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April 21, 2020

Whole-genome sequence of Arthrinium phaeospermum, a globally distributed pathogenic fungus.

Arthrinium phaeospermum (Corda) M.B. Ellis is a globally distributed pathogenic fungus with a wide host range; its hosts include not only plants, but also humans and animals. This study aimed to develop genomic resources for A. phaeospermum to provide solid data and a theoretical basis for further studies of its pathogenesis, transcriptomics, proteomics, metabolomics and RNA genomics. The genome was obtained from the mycelia of the strain AP-Z13 using a combination of analyses with the high-throughput Illumina HiSeq 4000 system and PacBio RSII LongRead sequencing platform. Functional annotation was performed by BLASTing protein sequences against those in different publicly available databases to obtain their corresponding annotations. The genome is 48.45?Mb in size, with an N90 scaffold size of 1,931,147?bp, and encodes 19,836 putative predicted genes. This is the first report of the genome-scale assembly and annotation for A. phaeospermum, the first species in the genus Arthrinium to be subjected to whole genome sequencing. Copyright © 2019 Elsevier Inc. All rights reserved.


April 21, 2020

Generating amplicon reads for microbial community assessment with next-generation sequencing.

Marker gene amplicon sequencing is often preferred over whole genome sequencing for microbial community characterization, due to its lower cost while still enabling assessment of uncultivable organisms. This technique involves many experimental steps, each of which can be a source of errors and bias. We present an up-to-date overview of the whole experimental pipeline, from sampling to sequencing reads, and give information allowing for informed choices at each step of both planning and execution of a microbial community assessment study. When applicable, we also suggest ways of avoiding inherent pitfalls in amplicon sequencing. © 2019 The Society for Applied Microbiology.


April 21, 2020

Rational development of transformation in Clostridium thermocellum ATCC 27405 via complete methylome analysis and evasion of native restriction-modification systems.

A major barrier to both metabolic engineering and fundamental biological studies is the lack of genetic tools in most microorganisms. One example is Clostridium thermocellum ATCC 27405T, where genetic tools are not available to help validate decades of hypotheses. A significant barrier to DNA transformation is restriction-modification systems, which defend against foreign DNA methylated differently than the host. To determine the active restriction-modification systems in this strain, we performed complete methylome analysis via single-molecule, real-time sequencing to detect 6-methyladenine and 4-methylcytosine and the rarely used whole-genome bisulfite sequencing to detect 5-methylcytosine. Multiple active systems were identified, and corresponding DNA methyltransferases were expressed from the Escherichia coli chromosome to mimic the C. thermocellum methylome. Plasmid methylation was experimentally validated and successfully electroporated into C. thermocellum ATCC 27405. This combined approach enabled genetic modification of the C. thermocellum-type strain and acts as a blueprint for transformation of other non-model microorganisms.


April 21, 2020

Comparative genomics reveals unique wood-decay strategies and fruiting body development in the Schizophyllaceae.

Agaricomycetes are fruiting body-forming fungi that produce some of the most efficient enzyme systems to degrade wood. Despite decades-long interest in their biology, the evolution and functional diversity of both wood-decay and fruiting body formation are incompletely known. We performed comparative genomic and transcriptomic analyses of wood-decay and fruiting body development in Auriculariopsis ampla and Schizophyllum commune (Schizophyllaceae), species with secondarily simplified morphologies, an enigmatic wood-decay strategy and weak pathogenicity to woody plants. The plant cell wall-degrading enzyme repertoires of Schizophyllaceae are transitional between those of white rot species and less efficient wood-degraders such as brown rot or mycorrhizal fungi. Rich repertoires of suberinase and tannase genes were found in both species, with tannases restricted to Agaricomycetes that preferentially colonize bark-covered wood, suggesting potential complementation of their weaker wood-decaying abilities and adaptations to wood colonization through the bark. Fruiting body transcriptomes revealed a high rate of divergence in developmental gene expression, but also several genes with conserved expression patterns, including novel transcription factors and small-secreted proteins, some of the latter which might represent fruiting body effectors. Taken together, our analyses highlighted novel aspects of wood-decay and fruiting body development in an important family of mushroom-forming fungi. © 2019 The Authors. New Phytologist © 2019 New Phytologist Trust.


