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bioRxiv2018

Hybrid de novo assembly of the draft genome of the freshwater mussel Venustaconcha ellipsiformis (Bivalvia: Unionida).

Freshwater mussels (Bivalvia: Unionida) serve an important role as aquatic ecosystem engineers but are one of the most critically imperilled groups of animals. Here, we used a combination of sequencing strategies to assemble and annotate a draft genome of Venustaconcha ellipsiformis, which will serve as a valuable genomic resource given the ecological value and unique \"doubly uniparental inheritance\" mode of mitochondrial DNA transmission of freshwater mussels. The genome described here was obtained by combining high coverage short reads (65X genome coverage of Illumina paired-end and 11X genome coverage of mate-pairs sequences) with low coverage Pacific Biosciences long reads (0.3X genome coverage). Briefly, the final scaffold assembly accounted for a total size of 1.54Gb (366,926 scaffolds, N50 = 6.5Kb, with 2.3% of \"N\" nucleotides), representing 86% of the predicted genome size of 1.80Gb, while over one third of the genome (37.5%) consisted of repeated elements and more than 85% of the core eukaryotic genes were recovered. Given the repeated genetic bottlenecks of V. ellipsiformis populations as a result of glaciations events, heterozygosity was also found to be remarkably low (0.6%), in contrast to most other sequenced bivalve species. Finally, we reassembled the full mitochondrial genome and found six polymorphic sites with respect to the previously published reference. This resource opens the way to comparative genomics studies to identify genes related to the unique adaptations of freshwater mussels and their distinctive mitochondrial inheritance mechanism.

genomics↗

bioRxiv2018

High-quality genome assemblies of 15 Drosophila species generated using Nanopore sequencing

The Drosophila genus is a unique group containing a wide range of species that occupy diverse ecosystems. In addition to the most widely studied species, Drosophila melanogaster, many other members in this genus also possess a well-developed set of genetic tools. Indeed, high-quality genomes exist for several species within the genus, facilitating studies of the function and evolution of cis-regulatory regions and proteins by allowing comparisons across at least 50 million years of evolution. Yet, the available genomes still fail to capture much of the substantial genetic diversity within the Drosophila genus. We have therefore tested protocols to rapidly and inexpensively sequence and assemble the genome from any Drosophila species using single-molecule sequencing technology from Oxford Nanopore. Here, we use this technology to present high-quality genome assemblies of 15 Drosophila species: 10 of the 12 originally sequenced Drosophila species (ananassae, erecta, mojavensis, persimilis, pseudoobscura, sechellia, simulans, virilis, willistoni, and yakuba), four additional species that had previously reported assemblies (biarmipes, bipectinata, eugracilis, and mauritiana), and one novel assembly (triauraria). Genomes were generated from an average of 29x depth-of-coverage data that after assembly resulted in an average contig N50 of 4.4 Mb. Subsequent alignment of contigs from the published reference genomes demonstrates that our assemblies could be used to close over 60% of the gaps present in the currently published reference genomes. Importantly, the materials and reagents cost for each genome was approximately $1,000 (USD). This study demonstrates the power and cost-effectiveness of long-read sequencing for genome assembly in Drosophila and provides a framework for the affordable sequencing and assembly of additional Drosophila genomes.

genomics↗

bioRxiv2018

GET_PHYLOMARKERS, a software package to select optimal orthologous clusters for phylogenomics and inferring pan-genome phylogenies, used for a critical geno-taxonomic revision of the genus Stenotrophomonas

