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Auto Tag: Long-read assembly

September 22, 2019  |  

Complete genome sequence of Bacillus velezensis ZY-1-1 reveals the genetic basis for its hemicellulosic/cellulosic substrate-inducible xylanase and cellulase activities.

Bacillus velezensis ZY-1-1 was isolated from the larval gut of the lignocellulose-rich diet-fed scarab beetle, Holotrichia parallela, and confirmed to possess extremely high xylanase (48153.8?±?412.1 U/L) and relatively moderate cellulase activity (610.1?±?8.2 U/L). Notably, these xylanase and cellulase activities were enhanced by xylan (1.4 and 5.8-fold, respectively) and cellulose (1.1 and 3.5-fold, respectively), which indicated the hemicellulosic/cellulosic substrate-inducible lignocellulolytic activities of this strain. The complete genome of B. velezensis ZY-1-1 comprises of 3,899,251 bp in a circular chromosome with a G?+?C content of 46.6%. Among the predicted 3688 protein-coding genes, 24 genes are involved in the degradation of lignocellulose and other polysaccharides, including 8, 7 and 2 critical genes for the degradation of xylan, cellulose and lignin, respectively. This genome-based analysis will facilitate our understanding of the mechanism underlying the biodegradation of lignocellulose and the biotechnological application of this novel lignocellulolytic bacteria or related enzymes.

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September 22, 2019  |  

Acquired interbacterial defense systems protect against interspecies antagonism in the human gut microbiome

The genomes of bacteria derived from the gut microbiota are replete with pathways that mediate contact-dependent interbacterial antagonism. However, the role of direct interactions between co-resident microbes in driving microbiome composition is not well understood. Here we report the widespread occurrence of acquired interbacterial defense (AID) gene clusters in the human gut microbiome. These clusters are found on predicted mobile elements and encode arrays of immunity genes that confer protection against interbacterial toxin-mediated antagonism in vitro and in gnotobiotic mice. We find that Bacteroides ovatus strains containing AID systems that inactivate B. fragilis toxins delivered between cells by the type VI secretion system are enriched in samples lacking detectable B. fragilis. Moreover, these strains display significantly higher abundance in gut metagenomes than strains without AID systems. Finally, we identify a recombinase-associated AID subtype present broadly in Bacteroidales genomes with features suggestive of active gene acquisition. Our data suggest that neutralization of contact-dependent interbacterial antagonism via AID systems plays an important role in shaping human gut microbiome ecology.

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September 22, 2019  |  

Genomic Tandem Quadruplication is Associated with Ketoconazole Resistance in Malassezia pachydermatis.

Malassezia pachydermatis is a commensal yeast found on the skin of dogs. However, M. pachydermatis is also considered an opportunistic pathogen and is associated with various canine skin diseases including otitis externa and atopic dermatitis, which usually require treatment using an azole antifungal drug, such as ketoconazole. In this study, we isolated a ketoconazole-resistant strain of M. pachydermatis, designated “KCTC 27587,” from the external ear canal of a dog with otitis externa and analyzed its resistance mechanism. To understand the mechanism underlying ketoconazole resistance of the clinical isolate M. pachydermatis KCTC 27587, the whole genome of the yeast was sequenced using the PacBio platform and was compared with M. pachydermatis type strain CBS 1879. We found that a ~84-kb region in chromosome 4 of M. pachydermatis KCTC 27587 was tandemly quadruplicated. The quadruplicated region contains 52 protein coding genes, including the homologs of ERG4 and ERG11, whose overexpression is known to be associated with azole resistance. Our data suggest that the quadruplication of the ~84-kb region may be the cause of the ketoconazole resistance in M. pachydermatis KCTC 27587.

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September 22, 2019  |  

Hybrid correction of highly noisy long reads using a variable-order de Bruijn graph.

