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July 7, 2019

Whole-genome de novo sequencing of wood rot fungus Fomitopsis palustris (ATCC62978) with both a cellulolytic and ligninolytic enzyme system.

Fomitopsis palustris is a model brown rot fungus causing destructive wood decay based on the cellulase system. Endoglucanase secreted by F. palustris hydrolyzes cellulose in both the crystalline and amorphous form. In this study, whole-genome sequencing was conducted to identify genes related to F. palustris cellulose degradation and their functions. We determined the 43-Mb complete draft genome of F. palustris (ATCC 62978), comprising 14,592 predicted gene models. Gene annotation provided crucial information about the location and function of protein-encoding genes. Three types of endoglucanases were expressed: endo-1,3-beta-glucanase, endo-1,4-beta-d-glucanase, and endoglucanase. In addition, various ligninolytic enzymes such as laccase, aromatic compound dioxygenase, and aryl alcohol dehydrogenase were expressed in F. palustris (ATCC 62978). Colony polymerase chain reaction (PCR) indicated that the endo-1,4-beta-d-glucanase gene comprises 732bp. Optimization of the expression conditions of endoglucanase by real-time PCR revealed that endoglucanase was highly expressed after 7days in all conditions, which was secreted during the secondary metabolism. Studies for large-scale cellulase production from this fungus and investigation of its ligninolytic system will promote its extensive use in various applications. The genomic information determined herein provides a basis for molecular genetics studies to understand the genome functions of F. palustris (ATCC 62978). Copyright © 2017 Elsevier B.V. All rights reserved.

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July 7, 2019

Chromosome-level genome assembly and transcriptome of the green alga Chromochloris zofingiensis illuminates astaxanthin production.

Microalgae have potential to help meet energy and food demands without exacerbating environmental problems. There is interest in the unicellular green alga Chromochloris zofingiensis, because it produces lipids for biofuels and a highly valuable carotenoid nutraceutical, astaxanthin. To advance understanding of its biology and facilitate commercial development, we present a C. zofingiensis chromosome-level nuclear genome, organelle genomes, and transcriptome from diverse growth conditions. The assembly, derived from a combination of short- and long-read sequencing in conjunction with optical mapping, revealed a compact genome of ~58 Mbp distributed over 19 chromosomes containing 15,274 predicted protein-coding genes. The genome has uniform gene density over chromosomes, low repetitive sequence content (~6%), and a high fraction of protein-coding sequence (~39%) with relatively long coding exons and few coding introns. Functional annotation of gene models identified orthologous families for the majority (~73%) of genes. Synteny analysis uncovered localized but scrambled blocks of genes in putative orthologous relationships with other green algae. Two genes encoding beta-ketolase (BKT), the key enzyme synthesizing astaxanthin, were found in the genome, and both were up-regulated by high light. Isolation and molecular analysis of astaxanthin-deficient mutants showed that BKT1 is required for the production of astaxanthin. Moreover, the transcriptome under high light exposure revealed candidate genes that could be involved in critical yet missing steps of astaxanthin biosynthesis, including ABC transporters, cytochrome P450 enzymes, and an acyltransferase. The high-quality genome and transcriptome provide insight into the green algal lineage and carotenoid production.

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July 7, 2019

Sequencing a piece of history: complete genome sequence of the original Escherichia coli strain.

In 1885, Theodor Escherich first described the Bacillus coli commune, which was subsequently renamed Escherichia coli. We report the complete genome sequence of this original strain (NCTC 86). The 5?144?392?bp circular chromosome encodes the genes for 4805 proteins, which include antigens, virulence factors, antimicrobial-resistance factors and secretion systems, of a commensal organism from the pre-antibiotic era. It is located in the E. coli A subgroup and is closely related to E. coli K-12 MG1655. E. coli strain NCTC 86 and the non-pathogenic K-12, C, B and HS strains share a common backbone that is largely co-linear. The exception is a large 2?803?932?bp inversion that spans the replication terminus from gmhB to clpB. Comparison with E. coli K-12 reveals 41 regions of difference (577?351?bp) distributed across the chromosome. For example, and contrary to current dogma, E. coli NCTC 86 includes a nine gene sil locus that encodes a silver-resistance efflux pump acquired before the current widespread use of silver nanoparticles as an antibacterial agent, possibly resulting from the widespread use of silver utensils and currency in Germany in the 1800s. In summary, phylogenetic comparisons with other E. coli strains confirmed that the original strain isolated by Escherich is most closely related to the non-pathogenic commensal strains. It is more distant from the root than the pathogenic organisms E. coli 042 and O157?:?H7; therefore, it is not an ancestral state for the species.