The massive accumulation of genome-sequences in public databases promoted the proliferation of genome-level phylogenetic analyses in many areas of biological research. However, due to diverse evolutionary and genetic processes, many loci have undesirable properties for phylogenetic reconstruction. These, if undetected, can result in erroneous or biased estimates, particularly when estimating species trees from concatenated datasets. To deal with these problems, we developed GET_PHYLOMARKERS, a pipeline designed to identify high-quality markers to estimate robust genome phylogenies from the orthologous clusters, or the pan-genome matrix (PGM), computed by GET_HOMOLOGUES. In the first context, a set of sequential filters are applied to exclude recombinant alignments and those producing anomalous or poorly resolved trees. Multiple sequence alignments and maximum likelihood (ML) phylogenies are computed in parallel on multi-core computers. A ML species tree is estimated from the concatenated set of top-ranking alignments at the DNA or protein levels, using either FastTree or IQ-TREE (IQT). The latter is used by default due to its superior performance revealed in an extensive benchmark analysis. In addition, parsimony and ML phylogenies can be estimated from the PGM.\n\nWe demonstrate the practical utility of the software by analyzing 170 Stenotrophomonas genome sequences available in RefSeq and 10 new complete genomes of environmental S. maltophilia complex (Smc) isolates reported herein. A combination of core-genome and PGM analyses was used to revise the molecular systematics of the genus. An unsupervised learning approach that uses a goodness of clustering statistic identified 20 groups within the Smc at a core-genome average nucleotide identity of 95.9% that are perfectly consistent with strongly supported clades on the core- and pan-genome trees. In addition, we identified 14 misclassified RefSeq genome sequences, 12 of them labeled as S. maltophilia, demonstrating the broad utility of the software for phylogenomics and geno-taxonomic studies. The code, a detailed manual and tutorials are freely available for Linux/UNIX servers under the GNU GPLv3 license at https://github.com/vinuesa/get_phylomarkers. A docker image bundling GET_PHYLOMARKERS with GET_HOMOLOGUES is available at https://hub.docker.com/r/csicunam/get_homologues/, which can be easily run on any platform.

genomics↗

bioRxiv2018

Genomic resources for Goniozus legneri, Aleochara bilineata and Paykullia maculata, representing three independent origins of the parasitoid lifestyle in insects

Parasitoid insects are important model systems for a multitude of biological research topics and widely used as biological control agents against insect pests. While the parasitoid lifestyle has evolved numerous times in different insect groups, research has focused almost exclusively on Hymenoptera from the parasitica clade. The genomes of several members of this group have been sequenced, but no genomic resources are available from any of the other, independent evolutionary origins of the parasitoid lifestyle. Our aim here was to develop genomic resources for three parasitoid insects outside the parasitica. We present draft genome assemblies for Goniozus legneri, a parasitoid Hymenopteran more closely related to the non-parasitoid wasps and bees than to the parasitica wasps, the Coleopteran parasitoid Aleochara bilineata and the Dipteran parasitoid Paykullia maculata. The genome assemblies are fragmented, but complete in terms of gene content. We also provide preliminary structural annotations. We anticipate that these genomic resources will be valuable for testing the generality of findings obtained from parasitica wasps in future comparative studies.\n\nData availabilityThe Whole Genome Shotgun projects have been deposited at DDBJ/ENA/GenBank under the accessions NCVS00000000 (G. legneri), NBZA00000000 (A. bilineata) and NDXZ00000000 (P. maculata). The versions described in this paper are versions NCVS01000000, NBZA01000000 and NDXZ01000000, respectively. Mapped reads and genome annotations are available through http://parasitoids.labs.vu.nl/parasitoids/. This website also includes genome browsers and viroblast instances for each genome.

genomics↗

bioRxiv2018

Assembly of chloroplast genomes with long- and short-read data: a comparison of approaches using Eucalyptus pauciflora as a test case

