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July 7, 2019

Evolutionary genomics of the cold-adapted diatom Fragilariopsis cylindrus.

The Southern Ocean houses a diverse and productive community of organisms. Unicellular eukaryotic diatoms are the main primary producers in this environment, where photosynthesis is limited by low concentrations of dissolved iron and large seasonal fluctuations in light, temperature and the extent of sea ice. How diatoms have adapted to this extreme environment is largely unknown. Here we present insights into the genome evolution of a cold-adapted diatom from the Southern Ocean, Fragilariopsis cylindrus, based on a comparison with temperate diatoms. We find that approximately 24.7 per cent of the diploid F. cylindrus genome consists of genetic loci with alleles that are highly divergent (15.1 megabases of the total genome size of 61.1 megabases). These divergent alleles were differentially expressed across environmental conditions, including darkness, low iron, freezing, elevated temperature and increased CO2. Alleles with the largest ratio of non-synonymous to synonymous nucleotide substitutions also show the most pronounced condition-dependent expression, suggesting a correlation between diversifying selection and allelic differentiation. Divergent alleles may be involved in adaptation to environmental fluctuations in the Southern Ocean.

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July 7, 2019

Identification of mutant genes and introgressed tiger salamander DNA in the laboratory axolotl, Ambystoma mexicanum.

The molecular genetic toolkit of the Mexican axolotl, a classic model organism, has matured to the point where it is now possible to identify genes for mutant phenotypes. We used a positional cloning-candidate gene approach to identify molecular bases for two historic axolotl pigment phenotypes: white and albino. White (d/d) mutants have defects in pigment cell morphogenesis and differentiation, whereas albino (a/a) mutants lack melanin. We identified in white mutants a transcriptional defect in endothelin 3 (edn3), encoding a peptide factor that promotes pigment cell migration and differentiation in other vertebrates. Transgenic restoration of Edn3 expression rescued the homozygous white mutant phenotype. We mapped the albino locus to tyrosinase (tyr) and identified polymorphisms shared between the albino allele (tyr (a) ) and tyr alleles in a Minnesota population of tiger salamanders from which the albino trait was introgressed. tyr (a) has a 142?bp deletion and similar engineered alleles recapitulated the albino phenotype. Finally, we show that historical introgression of tyr (a) significantly altered genomic composition of the laboratory axolotl, yielding a distinct, hybrid strain of ambystomatid salamander. Our results demonstrate the feasibility of identifying genes for traits in the laboratory Mexican axolotl.

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July 7, 2019

The evolution and population diversity of human-specific segmental duplications

Segmental duplications contribute to human evolution, adaptation and genomic instability but are often poorly characterized. We investigate the evolution, genetic variation and coding potential of human-specific segmental duplications (HSDs). We identify 218 HSDs based on analysis of 322 deeply sequenced archaic and contemporary hominid genomes. We sequence 550 human and nonhuman primate genomic clones to reconstruct the evolution of the largest, most complex regions with protein-coding potential (N?=?80 genes from 33 gene families). We show that HSDs are non-randomly organized, associate preferentially with ancestral ape duplications termed ‘core duplicons’ and evolved primarily in an interspersed inverted orientation. In addition to Homo sapiens-specific gene expansions (such as TCAF1/TCAF2), we highlight ten gene families (for example, ARHGAP11B and SRGAP2C) where copy number never returns to the ancestral state, there is evidence of mRNA splicing and no common gene-disruptive mutations are observed in the general population. Such duplicates are candidates for the evolution of human-specific adaptive traits.

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July 7, 2019

Epigenetic origin of evolutionary novel centromeres.

Most evolutionary new centromeres (ENC) are composed of large arrays of satellite DNA and surrounded by segmental duplications. However, the hypothesis is that ENCs are seeded in an anonymous sequence and only over time have acquired the complexity of “normal” centromeres. Up to now evidence to test this hypothesis was lacking. We recently discovered that the well-known polymorphism of orangutan chromosome 12 was due to the presence of an ENC. We sequenced the genome of an orangutan homozygous for the ENC, and we focused our analysis on the comparison of the ENC domain with respect to its wild type counterpart. No significant variations were found. This finding is the first clear evidence that ENC seedings are epigenetic in nature. The compaction of the ENC domain was found significantly higher than the corresponding WT region and, interestingly, the expression of the only gene embedded in the region was significantly repressed.