April 21, 2020

Transcriptional initiation of a small RNA, not R-loop stability, dictates the frequency of pilin antigenic variation in Neisseria gonorrhoeae.

Neisseria gonorrhoeae, the sole causative agent of gonorrhea, constitutively undergoes diversification of the Type IV pilus. Gene conversion occurs between one of the several donor silent copies located in distinct loci and the recipient pilE gene, encoding the major pilin subunit of the pilus. A guanine quadruplex (G4) DNA structure and a cis-acting sRNA (G4-sRNA) are located upstream of the pilE gene and both are required for pilin antigenic variation (Av). We show that the reduced sRNA transcription lowers pilin Av frequencies. Extended transcriptional elongation is not required for Av, since limiting the transcript to 32 nt allows for normal Av frequencies. Using chromatin immunoprecipitation (ChIP) assays, we show that cellular G4s are less abundant when sRNA transcription is lower. In addition, using ChIP, we demonstrate that the G4-sRNA forms a stable RNA:DNA hybrid (R-loop) with its template strand. However, modulating R-loop levels by controlling RNase HI expression does not alter G4 abundance quantified through ChIP. Since pilin Av frequencies were not altered when modulating R-loop levels by controlling RNase HI expression, we conclude that transcription of the sRNA is necessary, but stable R-loops are not required to promote pilin Av. © 2019 John Wiley & Sons Ltd.


April 21, 2020

How Genomics Is Changing What We Know About the Evolution and Genome of Bordetella pertussis.

The evolution of Bordetella pertussis from a common ancestor similar to Bordetella bronchiseptica has occurred through large-scale gene loss, inactivation and rearrangements, largely driven by the spread of insertion sequence element repeats throughout the genome. B. pertussis is widely considered to be monomorphic, and recent evolution of the B. pertussis genome appears to, at least in part, be driven by vaccine-based selection. Given the recent global resurgence of whooping cough despite the wide-spread use of vaccination, a more thorough understanding of B. pertussis genomics could be highly informative. In this chapter we discuss the evolution of B. pertussis, including how vaccination is changing the circulating B. pertussis population at the gene-level, and how new sequencing technologies are revealing previously unknown levels of inter- and intra-strain variation at the genome-level.


April 21, 2020

Early emergence of mcr-1-positive Enterobacteriaceae in gulls from Spain and Portugal.

We tested extended-spectrum ß-lactamase producing bacteria from wild gulls (Larus spp.) sampled in 2009 for the presence of mcr-1. We report the detection of mcr-1 and describe genome characteristics of four Escherichia coli and one Klebsiella pneumoniae isolate from Spain and Portugal that also exhibited colistin resistance. Results represent the earliest evidence for colistin-resistant bacteria in European wildlife.Published 2019. This article is a U.S. Government work and is in the public domain in the USA.


April 21, 2020

Cupriavidus sp. strain Ni-2 resistant to high concentration of nickel and its genes responsible for the tolerance by genome comparison.

The widespread use of metals influenced many researchers to examine the relationship between heavy metal toxicity and bacterial resistance. In this study, we have inoculated heavy metal-contaminated soil from Janghang region of South Korea in the nickel-containing media (20 mM Ni2+) for the enrichment. Among dozens of the colonies acquired from the several transfers and serial dilutions with the same concentrations of Ni, the strain Ni-2 was chosen for further studies. The isolates were identified for their phylogenetic affiliations using 16S rRNA gene analysis. The strain Ni-2 was close to Cupriavidus metallidurans and was found to be resistant to antibiotics of vancomycin, erythromycin, chloramphenicol, ampicillin, gentamicin, streptomycin, and kanamycin by disk diffusion method. Of the isolated strains, Ni-2 was sequenced for the whole genome, since the Ni-resistance seemed to be better than the other strains. From the genome sequence we have found that there was a total of 89 metal-resistance-related genes including 11 Ni-resistance genes, 41 heavy metal (As, Cd, Zn, Hg, Cu, and Co)-resistance genes, 22 cation-efflux genes, 4 metal pumping ATPase genes, and 11 metal transporter genes.


April 21, 2020

Paenibacillus albus sp. nov., a UV radiation-resistant bacterium isolated from soil in Korea.

