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Asset Tag: Human Biomedical Research,

July 7, 2019

CHOgenome.org 2.0: Genome resources and website updates.

Chinese hamster ovary (CHO) cells are a major host cell line for the production of therapeutic proteins, and CHO cell and Chinese hamster (CH) genomes have recently been sequenced using next-generation sequencing methods. CHOgenome.org was launched in 2011 (version 1.0) to serve as a database repository and to provide bioinformatics tools for the CHO community. CHOgenome.org (version 1.0) maintained GenBank CHO-K1 genome data, identified CHO-omics literature, and provided a CHO-specific BLAST service. Recent major updates to CHOgenome.org (version 2.0) include new sequence and annotation databases for both CHO and CH genomes, a more user-friendly website, and new research tools, including a proteome browser and a genome viewer. CHO cell-line specific sequences and annotations facilitate cell line development opportunities, several of which are discussed. Moving forward, CHOgenome.org will host the increasing amount of CHO-omics data and continue to make useful bioinformatics tools available to the CHO community. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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July 7, 2019

Insights on virulence from the complete genome of Staphylococcus capitis.

Staphylococcus capitis is an opportunistic pathogen of the coagulase negative staphylococci (CoNS). Functional genomic studies of S. capitis have thus far been limited by a lack of available complete genome sequences. Here, we determined the closed S. capitis genome and methylome using Single Molecule Real Time (SMRT) sequencing. The strain, AYP1020, harbors a single circular chromosome of 2.44 Mb encoding 2304 predicted proteins, which is the smallest of all complete staphylococcal genomes sequenced to date. AYP1020 harbors two large mobile genetic elements; a plasmid designated pAYP1020 (59.6 Kb) and a prophage, FAYP1020 (48.5 Kb). Methylome analysis identified significant adenine methylation across the genome involving two distinct methylation motifs (1972 putative 6-methyladenine (m6A) residues identified). Putative adenine methyltransferases were also identified. Comparative analysis of AYP1020 and the closely related CoNS, S. epidermidis RP62a, revealed a host of virulence factors that likely contribute to S. capitis pathogenicity, most notably genes important for biofilm formation and a suite of phenol soluble modulins (PSMs); the expression/production of these factors were corroborated by functional assays. The complete S. capitis genome will aid future studies on the evolution and pathogenesis of the coagulase negative staphylococci.

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July 7, 2019

Jitterbug: somatic and germline transposon insertion detection at single-nucleotide resolution.

Transposable elements are major players in genome evolution. Transposon insertion polymorphisms can translate into phenotypic differences in plants and animals and are linked to different diseases including human cancer, making their characterization highly relevant to the study of genome evolution and genetic diseases. Here we present Jitterbug, a novel tool that identifies transposable element insertion sites at single-nucleotide resolution based on the pairedend mapping and clipped-read signatures produced by NGS alignments. Jitterbug can be easily integrated into existing NGS analysis pipelines, using the standard BAM format produced by frequently applied alignment tools (e.g. bwa, bowtie2), with no need to realign reads to a set of consensus transposon sequences. Jitterbug is highly sensitive and able to recall transposon insertions with a very high specificity, as demonstrated by benchmarks in the human and Arabidopsis genomes, and validation using long PacBio reads. In addition, Jitterbug estimates the zygosity of transposon insertions with high accuracy and can also identify somatic insertions. We demonstrate that Jitterbug can identify mosaic somatic transposon movement using sequenced tumor-normal sample pairs and allows for estimating the cancer cell fraction of clones containing a somatic TE insertion. We suggest that the independent methods we use to evaluate performance are a step towards creating a gold standard dataset for benchmarking structural variant prediction tools.

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July 7, 2019

Complete genome sequence of the Clostridium difficile type strain DSM 1296T.

In this study, we sequenced the complete genome of the Clostridium difficile type strain DSM 1296(T). A combination of single-molecule real-time (SMRT) and Illumina sequencing technology revealed the presence of one chromosome and two extrachromosomal elements, the bacteriophage phiCDIF1296T and a putative plasmid-like structure harboring genes of another bacteriophage. Copyright © 2015 Riedel et al.

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July 7, 2019

Completing the human genome: the progress and challenge of satellite DNA assembly.

Genomic studies rely on accurate chromosome assemblies to explore sequence-based models of cell biology, evolution and biomedical disease. However, even the extensively studied human genome has not yet reached a complete, ‘telomere-to-telomere’, chromosome assembly. The largest assembly gaps remain in centromeric regions and acrocentric short arms, sites known to contain megabase-sized arrays of tandem repeats, or satellite DNAs. This review aims to briefly address the progress and challenges of generating correct assemblies of satellite DNA arrays. Although the focus is placed on the human genome, many concepts presented here are applicable to other genomes.

