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Tuesday, April 21, 2020

Reconstruction of the genomes of drug-resistant pathogens for outbreak investigation through metagenomic sequencing

Culture-independent methods that target genome fragments have shown promise in identifying certain pathogens, but the holy grail of comprehensive pathogen genome detection from microbiologically complex samples for subsequent forensic analyses remains a challenge. In the context of an investigation of a nosocomial outbreak, we used shotgun metagenomic sequencing of a human fecal sample and a neural network algorithm based on tetranucleotide frequency profiling to reconstruct microbial genomes and tested the same approach using rectal swabs from a second patient. The approach rapidly and readily detected the genome of Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae in the patient fecal specimen and…

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Tuesday, April 21, 2020

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…

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Tuesday, April 21, 2020

Complete Genome Sequence of Actinosynnema pretiosum X47, An Industrial Strain that Produces the Antibiotic Ansamitocin AP-3.

Ansamitocins are extraordinarily potent antitumor agents. Ansamitocin P-3 (AP-3), which is produced by Actinosynnema pretiosum, has been developed as a cytotoxic drug for breast cancer. Despite its importance, AP-3 is of limited applicability because of the low production yield. A. pretiosum strain X47 was developed from A. pretiosum ATCC 31565 by mutation breeding and shows a relatively high AP-3 yield. Here, we analyzed the A. pretiosum X47 genome, which is ~8.13 Mb in length with 6693 coding sequences, 58 tRNA genes, and 15 rRNA genes. The DNA sequence of the ansamitocin biosynthetic gene cluster is highly similar to that of the…

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Tuesday, April 21, 2020

Long-read sequence and assembly of segmental duplications.

We have developed a computational method based on polyploid phasing of long sequence reads to resolve collapsed regions of segmental duplications within genome assemblies. Segmental Duplication Assembler (SDA; https://github.com/mvollger/SDA ) constructs graphs in which paralogous sequence variants define the nodes and long-read sequences provide attraction and repulsion edges, enabling the partition and assembly of long reads corresponding to distinct paralogs. We apply it to single-molecule, real-time sequence data from three human genomes and recover 33-79 megabase pairs (Mb) of duplications in which approximately half of the loci are diverged (99.9%) and that the diverged sequence corresponds to copy-number-variable paralogs that…

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Tuesday, April 21, 2020

Comprehensive evaluation of non-hybrid genome assembly tools for third-generation PacBio long-read sequence data.

Long reads obtained from third-generation sequencing platforms can help overcome the long-standing challenge of the de novo assembly of sequences for the genomic analysis of non-model eukaryotic organisms. Numerous long-read-aided de novo assemblies have been published recently, which exhibited superior quality of the assembled genomes in comparison with those achieved using earlier second-generation sequencing technologies. Evaluating assemblies is important in guiding the appropriate choice for specific research needs. In this study, we evaluated 10 long-read assemblers using a variety of metrics on Pacific Biosciences (PacBio) data sets from different taxonomic categories with considerable differences in genome size. The results allowed…

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Tuesday, April 21, 2020

Tools and Strategies for Long-Read Sequencing and De Novo Assembly of Plant Genomes.

The commercial release of third-generation sequencing technologies (TGSTs), giving long and ultra-long sequencing reads, has stimulated the development of new tools for assembling highly contiguous genome sequences with unprecedented accuracy across complex repeat regions. We survey here a wide range of emerging sequencing platforms and analytical tools for de novo assembly, provide background information for each of their steps, and discuss the spectrum of available options. Our decision tree recommends workflows for the generation of a high-quality genome assembly when used in combination with the specific needs and resources of a project.Copyright © 2019 Elsevier Ltd. All rights reserved.

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Tuesday, April 21, 2020

Characterizing the major structural variant alleles of the human genome.

In order to provide a comprehensive resource for human structural variants (SVs), we generated long-read sequence data and analyzed SVs for fifteen human genomes. We sequence resolved 99,604 insertions, deletions, and inversions including 2,238 (1.6 Mbp) that are shared among all discovery genomes with an additional 13,053 (6.9 Mbp) present in the majority, indicating minor alleles or errors in the reference. Genotyping in 440 additional genomes confirms the most common SVs in unique euchromatin are now sequence resolved. We report a ninefold SV bias toward the last 5 Mbp of human chromosomes with nearly 55% of all VNTRs (variable number…

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Tuesday, April 21, 2020

Complete genome sequence of Streptomyces spongiicola HNM0071T, a marine sponge-associated actinomycete producing staurosporine and echinomycin

Streptomyes spongiicola HNM0071T is a novel marine sponge-associated actinomycete with potential to produce antitumor agents including staurosporine and echinomycin. Here, we present the complete genome sequence of S. spongiicola HNM0071, which consists of a linear chromosome of 7,180,417?bp, 5669 protein coding genes, 18 rRNA genes, and 66 tRNA genes. Twenty-seven putative secondary metabolite biosynthetic gene clusters were found in the genome. Among them, the staurosporine and echinomycin gene clusters have been described completely. The complete genome information presented here will enable us to investigate the biosynthetic mechanism of two well-known antitumor antibiotics and to discover novel secondary metabolites with potential…

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Tuesday, April 21, 2020

Potential KPC-2 carbapenemase reservoir of environmental Aeromonas hydrophila and Aeromonas caviae isolates from the effluent of an urban wastewater treatment plant in Japan.