A novel Gram-stain-positive, motile, white color and endospore-forming bacterium, designated 18JY67-1T, was isolated from soil in Jeju Island, Korea. The strain grow at 15-42 °C (optimum 30 °C) in R2A medium at pH (6.0-9.5) (optimum 7.5). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain 18JY67-1T formed a distinct lineage within the family Paenibacillaceae (order Bacillales, class Bacilli), and was closely related to Paenibacillus rhizoryzae (KP675984; 96.9% 16S rRNA gene sequence similarity). The major cellular fatty acids of the strain 18JY67-1T were C16:0 and anteiso-C15:0. The predominant respiratory quinones were MK-7. The major polar lipid was identified as diphosphatidylglycerol. On the basis of phenotypic, chemotaxonomic and genotypic properties clearly indicated that isolate 18JY67-1T represents a novel species within the genus Paenibacillus, for which the name Paenibacillus flavus sp. nov. is proposed. The type strain of Paenibacillus flavus is 18JY67-1T (=?KCTC 33959T =?JCM 33184T).


April 21, 2020

Complete genome sequence of Paracoccus sp. Arc7-R13, a silver nanoparticles synthesizing bacterium isolated from Arctic Ocean sediments

Paracoccus sp. Arc7-R13, a silver nanoparticles (AgNPs) synthesizing bacterium, was isolated from Arctic Ocean sediment. Here we describe the complete genome of Paracoccus sp. Arc7-R13. The complete genome contains 4,040,012?bp with 66.66?mol%?G?+?C content, including one circular chromosome of 3,231,929?bp (67.45?mol%?G?+?C content), and eight plasmids with length ranging from 24,536?bp to 199,685?bp. The genome contains 3835 protein-coding genes (CDSs), 49 tRNA genes, as well as 3 rRNA operons as 16S-23S-5S rRNA. Based on the gene annotation and Swiss-Prot analysis, a total of 15 genes belonging to 11 kinds, including silver exporting P-type ATPase (SilP), alkaline phosphatase, nitroreductase, thioredoxin reductase, NADPH dehydrogenase and glutathione peroxidase, might be related to the synthesis of AgNPs. Meanwhile, many additional genes associated with synthesis of AgNPs such as protein-disulfide isomerase, c-type cytochrome, glutathione synthase and dehydrogenase reductase were also identified.


April 21, 2020

Whole genome sequence of first Candida auris strain, isolated in Russia.

Candida auris is an emergent yeast pathogen, easily transmissible between patients and with high percent of multidrug resistant strains. Here we present a draft genome sequence of the first known Russian strain of C. auris, isolated from a case of candidemia. The strain clustered within South Asian C. auris clade and seemingly represented an independent event of dissemination from the original species range. Observed fluconazole resistance was probably due to F105L and K143R mutations in ERG11. © The Author(s) 2019. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology.


April 21, 2020

Updated assembly resource of Phytophthora ramorum Pr102 isolate incorporating long reads from PacBio sequencing.

The NA1 clonal lineage of Phytophthora ramorum is responsible for Sudden Oak Death, an epidemic that has devastated California’s coastal forest ecosystems. An NA1 isolate Pr102 derived from coast live oak in California was previously sequenced and reported with 65 Mb assembly containing 12 Mb gaps in 2576 scaffolds. Here we report an improved 70 Mb genome in 1512 scaffolds with 6752 bp gaps after incorporating PacBio P5-C3 longreads. This assembly contains 19494 gene models (average gene length 2515 bp) compared to 16134 genes (average gene length of 1673 bp) in the previous version. We predicted 29 new RXLRs and 76 new paralogs of a total 392 RXLRs from this assembly. We predicted 35 CRNs compared to 19 in earlier version with six paralogs. Our lncRNAs prediction identified 255 candidates. This new resource will be invaluable for future evolution studies on the invasive plant pathogen.


April 21, 2020

Comparative Genomic Analysis of Virulence, Antimicrobial Resistance, and Plasmid Profiles of Salmonella Dublin Isolated from Sick Cattle, Retail Beef, and Humans in the United States.

