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Asset Tag: de novo assembly

April 21, 2020

The vaginal microbiome and preterm birth.

The incidence of preterm birth exceeds 10% worldwide. There are significant disparities in the frequency of preterm birth among populations within countries, and women of African ancestry disproportionately bear the burden of risk in the United States. In the present study, we report a community resource that includes ‘omics’ data from approximately 12,000 samples as part of the integrative Human Microbiome Project. Longitudinal analyses of 16S ribosomal RNA, metagenomic, metatranscriptomic and cytokine profiles from 45 preterm and 90 term birth controls identified harbingers of preterm birth in this cohort of women predominantly of African ancestry. Women who delivered preterm exhibited significantly lower vaginal levels of Lactobacillus crispatus and higher levels of BVAB1, Sneathia amnii, TM7-H1, a group of Prevotella species and nine additional taxa. The first representative genomes of BVAB1 and TM7-H1 are described. Preterm-birth-associated taxa were correlated with proinflammatory cytokines in vaginal fluid. These findings highlight new opportunities for assessment of the risk of preterm birth.

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April 21, 2020

Recompleting the Caenorhabditis elegans genome.

Caenorhabditis elegans was the first multicellular eukaryotic genome sequenced to apparent completion. Although this assembly employed a standard C. elegans strain (N2), it used sequence data from several laboratories, with DNA propagated in bacteria and yeast. Thus, the N2 assembly has many differences from any C. elegans available today. To provide a more accurate C. elegans genome, we performed long-read assembly of VC2010, a modern strain derived from N2. Our VC2010 assembly has 99.98% identity to N2 but with an additional 1.8 Mb including tandem repeat expansions and genome duplications. For 116 structural discrepancies between N2 and VC2010, 97 structures matching VC2010 (84%) were also found in two outgroup strains, implying deficiencies in N2. Over 98% of N2 genes encoded unchanged products in VC2010; moreover, we predicted =53 new genes in VC2010. The recompleted genome of C. elegans should be a valuable resource for genetics, genomics, and systems biology. © 2019 Yoshimura et al.; Published by Cold Spring Harbor Laboratory Press.

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April 21, 2020

Detection of Fusarium oxysporum f. sp. fragariae from Infected Strawberry Plants.

Isolates of the Fusarium oxysporum species complex have been characterized as plant pathogens that commonly cause vascular wilt, stunting, and yellowing of the leaves in a variety of hosts. F. oxysporum species complex isolates have been grouped into formae speciales based on their ability to cause disease on a specific host. F. oxysporum f. sp. fragariae is the causal agent of Fusarium wilt of strawberry and has become a threat to production as fumigation practices have changed in California. F. oxysporum f. sp. fragariae is polyphyletic and limited genetic markers are available for its detection. In this study, next-generation sequencing and comparative genomics were used to identify a unique genetic locus that can detect all of the somatic compatibility groups of F. oxysporum f. sp. fragariae identified in California. This locus was used to develop a TaqMan quantitative polymerase chain reaction assay and an isothermal recombinase polymerase amplification (RPA) assay that have very high sensitivity and specificity for more than 180 different isolates of the pathogen tested. RPA assay results from multiple field samples were validated with pathogenicity tests of recovered isolates.

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April 21, 2020

SMRT long reads and Direct Label and Stain optical maps allow the generation of a high-quality genome assembly for the European barn swallow (Hirundo rustica rustica).

The barn swallow (Hirundo rustica) is a migratory bird that has been the focus of a large number of ecological, behavioral, and genetic studies. To facilitate further population genetics and genomic studies, we present a reference genome assembly for the European subspecies (H. r. rustica).As part of the Genome10K effort on generating high-quality vertebrate genomes (Vertebrate Genomes Project), we have assembled a highly contiguous genome assembly using single molecule real-time (SMRT) DNA sequencing and several Bionano optical map technologies. We compared and integrated optical maps derived from both the Nick, Label, Repair, and Stain technology and from the Direct Label and Stain (DLS) technology. As proposed by Bionano, DLS more than doubled the scaffold N50 with respect to the nickase. The dual enzyme hybrid scaffold led to a further marginal increase in scaffold N50 and an overall increase of confidence in the scaffolds. After removal of haplotigs, the final assembly is approximately 1.21 Gbp in size, with a scaffold N50 value of more than 25.95 Mbp.This high-quality genome assembly represents a valuable resource for future studies of population genetics and genomics in the barn swallow and for studies concerning the evolution of avian genomes. It also represents one of the very first genomes assembled by combining SMRT long-read sequencing with the new Bionano DLS technology for scaffolding. The quality of this assembly demonstrates the potential of this methodology to substantially increase the contiguity of genome assemblies.

