Goats are specialized in dairy, meat and fiber production, being adapted to a wide range of environmental conditions and having a large economic impact in developing countries. In the last years, there have been dramatic advances in the knowledge of the structure and diversity of the goat genome/transcriptome and in the development of genomic tools, rapidly narrowing the gap between goat and related species such as cattle and sheep. Major advances are: 1) publication of a de novo goat genome reference sequence; 2) Development of whole genome high density RH maps, and; 3) Design of a commercial 50K SNP array. Moreover, there are currently several projects aiming at improving current genomic tools and resources. An improved assembly of the goat genome using PacBio reads is being produced, and the design of new SNP arrays is being studied to accommodate the specific needs of this species in the context of very large scale genotyping projects (i.e. breed characterization at an international scale and genomic selection) and parentage analysis. As in other species, the focus has now turned to the identification of causative mutations underlying the phenotypic variation of traits. In addition, since 2014, the ADAPTmap project (www.goatadaptmap.org) has gathered data to explore the diversity of caprine populations at a worldwide scale by using a wide variety of approaches and data.
Numerous whole genome sequencing projects already achieved or ongoing have highlighted the fact that obtaining a high quality genome sequence is necessary to address comparative genomics questions such as structural variations among genotypes and gain or loss of specific function. Despite the spectacular progress that has been done regarding sequencing technologies, accurate and reliable data are still challenging, at the whole genome scale but also when targeting specific genomic regions. These issues are even more noticeable for complex plant genomes. Most plant genomes are known to be particularly challenging due to their size, high density of repetitive elements and various levels of ploidy. To overcome these issues, we have developed a strategy in order to reduce the genome complexity by using the large insert BAC libraries combined with next generation sequencing technologies. We have compared two different technologies (Roche-454 and Pacific Biosciences PacBio RS II) to sequence pools of BAC clones in order to obtain the best quality sequence. We targeted nine BAC clones from different species (maize, wheat, strawberry, barley, sugarcane and sunflower) known to be complex in terms of sequence assembly. We sequenced the pools of the nine BAC clones with both technologies. We have compared results of assembly and highlighted differences due to the sequencing technologies used. We demonstrated that the long reads obtained with the PacBio RS II technology enables to obtain a better and more reliable assembly notably by preventing errors due to duplicated or repetitive sequences in the same region.
It is a common knowledge that sex chromosome mutations are better tolerated and more viable compared to changes in autosomes. This is explained by relatively low gene density in both the X and the Y chromosome and by random X chromosome inactivation in mammalian females buffering the effect of X-aneuploidies. However, it is not well understood why apparently similar sex chromosome abnormalities, such as X-monosomy or certain Y chromosome rearrangements, result in different phenotypic effects in different species. It is thought that this is due to species differences in the organization of the Y chromosome, differences in the set of genes escaping X-inactivation, and the presence of species/lineage specific sex-linked genes with functions in development and reproduction. Current knowledge about the species differences in sex chromosome organization and function is limited, this despite the availability of reference genome assemblies for most domestic species. It appears that sequence assembly of the X chromosome in most species is rather patchy containing multiple gaps and possible misassemblies, being the poorest in the pseudoautosomal region and in regions containing putative lineage-specific sequences. The Y chromosome, on the other hand, is typically not included in the reference genome and is studied separately, whereas complete sequence assembly of the male-specific portion of the Y is not yet available for any domestic species. In this talk I will discuss comparative organization and function of animal sex chromosomes and related phenotypes proceeding from our research in horses.
Grant Cramer from the University of Nevada, Reno, and Dario Cantu from the Univeristy of Callifornia, Davis, discuss past challenges with sequencing Clone 8 of Cabernet Sauvignon (Vitis vinifera). An…
Domestication of clonally propagated crops such as pineapple from South America was hypothesized to be a ‘one-step operation’. We sequenced the genome of Ananas comosus var. bracteatus CB5 and assembled 513?Mb into 25 chromosomes with 29,412 genes. Comparison of the genomes of CB5, F153 and MD2 elucidated the genomic basis of fiber production, color formation, sugar accumulation and fruit maturation. We also resequenced 89 Ananas genomes. Cultivars ‘Smooth Cayenne’ and ‘Queen’ exhibited ancient and recent admixture, while ‘Singapore Spanish’ supported a one-step operation of domestication. We identified 25 selective sweeps, including a strong sweep containing a pair of tandemly duplicated bromelain inhibitors. Four candidate genes for self-incompatibility were linked in F153, but were not functional in self-compatible CB5. Our findings support the coexistence of sexual recombination and a one-step operation in the domestication of clonally propagated crops. This work guides the exploration of sexual and asexual domestication trajectories in other clonally propagated crops.