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July 7, 2019

Complete genome sequence of Thermus brockianus GE-1 reveals key enzymes of xylan/xylose metabolism.

Thermus brockianus strain GE-1 is a thermophilic, Gram-negative, rod-shaped and non-motile bacterium that was isolated from the Geysir geothermal area, Iceland. Like other thermophiles, Thermus species are often used as model organisms to understand the mechanism of action of extremozymes, especially focusing on their heat-activity and thermostability. Genome-specific features of T. brockianus GE-1 and their properties further help to explain processes of the adaption of extremophiles at elevated temperatures. Here we analyze the first whole genome sequence of T. brockianus strain GE-1. Insights of the genome sequence and the methodologies that were applied during de novo assembly and annotation are given in detail. The finished genome shows a phred quality value of QV50. The complete genome size is 2.38 Mb, comprising the chromosome (2,035,182 bp), the megaplasmid pTB1 (342,792 bp) and the smaller plasmid pTB2 (10,299 bp). Gene prediction revealed 2,511 genes in total, including 2,458 protein-encoding genes, 53 RNA and 66 pseudo genes. A unique genomic region on megaplasmid pTB1 was identified encoding key enzymes for xylan depolymerization and xylose metabolism. This is in agreement with the growth experiments in which xylan is utilized as sole source of carbon. Accordingly, we identified sequences encoding the xylanase Xyn10, an endoglucanase, the membrane ABC sugar transporter XylH, the xylose-binding protein XylF, the xylose isomerase XylA catalyzing the first step of xylose metabolism and the xylulokinase XylB, responsible for the second step of xylose metabolism. Our data indicate that an ancestor of T. brockianus obtained the ability to use xylose as alternative carbon source by horizontal gene transfer.

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July 7, 2019

Neurotrophin biology at NGF 2016: From fundamental science to clinical applications.

In 1986, members of the growing neurotrophin community came together to honor the scientific contributions (and 77th birth- day) of Dr. Rita Levi-Montalcini. The celebration took the form of a conference dedicated to the field birthed by Dr. Levi-Montalcini’s discovery of nerve growth factor (NGF), for which she shared the Nobel Prize later that year with Stanley Cohen. The meeting proved to be a great success, and eventually became an ongoing series. The NGF 2016 meeting, held at the beautiful Asilomar conference cen- ter in Monterey, California, was the 13th meeting in this series, and marked the 30th anniversary of the original meeting. A diverse col- lection of investigators, representing academia and industry across 4 continents, gathered to celebrate the past 30 years, discuss the current state of the art, and share in the excitement of envisioning the next 30 years of neurotrophic factor research and applications.

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July 7, 2019

Evaluation of GRCh38 and de novo haploid genome assemblies demonstrates the enduring quality of the reference assembly.

The human reference genome assembly plays a central role in nearly all aspects of today’s basic and clinical research. GRCh38 is the first coordinate-changing assembly update since 2009; it reflects the resolution of roughly 1000 issues and encompasses modifications ranging from thousands of single base changes to megabase-scale path reorganizations, gap closures, and localization of previously orphaned sequences. We developed a new approach to sequence generation for targeted base updates and used data from new genome mapping technologies and single haplotype resources to identify and resolve larger assembly issues. For the first time, the reference assembly contains sequence-based representations for the centromeres. We also expanded the number of alternate loci to create a reference that provides a more robust representation of human population variation. We demonstrate that the updates render the reference an improved annotation substrate, alter read alignments in unchanged regions, and impact variant interpretation at clinically relevant loci. We additionally evaluated a collection of new de novo long-read haploid assemblies and conclude that although the new assemblies compare favorably to the reference with respect to continuity, error rate, and gene completeness, the reference still provides the best representation for complex genomic regions and coding sequences. We assert that the collected updates in GRCh38 make the newer assembly a more robust substrate for comprehensive analyses that will promote our understanding of human biology and advance our efforts to improve health. © 2017 Schneider et al.; Published by Cold Spring Harbor Laboratory Press.