The recent rise of long read sequencing technologies such as Pacific Biosciences and Oxford Nanopore allows to solve assembly problems for larger and more complex genomes than what allowed short reads technologies. However, these long reads are very noisy, reaching an error rate of around 10-15% for Pacific Biosciences, and up to 30% for Oxford Nanopore. The error correction problem has been tackled by either self-correcting the long reads, or using complementary short reads in a hybrid approach. However, even though sequencing technologies promise to lower the error rate of the long reads below 10%, it is still higher in practice, and correcting such noisy long reads remains an issue.We present HG-CoLoR, a hybrid error correction method that focuses on a seed-and-extend approach based on the alignment of the short reads to the long reads, followed by the traversal of a variable-order de Bruijn graph, built from the short reads. Our experiments show that HG-CoLoR manages to efficiently correct highly noisy long reads that display an error rate as high as 44%. When compared to other state-of-the-art long read error correction methods, our experiments also show that HG-CoLoR provides the best trade-off between runtime and quality of the results, and is the only method able to efficiently scale to eukaryotic genomes.HG-CoLoR is implemented is C++, supported on Linux platforms and freely available at https://github.com/morispi/HG-CoLoR.Supplementary data are available at Bioinformatics online.

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September 22, 2019  |  

Enterobacter cloacae Complex Sequence Type 171 Isolates Expressing KPC-4 Carbapenemase Recovered from Canine Patients in Ohio.

Companion animals are likely relevant in the transmission of antimicrobial-resistant bacteria. Enterobacter xiangfangensis sequence type 171 (ST171), a clone that has been implicated in clusters of infections in humans, was isolated from two dogs with clinical disease in Ohio. The canine isolates contained IncHI2 plasmids encoding blaKPC-4 Whole-genome sequencing was used to put the canine isolates in phylogenetic context with available human ST171 sequences, as well as to characterize their blaKPC-4 plasmids. Copyright © 2018 American Society for Microbiology.

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September 22, 2019  |  

Whole-genome landscape of Medicago truncatula symbiotic genes.

Advances in deciphering the functional architecture of eukaryotic genomes have been facilitated by recent breakthroughs in sequencing technologies, enabling a more comprehensive representation of genes and repeat elements in genome sequence assemblies, as well as more sensitive and tissue-specific analyses of gene expression. Here we show that PacBio sequencing has led to a substantially improved genome assembly of Medicago truncatula A17, a legume model species notable for endosymbiosis studies1, and has enabled the identification of genome rearrangements between genotypes at a near-base-pair resolution. Annotation of the new M. truncatula genome sequence has allowed for a thorough analysis of transposable elements and their dynamics, as well as the identification of new players involved in symbiotic nodule development, in particular 1,037 upregulated long non-coding RNAs (lncRNAs). We have also discovered that a substantial proportion (~35% and 38%, respectively) of the genes upregulated in nodules or expressed in the nodule differentiation zone colocalize in genomic clusters (270 and 211, respectively), here termed symbiotic islands. These islands contain numerous expressed lncRNA genes and display differentially both DNA methylation and histone marks. Epigenetic regulations and lncRNAs are therefore attractive candidate elements for the orchestration of symbiotic gene expression in the M. truncatula genome.

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September 22, 2019  |  

Desiccation Tolerance Evolved through Gene Duplication and Network Rewiring in Lindernia.

Although several resurrection plant genomes have been sequenced, the lack of suitable dehydration-sensitive outgroups has limited genomic insights into the origin of desiccation tolerance. Here, we utilized a comparative system of closely related desiccation-tolerant (Lindernia brevidens) and -sensitive (Lindernia subracemosa) species to identify gene- and pathway-level changes associated with the evolution of desiccation tolerance. The two high-quality Lindernia genomes we assembled are largely collinear, and over 90% of genes are conserved. L. brevidens and L. subracemosa have evidence of an ancient, shared whole-genome duplication event, and retained genes have neofunctionalized, with desiccation-specific expression in L. brevidens Tandem gene duplicates also are enriched in desiccation-associated functions, including a dramatic expansion of early light-induced proteins from 4 to 26 copies in L. brevidens A comparative differential gene coexpression analysis between L. brevidens and L. subracemosa supports extensive network rewiring across early dehydration, desiccation, and rehydration time courses. Many LATE EMBRYOGENESIS ABUNDANT genes show significantly higher expression in L. brevidens compared with their orthologs in L. subracemosa Coexpression modules uniquely upregulated during desiccation in L. brevidens are enriched with seed-specific and abscisic acid-associated cis-regulatory elements. These modules contain a wide array of seed-associated genes that have no expression in the desiccation-sensitive L. subracemosa Together, these findings suggest that desiccation tolerance evolved through a combination of gene duplications and network-level rewiring of existing seed desiccation pathways.© 2018 American Society of Plant Biologists. All rights reserved.