Forest tree species are increasingly subject to severe mortalities from exotic pests, diseases, and invasive organisms, accelerated by climate change. Forest health issues are threatening multiple species and ecosystem sustainability globally. While sources of resistance may be available in related species, or among surviving trees, introgression of resistance genes into threatened tree species in reasonable time frames requires genome-wide breeding tools. Asian species of chestnut (Castanea spp.) are being employed as donors of disease resistance genes to restore native chestnut species in North America and Europe. To aid in the restoration of threatened chestnut species, we present the assembly of a reference genome with chromosome-scale sequences for Chinese chestnut (C. mollissima), the disease-resistance donor for American chestnut restoration. We also demonstrate the value of the genome as a platform for research and species restoration, including new insights into the evolution of blight resistance in Asian chestnut species, the locations in the genome of ecologically important signatures of selection differentiating American chestnut from Chinese chestnut, the identification of candidate genes for disease resistance, and preliminary comparisons of genome organization with related species.
Intercellular communication is required for trap formation in the nematode-trapping fungus Duddingtonia flagrans.
Nematode-trapping fungi (NTF) are a large and diverse group of fungi, which may switch from a saprotrophic to a predatory lifestyle if nematodes are present. Different fungi have developed different trapping devices, ranging from adhesive cells to constricting rings. After trapping, fungal hyphae penetrate the worm, secrete lytic enzymes and form a hyphal network inside the body. We sequenced the genome of Duddingtonia flagrans, a biotechnologically important NTF used to control nematode populations in fields. The 36.64 Mb genome encodes 9,927 putative proteins, among which are more than 638 predicted secreted proteins. Most secreted proteins are lytic enzymes, but more than 200 were classified as small secreted proteins (< 300 amino acids). 117 putative effector proteins were predicted, suggesting interkingdom communication during the colonization. As a first step to analyze the function of such proteins or other phenomena at the molecular level, we developed a transformation system, established the fluorescent proteins GFP and mCherry, adapted an assay to monitor protein secretion, and established gene-deletion protocols using homologous recombination or CRISPR/Cas9. One putative virulence effector protein, PefB, was transcriptionally induced during the interaction. We show that the mature protein is able to be imported into nuclei in Caenorhabditis elegans cells. In addition, we studied trap formation and show that cell-to-cell communication is required for ring closure. The availability of the genome sequence and the establishment of many molecular tools will open new avenues to studying this biotechnologically relevant nematode-trapping fungus.
A whole genome scan of SNP data suggests a lack of abundant hard selective sweeps in the genome of the broad host range plant pathogenic fungus Sclerotinia sclerotiorum.
The pathogenic fungus Sclerotinia sclerotiorum infects over 600 species of plant. It is present in numerous environments throughout the world and causes significant damage to many agricultural crops. Fragmentation and lack of gene flow between populations may lead to population sub-structure. Within discrete recombining populations, positive selection may lead to a ‘selective sweep’. This is characterised by an increase in frequency of a favourable allele leading to reduction in genotypic diversity in a localised genomic region due to the phenomenon of genetic hitchhiking. We aimed to assess whether isolates of S. sclerotiorum from around the world formed genotypic clusters associated with geographical origin and to determine whether signatures of population-specific positive selection could be detected. To do this, we sequenced the genomes of 25 isolates of S. sclerotiorum collected from four different continents-Australia, Africa (north and south), Europe and North America (Canada and the northen United States) and conducted SNP based analyses of population structure and selective sweeps. Among the 25 isolates, there was evidence for two major population clusters. One of these consisted of 11 isolates from Canada, the USA and France (population 1), and the other consisted of nine isolates from Australia and one from Morocco (population 2). The rest of the isolates were genotypic outliers. We found that there was evidence of outcrossing in these two populations based on linkage disequilibrium decay. However, only a single candidate selective sweep was observed, and it was present in population 2. This sweep was close to a Major Facilitator Superfamily transporter gene, and we speculate that this gene may have a role in nutrient uptake from the host. The low abundance of selective sweeps in the S. sclerotiorum genome contrasts the numerous examples in the genomes of other fungal pathogens. This may be a result of its slow rate of evolution and low effective recombination rate due to self-fertilisation and vegetative reproduction.
