMR-AGAP Equipe DAVV (Diversité, adaptation et amélioration de la vigne) ; équipe ID (Intégration de Données)International audienceAbstractBackgroundAs for many crops, new high-quality grapevine varieties requiring less pesticide and adapted to climate change are needed. In perennial species, breeding is a long process which can be speeded up by gaining knowledge about quantitative trait loci linked to agronomic traits variation. However, due to the long juvenile period of these species, establishing numerous highly recombinant populations for high resolution mapping is both costly and time-consuming. Genome wide association studies in germplasm panels is an alternative method of choice, since it allows identifying the main quantitative trait loci with high resolution by exploiting past recombination events between cultivars. Such studies require adequate panel design to represent most of the available genetic and phenotypic diversity. Assessing linkage disequilibrium extent and panel power is also needed to determine the marker density required for association studies.ResultsStarting from the largest grapevine collection worldwide maintained in Vassal (France), we designed a diversity panel of 279 cultivars with limited relatedness, reflecting the low structuration in three genetic pools resulting from different uses (table vs wine) and geographical origin (East vs West), and including the major founders of modern cultivars. With 20 simple sequence repeat markers and five quantitative traits, we showed that our panel adequately captured most of the genetic and phenotypic diversity existing within the entire Vassal collection. To assess linkage disequilibrium extent and panel power, we genotyped single nucleotide polymorphisms: 372 over four genomic regions and 129 distributed over the whole genome. Linkage disequilibrium, measured by correlation corrected for kinship, reached 0.2 for a physical distance between 9 and 458 Kb depending on genetic pool and genomic region, with varying size of linkage disequilibrium blocks. This panel achieved reasonable power to detect associations between traits with high broad-sense heritability (> 0.7) and causal loci with intermediate allelic frequency and strong effect (explaining > 10 % of total variance).ConclusionsOur association panel constitutes a new, highly valuable resource for genetic association studies in grapevine, and deserves dissemination to diverse field and greenhouse trials to gain more insight into the genetic control of many agronomic traits and their interaction with the environment

Patterns of sequence polymorphism in the fleshless berry locus in cultivated and wild Vitis vinifera accessions

<p>Abstract</p> <p>Background</p> <p>Unlike in tomato, little is known about the genetic and molecular control of fleshy fruit development of perennial fruit trees like grapevine (<it>Vitis vinifera </it>L.). Here we present the study of the sequence polymorphism in a 1 Mb grapevine genome region at the top of chromosome 18 carrying the <it>fleshless berry </it>mutation (<it>flb</it>) in order, first to identify SNP markers closely linked to the gene and second to search for possible signatures of domestication.</p> <p>Results</p> <p>In total, 62 regions (17 SSR, 3 SNP, 1 CAPS and 41 re-sequenced gene fragments) were scanned for polymorphism along a 3.4 Mb interval (85,127-3,506,060 bp) at the top of the chromosome 18, in both <it>V. vinifera cv</it>. Chardonnay and a genotype carrying the <it>flb </it>mutation, <it>V. vinifera cv</it>. Ugni Blanc mutant. A nearly complete homozygosity in Ugni Blanc (wild and mutant forms) and an expected high level of heterozygosity in Chardonnay were revealed. Experiments using qPCR and BAC FISH confirmed the observed homozygosity. Under the assumption that <it>flb </it>could be one of the genes involved into the domestication syndrome of grapevine, we sequenced 69 gene fragments, spread over the <it>flb </it>region, representing 48,874 bp in a highly diverse set of cultivated and wild <it>V. vinifera </it>genotypes, to identify possible signatures of domestication in the cultivated <it>V. vinifera </it>compartment. We identified eight gene fragments presenting a significant deviation from neutrality of the Tajima's D parameter in the cultivated pool. One of these also showed higher nucleotide diversity in the wild compartments than in the cultivated compartments. In addition, SNPs significantly associated to berry weight variation were identified in the <it>flb </it>region.</p> <p>Conclusions</p> <p>We observed the occurrence of a large homozygous region in a non-repetitive region of the grapevine otherwise highly-heterozygous genome and propose a hypothesis for its formation. We demonstrated the feasibility to apply BAC FISH on the very small grapevine chromosomes and provided a specific probe for the identification of chromosome 18 on a cytogenetic map. We evidenced genes showing putative signatures of selection and SNPs significantly associated with berry weight variation in the <it>flb </it>region. In addition, we provided to the community 554 SNPs at the top of chromosome 18 for the development of a genotyping chip for future fine mapping of the <it>flb </it>gene in a F2 population when available.</p

Effet de l'Ustilago scitaminea Syd. sur le développement, le contenu en sucres et l'activité invertasique de la canne à sucre

Avant l'apparition du fouet, la maladie du charbon de la canne à sucre se caractérise par un allongement accéléré des tiges infectées. Il en résulte une diminution de leur diamètre et une différenciation précoce des feuilles, de taille réduite, correspondant aux entre-noeuds subissant l'élongation. Ces changements morphologiques sont associés à une perturbation des contenus en sucres : le rapport sucres réducteurs sur sucres totaux solubles (SR/STS) augmente plus rapidement chez les plantes malades que chez les plantes saines. Par ailleurs, l'activité invertasique (AI) du champignon augmente considérablement dans le fouet et l'accroissement de l'AI de la plante malade apparait lié au processus d'élongation. Ustilago scitaminea produit in vitro l'érythritol et le mannitol et ces polyols ont été détectés in vivo dans le fouet. L'application exogène d'acide gibberellique (GA3) provoque également une croissance accélérée de la tige et une augmentation du rapport SR/STS, mais les sensibilités au charbon et GA3 ne semblent pas être directement reliées. Le rôle du GA3 en relation avec les équilibres hormonaux est discuté et les événements physiologiques survenant lors de l'infection sont mis en parallèle avec la mise à fleu