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July 7, 2019

Metabolic diversity of the emerging pathogenic lineages of Klebsiella pneumoniae.

Multidrug resistant and hypervirulent clones of Klebsiella pneumoniae are emerging pathogens. To understand the association between genotypic and phenotypic diversity in this process, we combined genomic, phylogenomic and phenotypic analysis of a diverse set of K. pneumoniae and closely related species. These species were able to use an unusually large panel of metabolic substrates for growth, many of which were shared between all strains. We analysed the substrates used by only a fraction of the strains, identified some of their genetic basis, and found that many could not be explained by the phylogeny of the strains. Puzzlingly, few traits were associated with the ecological origin of the strains. One noticeable exception was the ability to use D-arabinose, which was much more frequent in hypervirulent strains. The broad carbon and nitrogen core metabolism of K. pneumoniae might contribute to its ability to thrive in diverse environments. Accordingly, even the hypervirulent and multidrug resistant clones have the metabolic signature of ubiquitous bacteria. The apparent few metabolic differences between hypervirulent, multi-resistant and environmental strains may favour the emergence of dual-risk strains that combine resistance and hypervirulence.© 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

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July 7, 2019

Genetic control of plasticity of oil yield for combined abiotic stresses using a joint approach of crop modelling and genome-wide association.

Understanding the genetic basis of phenotypic plasticity is crucial for predicting and managing climate change effects on wild plants and crops. Here, we combined crop modelling and quantitative genetics to study the genetic control of oil yield plasticity for multiple abiotic stresses in sunflower. First, we developed stress indicators to characterize 14 environments for three abiotic stresses (cold, drought and nitrogen) using the SUNFLO crop model and phenotypic variations of three commercial varieties. The computed plant stress indicators better explain yield variation than descriptors at the climatic or crop levels. In those environments, we observed oil yield of 317 sunflower hybrids and regressed it with three selected stress indicators. The slopes of cold stress norm reaction were used as plasticity phenotypes in the following genome-wide association study. Among the 65 534 tested Single Nucleotide Polymorphisms (SNPs), we identified nine quantitative trait loci controlling oil yield plasticity to cold stress. Associated single nucleotide polymorphisms are localized in genes previously shown to be involved in cold stress responses: oligopeptide transporters, lipid transfer protein, cystatin, alternative oxidase or root development. This novel approach opens new perspectives to identify genomic regions involved in genotype-by-environment interaction of a complex traits to multiple stresses in realistic natural or agronomical conditions.© 2017 John Wiley & Sons Ltd.

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July 7, 2019

A small secreted protein in Zymoseptoria tritici is responsible for avirulence on wheat cultivars carrying the Stb6 resistance gene.

Zymoseptoria tritici is the causal agent of Septoria tritici blotch, a major pathogen of wheat globally and the most damaging pathogen of wheat in Europe. A gene-for-gene (GFG) interaction between Z. tritici and wheat cultivars carrying the Stb6 resistance gene has been postulated for many years, but the genes have not been identified. We identified AvrStb6 by combining quantitative trait locus mapping in a cross between two Swiss strains with a genome-wide association study using a natural population of c. 100 strains from France. We functionally validated AvrStb6 using ectopic transformations. AvrStb6 encodes a small, cysteine-rich, secreted protein that produces an avirulence phenotype on wheat cultivars carrying the Stb6 resistance gene. We found 16 nonsynonymous single nucleotide polymorphisms among the tested strains, indicating that AvrStb6 is evolving very rapidly. AvrStb6 is located in a highly polymorphic subtelomeric region and is surrounded by transposable elements, which may facilitate its rapid evolution to overcome Stb6 resistance. AvrStb6 is the first avirulence gene to be functionally validated in Z. tritici, contributing to our understanding of avirulence in apoplastic pathogens and the mechanisms underlying GFG interactions between Z. tritici and wheat. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

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July 7, 2019

Strategies for optimizing BioNano and Dovetail explored through a second reference quality assembly for the legume model, Medicago truncatula.

