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Asset Tag: Whole Genome Sequencing,

April 21, 2020

Giant tortoise genomes provide insights into longevity and age-related disease.

Giant tortoises are among the longest-lived vertebrate animals and, as such, provide an excellent model to study traits like longevity and age-related diseases. However, genomic and molecular evolutionary information on giant tortoises is scarce. Here, we describe a global analysis of the genomes of Lonesome George-the iconic last member of Chelonoidis abingdonii-and the Aldabra giant tortoise (Aldabrachelys gigantea). Comparison of these genomes with those of related species, using both unsupervised and supervised analyses, led us to detect lineage-specific variants affecting DNA repair genes, inflammatory mediators and genes related to cancer development. Our study also hints at specific evolutionary strategies linked to increased lifespan, and expands our understanding of the genomic determinants of ageing. These new genome sequences also provide important resources to help the efforts for restoration of giant tortoise populations.

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April 21, 2020

The genome sequence of segmental allotetraploid peanut Arachis hypogaea.

Like many other crops, the cultivated peanut (Arachis hypogaea L.) is of hybrid origin and has a polyploid genome that contains essentially complete sets of chromosomes from two ancestral species. Here we report the genome sequence of peanut and show that after its polyploid origin, the genome has evolved through mobile-element activity, deletions and by the flow of genetic information between corresponding ancestral chromosomes (that is, homeologous recombination). Uniformity of patterns of homeologous recombination at the ends of chromosomes favors a single origin for cultivated peanut and its wild counterpart A. monticola. However, through much of the genome, homeologous recombination has created diversity. Using new polyploid hybrids made from the ancestral species, we show how this can generate phenotypic changes such as spontaneous changes in the color of the flowers. We suggest that diversity generated by these genetic mechanisms helped to favor the domestication of the polyploid A. hypogaea over other diploid Arachis species cultivated by humans.

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April 21, 2020

Structural variants in 3000 rice genomes.

Investigation of large structural variants (SVs) is a challenging yet important task in understanding trait differences in highly repetitive genomes. Combining different bioinformatic approaches for SV detection, we analyzed whole-genome sequencing data from 3000 rice genomes and identified 63 million individual SV calls that grouped into 1.5 million allelic variants. We found enrichment of long SVs in promoters and an excess of shorter variants in 5′ UTRs. Across the rice genomes, we identified regions of high SV frequency enriched in stress response genes. We demonstrated how SVs may help in finding causative variants in genome-wide association analysis. These new insights into rice genome biology are valuable for understanding the effects SVs have on gene function, with the prospect of identifying novel agronomically important alleles that can be utilized to improve cultivated rice. © 2019 Fuentes et al.; Published by Cold Spring Harbor Laboratory Press.

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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 us to narrow down the list to a few assemblers that can be effectively applied to eukaryotic assembly projects. Moreover, we highlight how best to use limited genomic resources for effectively evaluating the genome assemblies of non-model organisms. © The Author 2017. Published by Oxford University Press.

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April 21, 2020

Whole genome sequencing of NDM-1-producing serotype K1 ST23 hypervirulent Klebsiella pneumoniae in China.

The emergence and spread of carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKP) is causing worldwide concern, whereas NDM-producing hvKP is still rare. Here we report the complete genome sequence characteristics of an NDM-1-producing ST23 type clinical hvKP in PR China.Capsular polysaccharide serotyping was performed by PCR. The complete genome sequence of isolate 3214 was obtained using both the Illumina Hiseq platform and Pacbio RS platform. Multilocus sequence type was identified by submitting the genome sequence to mlst 2.0 and the antimicrobial resistance genes and plasmid replicons were identified using ResFinder and PlasmidFinder, respectively. Transferability of the blaNDM-1-bearing plasmid was determined by conjugation experiment, S1 pulsed-field gel electrophoresis and Southern hybridization.Isolate 3214 was classified to ST23 and belonged to the K1 capsular serotype. The isolate’s total genome size was 6 171 644?bp with a G+C content of 56.39 %, consisting of a 5 448 209?bp chromosome and seven plasmids. The resistome included 18 types of antibiotic resistance genes. Fourteen resistance genes including blaNDM-1 and blaCTX-M-14 were located on plasmids and five also including blaCTX-M-14 were in the chromosome. Plasmid pNDM_3214 carrying blaNDM-1 harboured six types of resistance genes surrounded by insertion sequences and was conjugative. The worldwide pLVPK-like virulence plasmid harbouring rmpA2 and rmpA was also found in this isolate.This study provides basic information of phenotypic and genomic features of ST23 CR-hvKP isolate 3214. Our data highlights the potential risk of spread of NDM-1-producing ST23 hvKP.

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April 21, 2020

The genome of cultivated peanut provides insight into legume karyotypes, polyploid evolution and crop domestication.