BackgroundChloroplasts are organelles that conduct photosynthesis in plant and algal cells. Chloroplast genomes code for around 130 genes, and the information they contain is widely used in agriculture and studies of evolution and ecology. Correctly assembling complete chloroplast genomes can be challenging because the chloroplast genome contains a pair of long inverted repeats (10-30 kb). The advent of long-read sequencing technologies should alleviate this problem by providing sufficient information to completely span the inverted repeat regions. Yet, long-reads tend to have higher error rates than short-reads, and relatively little is known about the best way to combine long- and short-reads to obtain the most accurate chloroplast genome assemblies. Using Eucalyptus pauciflora, the snow gum, as a test case, we evaluated the effect of multiple parameters, such as different coverage of long (Oxford nanopore) and short (Illumina) reads, different long-read lengths, different assembly pipelines, and different genome polishing steps, with a view to determining the most accurate and efficient approach to chloroplast genome assembly.\n\nResultsHybrid assemblies combining at least 20x coverage of both long-reads and short-reads generated a single contig spanning the entire chloroplast genome with few or no detectable errors. Short-read-only assemblies generated three contigs representing the long single copy, short single copy and inverted repeat regions of the chloroplast genome. These contigs contained few single-base errors but tended to exclude several bases at the beginning or end of each contig. Long-read-only assemblies tended to create multiple contigs with a much higher single-base error rate, even after polishing. The chloroplast genome of Eucalyptus pauciflora is 159,942 bp, contains 131 genes of known function, and confirms the phylogenetic position of Eucalyptus pauciflora as a close relative of Eucalyptus regnans.\n\nConclusionsOur results suggest that very accurate assemblies of chloroplast genomes can be achieved using a combination of at least 20x coverage of long- and short-reads respectively, provided that the long-reads contain at least ~5x coverage of reads longer than the inverted repeat region. We show that further increases in coverage give little or no improvement in accuracy, and that hybrid assemblies are more accurate than long-read-only or short-read-only assemblies.

genomics↗

bioRxiv2018

Comparative Genomic Analysis of Bacillus thuringiensis Reveals Molecular Adaptation to Copper Tolerance

Bacillus thuringiensis is a type of Gram positive and rod shaped bacterium that is found in a wide range of habitats. Despite the intensive studies conducted on this bacterium, most of the information available are related to its pathogenic characteristics, with only a limited number of publications mentioning its ability to survive in extreme environments. Recently, a B. thuringiensis MCMY1 strain was successfully isolated from a copper contaminated site in Mamut Copper Mine, Sabah. This study aimed to conduct a comparative genomic analysis by using the genome sequence of MCMY1 strain published in GenBank (PRJNA374601) as a target genome for comparison with other available B. thuringiensis genomes at the GenBank. Whole genome alignment, Fragment all-against-all comparison analysis, phylogenetic reconstruction and specific copper genes comparison were applied to all forty-five B. thuringiensis genomes to reveal the molecular adaptation to copper tolerance. The comparative results indicated that B. thuringiensis MCMY1 strain is closely related to strain Bt407 and strain IS5056. This strain harbors almost all available copper genes annotated from the forty-five B. thuringiensis genomes, except for the gene for Magnesium and cobalt efflux protein (CorC) which plays an indirect role in reducing the oxidative stress that caused by copper and other metal ions. Furthermore, the findings also showed that the Copper resistance gene family, CopABCDZ and its repressor (CsoR) are conserved in almost all sequenced genomes but the presence of the genes for Cytoplasmic copper homeostasis protein (CutC) and CorC across the sample genomes are highly inconsonant. The variation of these genes across the B. thuringiensis genomes suggests that each strain may have adapted to their specific ecological niche. However, further investigations will be need to support this preliminary hypothesis.

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bioRxiv2018

Evaluation of strategies for the assembly of diverse bacterial genomes using MinION long-read sequencing