Third generation sequencing technologies, with sequencing reads in the tens- of kilo-bases, facilitate genome assembly by spanning ambiguous regions and improving continuity. This has been critical for plant genomes, which are difficult to assemble due to high repeat content, gene family expansions, segmental and tandem duplications, and polyploidy. Recently, high-throughput mapping and scaffolding strategies have further improved continuity. Together, these long-range technologies enable quality draft assemblies of complex genomes in a cost-effective and timely manner.Here, we present high quality genome assemblies of the model legume plant, Medicago truncatula (R108) using PacBio, Dovetail Chicago (hereafter, Dovetail) and BioNano technologies. To test these technologies for plant genome assembly, we generated five assemblies using all possible combinations and ordering of these three technologies in the R108 assembly. While the BioNano and Dovetail joins overlapped, they also showed complementary gains in continuity and join numbers. Both technologies spanned repetitive regions that PacBio alone was unable to bridge. Combining technologies, particularly Dovetail followed by BioNano, resulted in notable improvements compared to Dovetail or BioNano alone. A combination of PacBio, Dovetail, and BioNano was used to generate a high quality draft assembly of R108, a M. truncatula accession widely used in studies of functional genomics. As a test for the usefulness of the resulting genome sequence, the new R108 assembly was used to pinpoint breakpoints and characterize flanking sequence of a previously identified translocation between chromosomes 4 and 8, identifying more than 22.7 Mb of novel sequence not present in the earlier A17 reference assembly.Adding Dovetail followed by BioNano data yielded complementary improvements in continuity over the original PacBio assembly. This strategy proved efficient and cost-effective for developing a quality draft assembly compared to traditional reference assemblies.

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July 7, 2019

Draft genome sequence of Grammothele lineata SDL-CO-2015-1, a jute endophyte with a potential for paclitaxel biosynthesis.

Grammothele lineata strain SDL-CO-2015-1, a basidiomycete fungus, was identified as an endophyte from a jute species, Corchorus olitorius var. 2015, and found to produce paclitaxel, a diterpenic polyoxygenated pseudoalkaloid with antitumor activity. Here, we report the draft genome sequence (42.8 Mb with 9,395 genes) of this strain. Copyright © 2017 Das et al.

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July 7, 2019

Whole-genome sequence and variant analysis of W303, a widely-used strain of Saccharomyces cerevisiae.

The yeast Saccharomyces cerevisiae has emerged as a superior model organism. Selection of distinct laboratory strains of S. cerevisiae with unique phenotypic properties, such as superior mating or sporulation efficiencies, has facilitated advancements in research. W303 is one such laboratory strain that is closely related to the first completely sequenced yeast strain, S288C. In this work, we provide a high-quality, annotated genome sequence for W303 for utilization in comparative analyses and genome-wide studies. Approximately 9500 variations exist between S288C and W303, affecting the protein sequences of ~700 genes. A listing of the polymorphisms and divergent genes is provided for researchers interested in identifying the genetic basis for phenotypic differences between W303 and S288C. Several divergent functional gene families were identified, including flocculation and sporulation genes, likely representing selection for desirable laboratory phenotypes. Interestingly, remnants of ancestor wine strains were found on several chromosomes. Finally, as a test of the utility of the high-quality reference genome, variant mapping revealed more accurate identification of accumulated mutations in passaged mismatch repair-defective strains. Copyright © 2017 Matheson et al.

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July 7, 2019

Novel pelagic iron-oxidizing Zetaproteobacteria from the Chesapeake Bay oxic-anoxic transition zone.