Salmonella enterica serovar Dublin is a host-adapted serotype associated with typhoidal disease in cattle. While rare in humans, it usually causes severe illness, including bacteremia. In the United States, Salmonella Dublin has become one of the most multidrug-resistant (MDR) serotypes. To understand the genetic elements that are associated with virulence and resistance, we sequenced 61 isolates of Salmonella Dublin (49 from sick cattle and 12 from retail beef) using the Illumina MiSeq and closed 5 genomes using the PacBio sequencing platform. Genomic data of eight human isolates were also downloaded from NCBI (National Center for Biotechnology Information) for comparative analysis. Fifteen Salmonella pathogenicity islands (SPIs) and a spv operon (spvRABCD), which encodes important virulence factors, were identified in all 69 (100%) isolates. The 15 SPIs were located on the chromosome of the 5 closed genomes, with each of these isolates also carrying 1 or 2 plasmids with sizes between 36 and 329?kb. Multiple antimicrobial resistance genes (ARGs), including blaCMY-2, blaTEM-1B, aadA12, aph(3′)-Ia, aph(3′)-Ic, strA, strB, floR, sul1, sul2, and tet(A), along with spv operons were identified on these plasmids. Comprehensive antimicrobial resistance genotypes were determined, including 17 genes encoding resistance to 5 different classes of antimicrobials, and mutations in the housekeeping gene (gyrA) associated with resistance or decreased susceptibility to fluoroquinolones. Together these data revealed that this panel of Salmonella Dublin commonly carried 15 SPIs, MDR/virulence plasmids, and ARGs against several classes of antimicrobials. Such genomic elements may make important contributions to the severity of disease and treatment failures in Salmonella Dublin infections in both humans and cattle.


April 21, 2020

Convergent horizontal gene transfer and cross-talk of mobile nucleic acids in parasitic plants.

Horizontal gene transfer (HGT), the movement and genomic integration of DNA across species boundaries, is commonly associated with bacteria and other microorganisms, but functional HGT (fHGT) is increasingly being recognized in heterotrophic parasitic plants that obtain their nutrients and water from their host plants through direct haustorial feeding. Here, in the holoparasitic stem parasite Cuscuta, we identify 108?transcribed and probably functional HGT events in Cuscuta campestris and related species, plus 42?additional regions with host-derived transposon, pseudogene and non-coding sequences. Surprisingly, 18?Cuscuta fHGTs were acquired from the same gene families by independent HGT events in Orobanchaceae parasites, and the majority are highly expressed in the haustorial feeding structures in both lineages. Convergent retention and expression of HGT sequences suggests an adaptive role for specific additional genes in parasite biology. Between 16 and 20 of the transcribed HGT events are inferred as ancestral in Cuscuta based on transcriptome sequences from species across the phylogenetic range of the genus, implicating fHGT in the successful radiation of Cuscuta parasites. Genome sequencing of C. campestris supports transfer of genomic DNA-rather than retroprocessed RNA-as the mechanism of fHGT. Many of the C. campestris genes horizontally acquired are also frequent sources of 24-nucleotide small RNAs that are typically associated with RNA-directed DNA methylation. One HGT encoding a leucine-rich repeat protein kinase overlaps with a microRNA that has been shown to regulate host gene expression, suggesting that HGT-derived parasite small RNAs may function in the parasite-host interaction. This study enriches our understanding of HGT by describing a parasite-host system with unprecedented gene exchange that points to convergent evolution of HGT events and the functional importance of horizontally transferred coding and non-coding sequences.


April 21, 2020

Phylogenetic reconciliation reveals the natural history of glycopeptide antibiotic biosynthesis and resistance.

Glycopeptide antibiotics are produced by Actinobacteria through biosynthetic gene clusters that include genes supporting their regulation, synthesis, export and resistance. The chemical and biosynthetic diversities of glycopeptides are the product of an intricate evolutionary history. Extracting this history from genome sequences is difficult as conservation of the individual components of these gene clusters is variable and each component can have a different trajectory. We show that glycopeptide biosynthesis and resistance in Actinobacteria maps to approximately 150-400 million years ago. Phylogenetic reconciliation reveals that the precursors of glycopeptide biosynthesis are far older than other components, implying that these clusters arose from a pre-existing pool of genes. We find that resistance appeared contemporaneously with biosynthetic genes, raising the possibility that the mechanism of action of glycopeptides was a driver of diversification in these gene clusters. Our results put antibiotic biosynthesis and resistance into an evolutionary context and can guide the future discovery of compounds possessing new mechanisms of action, which are especially needed as the usefulness of the antibiotics available at present is imperilled by human activity.


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