High oil and protein content make tetraploid peanut a leading oil and food legume. Here we report a high-quality peanut genome sequence, comprising 2.54?Gb with 20 pseudomolecules and 83,709 protein-coding gene models. We characterize gene functional groups implicated in seed size evolution, seed oil content, disease resistance and symbiotic nitrogen fixation. The peanut B subgenome has more genes and general expression dominance, temporally associated with long-terminal-repeat expansion in the A subgenome that also raises questions about the A-genome progenitor. The polyploid genome provided insights into the evolution of Arachis hypogaea and other legume chromosomes. Resequencing of 52 accessions suggests that independent domestications formed peanut ecotypes. Whereas 0.42-0.47 million years ago (Ma) polyploidy constrained genetic variation, the peanut genome sequence aids mapping and candidate-gene discovery for traits such as seed size and color, foliar disease resistance and others, also providing a cornerstone for functional genomics and peanut improvement.

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April 21, 2020

Transmission of ciprofloxacin resistance in Salmonella mediated by a novel type of conjugative helper plasmids.

Ciprofloxacin resistance in Salmonella has been increasingly reported due to the emergence and dissemination of multiple Plasmid-Mediated Quinolone Resistance (PMQR) determinants, which are mainly located in non-conjugative plasmids or chromosome. In this study, we aimed to depict the molecular mechanisms underlying the rare phenomenon of horizontal transfer of ciprofloxacin resistance phenotype in Salmonella by conjugation experiments, S1-PFGE and complete plasmid sequencing. Two types of non-conjugative plasmids, namely an IncX1 type carrying a qnrS1 gene, and an IncH1 plasmid carrying the oqxAB-qnrS gene, both ciprofloxacin resistance determinants in Salmonella, were recovered from two Salmonella strains. Importantly, these non-conjugative plasmids could be fused with a novel Incl1 type conjugative helper plasmid, which could target insertion sequence (IS) elements located in the non-conjugative, ciprofloxacin-resistance-encoding plasmid through replicative transcription, eventually forming a hybrid conjugative plasmid transmissible among members of Enterobacteriaceae. Since our data showed that such conjugative helper plasmids are commonly detectable among clinical Salmonella strains, particularly S. Typhimurium, fusion events leading to generation and enhanced dissemination of conjugative ciprofloxacin resistance-encoding plasmids in Salmonella are expected to result in a sharp increase in the incidence of resistance to fluoroquinolone, the key choice for treating life-threatening Salmonella infections, thereby posing a serious public health threat.

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April 21, 2020

Interspecies association mapping links reduced CG to TG substitution rates to the loss of gene-body methylation.

Comparative genomics can unravel the genetic basis of species differences; however, successful reports on quantitative traits are still scarce. Here we present genome assemblies of 31 so-far unassembled Brassicaceae plant species and combine them with 16 previously published assemblies to establish the Brassicaceae Diversity Panel. Using a new interspecies association strategy for quantitative traits, we found a so-far unknown association between the unexpectedly high variation in CG to TG substitution rates in genes and the absence of CHROMOMETHYLASE3 (CMT3) orthologues. Low substitution rates were associated with the loss of CMT3, while species with conserved CMT3 orthologues showed high substitution rates. Species without CMT3 also lacked gene-body methylation (gbM), suggesting an evolutionary trade-off between the unknown function of gbM and low substitution rates in Brassicaceae, possibly due to low mutability of non-methylated cytosines.

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April 21, 2020

Genomic and transcriptomic insights into the survival of the subaerial cyanobacterium Nostoc flagelliforme in arid and exposed habitats.

The cyanobacterium Nostoc flagelliforme is an extremophile that thrives under extraordinary desiccation and ultraviolet (UV) radiation conditions. To investigate its survival strategies, we performed whole-genome sequencing of N. flagelliforme CCNUN1 and transcriptional profiling of its field populations upon rehydration in BG11 medium. The genome of N. flagelliforme is 10.23 Mb in size and contains 10 825 predicted protein-encoding genes, making it one of the largest complete genomes of cyanobacteria reported to date. Comparative genomics analysis among 20 cyanobacterial strains revealed that genes related to DNA replication, recombination and repair had disproportionately high contributions to the genome expansion. The ability of N. flagelliforme to thrive under extreme abiotic stresses is supported by the acquisition of genes involved in the protection of photosynthetic apparatus, the formation of monounsaturated fatty acids, responses to UV radiation, and a peculiar role of ornithine metabolism. Transcriptome analysis revealed a distinct acclimation strategy to rehydration, including the strong constitutive expression of genes encoding photosystem I assembly factors and the involvement of post-transcriptional control mechanisms of photosynthetic resuscitation. Our results provide insights into the adaptive mechanisms of subaerial cyanobacteria in their harsh habitats and have important implications to understand the evolutionary transition of cyanobacteria from aquatic environments to terrestrial ecosystems. © 2019 Society for Applied Microbiology and John Wiley & Sons Ltd.