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September 22, 2019  |  

Regulation of yeast-to-hyphae transition in Yarrowia lipolytica.

The yeast Yarrowia lipolytica undergoes a morphological transition from yeast-to-hyphal growth in response to environmental conditions. A forward genetic screen was used to identify mutants that reliably remain in the yeast phase, which were then assessed by whole-genome sequencing. All the smooth mutants identified, so named because of their colony morphology, exhibit independent loss of DNA at a repetitive locus made up of interspersed ribosomal DNA and short 10- to 40-mer telomere-like repeats. The loss of repetitive DNA is associated with downregulation of genes with stress response elements (5′-CCCCT-3′) and upregulation of genes with cell cycle box (5′-ACGCG-3′) motifs in their promoter region. The stress response element is bound by the transcription factor Msn2p in Saccharomyces cerevisiae We confirmed that the Y. lipolyticamsn2 (Ylmsn2) ortholog is required for hyphal growth and found that overexpression of Ylmsn2 enables hyphal growth in smooth strains. The cell cycle box is bound by the Mbp1p/Swi6p complex in S. cerevisiae to regulate G1-to-S phase progression. We found that overexpression of either the Ylmbp1 or Ylswi6 homologs decreased hyphal growth and that deletion of either Ylmbp1 or Ylswi6 promotes hyphal growth in smooth strains. A second forward genetic screen for reversion to hyphal growth was performed with the smooth-33 mutant to identify additional genetic factors regulating hyphal growth in Y. lipolytica Thirteen of the mutants sequenced from this screen had coding mutations in five kinases, including the histidine kinases Ylchk1 and Ylnik1 and kinases of the high-osmolarity glycerol response (HOG) mitogen-activated protein (MAP) kinase cascade Ylssk2, Ylpbs2, and Ylhog1 Together, these results demonstrate that Y. lipolytica transitions to hyphal growth in response to stress through multiple signaling pathways.IMPORTANCE Many yeasts undergo a morphological transition from yeast-to-hyphal growth in response to environmental conditions. We used forward and reverse genetic techniques to identify genes regulating this transition in Yarrowia lipolytica We confirmed that the transcription factor Ylmsn2 is required for the transition to hyphal growth and found that signaling by the histidine kinases Ylchk1 and Ylnik1 as well as the MAP kinases of the HOG pathway (Ylssk2, Ylpbs2, and Ylhog1) regulates the transition to hyphal growth. These results suggest that Y. lipolytica transitions to hyphal growth in response to stress through multiple kinase pathways. Intriguingly, we found that a repetitive portion of the genome containing telomere-like and rDNA repeats may be involved in the transition to hyphal growth, suggesting a link between this region and the general stress response. Copyright © 2018 Pomraning et al.

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September 22, 2019  |  

Genomic insights into multidrug-resistance, mating and virulence in Candida auris and related emerging species.

Candida auris is an emergent multidrug-resistant fungal pathogen causing increasing reports of outbreaks. While distantly related to C. albicans and C. glabrata, C. auris is closely related to rarely observed and often multidrug-resistant species from the C. haemulonii clade. Here, we analyze near complete genome assemblies for the four C. auris clades and three related species, and map intra- and inter-species rearrangements across the seven chromosomes. Using RNA-Seq-guided gene predictions, we find that most mating and meiosis genes are conserved and that clades contain either the MTLa or MTLa mating loci. Comparing the genomes of these emerging species to those of other Candida species identifies genes linked to drug resistance and virulence, including expanded families of transporters and lipases, as well as mutations and copy number variants in ERG11. Gene expression analysis identifies transporters and metabolic regulators specific to C. auris and those conserved with related species which may contribute to differences in drug response in this emerging fungal clade.