BackgroundShort-read sequencing technologies have made microbial genome sequencing cheap and accessible. However, closing genomes is often costly and assembling short reads from genomes that are repetitive and/or have extreme %GC content remains challenging. Long-read, single-molecule sequencing technologies such as the Oxford Nanopore MinION have the potential to overcome these difficulties, although the best approach for harnessing their potential remains poorly evaluated.\n\nResultsWe sequenced nine bacterial genomes spanning a wide range of GC contents using Illumina MiSeq and Oxford Nanopore MinION sequencing technologies to determine the advantages of each approach, both individually and combined. Assemblies using only MiSeq reads were highly accurate but lacked contiguity, a deficiency that was partially overcome by adding MinION reads to these assemblies. Even more contiguous genome assemblies were generated by using MinION reads for initial assembly, but these were more error-prone and required further polishing. This was especially pronounced when Illumina libraries were biased, as was the case for our strains with both high and low GC content. Increased genome contiguity dramatically improved the annotation of insertion sequences and secondary metabolite biosynthetic gene clusters, likely because long-reads can disambiguate these highly repetitive but biologically important genomic regions.\n\nConclusionsGenome assembly using short-reads is challenged by repetitive sequences and extreme GC contents. Our results indicate that these difficulties can be largely overcome by using single-molecule, long-read sequencing technologies such as the Oxford Nanopore MinION. Using MinION reads for assembly followed by polishing with Illumina reads generated the most contiguous genomes and enabled the accurate annotation of important but difficult to sequence genomic features such as insertion sequences and secondary metabolite biosynthetic gene clusters. The combination of Oxford Nanopore and Illumina sequencing is cost effective and dramatically advances studies of microbial evolution and genome-driven drug discovery.

genomics↗

bioRxiv2018

A Computational Framework To Assess Genome-Wide Distribution Of Polymorphic Human Endogenous Retrovirus-K In Human Populations

Human Endogenous Retrovirus type K (HERV-K) is the only HERV known to be insertionally polymorphic. It is possible that HERV-Ks contribute to human disease because people differ in both number and genomic location of these retroviruses. Indeed viral transcripts, proteins, and antibody against HERV-K are detected in cancers, auto-immune, and neurodegenerative diseases. However, attempts to link a polymorphic HERV-K with any disease have been frustrated in part because population frequency of HERV-K provirus at each site is lacking and it is challenging to identify closely related elements such as HERV-K from short read sequence data. We present an integrated and computationally robust approach that uses whole genome short read data to determine the occupation status at all sites reported to contain a HERV-K provirus. Our method estimates the proportion of fixed length genomic sequence (k-mers) from whole genome sequence data matching a reference set of k-mers unique to each HERV-K loci and applies mixture model-based clustering to account for low depth sequence data. Our analysis of 1000 Genomes Project Data (KGP) reveals numerous differences among the five KGP super-populations in the frequency of individual and co-occurring HERV-K proviruses; we provide a visualization tool to easily depict the prevalence of any combination of HERV-K among KGP populations. Further, the genome burden of polymorphic HERV-K is variable in humans, with East Asian (EAS) individuals having the fewest integration sites. Our study identifies population-specific sequence variation for several HERV-K proviruses. We expect these resources will advance research on HERV-K contributions to human diseases.\n\nAuthor summaryHuman Endogenous Retrovirus type K (HERV-K) is the youngest of retrovirus families in the human genome and is the only group that is polymorphic; a HERV-K can be present in one individual but absent from others. HERV-Ks could contribute to disease risk but establishing a link of a polymorphic HERV-K to a specific disease has been difficult. We develop an easy to use method that reveals the considerable variation existing among global populations in the frequency of individual and co-occurring polymorphic HERV-K, and in the total number of HERV-K that any individual has in their genome. Our study provides a global reference set of HERV-K genomic diversity and tools needed to determine the genomic landscape of HERV-K in any patient population.