Complete Genome Sequence of Bradyrhizobium sp. Strain ORS3257, an Efficient Nitrogen-Fixing Bacterium Isolated from Cowpea in Senegal.
Here, we report the complete genome sequence of Bradyrhizobium sp. strain ORS3257, which forms efficient symbioses with cowpea, peanut, or groundnut. These genomic data will be useful to identify genes associated with symbiotic performance and host compatibility on several legumes, including Aeschynomene species, with which a Nod-independent type III secretion system (T3SS)-dependent symbiosis can be established.
A High-Quality Grapevine Downy Mildew Genome Assembly Reveals Rapidly Evolving and Lineage-Specific Putative Host Adaptation Genes.
Downy mildews are obligate biotrophic oomycete pathogens that cause devastating plant diseases on economically important crops. Plasmopara viticola is the causal agent of grapevine downy mildew, a major disease in vineyards worldwide. We sequenced the genome of Pl. viticola with PacBio long reads and obtained a new 92.94?Mb assembly with high contiguity (359 scaffolds for a N50 of 706.5?kb) due to a better resolution of repeat regions. This assembly presented a high level of gene completeness, recovering 1,592 genes encoding secreted proteins involved in plant-pathogen interactions. Plasmopara viticola had a two-speed genome architecture, with secreted protein-encoding genes preferentially located in gene-sparse, repeat-rich regions and evolving rapidly, as indicated by pairwise dN/dS values. We also used short reads to assemble the genome of Plasmopara muralis, a closely related species infecting grape ivy (Parthenocissus tricuspidata). The lineage-specific proteins identified by comparative genomics analysis included a large proportion of RxLR cytoplasmic effectors and, more generally, genes with high dN/dS values. We identified 270 candidate genes under positive selection, including several genes encoding transporters and components of the RNA machinery potentially involved in host specialization. Finally, the Pl. viticola genome assembly generated here will allow the development of robust population genomics approaches for investigating the mechanisms involved in adaptation to biotic and abiotic selective pressures in this species. © The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.
Extracellular RNA has been proposed to mediate communication between cells and organisms however relatively little is understood regarding how specific sequences are selected for export. Here, we describe a specific Argonaute protein (exWAGO) that is secreted in extracellular vesicles (EVs) released by the gastrointestinal nematode Heligmosomoides bakeri, at multiple copies per EV. Phylogenetic and gene expression analyses demonstrate exWAGO orthologues are highly conserved and abundantly expressed in related parasites but highly diverged in free-living genus Caenorhabditis. We show that the most abundant small RNAs released from the nematode parasite are not microRNAs as previously thought, but rather secondary small interfering RNAs (siRNAs) that are produced by RNA-dependent RNA Polymerases. The siRNAs that are released in EVs have distinct evolutionary properties compared to those resident in free-living or parasitic nematodes. Immunoprecipitation of exWAGO demonstrates that it specifically associates with siRNAs from transposons and newly evolved repetitive elements that are packaged in EVs and released into the host environment. Together this work demonstrates molecular and evolutionary selectivity in the small RNA sequences that are released in EVs into the host environment and identifies a novel Argonaute protein as the mediator of this. © The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.
Genetic characterization and potential molecular dissemination mechanism of tet(31) gene in Aeromonas caviae from an oxytetracycline wastewater treatment system.
Recently, the rarely reported tet(31) tetracycline resistance determinant was commonly found in Aeromonas salmonicida, Gallibacterium anatis, and Oblitimonas alkaliphila isolated from farming animals and related environment. However, its distribution in other bacteria and potential molecular dissemination mechanism in environment are still unknown. The purpose of this study was to investigate the potential mechanism underlying dissemination of tet(31) by analysing the tet(31)-carrying fragments in A. caviae strains isolated from an aerobic biofilm reactor treating oxytetracycline bearing wastewater. Twenty-three A. caviae strains were screened for the tet(31) gene by polymerase chain reaction (PCR). Three strains (two harbouring tet(31), one not) were subjected to whole genome sequencing using the PacBio RSII platform. Seventeen A. caviae strains carried the tet(31) gene and exhibited high resistance levels to oxytetracycline with minimum inhibitory concentrations (MICs) ranging from 256 to 512?mg/L. tet(31) was comprised of the transposon Tn6432 on the chromosome of A. caviae, and Tn6432 was also found in 15 additional tet(31)-positive A. caviae isolates by PCR. More important, Tn6432 was located on an integrative conjugative element (ICE)-like element, which could mediate the dissemination of the tet(31)-carrying transposon Tn6432 between bacteria. Comparative analysis demonstrated that Tn6432 homologs with the structure ISCR2-?phzF-tetR(31)-tet(31)-?glmM-sul2 were also carried by A. salmonicida, G. anatis, and O. alkaliphila, suggesting that this transposon can be transferred between species and even genera. This work provides the first report on the identification of the tet(31) gene in A. caviae, and will be helpful in exploring the dissemination mechanisms of tet(31) in water environment.Copyright © 2018. Published by Elsevier B.V.