Aeromonas hydrophila and Aeromonas caviae adapt to saline water environments and are the most predominant Aeromonas species isolated from estuaries. Here, we isolated antimicrobial-resistant (AMR) Aeromonas strains (A. hydrophila GSH8-2 and A. caviae GSH8M-1) carrying the carabapenemase blaKPC-2 gene from a wastewater treatment plant (WWTP) effluent in Tokyo Bay (Japan) and determined their complete genome sequences. GSH8-2 and GSH8M-1 were classified as newly assigned sequence types ST558 and ST13, suggesting no supportive evidence of clonal dissemination. The strains appear to have acquired blaKPC-2 -positive IncP-6-relative plasmids (pGSH8-2 and pGSH8M-1-2) that share a common backbone with plasmids in Aeromonas sp. ASNIH3…

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Tuesday, April 21, 2020

Assembly of long, error-prone reads using repeat graphs.

Accurate genome assembly is hampered by repetitive regions. Although long single molecule sequencing reads are better able to resolve genomic repeats than short-read data, most long-read assembly algorithms do not provide the repeat characterization necessary for producing optimal assemblies. Here, we present Flye, a long-read assembly algorithm that generates arbitrary paths in an unknown repeat graph, called disjointigs, and constructs an accurate repeat graph from these error-riddled disjointigs. We benchmark Flye against five state-of-the-art assemblers and show that it generates better or comparable assemblies, while being an order of magnitude faster. Flye nearly doubled the contiguity of the human genome…

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Tuesday, April 21, 2020

Genome sequence of Jatropha curcas L., a non-edible biodiesel plant, provides a resource to improve seed-related traits.

Jatropha curcas (physic nut), a non-edible oilseed crop, represents one of the most promising alternative energy sources due to its high seed oil content, rapid growth and adaptability to various environments. We report ~339 Mbp draft whole genome sequence of J. curcas var. Chai Nat using both the PacBio and Illumina sequencing platforms. We identified and categorized differentially expressed genes related to biosynthesis of lipid and toxic compound among four stages of seed development. Triacylglycerol (TAG), the major component of seed storage oil, is mainly synthesized by phospholipid:diacylglycerol acyltransferase in Jatropha, and continuous high expression of homologs of oleosin over seed development…

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Tuesday, April 21, 2020

Endogenous pararetrovirus sequences are widely present in Citrinae genomes.

Endogenous pararetroviruses (EPRVs) are characterized in several plant genomes and their biological effects have been reported. In this study, hundreds of EPRV segments were identified in six Citrinae genomes. A total of 1034 EPRV segments were identified in the genomes of sweet orange, 2036 in pummelo, 598 in clementine mandarin, 752 in Ichang papeda, 2060 in citron and 245 in atalantia. Genomic analysis indicated that EPRV segments tend to cluster as hot spots in the genomes, particularly on chromosome 2 and 5. Large numbers of simple repeats and transposable elements were identified in the 2-kb flanking regions of the EPRV…

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Tuesday, April 21, 2020

Nephromyces encodes a urate metabolism pathway and predicted peroxisomes, demonstrating that these are not ancient losses of apicomplexans.

The phylum Apicomplexa is a quintessentially parasitic lineage, whose members infect a broad range of animals. One exception to this may be the apicomplexan genus Nephromyces, which has been described as having a mutualistic relationship with its host. Here we analyze transcriptome data from Nephromyces and its parasitic sister taxon, Cardiosporidium, revealing an ancestral purine degradation pathway thought to have been lost early in apicomplexan evolution. The predicted localization of many of the purine degradation enzymes to peroxisomes, and the in silico identification of a full set of peroxisome proteins, indicates that loss of both features in other apicomplexans occurred…

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Tuesday, April 21, 2020

Nodule bacteria from the cultured legume Phaseolus dumosus (belonging to the Phaseolus vulgaris cross-inoculation group) with common tropici phenotypic characteristics and symbiovar but distinctive phylogenomic position and chromid.

Phaseolus dumosus is an endemic species from mountain tops in Mexico that was found in traditional agriculture areas in Veracruz, Mexico. P. dumosus plants were identified by ITS sequences and their nodules were collected from agricultural fields or from trap plant experiments in the laboratory. Bacteria from P. dumosus nodules were identified as belonging to the phaseoli-etli-leguminosarum (PEL) or to the tropici group by 16S rRNA gene sequences. We obtained complete closed genomes from two P. dumosus isolates CCGE531 and CCGE532 that were phylogenetically placed within the tropici group but with a distinctive phylogenomic position and low average nucleotide identity…

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Tuesday, April 21, 2020

Heterochromatin-enriched assemblies reveal the sequence and organization of the Drosophila melanogaster Y chromosome.

Heterochromatic regions of the genome are repeat-rich and poor in protein coding genes, and are therefore underrepresented in even the best genome assemblies. One of the most difficult regions of the genome to assemble are sex-limited chromosomes. The Drosophila melanogaster Y chromosome is entirely heterochromatic, yet has wide-ranging effects on male fertility, fitness, and genome-wide gene expression. The genetic basis of this phenotypic variation is difficult to study, in part because we do not know the detailed organization of the Y chromosome. To study Y chromosome organization in D. melanogaster, we develop an assembly strategy involving the in silico enrichment…

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