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bioRxiv2018

Comparative analysis of Streptococcus genomes

BackgroundGenome sequencing of multiple strains demonstrated high variability in gene content even in closely related strains of the same species and created a newly emerged object for genomic analysis, the pan-genome, that is, the complete set of genes observed in a given species or a higher level taxon. Here we analysed the pan-genome structure and the genome evolution of 25 strains of Streptococcus suis, 50 strains of Streptococcus pyogenes and 28 strains of Streptococcus pneumoniae.\n\nResultsFractions of the pan-genome, unique, periphery, and universal genes differ in size, functional composition, the level of nucleotide substitutions, and predisposition to horizontal gene transfer and genomic rearrangements. The density of substitutions in intergenic regions appears to be correlated with selection acting on adjacent genes, implying that more conserved genes tend to have more conserved regulatory regions. The total pan-genome of the genus is open, but only due to strain-specific genes, whereas other pan-genome fractions reach saturation. The strain-specific fraction is enriched with mobile elements and hypothetical proteins, but also contains a number of candidate virulence-related genes, so it may have a strong impact on adaptability and pathogenicity.\n\nAbout 7% of single-copy periphery genes have been found in different syntenic regions. More than a half of these genes are rare in all Streptococcus species; others are rare in at least one species. We have identified the set of genes with phylogenies inconsistent with species and non-conserved location in the chromosome; these genes are candidates for horizontal transfer between species.\n\nAn inversion of length 15 kB found in four independent branches of S. pneumoniae has breakpoints formed by genes encoding a surface antigen protein (PhtD). The observed parallelism may indicate the action of an antigen variation mechanism.\n\nConclusionsMembers of the genus Streptococcus have a highly dynamic, open pan-genome, that potentially confers them with the ability to adapt to changing environmental conditions, i.e. antibiotic resistance or transmission between different hosts. Hence, understanding of genome evolution is important for the identification of potential pathogens and design of drugs and vaccines.

genomics↗

bioRxiv2018

Rhopalocnemis phalloides has one of the most reduced and mutated plastid genomes known

Although most plant species are photosynthetic, several hundred species have lost the ability to photosynthesize and instead obtain nutrients via various types of heterotrophic feeding. Their genomes, especially plastid genomes, markedly differ from the genomes of photosynthetic plants. In this work, we describe the sequenced plastid genome of the heterotrophic plant Rhopalocnemis phalloides, which belongs to the family Balanophoraceae and feeds by parasitizing on other plants. The genome is highly reduced (18 622 base pairs versus approximately 150 kilobase pairs in autotrophic plants) and possesses an outstanding AT content, 86.8%, the highest of all sequenced plant plastid genomes. The gene content of this genome is quite typical of heterotrophic plants, with all of the genes related to photosynthesis having been lost. The remaining genes are notably distorted by a high mutation rate and the aforementioned AT content. The high AT content has led to sequence convergence between some of the remaining genes and their homologues from AT-rich plastid genomes of protists. Overall, the plastid genome of R. phalloides is one of the most unusual plastid genomes known.

genomics↗

bioRxiv2018

Gene erosion and genome expansion in a group of highly host-specialized fungal phytopathogens

Due to their comparatively small genome size and short generation time, fungi are exquisite model systems to study eukaryotic genome evolution. Powdery mildew (PM) fungi present an exceptional case where their strict host dependency (a lifestyle termed obligate biotrophy) is associated with some of the largest fungal genomes sequenced so far (>100 Mbp). This size expansion is largely due to the pervasiveness of transposable elements (TEs), which can cover more than 70% of these genomes, and is associated with the loss of multiple conserved ascomycete genes (CAGs) required for a free-living lifestyle. To date, little is known about the mechanisms that drove this expansion, and information on ancestral PM genomes is lacking. We report the genome analysis of the early-diverged PM species Parauncinula polyspora that in contrast to most other PMs reproduces exclusively sexually. The P. polyspora genome is surprisingly small (<30 Mb) and sparsely equipped with TEs (<10%), despite the conserved absence of a common defense mechanism (RIP) involved in constraining repetitive elements. The genome still harbors the majority of the CAGs that are absent in the genomes of the recently evolved PMs. We speculate that TE spread might have been limited by its unique reproduction strategy and host features and further hypothesize that the loss of CAGs may promote the evolutionary isolation and host niche specialization of PM fungi. Limitations associated with this evolutionary trajectory might have been in part counteracted by the evolution of plastic, TE-rich genomes and/or the expansion of gene families encoding secreted virulence proteins.

genomics↗

bioRxiv2018

A High-Quality De Novo Genome Assembly from a Single Mosquito using PacBio Sequencing