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April 21, 2020

The history, genome and biology of NCTC 30: a non-pandemic Vibrio cholerae isolate from World War One.

The sixth global cholera pandemic lasted from 1899 to 1923. However, despite widespread fear of the disease and of its negative effects on troop morale, very few soldiers in the British Expeditionary Forces contracted cholera between 1914 and 1918. Here, we have revived and sequenced the genome of NCTC 30, a 102-year-old Vibrio cholerae isolate, which we believe is the oldest publicly available live V. cholerae strain in existence. NCTC 30 was isolated in 1916 from a British soldier convalescent in Egypt. We found that this strain does not encode cholera toxin, thought to be necessary to cause cholera, and is not part of V. cholerae lineages responsible for the pandemic disease. We also show that NCTC 30, which predates the introduction of penicillin-based antibiotics, harbours a functional ß-lactamase antibiotic resistance gene. Our data corroborate and provide molecular explanations for previous phenotypic studies of NCTC 30 and provide a new high-quality genome sequence for historical, non-pandemic V. cholerae.

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April 21, 2020

Competition between mobile genetic elements drives optimization of a phage-encoded CRISPR-Cas system: insights from a natural arms race.

CRISPR-Cas systems function as adaptive immune systems by acquiring nucleotide sequences called spacers that mediate sequence-specific defence against competitors. Uniquely, the phage ICP1 encodes a Type I-F CRISPR-Cas system that is deployed to target and overcome PLE, a mobile genetic element with anti-phage activity in Vibrio cholerae. Here, we exploit the arms race between ICP1 and PLE to examine spacer acquisition and interference under laboratory conditions to reconcile findings from wild populations. Natural ICP1 isolates encode multiple spacers directed against PLE, but we find that single spacers do not interfere equally with PLE mobilization. High-throughput sequencing to assay spacer acquisition reveals that ICP1 can also acquire spacers that target the V. cholerae chromosome. We find that targeting the V. cholerae chromosome proximal to PLE is sufficient to block PLE and is dependent on Cas2-3 helicase activity. We propose a model in which indirect chromosomal spacers are able to circumvent PLE by Cas2-3-mediated processive degradation of the V. cholerae chromosome before PLE mobilization. Generally, laboratory-acquired spacers are much more diverse than the subset of spacers maintained by ICP1 in nature, showing how evolutionary pressures can constrain CRISPR-Cas targeting in ways that are often not appreciated through in vitro analyses. This article is part of a discussion meeting issue ‘The ecology and evolution of prokaryotic CRISPR-Cas adaptive immune systems’.

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April 21, 2020

Comparative Genome Characterization of a Petroleum-Degrading Bacillus subtilis Strain DM2.

The complete genome sequence of Bacillus subtilis strain DM2 isolated from petroleum-contaminated soil on the Tibetan Plateau was determined. The genome of strain DM2 consists of a circular chromosome of 4,238,631 bp for 4458 protein-coding genes and a plasmid of 84,240 bp coding for 103 genes. Thirty-four genomic islands coding for 330 proteins and 5 prophages are found in the genome. The DDH value shows that strain DM2 belongs to B. subtilis subsp. subtilis subspecies, but significant variations of the genome are also present. Comparative analysis showed that the genome of strain DM2 encodes some strain-specific proteins in comparison with B. subtilis subsp. subtilis str. 168, such as carboxymuconolactone decarboxylase family protein, gfo/Idh/MocA family oxidoreductases, GlsB/YeaQ/YmgE family stress response membrane protein, HlyC/CorC family transporters, LLM class flavin-dependent oxidoreductase, and LPXTG cell wall anchor domain-containing protein. Most of the common strain-specific proteins in DM2 and MJ01 strains, or proteins unique to DM2 strain, are involved in the pathways related to stress response, signaling, and hydrocarbon degradation. Furthermore, the strain DM2 genome contains 122 genes coding for developed two-component systems and 138 genes coding for ABC transporter systems. The prominent features of the strain DM2 genome reflect the evolutionary fitness of this strain to harsh conditions and hydrocarbon utilization.