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July 7, 2019

HySA: a Hybrid Structural variant Assembly approach using next-generation and single-molecule sequencing technologies.

Achieving complete, accurate, and cost-effective assembly of human genomes is of great importance for realizing the promise of precision medicine. The abundance of repeats and genetic variations in human genomes and the limitations of existing sequencing technologies call for the development of novel assembly methods that can leverage the complementary strengths of multiple technologies. We propose a Hybrid Structural variant Assembly (HySA) approach that integrates sequencing reads from next-generation sequencing and single-molecule sequencing technologies to accurately assemble and detect structural variants (SVs) in human genomes. By identifying homologous SV-containing reads from different technologies through a bipartite-graph-based clustering algorithm, our approach turns a whole genome assembly problem into a set of independent SV assembly problems, each of which can be effectively solved to enhance the assembly of structurally altered regions in human genomes. We used data generated from a haploid hydatidiform mole genome (CHM1) and a diploid human genome (NA12878) to test our approach. The result showed that, compared with existing methods, our approach had a low false discovery rate and substantially improved the detection of many types of SVs, particularly novel large insertions, small indels (10-50 bp), and short tandem repeat expansions and contractions. Our work highlights the strengths and limitations of current approaches and provides an effective solution for extending the power of existing sequencing technologies for SV discovery.© 2017 Fan et al.; Published by Cold Spring Harbor Laboratory Press.

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July 7, 2019

Phenotypic and genomic comparison of Mycobacterium aurum and surrogate model species to Mycobacterium tuberculosis: implications for drug discovery.

Tuberculosis (TB) is caused by Mycobacterium tuberculosis and represents one of the major challenges facing drug discovery initiatives worldwide. The considerable rise in bacterial drug resistance in recent years has led to the need of new drugs and drug regimens. Model systems are regularly used to speed-up the drug discovery process and circumvent biosafety issues associated with manipulating M. tuberculosis. These include the use of strains such as Mycobacterium smegmatis and Mycobacterium marinum that can be handled in biosafety level 2 facilities, making high-throughput screening feasible. However, each of these model species have their own limitations.We report and describe the first complete genome sequence of Mycobacterium aurum ATCC23366, an environmental mycobacterium that can also grow in the gut of humans and animals as part of the microbiota. This species shows a comparable resistance profile to that of M. tuberculosis for several anti-TB drugs. The aims of this study were to (i) determine the drug resistance profile of a recently proposed model species, Mycobacterium aurum, strain ATCC23366, for anti-TB drug discovery as well as Mycobacterium smegmatis and Mycobacterium marinum (ii) sequence and annotate the complete genome sequence of this species obtained using Pacific Bioscience technology (iii) perform comparative genomics analyses of the various surrogate strains with M. tuberculosis (iv) discuss how the choice of the surrogate model used for drug screening can affect the drug discovery process.We describe the complete genome sequence of M. aurum, a surrogate model for anti-tuberculosis drug discovery. Most of the genes already reported to be associated with drug resistance are shared between all the surrogate strains and M. tuberculosis. We consider that M. aurum might be used in high-throughput screening for tuberculosis drug discovery. We also highly recommend the use of different model species during the drug discovery screening process.

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July 7, 2019

Complete genome sequence of Leuconostoc suionicum DSM 20241(T) provides insights into its functional and metabolic features.

The genome of Leuconostoc suionicum DSM 20241(T) (=ATCC 9135(T) = LMG 8159(T) = NCIMB 6992(T)) was completely sequenced and its fermentative metabolic pathways were reconstructed to investigate the fermentative properties and metabolites of strain DSM 20241(T) during fermentation. The genome of L. suionicum DSM 20241(T) consists of a circular chromosome (2026.8 Kb) and a circular plasmid (21.9 Kb) with 37.58% G + C content, encoding 997 proteins, 12 rRNAs, and 72 tRNAs. Analysis of the metabolic pathways of L. suionicum DSM 20241(T) revealed that strain DSM 20241(T) performs heterolactic acid fermentation and can metabolize diverse organic compounds including glucose, fructose, galactose, cellobiose, mannose, sucrose, trehalose, arbutin, salcin, xylose, arabinose and ribose.