A high-quality reference genome is a fundamental resource for functional genetics, comparative genomics, and population genomics, and is increasingly important for conservation biology. PacBio Single Molecule, Real-Time (SMRT) sequencing generates long reads with uniform coverage and high consensus accuracy, making it a powerful technology for de novo genome assembly. Improvements in throughput and concomitant reductions in cost have made PacBio an attractive core technology for many large genome initiatives, however, relatively high DNA input requirements ([~]5 {micro}g for standard library protocol) have placed PacBio out of reach for many projects on small organisms that have lower DNA content, or on projects with limited input DNA for other reasons. Here we present a high-quality de novo genome assembly from a single Anopheles coluzzii mosquito. A modified SMRTbell library construction protocol without DNA shearing and size selection was used to generate a SMRTbell library from just 100 ng of starting genomic DNA. The sample was run on the Sequel System with chemistry 3.0 and software v6.0, generating, on average, 25 Gb of sequence per SMRT Cell with 20 hour movies, followed by diploid de novo genome assembly with FALCON-Unzip. The resulting curated assembly had high contiguity (contig N50 3.5 Mb) and completeness (more than 98% of conserved genes are present and full-length). In addition, this single-insect assembly now places 667 (>90%) of formerly unplaced genes into their appropriate chromosomal contexts in the AgamP4 PEST reference. We were also able to resolve maternal and paternal haplotypes for over 1/3 of the genome. By sequencing and assembling material from a single diploid individual, only two haplotypes are present, simplifying the assembly process compared to samples from multiple pooled individuals. The method presented here can be applied to samples with starting DNA amounts as low as 100 ng per 1 Gb genome size. This new low-input approach puts PacBio-based assemblies in reach for small highly heterozygous organisms that comprise much of the diversity of life.

genomics↗

bioRxiv2019

Pericentromeric heterochromatin is hierarchically organized and spatially contacts H3K9me2/3 islands located in euchromatic genome

Membraneless pericentromeric heterochromatin (PCH) domains play vital roles in chromosome dynamics and genome stability. However, our current understanding of 3D genome organization does not include PCH domains because of technical challenges associated with repetitive sequences enriched in PCH genomic regions. We investigated the 3D architecture of Drosophila melanogaster PCH domains and their spatial associations with euchromatic genome by developing a novel analysis method that incorporates genome-wide Hi-C reads originating from PCH DNA. Combined with cytogenetic analysis, we reveal a hierarchical organization of the PCH domains into distinct \"territories.\" Strikingly, H3K9me2/3-enriched regions embedded in the euchromatic genome show prevalent 3D interactions with the PCH domain. These spatial contacts require H3K9me2/3 enrichment, are likely mediated by liquid-liquid phase separation, and may influence organismal fitness. Our findings have important implications for how PCH architecture influences the function and evolution of both repetitive heterochromatin and the gene-rich euchromatin.\n\nAuthor summaryThe three dimensional (3D) organization of genomes in cell nuclei can influence a wide variety of genome functions. However, most of our understanding of this critical architecture has been limited to the gene-rich euchromatin, and largely ignores the gene-poor and repeat-rich pericentromeric heterochromatin, or PCH. PCH comprises large part of most eukaryotic genomes, forms 3D PCH domains in nuclei, and plays vital role in chromosome dynamics and genome stability. In this study, we developed a new method that overcomes the technical challenges imposed by the highly repetitive PCH DNA, and generated a comprehensive picture of its 3D organization. Combined with image analyses, we revealed a hierarchical organization of the PCH domains. Surprisingly, we showed that distant euchromatic regions enriched for repressive epigenetic marks also dynamically interact with the main PCH domains. These 3D interactions are mediated by liquid-liquid phase separation mechanisms, similar to how oil and vinegar separate in salad dressing, and can influence the fitness of individuals. Our discoveries have strong implications for how seemingly \"junk\" DNA could impact functions in the gene-rich euchromatin.