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September 22, 2019  |  

Investigation of a cluster of Sphingomonas koreensis infections.

Plumbing systems are an infrequent but known reservoir for opportunistic microbial pathogens that can infect hospitalized patients. In 2016, a cluster of clinical sphingomonas infections prompted an investigation.We performed whole-genome DNA sequencing on clinical isolates of multidrug-resistant Sphingomonas koreensis identified from 2006 through 2016 at the National Institutes of Health (NIH) Clinical Center. We cultured S. koreensis from the sinks in patient rooms and performed both whole-genome and shotgun metagenomic sequencing to identify a reservoir within the infrastructure of the hospital. These isolates were compared with clinical and environmental S. koreensis isolates obtained from other institutions.The investigation showed that two isolates of S. koreensis obtained from the six patients identified in the 2016 cluster were unrelated, but four isolates shared more than 99.92% genetic similarity and were resistant to multiple antibiotic agents. Retrospective analysis of banked clinical isolates of sphingomonas from the NIH Clinical Center revealed the intermittent recovery of a clonal strain over the past decade. Unique single-nucleotide variants identified in strains of S. koreensis elucidated the existence of a reservoir in the hospital plumbing. Clinical S. koreensis isolates from other facilities were genetically distinct from the NIH isolates. Hospital remediation strategies were guided by results of microbiologic culturing and fine-scale genomic analyses.This genomic and epidemiologic investigation suggests that S. koreensis is an opportunistic human pathogen that both persisted in the NIH Clinical Center infrastructure across time and space and caused health care-associated infections. (Funded by the NIH Intramural Research Programs.).

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September 22, 2019  |  

Genomic characterization of ß-glucuronidase-positive Escherichia coli O157:H7 producing Stx2a.

Among Shiga toxin (Stx)-producing Escherichia coli (STEC) O157:H7 strains, those producing Stx2a cause more severe diseases. Atypical STEC O157:H7 strains showing a ß-glucuronidase-positive phenotype (GP STEC O157:H7) have rarely been isolated from humans, mostly from persons with asymptomatic or mild infections; Stx2a-producing strains have not been reported. We isolated, from a patient with bloody diarrhea, a GP STEC O157:H7 strain (PV15-279) that produces Stx2a in addition to Stx1a and Stx2c. Genomic comparison with other STEC O157 strains revealed that PV15-279 recently emerged from the stx1a/stx2c-positive GP STEC O157:H7 clone circulating in Japan. Major virulence genes are shared between typical (ß-glucuronidase-negative) and GP STEC O157:H7 strains, and the Stx2-producing ability of PV15-279 is comparable to that of typical STEC O157:H7 strains; therefore, PV15-279 presents a virulence potential similar to that of typical STEC O157:H7. This study reveals the importance of GP O157:H7 as a source of highly pathogenic STEC clones.

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September 22, 2019  |  

A novel probiotic, Lactobacillus johnsonii 456, resists acid and can persist in the human gut beyond the initial ingestion period.

Probiotics are considered to have multiple beneficial effects on the human gastrointestinal tract, including immunomodulation, pathogen inhibition, and improved host nutrient metabolism. However, extensive characterization of these properties is needed to define suitable clinical applications for probiotic candidates. Lactobacillus johnsonii 456 (LBJ 456) was previously demonstrated to have anti-inflammatory and anti-genotoxic effects in a mouse model. Here, we characterize its resistance to gastric and bile acids as well as its ability to inhibit gut pathogens and adhere to host mucosa. While bile resistance and in vitro host attachment properties of LBJ 456 were comparable to other tested probiotics, LBJ 456 maintained higher viability at lower pH conditions compared to other tested strains. LBJ 456 also altered pathogen adhesion to LS 174T monolayers and demonstrated contact-dependent and independent inhibition of pathogen growth. Genome analyses further revealed possible genetic elements involved in host attachment and pathogen inhibition. Importantly, we show that ingestion of Lactobacillus johnsonii 456 over a one week yogurt course leads to persistent viable bacteria detectable even beyond the period of initial ingestion, unlike many other previously described probiotic species of lactic acid bacteria.