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April 21, 2020

The genome of the medicinal plant Andrographis paniculata provides insight into the biosynthesis of the bioactive diterpenoid neoandrographolide.

Andrographis paniculata is a herbaceous dicot plant widely used for its anti-inflammatory and anti-viral properties across its distribution in China, India and other Southeast Asian countries. A. paniculata was used as a crucial therapeutic treatment during the influenza epidemic of 1919 in India, and is still used for the treatment of infectious disease in China. A. paniculata produces large quantities of the anti-inflammatory diterpenoid lactones andrographolide and neoandrographolide, and their analogs, which are touted to be the next generation of natural anti-inflammatory medicines for lung diseases, hepatitis, neurodegenerative disorders, autoimmune disorders and inflammatory skin diseases. Here, we report a chromosome-scale A. paniculata genome sequence of 269 Mb that was assembled by Illumina short reads, PacBio long reads and high-confidence (Hi-C) data. Gene annotation predicted 25 428 protein-coding genes. In order to decipher the genetic underpinning of diterpenoid biosynthesis, transcriptome data from seedlings elicited with methyl jasmonate were also obtained, which enabled the identification of genes encoding diterpenoid synthases, cytochrome P450 monooxygenases, 2-oxoglutarate-dependent dioxygenases and UDP-dependent glycosyltransferases potentially involved in diterpenoid lactone biosynthesis. We further carried out functional characterization of pairs of class-I and -II diterpene synthases, revealing the ability to produce diversified labdane-related diterpene scaffolds. In addition, a glycosyltransferase able to catalyze O-linked glucosylation of andrograpanin, yielding the major active product neoandrographolide, was also identified. Thus, our results demonstrate the utility of the combined genomic and transcriptomic data set generated here for the investigation of the production of the bioactive diterpenoid lactone constituents of the important medicinal herb A. paniculata. © 2018 The Authors The Plant Journal © 2018 John Wiley & Sons Ltd.

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April 21, 2020

Streptococcus periodonticum sp. nov., Isolated from Human Subgingival Dental Plaque of Periodontitis Lesion.

A novel facultative anaerobic and Gram-stain-positive coccus, designated strain ChDC F135T, was isolated from human subgingival dental plaque of periodontitis lesion and was characterized by polyphasic taxonomic analysis. The 16S rRNA gene (16S rDNA) sequence of strain ChDC F135T was closest to that of Streptococcus sinensis HKU4T (98.2%), followed by Streptococcus intermedia SK54T (97.0%), Streptococcus constellatus NCTC11325T (96.0%), and Streptococcus anginosus NCTC 10713T (95.7%). In contrast, phylogenetic analysis based on the superoxide dismutase gene (sodA) and the RNA polymerase beta-subunit gene (rpoB) showed that the nucleotide sequence similarities of strain ChDC F135T were highly similar to the corresponding genes of S. anginosus NCTC 10713T (99.2% and 97.6%, respectively), S. constellatus NCTC11325T (87.8% and 91.4%, respectively), and S. intermedia SK54T (85.8% and 91.2%, respectively) rather than those of S. sinensis HKU4T (80.5% and 82.6%). The complete genome of strain ChDC F135T consisted of 1,901,251 bp and the G+C content was 38.9 mol %. Average nucleotide identity value between strain ChDC F135T and S. sinensis HKU4T or S. anginosus NCTC 10713T were 75.7% and 95.6%, respectively. The C14:0 composition of the cellular fatty acids of strain ChDC F135T (32.8%) was different from that of S. intermedia (6-8%), S. constellatus (6-13%), and S. anginosus (13-20%). Based on the results of phylogenetic and phenotypic analysis, strain ChDC F135T (=?KCOM 2412T?=?JCM 33300T) was classified as a type strain of a novel species of the genus Streptococcus, for which we proposed the name Streptococcus periodonticum sp. nov.

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April 21, 2020

Assembly of allele-aware, chromosomal-scale autopolyploid genomes based on Hi-C data.