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July 7, 2019

Phylogenomic analysis supports multiple instances of polyphyly in the oomycete peronosporalean lineage.

The study of biological diversification of oomycetes has been a difficult task for more than a century. Pioneer researchers used morphological characters to describe this heterogeneous group, and physiological and genetic tools expanded knowledge of these microorganisms. However, research on oomycete diversification is limited by conflicting phylogenies. Using whole genomic data from 17 oomycete taxa, we obtained a dataset of 277 core orthologous genes shared among these genomes. Analyses of this dataset resulted in highly congruent and strongly supported estimates of oomycete phylogeny when we used concatenated maximum likelihood and coalescent-based methods; the one important exception was the position of Albugo. Our results supported the position of Phytopythium vexans (formerly in Pythium clade K) as a sister clade to the Phytophthora-Hyaloperonospora clade. The remaining clades comprising Pythium sensu lato formed two monophyletic groups. One group was composed of three taxa that correspond to Pythium clades A, B and C, and the other group contained taxa representing clades F, G and I, in agreement with previous Pythium phylogenies. However, the group containing Pythium clades F, G and I was placed as sister to the Phytophthora-Hyaloperonospora-Phytopythium clade, thus confirming the lack of monophyly of Pythium sensu lato. Multispecies coalescent methods revealed that the white blister rust, Albugo laibachii, could not be placed with a high degree of confidence. Our analyses show that genomic data can resolve the oomycete phylogeny and provide a phylogenetic framework to study the evolution of oomycete lifestyles. Copyright © 2017 Elsevier Inc. All rights reserved.

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July 7, 2019

The dynamic three-dimensional organization of the diploid yeast genome.

The budding yeast Saccharomyces cerevisiae is a long-standing model for the three-dimensional organization of eukaryotic genomes. However, even in this well-studied model, it is unclear how homolog pairing in diploids or environmental conditions influence overall genome organization. Here, we performed high-throughput chromosome conformation capture on diverged Saccharomyces hybrid diploids to obtain the first global view of chromosome conformation in diploid yeasts. After controlling for the Rabl-like orientation using a polymer model, we observe significant homolog proximity that increases in saturated culture conditions. Surprisingly, we observe a localized increase in homologous interactions between the HAS1-TDA1 alleles specifically under galactose induction and saturated growth. This pairing is accompanied by relocalization to the nuclear periphery and requires Nup2, suggesting a role for nuclear pore complexes. Together, these results reveal that the diploid yeast genome has a dynamic and complex 3D organization.

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July 7, 2019

Genomic analysis of factors associated with low prevalence of antibiotic resistance in extraintestinal pathogenic Escherichia coli sequence type 95 strains.

Extraintestinal pathogenic Escherichia coli (ExPEC) strains belonging to multilocus sequence type 95 (ST95) are globally distributed and a common cause of infections in humans and domestic fowl. ST95 isolates generally show a lower prevalence of acquired antimicrobial resistance than other pandemic ExPEC lineages. We took a genomic approach to identify factors that may underlie reduced resistance. We fully assembled genomes for four ST95 isolates representing the four major fimH-based lineages within ST95 and also analyzed draft-level genomes from another 82 ST95 isolates, largely from the western United States. The fully assembled genomes of antibiotic-resistant isolates carried resistance genes exclusively on large (>90-kb) IncFIB/IncFII plasmids. These replicons were common in the draft genomes as well, particularly in antibiotic-resistant isolates, but we also observed multiple instances of a smaller (8.3-kb) ampicillin resistance plasmid that had been previously identified in Salmonella enterica. Among ST95 isolates, pansusceptibility to antibiotics was significantly associated with the fimH6 lineage and the presence of homologs of the previously identified 114-kb IncFIB/IncFII plasmid pUTI89, both of which were also associated with reduced carriage of other plasmids. Potential mechanistic explanations for lineage- and plasmid-specific effects on the prevalence of antibiotic resistance within the ST95 group are discussed. IMPORTANCE Antibiotic resistance in bacterial pathogens is a major public health concern. This work was motivated by the observation that only a small proportion of ST95 isolates, a major pandemic lineage of extraintestinal pathogenic E. coli, have acquired antibiotic resistance, in contrast to many other pandemic lineages. Understanding bacterial genetic factors that may prevent acquisition of resistance could contribute to the development of new biological, medical, or public health strategies to reduce antibiotic-resistant infections.