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bioRxiv2019

Retrovirus insertion site analysis of LGL leukemia patient genomes

BackgroundLarge granular lymphocyte (LGL) leukemia is an uncommon cancer characterized by a sustained clonal proliferation of LGL cells. Antibodies reactive to retroviruses have been documented in the serum of patients with LGL leukemia. Culture or molecular approaches have to date not been successful in identifying a retrovirus. MethodsBecause a retrovirus must integrate into the genome of an infected cell, we focused our efforts on detecting a novel retrovirus integration site in the clonally expanded LGL cells. We present a new computational tool that uses long-insert mate pair sequence data to search the genome of LGL leukemia cells for retrovirus integration sites. We also utilize recently published methods to interrogate the status of polymorphic human endogenous retrovirus type K (HERV-K) provirus in patient genomes. ResultsWhile our analysis did not reveal any new retrovirus insertions in LGL genomes from LGL leukemia patients, we did identify four HERV-K provirus integration sites that are polymorphic in the human population and absent from the human reference genome, hg19. To determine if the prevalence of these or other polymorphic proviral HERV-Ks differed between LGL leukemia patients and the general population, we applied a recently developed approach that reports all sites in the human genome occupied by a proviral HERV-K. Using the 1000 genomes project (KGP) data as a reference database for HERV-K proviral prevalence at each polymorphic site, we show that there are significant differences in the number of polymorphic HERV-Ks in the genomes of LGL leukemia patients of European origin compared to individuals with European ancestry in the KGP data. ConclusionsOur study confirms that the integration of a new infectious or endogenous retrovirus does not cause the clonal expansion of LGL cells in LGL leukemia, although we do not rule out that these cells could be responding to retroviral antigens produced in other cell types. However, it is of interest that the burden of polymorphic proviral HERV-K is elevated in LGL leukemia patient genomes. Our research emphasizes the merits of comprehensive genomic assessment of HERV-K in cancer samples and suggests that further analyses to determine contributions of HERV-K to LGL leukemia are warranted.

genomics↗

bioRxiv2019

Genome improvement and genetic map construction for Aethionema arabicum, the first divergent branch in the Brassicaceae family

BackgroundThe genus Aethionema is a sister-group to the core-group of the Brassicaceae family that includes Arabidopsis thaliana and the Brassica crops. Thus, Aethionema is phylogenetically well-placed for the investigation and understanding of genome and trait evolution across the family. We aimed to improve the quality of the reference genome draft version of the annual species Aethionema arabicum. Secondly, we constructed the first Ae. arabicum genetic map. The improved reference genome and genetic map enabled the development of each other.\n\nResultsWe started with the initially published genome (version 2.5). PacBio and MinION sequencing together with genetic map v2.5 were incorporated to produce the new reference genome v3.0. The improved genome contains 203 MB of sequence, with approximately 94% of the assembly made up of called bases, assembled into 2,883 scaffolds. The N50 (10.3 MB) represents an 80-fold over the initial genome release. We generated a Recombinant Inbred Line (RIL) population that was derived from two ecotypes: Cyprus and Turkey (the reference genotype. Using a Genotyping by Sequencing (GBS) approach, we generated a high-density genetic map with 749 (v2.5) and then 632 SNPs (v3.0) was generated. The genetic map and reference genome were integrated, thus greatly improving the scaffolding of the reference genome into 11 linkage groups.\n\nConclusionsWe show that long-read sequencing data and genetics are complementary, resulting in an improved genome assembly in Ae. arabicum. They will facilitate comparative genetic mapping work for the Brassicaceae family and are also valuable resources to investigate wide range of life history traits in Aethionema.