Population Genome Sequencing of the Scab Fungal Species Venturia inaequalis, Venturia pirina, Venturia aucupariae and Venturia asperata.
The Venturia genus comprises fungal species that are pathogens on Rosaceae host plants, including V. inaequalis and V. asperata on apple, V. aucupariae on sorbus and V. pirina on pear. Although the genetic structure of V. inaequalis populations has been investigated in detail, genomic features underlying these subdivisions remain poorly understood. Here, we report whole genome sequencing of 87 Venturia strains that represent each species and each population within V. inaequalis We present a PacBio genome assembly for the V. inaequalis EU-B04 reference isolate. The size of selected genomes was determined by flow cytometry, and varied from 45 to 93 Mb. Genome assemblies of V. inaequalis and V. aucupariae contain a high content of transposable elements (TEs), most of which belong to the Gypsy or Copia LTR superfamilies and have been inactivated by Repeat-Induced Point mutations. The reference assembly of V. inaequalis presents a mosaic structure of GC-equilibrated regions that mainly contain predicted genes and AT-rich regions, mainly composed of TEs. Six pairs of strains were identified as clones. Single-Nucleotide Polymorphism (SNP) analysis between these clones revealed a high number of SNPs that are mostly located in AT-rich regions due to misalignments and allowed determining a false discovery rate. The availability of these genome sequences is expected to stimulate genetics and population genomics research of Venturia pathogens. Especially, it will help understanding the evolutionary history of Venturia species that are pathogenic on different hosts, a history that has probably been substantially influenced by TEs.Copyright © 2019 Le Cam et al.
We report the first annotated chromosome-level reference genome assembly for pea, Gregor Mendel’s original genetic model. Phylogenetics and paleogenomics show genomic rearrangements across legumes and suggest a major role for repetitive elements in pea genome evolution. Compared to other sequenced Leguminosae genomes, the pea genome shows intense gene dynamics, most likely associated with genome size expansion when the Fabeae diverged from its sister tribes. During Pisum evolution, translocation and transposition differentially occurred across lineages. This reference sequence will accelerate our understanding of the molecular basis of agronomically important traits and support crop improvement.
Whole-genome sequencing reveals recent and frequent genetic recombination between clonal lineages of Cryphonectria parasitica in western Europe.
Changes in the mode of reproduction are frequently observed in invasive fungal populations. The ascomycete Cryphonectria parasitica, which causes Chestnut Blight, was introduced to Europe from North America and Asia in the 20th century. Previous genotyping studies based on ten microsatellite markers have identified several clonal lineages which have spread throughout western Europe, suggesting that asexuality was the main reproductive mode of this species during colonization, although occasional sexual reproduction is not excluded. Based on the whole-genome sequences alignment of 46 C. parasitica isolates from France, North America and Asia, genealogy and population structure analyses mostly confirmed these lineages as clonal. However, one of these clonal lineages showed a signal of strong recombination, suggesting different strategies of reproduction in western Europe. Signatures of several recent recombination events within all the French clonal lineages studied here were also identified, indicating that gene flow is regular between these lineages. In addition, haplotype identification of seven French clonal lineages revealed that emergences of new clonal lineages during colonization were the result of hybridization between the main expanding clonal lineages and minor haplotypes non-sequenced in the present study. This whole-genome sequencing study underlines the importance of recombination events in the invasive success of these clonal populations, and suggests that sexual reproduction may be more frequent within and between the western European clonal lineages of C. parasitica than previously assumed using few genetic markers.Copyright © 2019 Elsevier Inc. All rights reserved.