Chemolithotrophic iron-oxidizing bacteria (FeOB) could theoretically inhabit any environment where Fe(II) and O2 (or nitrate) coexist. Until recently, marine Fe-oxidizing Zetaproteobacteria had primarily been observed in benthic and subsurface settings, but not redox-stratified water columns. This may be due to the challenges that a pelagic lifestyle would pose for Zetaproteobacteria, given low Fe(II) concentrations in modern marine waters and the possibility that Fe oxyhydroxide biominerals could cause cells to sink. However, we recently cultivated Zetaproteobacteria from the Chesapeake Bay oxic-anoxic transition zone, suggesting that they can survive and contribute to biogeochemical cycling in a stratified estuary. Here we describe the isolation, characterization, and genomes of two new species, Mariprofundus aestuarium CP-5 and Mariprofundus ferrinatatus CP-8, which are the first Zetaproteobacteria isolates from a pelagic environment. We looked for adaptations enabling strains CP-5 and CP-8 to overcome the challenges of living in a low Fe redoxcline with frequent O2 fluctuations due to tidal mixing. We found that the CP strains produce distinctive dreadlock-like Fe oxyhydroxide structures that are easily shed, which would help cells maintain suspension in the water column. These oxides are by-products of Fe(II) oxidation, likely catalyzed by the putative Fe(II) oxidase encoded by the cyc2 gene, present in both CP-5 and CP-8 genomes; the consistent presence of cyc2 in all microaerophilic FeOB and other FeOB genomes supports its putative role in Fe(II) oxidation. The CP strains also have two gene clusters associated with biofilm formation (Wsp system and the Widespread Colonization Island) that are absent or rare in other Zetaproteobacteria. We propose that biofilm formation enables the CP strains to attach to FeS particles and form flocs, an advantageous strategy for scavenging Fe(II) and developing low [O2] microenvironments within more oxygenated waters. However, the CP strains appear to be adapted to somewhat higher concentrations of O2, as indicated by the presence of genes encoding aa3-type cytochrome c oxidases, but not the cbb3-type found in all other Zetaproteobacteria isolate genomes. Overall, our results reveal adaptations for life in a physically dynamic, low Fe(II) water column, suggesting that niche-specific strategies can enable Zetaproteobacteria to live in any environment with Fe(II).

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July 7, 2019

Improved annotation of the insect vector of citrus greening disease: biocuration by a diverse genomics community.

The Asian citrus psyllid (Diaphorina citri Kuwayama) is the insect vector of the bacterium Candidatus Liberibacter asiaticus (CLas), the pathogen associated with citrus Huanglongbing (HLB, citrus greening). HLB threatens citrus production worldwide. Suppression or reduction of the insect vector using chemical insecticides has been the primary method to inhibit the spread of citrus greening disease. Accurate structural and functional annotation of the Asian citrus psyllid genome, as well as a clear understanding of the interactions between the insect and CLas, are required for development of new molecular-based HLB control methods. A draft assembly of the D. citri genome has been generated and annotated with automated pipelines. However, knowledge transfer from well-curated reference genomes such as that of Drosophila melanogaster to newly sequenced ones is challenging due to the complexity and diversity of insect genomes. To identify and improve gene models as potential targets for pest control, we manually curated several gene families with a focus on genes that have key functional roles in D. citri biology and CLas interactions. This community effort produced 530 manually curated gene models across developmental, physiological, RNAi regulatory and immunity-related pathways. As previously shown in the pea aphid, RNAi machinery genes putatively involved in the microRNA pathway have been specifically duplicated. A comprehensive transcriptome enabled us to identify a number of gene families that are either missing or misassembled in the draft genome. In order to develop biocuration as a training experience, we included undergraduate and graduate students from multiple institutions, as well as experienced annotators from the insect genomics research community. The resulting gene set (OGS v1.0) combines both automatically predicted and manually curated gene models.

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July 7, 2019

Centrochromatin of fungi.