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September 22, 2019  |  

Phototaxis in a wild isolate of the cyanobacterium Synechococcus elongatus.

Many cyanobacteria, which use light as an energy source via photosynthesis, have evolved the ability to guide their movement toward or away from a light source. This process, termed “phototaxis,” enables organisms to localize in optimal light environments for improved growth and fitness. Mechanisms of phototaxis have been studied in the coccoid cyanobacterium Synechocystis sp. strain PCC 6803, but the rod-shaped Synechococcus elongatus PCC 7942, studied for circadian rhythms and metabolic engineering, has no phototactic motility. In this study we report a recent environmental isolate of S. elongatus, the strain UTEX 3055, whose genome is 98.5% identical to that of PCC 7942 but which is motile and phototactic. A six-gene operon encoding chemotaxis-like proteins was confirmed to be involved in phototaxis. Environmental light signals are perceived by a cyanobacteriochrome, PixJSe (Synpcc7942_0858), which carries five GAF domains that are responsive to blue/green light and resemble those of PixJ from Synechocystis Plate-based phototaxis assays indicate that UTEX 3055 uses PixJSe to sense blue and green light. Mutation of conserved functional cysteine residues in different GAF domains indicates that PixJSe controls both positive and negative phototaxis, in contrast to the multiple proteins that are employed for implementing bidirectional phototaxis in Synechocystis.

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September 22, 2019  |  

Comparative Analysis of the Genetic Basis of Branched Nonylphenol Degradation by Sphingobium amiense DSM 16289T and Sphingobium cloacae JCM 10874T.

Branched nonylphenol (BNP), a degradation product of nonylphenol polyethoxylates, exerts estrogenic effects on various organisms. The genes underlying BNP degradation by Sphingobium amiense DSM 16289T were analyzed by complete genome sequencing and compared with those of the versatile BNP-degrading Sphingobium cloacae JCM 10874T. An opdA homolog (opdADSM16289) encoding BNP degradation activity was identified in DSM 16289T, in contrast with JCM 10874T, possessing both the opdA homolog and nmoA. The degradation profile of different BNP isomers was examined by Escherichia coli transformants harboring opdADSM16289, opdAJCM10874, and nmoAJCM10874 to characterize and compare the expression activities of these genes.

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September 21, 2019  |  

in silico Whole Genome Sequencer & Analyzer (iWGS): a computational pipeline to guide the design and analysis of de novo genome sequencing studies.

The availability of genomes across the tree of life is highly biased toward vertebrates, pathogens, human disease models, and organisms with relatively small and simple genomes. Recent progress in genomics has enabled the de novo decoding of the genome of virtually any organism, greatly expanding its potential for understanding the biology and evolution of the full spectrum of biodiversity. The increasing diversity of sequencing technologies, assays, and de novo assembly algorithms have augmented the complexity of de novo genome sequencing projects in non-model organisms. To reduce the costs and challenges in de novo genome sequencing projects and streamline their experimental design and analysis, we developed iWGS (in silico Whole Genome Sequencer and Analyzer), an automated pipeline for guiding the choice of appropriate sequencing strategy and assembly protocols. iWGS seamlessly integrates the four key steps of a de novo genome sequencing project: data generation (through simulation), data quality control, de novo assembly, and assembly evaluation and validation. The last three steps can also be applied to the analysis of real data. iWGS is designed to enable the user to have great flexibility in testing the range of experimental designs available for genome sequencing projects, and supports all major sequencing technologies and popular assembly tools. Three case studies illustrate how iWGS can guide the design of de novo genome sequencing projects and evaluate the performance of a wide variety of user-specified sequencing strategies and assembly protocols on genomes of differing architectures. iWGS, along with a detailed documentation, is freely available at https://github.com/zhouxiaofan1983/iWGS. Copyright © 2016 Author et al.

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