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April 21, 2020

Complete Genome Sequence of Photobacterium damselae Subsp. damselae Strain SSPD1601 Isolated from Deep-Sea Cage-Cultured Sebastes schlegelii with Septic Skin Ulcer.

Photobacterium damselae subsp. damselae (PDD) is a Gram-negative bacterium that can infect a variety of aquatic organisms and humans. Based on an epidemiological investigation conducted over the past 3 years, PDD is one of the most important pathogens causing septic skin ulcer in deep-sea cage-cultured Sebastes schlegelii in the Huang-Bohai Sea area and present throughout the year with high abundance. To further understand the pathogenicity of this species, the pathogenic properties and genome of PDD strain SSPD1601 were analyzed. The results revealed that PDD strain SSPD1601 is a rod-shaped cell with a single polar flagellum, and the clinical symptoms were replicated during artificial infection. The SSPD1601 genome consists of two chromosomes and two plasmids, totaling 4,252,294?bp with 3,751 coding sequences (CDSs), 196 tRNA genes, and 47 rRNA genes. Common virulence factors including flagellin, Fur, RstB, hcpA, OMPs, htpB-Hsp60, VasK, and vgrG were found in strain SSPD1601. Furthermore, SSPD1601 is a pPHDD1-negative strain containing the hemolysin gene hlyAch and three putative hemolysins (emrA, yoaF, and VPA0226), which are likely responsible for the pathogenicity of SSPD1601. The phylogenetic analysis revealed SSPD1601 to be most closely related to Phdp Wu-1. In addition, the antibiotic resistance phenotype indicated that SSPD1601 was not sensitive to ceftazidime, pipemidic, streptomycin, cefalexin, bacitracin, cefoperazone sodium, acetylspiramycin, clarithromycin, amikacin, gentamycin, kanamycin, oxacillin, ampicillin, and trimethoprim-sulfamethoxazole, but only the bacitracin resistance gene bacA was detected based on Antibiotic Resistance Genes Database. These results expand our understanding of PDD, setting the stage for further studies of its pathogenesis and disease prevention.

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April 21, 2020

Analysis of the Complete Genome Sequence of a Novel, Pseudorabies Virus Strain Isolated in Southeast Europe.

Pseudorabies virus (PRV) is the causative agent of Aujeszky’s disease giving rise to significant economic losses worldwide. Many countries have implemented national programs for the eradication of this virus. In this study, long-read sequencing was used to determine the nucleotide sequence of the genome of a novel PRV strain (PRV-MdBio) isolated in Serbia.In this study, a novel PRV strain was isolated and characterized. PRV-MdBio was found to exhibit similar growth properties to those of another wild-type PRV, the strain Kaplan. Single-molecule real-time (SMRT) sequencing has revealed that the new strain differs significantly in base composition even from strain Kaplan, to which it otherwise exhibits the highest similarity. We compared the genetic composition of PRV-MdBio to strain Kaplan and the China reference strain Ea and obtained that radical base replacements were the most common point mutations preceding conservative and silent mutations. We also found that the adaptation of PRV to cell culture does not lead to any tendentious genetic alteration in the viral genome.PRV-MdBio is a wild-type virus, which differs in base composition from other PRV strains to a relatively large extent.

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April 21, 2020

A draft genome assembly of the solar-powered sea slug Elysia chlorotica.