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bioRxiv2019

Comparative genome analysis of a multidrug-resistant Pseudomonas aeruginosa sequence type 277 clone that harbours two copies of the blaSPM-1 gene and multiple single nucleotide polymorphisms in other resistance-associated genes

Pseudomonas aeruginosa is one of the most common pathogens related to healthcare-associated infections. The Brazilian isolate, named CCBH4851, is a multidrug-resistant clone belonging to the sequence type 277. The antimicrobial resistance mechanisms of the CCBH4851 strain are associated with the presence of blaSPM-1 gene, encoding a metallo-beta-lactamase, in addition to other exogenously acquired genes. Whole-genome sequencing studies focusing on emerging pathogens are essential to identify physiological key aspects that may lead to the exposure of new targets for therapy. This study was designed to characterize the genome of Pseudomonas aeruginosa CCBH4851 through the detection of genomic features and genome comparison with other Pseudomonas aeruginosa strains. The CCBH4851 closed genome showed features that were consistent with data reported for the specie. However, comparative genomics revealed the absence of genes important for pathogenesis. On the other hand, CCBH4851 genome contained acquired genomic islands that carry additional virulence and antimicrobial resistance-related genes. The presence of single nucleotide polymorphisms in the core genome, mainly those located in resistance-associated genes, suggests that these mutations could influence the multidrug-resistant behavior of CCBH4851. Overall, the characterization of Pseudomonas aeruginosa CCBH4851 complete genome revealed several features that could directly impact the profile of virulence and antibiotic resistance of this pathogen in infectious outbreaks.

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bioRxiv2019

Relevance of genomic evaluation for egg quality traits in layers

BackgroundGenomic evaluation, based on thousands of genetic markers, has become the standard evaluation methodology in dairy cattle breeding programs over the past few years. Despite the many differences between dairy cattle breeding and poultry breeding, genomic selection seems very promising for the avian sector, and studies are currently being conducted to optimize avian selection schemes. In this optimization perspective, one of the key parameters is to properly predict the accuracy of genomic evaluation in pure line layers.\n\nMethodsBoth genetic evaluation and genomic evaluation were performed on three candidate populations (male and female), using different sizes of phenotypic records on five egg quality traits and at two different ages. The methodologies used were BLUP & ssGBLUP, and variance-covariance matrices were estimated through REML. To estimate evaluation accuracy, the LR method was implemented. Four statistics were used to assess the relative accuracy of the estimated breeding values of candidates, their bias and dispersion, as well as the differences between genetic evaluation and genomic evaluation.\n\nResultsIt was observed that genomic evaluation, whether performed on males or females, always proved more accurate than genetic evaluation. The gain was higher when phenotypic information was narrowed and an augmentation of the size of the reference population led to an increase in accuracy prediction, for what regards genomic evaluation. By taking into account the increase of selection intensity and the decrease of the generation interval induced by genomic selection, the expected annual genetic gain would be higher with ancestry-based genomic evaluation of male candidates than with genetic evaluation based on collaterals. This advantage of genomic selection over genetic selection requires to be studied in more details for female candidates.\n\nConclusionsIn conclusion, in the population studied, genomic evaluation for egg quality traits of breeding birds at birth seems a promising strategy, at least for what regards males selection.

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bioRxiv2019

What can we learn from over 100,000 Escherichia coli genomes?

The explosion of microbial genome sequences in public databases allows for large-scale population genomic studies of bacterial species, such as Escherichia coli. In this study, we examine and classify more than one hundred thousand E. coli and Shigella genomes. After removing outliers, a semi-automated Mash-based analysis of 10,667 assembled genomes reveals 14 distinct phylogroups. A representative genome or medoid identified for each phylogroup serves as a proxy to classify more than 95,000 unassembled genomes. This analysis shows that most sequenced E. coli genomes belong to 4 phylogroups (A, C, B1 and E2(O157)). Authenticity of the 14 phylogroups described is supported by pangenomic and phylogenetic analyses, which show differences in gene preservation between phylogroups. A phylogenetic tree constructed with 2,613 single copy core genes along with a matrix of phylogenetic profiles is used to confirm that the 14 phylogroups change at different rates of gene gain/loss/duplication. The methodology used in this work is able to identify previously uncharacterized phylogroups in E. coli species. Some of these new phylogroups harbor clonal strains that have undergone a process of genomic adaptation to the acquisition of new genomic elements related to virulence or antibiotic resistance. This is, to our knowledge, the largest E. coli genome dataset analyzed to date and provides valuable insights into the population structure of the species.

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