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July 7, 2019

Genomic identification of nitrogen-fixing Klebsiella variicola, K. pneumoniae and K. quasipneumoniae.

It was difficult to differentiate Klebsiella pneumoniae, K. quasipneumoniae and K. variicola by biochemical and phenotypic tests. Genomics increase the resolution and credibility of taxonomy for closely-related species. Here, we obtained the complete genome sequence of the K. variicola type strain DSM 15968(T) (=F2R9(T) ). The genome of the type strain is a circular chromosome of 5,521,203?bp with 57.56% GC content. From 540 Klebsiella strains whose genomes had been publicly available as at 3 March 2015, we identified 21 strains belonging to K. variicola and 8 strains belonging to K. quasipneumoniae based on the genome average nucleotide identities (ANI). All the K. variicola strains, one K. pneumoniae strain and five K. quasipneumoniae strains contained nitrogen-fixing genes. A phylogenomic analysis showed clear species demarcations for these nitrogen-fixing bacteria. In accordance with the key biochemical characteristics of K. variicola, the idnO gene encoding 5-keto-D-gluconate 5-reductase for utilization of 5-keto-D-gluconate and the sorCDFBAME operon for catabolism of L-sorbose were present whereas the rbtRDKT operon for catabolism of adonitol was absent in the genomes of K. variicola strains. Therefore, the genomic analyses supported the ANI-based species delineation; the genome sequence of the K. variicola type strain provides the reference genome for genomic identification of K. variicola, which is a nitrogen-fixing species. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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July 7, 2019

Transfer of scarlet fever-associated elements into the group A Streptococcus M1T1 clone.

The group A Streptococcus (GAS) M1T1 clone emerged in the 1980s as a leading cause of epidemic invasive infections worldwide, including necrotizing fasciitis and toxic shock syndrome. Horizontal transfer of mobile genetic elements has played a central role in the evolution of the M1T1 clone, with bacteriophage-encoded determinants DNase Sda1 and superantigen SpeA2 contributing to enhanced virulence and colonization respectively. Outbreaks of scarlet fever in Hong Kong and China in 2011, caused primarily by emm12 GAS, led to our investigation of the next most common cause of scarlet fever, emm1 GAS. Genomic analysis of 18 emm1 isolates from Hong Kong and 16 emm1 isolates from mainland China revealed the presence of mobile genetic elements associated with the expansion of emm12 scarlet fever clones in the M1T1 genomic background. These mobile genetic elements confer expression of superantigens SSA and SpeC, and resistance to tetracycline, erythromycin and clindamycin. Horizontal transfer of mobile DNA conferring multi-drug resistance and expression of a new superantigen repertoire in the M1T1 clone should trigger heightened public health awareness for the global dissemination of these genetic elements.

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July 7, 2019

Chromosomal rearrangements as barriers to genetic homogenization between archaic and modern humans.

Chromosomal rearrangements, which shuffle DNA throughout the genome, are an important source of divergence across taxa. Using a paired-end read approach with Illumina sequence data for archaic humans, I identify changes in genome structure that occurred recently in human evolution. Hundreds of rearrangements indicate genomic trafficking between the sex chromosomes and autosomes, raising the possibility of sex-specific changes. Additionally, genes adjacent to genome structure changes in Neanderthals are associated with testis-specific expression, consistent with evolutionary theory that new genes commonly form with expression in the testes. I identify one case of new-gene creation through transposition from the Y chromosome to chromosome 10 that combines the 5′-end of the testis-specific gene Fank1 with previously untranscribed sequence. This new transcript experienced copy number expansion in archaic genomes, indicating rapid genomic change. Among rearrangements identified in Neanderthals, 13% are transposition of selfish genetic elements, whereas 32% appear to be ectopic exchange between repeats. In Denisovan, the pattern is similar but numbers are significantly higher with 18% of rearrangements reflecting transposition and 40% ectopic exchange between distantly related repeats. There is an excess of divergent rearrangements relative to polymorphism in Denisovan, which might result from nonuniform rates of mutation, possibly reflecting a burst of transposable element activity in the lineage that led to Denisovan. Finally, loci containing genome structure changes show diminished rates of introgression from Neanderthals into modern humans, consistent with the hypothesis that rearrangements serve as barriers to gene flow during hybridization. Together, these results suggest that this previously unidentified source of genomic variation has important biological consequences in human evolution. © The Author 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

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July 7, 2019

svviz: a read viewer for validating structural variants.