Construction of chromosome-level assembly is a vital step in achieving the goal of a ‘Platinum’ genome, but it remains a major challenge to assemble and anchor sequences to chromosomes in autopolyploid or highly heterozygous genomes. High-throughput chromosome conformation capture (Hi-C) technology serves as a robust tool to dramatically advance chromosome scaffolding; however, existing approaches are mostly designed for diploid genomes and often with the aim of reconstructing a haploid representation, thereby having limited power to reconstruct chromosomes for autopolyploid genomes. We developed a novel algorithm (ALLHiC) that is capable of building allele-aware, chromosomal-scale assembly for autopolyploid genomes using Hi-C paired-end reads with innovative ‘prune’ and ‘optimize’ steps. Application on simulated data showed that ALLHiC can phase allelic contigs and substantially improve ordering and orientation when compared to other mainstream Hi-C assemblers. We applied ALLHiC on an autotetraploid and an autooctoploid sugar-cane genome and successfully constructed the phased chromosomal-level assemblies, revealing allelic variations present in these two genomes. The ALLHiC pipeline enables de novo chromosome-level assembly of autopolyploid genomes, separating each allele. Haplotype chromosome-level assembly of allopolyploid and heterozygous diploid genomes can be achieved using ALLHiC, overcoming obstacles in assembling complex genomes.

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April 21, 2020

Comparative genomics and genome biology of Campylobacter showae.

Campylobacter showae a bacterium historically linked to gingivitis and periodontitis, has recently been associated with inflammatory bowel disease and colorectal cancer. Our aim was to generate genome sequences for new clinical C. showae strains and identify functional properties explaining their pathogenic potential. Eight C. showae genomes were assessed, four strains isolated from inflamed gut tissues from paediatric Crohn’s disease patients, three strains from colonic adenomas, and one from a gastroenteritis patient stool. Genome assemblies were analyzed alongside the only 3 deposited C. showae genomes. The pangenome from these 11 strains consisted of 4686 unique protein families, and the core genome size was estimated at 1050?±?15 genes with each new genome contributing an additional 206?±?16 genes. Functional assays indicated that colonic strains segregated into 2 groups: adherent/invasive vs. non-adherent/non-invasive strains. The former possessed Type IV secretion machinery and S-layer proteins, while the latter contained Cas genes and other CRISPR associated proteins. Comparison of gene profiles with strains in Human Microbiome Project metagenomes showed that gut-derived isolates share genes specific to tongue dorsum and supragingival plaque counterparts. Our findings indicate that C. showae strains are phenotypically and genetically diverse and suggest that secretion systems may play an important role in virulence potential.

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April 21, 2020

Salmonella harbouring the mcr-1 gene isolated from food in China between 2012 and 2016.

In November 2015, plasmid-mediated transferable colistin resistance encoded by the mcr-1 gene in Escherichia coli and Klebsiella pneumonia isolates was reported in China with a high rate of in vitro horizontal transfer (10-1–10-3 cells per recipient cell by conjugation).1 At that time, the mcr-1 gene had already been identified in >30 countries across five continents, with novel mcr-2, mcr-3, mcr-4 and mcr-5 genes being reported subsequently.2–5 Recently, a surveillance study was performed on mainland China to investigate the prevalence of the mcr-1 gene in foodborne Salmonella isolates isolated from various food matrices and others collected…

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April 21, 2020

A coupled role for CsMYB75 and CsGSTF1 in anthocyanin hyperaccumulation in purple tea.

Cultivars of purple tea (Camellia sinensis) that accumulate anthocyanins in place of catechins are currently attracting global interest in their use as functional health beverages. RNA-seq of normal (LJ43) and purple Zijuan (ZJ) cultivars identified the transcription factor CsMYB75 and phi (F) class glutathione transferase CsGSTF1 as being associated with anthocyanin hyperaccumulation. Both genes mapped as a quantitative trait locus (QTL) to the purple bud leaf color (BLC) trait in F1 populations, with CsMYB75 promoting the expression of CsGSTF1 in transgenic tobacco (Nicotiana tabacum). Although CsMYB75 elevates the biosynthesis of both catechins and anthocyanins, only anthocyanins accumulate in purple tea, indicating selective downstream regulation. As glutathione transferases in other plants are known to act as transporters (ligandins) of flavonoids, directing them for vacuolar deposition, the role of CsGSTF1 in selective anthocyanin accumulation was investigated. In tea, anthocyanins accumulate in multiple vesicles, with the expression of CsGSTF1 correlated with BLC, but not with catechin content, in diverse germplasm. Complementation of the Arabidopsis tt19-8 mutant, which is unable to express the orthologous ligandin AtGSTF12, restored anthocyanin accumulation, but did not rescue the transparent testa phenotype, confirming that CsGSTF1 did not function in catechin accumulation. Consistent with a ligandin function, transient expression of CsGSTF1 in Nicotiana occurred in the nucleus, cytoplasm and membrane. Furthermore, RNA-Seq of the complemented mutants exposed to 2% sucrose as a stress treatment showed unexpected roles for anthocyanin accumulation in affecting the expression of genes involved in redox responses, phosphate homeostasis and the biogenesis of photosynthetic components, as compared with non-complemented plants. © 2018 The Authors The Plant Journal © 2018 John Wiley & Sons Ltd.

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