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July 7, 2019

Resolving multicopy duplications de novo using polyploid phasing

While the rise of single-molecule sequencing systems has enabled an unprecedented rise in the ability to assemble complex regions of the genome, long segmental duplications in the genome still remain a challenging frontier in assembly. Segmental duplications are at the same time both gene rich and prone to large structural rearrangements, making the resolution of their sequences important in medical and evolutionary studies. Duplicated sequences that are collapsed in mammalian de novo assemblies are rarely identical; after a sequence is duplicated, it begins to acquire paralog-specific variants. In this paper, we study the problem of resolving the variations in multicopy, long segmental duplications by developing and utilizing algorithms for polyploid phasing. We develop two algorithms: the first one is targeted at maximizing the likelihood of observing the reads given the underlying haplotypes using discrete matrix completion. The second algorithm is based on correlation clustering and exploits an assumption, which is often satisfied in these duplications, that each paralog has a sizable number of paralog-specific variants. We develop a detailed simulation methodology and demonstrate the superior performance of the proposed algorithms on an array of simulated datasets. We measure the likelihood score as well as reconstruction accuracy, i.e., what fraction of the reads are clustered correctly. In both the performance metrics, we find that our algorithms dominate existing algorithms on more than 93% of the datasets. While the discrete matrix completion performs better on likelihood score, the correlation-clustering algorithm performs better on reconstruction accuracy due to the stronger regularization inherent in the algorithm. We also show that our correlation-clustering algorithm can reconstruct on average 7.0 haplotypes in 10-copy duplication datasets whereas existing algorithms reconstruct less than one copy on average.

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July 7, 2019

Complete genome analysis of Thermus parvatiensis and comparative genomics of Thermus spp. provide insights into genetic variability and evolution of natural competence as strategic survival attributes.

Thermophilic environments represent an interesting niche. Among thermophiles, the genus Thermus is among the most studied genera. In this study, we have sequenced the genome of Thermus parvatiensis strain RL, a thermophile isolated from Himalayan hot water springs (temperature >96°C) using PacBio RSII SMRT technique. The small genome (2.01 Mbp) comprises a chromosome (1.87 Mbp) and a plasmid (143 Kbp), designated in this study as pTP143. Annotation revealed a high number of repair genes, a squeezed genome but containing highly plastic plasmid with transposases, integrases, mobile elements and hypothetical proteins (44%). We performed a comparative genomic study of the group Thermus with an aim of analysing the phylogenetic relatedness as well as niche specific attributes prevalent among the group. We compared the reference genome RL with 16 Thermus genomes to assess their phylogenetic relationships based on 16S rRNA gene sequences, average nucleotide identity (ANI), conserved marker genes (31 and 400), pan genome and tetranucleotide frequency. The core genome of the analyzed genomes contained 1,177 core genes and many singleton genes were detected in individual genomes, reflecting a conserved core but adaptive pan repertoire. We demonstrated the presence of metagenomic islands (chromosome:5, plasmid:5) by recruiting raw metagenomic data (from the same niche) against the genomic replicons of T. parvatiensis. We also dissected the CRISPR loci wide all genomes and found widespread presence of this system across Thermus genomes. Additionally, we performed a comparative analysis of competence loci wide Thermus genomes and found evidence for recent horizontal acquisition of the locus and continued dispersal among members reflecting that natural competence is a beneficial survival trait among Thermus members and its acquisition depicts unending evolution in order to accomplish optimal fitness.

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