Elysia chlorotica, a sacoglossan sea slug found off the East Coast of the United States, is well-known for its ability to sequester chloroplasts from its algal prey and survive by photosynthesis for up to 12 months in the absence of food supply. Here we present a draft genome assembly of E. chlorotica that was generated using a hybrid assembly strategy with Illumina short reads and PacBio long reads. The genome assembly comprised 9,989 scaffolds, with a total length of 557?Mb and a scaffold N50 of 442?kb. BUSCO assessment indicated that 93.3% of the expected metazoan genes were completely present in the genome assembly. Annotation of the E. chlorotica genome assembly identified 176?Mb (32.6%) of repetitive sequences and a total of 24,980 protein-coding genes. We anticipate that the annotated draft genome assembly of the E. chlorotica sea slug will promote the investigation of sacoglossan genetics, evolution, and particularly, the genetic signatures accounting for the long-term functioning of algal chloroplasts in an animal.

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April 21, 2020

The sequence and de novo assembly of Oxygymnocypris stewartii genome.

Animal genomes in the Qinghai-Tibetan Plateau provide valuable resources for scientists to understand the molecular mechanism of environmental adaptation. Tibetan fish species play essential roles in the local ecology; however, the genomic information for native fishes was still insufficient. Oxygymnocypris stewartii, belonging to Oxygymnocypris genus, Schizothoracinae subfamily, is a native fish in the Tibetan plateau living within the elevation from roughly 3,000?m to 4,200?m. In this report, PacBio and Illumina sequencing platform were used to generate ~385.3?Gb genomic sequencing data. A genome of about 1,849.2?Mb was obtained with a contig N50 length of 257.1?kb. More than 44.5% of the genome were identified as repetitive elements, and 46,400 protein-coding genes were annotated in the genome. The assembled genome can be used as a reference for future population genetic studies of O. stewartii and will improve our understanding of high altitude adaptation of fishes in the Qinghai-Tibetan Plateau.

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April 21, 2020

Identification of plasmid encoded osmoregulatory genes from halophilic bacteria isolated from the rhizosphere of halophytes.

Bacterial plasmids carry genes that code for additional traits such as osmoregulation, CO2 fixation, antibiotic and heavy metal resistance, root nodulation and nitrogen fixation. The main objective of the current study was to identify plasmid-conferring osmoregulatory genes in bacteria isolated from rhizospheric and non-rhizospheric soils of halophytes (Salsola stocksii and Atriplex amnicola). More than 55% of halophilic bacteria from the rhizosphere and 70% from non-rhizospheric soils were able to grow at 3?M salt concentrations. All the strains showed optimum growth at 1.5-3.0?M NaCl. Bacterial strains from the Salsola rhizosphere showed maximum (31%) plasmid elimination during curing experiments as compared to bacterial strains from the Atriplex rhizosphere and non-rhizospheric soils. Two plasmid cured strains Bacillus HL2HP6 and Oceanobacillus HL2RP7 lost their ability to grow in halophilic medium, but they grew well on LB medium. The plasmid cured strains also showed a change in sensitivity to specific antibiotics. These plasmids were isolated and transformed into E. coli strains and growth response of wild-type and transformed E. coli strains was compared at 1.5-4?M NaCl concentrations. Chromosomal DNA and plasmids from Bacillus filamentosus HL2HP6 were sequenced by using high throughput sequencing approach. Results of functional analysis of plasmid sequences showed different proteins and enzymes involved in osmoregulation of bacteria, such as trehalose, ectoine synthetase, porins, proline, alanine, inorganic ion transporters, dehydrogenases and peptidases. Our results suggested that plasmid conferring osmoregulatory genes play a vital role to maintain internal osmotic balance of bacterial cells and these genes can be used to develop salt tolerant transgenic crops.Copyright © 2019 Elsevier GmbH. All rights reserved.

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April 21, 2020

Bradyrhizobium nanningense sp. nov., Bradyrhizobium guangzhouense sp. nov. and Bradyrhizobium zhanjiangense sp. nov., isolated from effective nodules of peanut in Southeast China.