Visualizing read alignments is the most effective way to validate candidate structural variants (SVs) with existing data. We present svviz, a sequencing read visualizer for SVs that sorts and displays only reads relevant to a candidate SV. svviz works by searching input bam(s) for potentially relevant reads, realigning them against the inferred sequence of the putative variant allele as well as the reference allele and identifying reads that match one allele better than the other. Separate views of the two alleles are then displayed in a scrollable web browser view, enabling a more intuitive visualization of each allele, compared with the single reference genome-based view common to most current read browsers. The browser view facilitates examining the evidence for or against a putative variant, estimating zygosity, visualizing affected genomic annotations and manual refinement of breakpoints. svviz supports data from most modern sequencing platforms.svviz is implemented in python and freely available from http://svviz.github.io/. Published by Oxford University Press 2015. This work is written by US Government employees and is in the public domain in the US.

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July 7, 2019

A rebeccamycin analog provides plasmid-encoded niche defense.

Bacterial symbionts of fungus-growing ants occupy a highly specialized ecological niche and face the constant existential threat of displacement by another strain of ant-adapted bacteria. As part of a systematic study of the small molecules underlying this fraternal competition, we discovered an analog of the antitumor agent rebeccamycin, a member of the increasingly important indolocarbazole family. While several gene clusters consistent with this molecule’s newly reported modification had previously been identified in metagenomic studies, the metabolite itself has been cryptic. The biosynthetic gene cluster for 9-methoxyrebeccamycin is encoded on a plasmid in a manner reminiscent of plasmid-derived peptide antimicrobials that commonly mediate antagonism among closely related Gram-negative bacteria.

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July 7, 2019

Genome sequence of Salmonella enterica subsp. enterica serovar Typhi isolate PM016/13 from untreated well water associated with a Typhoid outbreak in Pasir Mas, Kelantan, Malaysia.

Salmonella enterica subsp. enterica serovar Typhi is a human-restricted pathogen that causes typhoid fever. Even though it is a human-restricted pathogen, the bacterium is also isolated from environments such as groundwater and pond water. Here, we describe the genome sequence of the Salmonella enterica subsp. enterica serovar Typhi PM016/13 which was isolated from well water during a typhoid outbreak in Kelantan, Malaysia, in 2013. Copyright © 2015 Muhamad Harish et al.

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July 7, 2019

Complete genome sequence of a human-invasive Salmonella enterica Serovar Typhimurium strain of the emerging sequence type 213 harboring a multidrug resistance IncA/C plasmid and a blaCMY-2-carrying IncF plasmid.

Salmonella enterica subsp. enterica serovar Typhimurium strain 33676 was isolated in Mexico City, Mexico, from a patient with a systemic infection, and its complete genome sequence was determined using PacBio single-molecule real-time technology. Strain 33676 harbors an IncF plasmid carrying the extended-spectrum cephalosporin gene blaCMY-2 and a multidrug resistance IncA/C plasmid. Copyright © 2015 Silva et al.

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July 7, 2019

Clonal dissemination of Enterobacter cloacae harboring blaKPC-3 in the upper midwestern United States.

Carbapenemase-producing, carbapenem-resistant Enterobacteriaceae, or CP-CRE, are an emerging threat to human and animal health, because they are resistant to many of the last-line antimicrobials available for disease treatment. Carbapenemase-producing Enterobacter cloacae harboring blaKPC-3 recently was reported in the upper midwestern United States and implicated in a hospital outbreak in Fargo, North Dakota (L. M. Kiedrowski, D. M. Guerrero, F. Perez, R. A. Viau, L. J. Rojas, M. F. Mojica, S. D. Rudin, A. M. Hujer, S. H. Marshall, and R. A. Bonomo, Emerg Infect Dis 20:1583-1585, 2014, http://dx.doi.org/10.3201/eid2009.140344). In early 2009, the Minnesota Department of Health began collecting and screening CP-CRE from patients throughout Minnesota. Here, we analyzed a retrospective group of CP-E. cloacae isolates (n = 34) collected between 2009 and 2013. Whole-genome sequencing and analysis revealed that 32 of the strains were clonal, belonging to the ST171 clonal complex and differing collectively by 211 single-nucleotide polymorphisms, and it revealed a dynamic clone under positive selection. The phylogeography of these strains suggests that this clone existed in eastern North Dakota and western Minnesota prior to 2009 and subsequently was identified in the Minneapolis and St. Paul metropolitan area. All strains harbored identical IncFIA-like plasmids conferring a CP-CRE phenotype and an additional IncX3 plasmid. In a single patient with multiple isolates submitted over several months, we found evidence that these plasmids had transferred from the E. cloacae clone to an Escherichia coli ST131 bacterium, rendering it as a CP-CRE. The spread of this clone throughout the upper midwestern United States is unprecedented for E. cloacae and highlights the importance of continued surveillance to identify such threats to human health. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

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