The centromere is an essential chromosomal locus that dictates the nucleation point for assembly of the kinetochore and subsequent attachment of spindle microtubules during chromosome segregation. Research over the last decades demonstrated that centromeres are defined by a combination of genetic and epigenetic factors. Recent work showed that centromeres are quite diverse and flexible and that many types of centromere sequences and centromeric chromatin (“centrochromatin”) have evolved. The kingdom of the fungi serves as an outstanding example of centromere plasticity, including organisms with centromeres as diverse as 0.15-300 kb in length, and with different types of chromatin states for most species examined thus far. Some of the species in the less familiar taxa provide excellent opportunities to help us better understand centromere biology in all eukaryotes, which may improve treatment options against fungal infection, and biotechnologies based on fungi. This review summarizes the current knowledge of fungal centromeres and centrochromatin, including an outlook for future research.

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July 7, 2019

Single-molecule sequencing and Hi-C-based proximity-guided assembly of amaranth (Amaranthus hypochondriacus) chromosomes provide insights into genome evolution.

Amaranth (Amaranthus hypochondriacus) was a food staple among the ancient civilizations of Central and South America that has recently received increased attention due to the high nutritional value of the seeds, with the potential to help alleviate malnutrition and food security concerns, particularly in arid and semiarid regions of the developing world. Here, we present a reference-quality assembly of the amaranth genome which will assist the agronomic development of the species.Utilizing single-molecule, real-time sequencing (Pacific Biosciences) and chromatin interaction mapping (Hi-C) to close assembly gaps and scaffold contigs, respectively, we improved our previously reported Illumina-based assembly to produce a chromosome-scale assembly with a scaffold N50 of 24.4 Mb. The 16 largest scaffolds contain 98% of the assembly and likely represent the haploid chromosomes (n?=?16). To demonstrate the accuracy and utility of this approach, we produced physical and genetic maps and identified candidate genes for the betalain pigmentation pathway. The chromosome-scale assembly facilitated a genome-wide syntenic comparison of amaranth with other Amaranthaceae species, revealing chromosome loss and fusion events in amaranth that explain the reduction from the ancestral haploid chromosome number (n?=?18) for a tetraploid member of the Amaranthaceae.The assembly method reported here minimizes cost by relying primarily on short-read technology and is one of the first reported uses of in vivo Hi-C for assembly of a plant genome. Our analyses implicate chromosome loss and fusion as major evolutionary events in the 2n?=?32 amaranths and clearly establish the homoeologous relationship among most of the subgenome chromosomes, which will facilitate future investigations of intragenomic changes that occurred post polyploidization.

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July 7, 2019

Complete genome sequence of the fruiting myxobacterium Myxococcus macrosporus strain DSM 14697, generated by PacBio sequencing.

Members of the Myxococcales order initiate a developmental program in response to starvation that culminates in formation of spore-filled fruiting bodies. To investigate the genetic basis for fruiting body formation, we present the complete 8.9-Mb genome sequence of Myxococcus macrosporus strain DSM 14697, generated using the PacBio sequencing platform. Copyright © 2017 Treuner-Lange et al.

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July 7, 2019

Multiple hybrid de novo genome assembly of finger millet, an orphan allotetraploid crop.

Finger millet (Eleusine coracana (L.) Gaertn) is an important crop for food security because of its tolerance to drought, which is expected to be exacerbated by global climate changes. Nevertheless, it is often classified as an orphan/underutilized crop because of the paucity of scientific attention. Among several small millets, finger millet is considered as an excellent source of essential nutrient elements, such as iron and zinc; hence, it has potential as an alternate coarse cereal. However, high-quality genome sequence data of finger millet are currently not available. One of the major problems encountered in the genome assembly of this species was its polyploidy, which hampers genome assembly compared with a diploid genome. To overcome this problem, we sequenced its genome using diverse technologies with sufficient coverage and assembled it via a novel multiple hybrid assembly workflow that combines next-generation with single-molecule sequencing, followed by whole-genome optical mapping using the Bionano Irys® system. The total number of scaffolds was 1,897 with an N50 length?>2.6?Mb and detection of 96% of the universal single-copy orthologs. The majority of the homeologs were assembled separately. This indicates that the proposed workflow is applicable to the assembly of other allotetraploid genomes.© The Author 2017. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

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