Nine slow-growing rhizobia isolated from effective nodules on peanut (Arachis hypogaea) were characterized to clarify the taxonomic status using a polyphasic approach. They were assigned to the genus Bradyrhizobium on the basis of 16S rRNA sequences. MLSA of concatenated glnII-recA-dnaK genes classified them into three species represented by CCBAU 53390T, CCBAU 51670T and CCBAU 51778T, which presented the closest similarity to B. guangxiense CCBAU 53363T, B. guangdongense CCBAU 51649T and B. manausense BR 3351T, B. vignae 7-2T and B. forestalis INPA 54BT, respectively. The dDDH (digital DNA-DNA hybridization) and ANI (Average Nucleotide Identity) between the genomes of the three representative strains and type strains for the closest Bradyrhizobium species were less than 42.1% and 91.98%, respectively, below the threshold of species circumscription. Effective nodules could be induced on peanut and Lablab purpureus by all representative strains, while Vigna radiata formed effective nodules only with CCBAU 53390T and CCBAU 51778T. Phenotypic characteristics including sole carbon sources and growth features supported the phylogenetic results. Based on the genotypic and phenotypic features, strains CCBAU 53390T, CCBAU 51670T and CCBAU 51778T are designated the type strains of three novel species, for which the names Bradyrhizobium nanningense sp. nov., Bradyrhizobium guangzhouense sp. nov. and Bradyrhizobium zhanjiangense sp. nov. are proposed, respectively.Copyright © 2019 Elsevier GmbH. All rights reserved.

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April 21, 2020

The complete genome sequence of Ethanoligenens harbinense reveals the metabolic pathway of acetate-ethanol fermentation: A novel understanding of the principles of anaerobic biotechnology.

Ethanol-type fermentation is one of three main fermentation types in the acidogenesis of anaerobic treatment systems. Non-spore-forming Ethanoligenens is as a typical genus capable of ethanol-type fermentation in mixed culture (i.e. acetate-ethanol fermentation). This genus can produce ethanol, acetate, CO2, and H2 using carbohydrates, and has application potential in anaerobic bioprocesses. Here, the complete genome sequences and methylome of Ethanoligenens harbinense strains with different autoaggregative and coaggregative abilities were obtained using the PacBio single-molecule real-time sequencing platform. The genome size of E. harbinense strains was about 2.97-3.10?Mb with 55.5% G+C content. 3020-3153 genes were annotated, most of which were methylated at specific sites or motifs. The methylation types included 6mA, 4mC, and unknown types. Comparative genomic analysis demonstrated low levels of genetic similarity between E. harbinense and other well-known hydrogen-producing bacteria (i.e., Clostridium and Thermoanaerobacter) in phylogenesis. Hydrogen production of E. harbinense was catalyzed by genes that encode [FeFe]-hydrogenases and that were synthesized by three maturases of [FeFe]-H2ase. The metabolic mechanism of H2-ethanol co-production fermentation, catalyzed by pyruvate ferredoxin oxidoreductase was proposed. This study provides genetic and evolutionary information of a model genus for the further investigation of the metabolic pathway and regulatory network of ethanol-type fermentation and anaerobic bioprocesses for waste or wastewater treatment.Copyright © 2019. Published by Elsevier Ltd.

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April 21, 2020

Complete genome sequence data of Flavobacterium anhuiense strain GSE09, a volatile-producing biocontrol bacterium isolated from cucumber (Cucumis sativus) root.

Flavobacterium anhuiense (previously identified as Flavobacterium johnsoniae) strain GSE09 is a volatile-producing bacterium that exhibits significant biocontrol activity against an oomycete pathogen, Phytophthora capsici, on pepper plants. Here, we report the complete genome sequence data of strain GSE09, isolated from surface-sterilized cucumber root. The genome consists of a circular 5,109,718-bp chromosome with a G + C content of 34.30%. A total of 4,138 complete coding sequences including 15 rRNA, 66 tRNA, 3 ncRNA, and 51 pseudogene sequences were retrieved. Thus, the genome sequence data of F. anhuiense GSE09 may facilitate the elucidation of many biological traits related to the biocontrol against plant pathogens.

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