Phylogenetic Placement of Isolates Within the Trans-Eurasian Clade A.Br.008/009 of Bacillus anthracis

Antwerpen, Markus; Beyer, Wolfgang; Bassy, Olga; García, Ma V. Ortega; Ramos, JC Cabria; Grass, Gregor; Wölfel, Roman

doi:10.3390/microorganisms7120689

Cited by 14 publications

(25 citation statements)

References 29 publications

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“…Data Set 1 was aimed at (i) investigating the overall cgMLST scheme typeability (defined as the percentage of genes assigned allele numbers per strain) and (ii) investigating the backward compatibility of the new scheme to standard canonical SNP grouping to classify B. anthracis into major clades and subclades as previously defined and accepted by anthrax laboratories around the globe ( 20 ). This data set was investigated in previous studies ( 7 , 20 – 22 ) and includes sequence data of 596 B. anthracis strains that cover the global population diversity of B. anthracis and were recovered over the last 110 years from 50 countries on six continents (Table S1).…”

Section: Methodsmentioning

confidence: 99%

“…Standard genotyping methods for B. anthracis include canonical single-nucleotide polymorphisms (SNPs) and multilocus variable-number tandem repeat (VNTR) analysis (MLVA). Canonical SNP typing employs a selection of representative branch-specific SNPs for phylogenetic branches, which can be used to define key phylogenetic positions for descendant strains ( 7 – 9 ). Based on canonical SNP analysis ( 8 – 10 ), B. anthracis can be divided into three major lineages—A, B, and C—that subdivide into sublineages with a typical geographical distribution ( 10 ).…”

Section: Introductionmentioning

confidence: 99%

“…However, MLVA is laborious and prone to homoplasy problems ( 5 ). Today, whole-genome sequencing (WGS) is the method of choice to identify genome-wide SNPs in B. anthracis , which allows the definition of new canonical genetic lineages ( 7 , 15 ), but also greatly enhances the resolution of phylogenetic analyses in outbreak settings. Core-genome-based multilocus sequence typing (cgMLST) is a genome-wide typing system for a high-resolution clustering that indexes strain genotyping results into allelic numbers that can be accessed via a central database.…”

Section: Introductionmentioning

confidence: 99%

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A Whole-Genome-Based Gene-by-Gene Typing System for Standardized High-Resolution Strain Typing of Bacillus anthracis

et al. 2021

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Whole-genome-sequencing (WGS) has been established for bacterial subtyping and is regularly used to study pathogen transmission, to investigate outbreaks, and to perform routine surveillance. Core genome multilocus sequence typing (cgMLST) is a bacterial subtyping method that uses WGS data to provide a high resolution strain characterisation. This study aimed at developing a novel cgMLST scheme for Bacillus anthracis, a notorious pathogen that causes anthrax in livestock and humans worldwide. The scheme comprises 3,803 genes that were conserved in 57 B. anthracis genomes spanning the whole phylogeny. The scheme has been evaluated and applied to 584 genomes from 50 countries. On average, 99.5% of the cgMLST targets were detected. The cgMLST results confirmed the classical canonical Single-Nucleotide-Polymorphisms (SNPs) grouping of B. anthracis into major clades and subclades. Genetic distances calculated based on cgMLST were comparable to distances from whole-genome-based SNP analysis with similar phylogenetic topology and comparable discriminatory power. Additionally, the application of the cgMLST scheme to anthrax outbreaks from Germany and Italy led to a definition of a cut-off threshold of five allele differences to trace epidemiologically linked strains for cluster typing and transmission analysis. Finally, the association of two clusters of B. anthracis to human cases of injectional anthrax in four European countries was confirmed using cgMLST. In summary, this study presents a novel cgMLST scheme that provides a high-resolution strain genotyping for B. anthracis. This scheme can be used in parallel with SNP typing methods to facilitate rapid and harmonised inter-laboratory comparisons, essential for global surveillance and outbreak analysis. The scheme is publicly-available for application from users including those with little bioinformatics knowledge.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A Whole-Genome-Based Gene-by-Gene Typing System for Standardized High-Resolution Strain Typing of Bacillus anthracis

et al. 2021

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“…To formally validate the sensitivity of the 16S rRNA SNP-PCR assay, a panel of 14 different B. anthracis DNAs was employed. These B. anthracis strains represent all major branches A, B, and C [27], including prominent sub-branches [28] of the global B. anthracis phylogeny (Table S10). All DNAs produced positive PCR results.…”

Section: Sensitivity and Specificity Of The 16s Rrna Snp-pcr Assaymentioning

confidence: 99%

Ultrasensitive Detection of Bacillus anthracis by Real-Time PCR Targeting a Polymorphism in Multi-Copy 16S rRNA Genes and Their Transcripts

Braun

Nguyen

Walter

et al. 2021

IJMS

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The anthrax pathogen Bacillus anthracis poses a significant threat to human health. Identification of B. anthracis is challenging because of the bacterium’s close genetic relationship to other Bacillus cereus group species. Thus, molecular detection is founded on species-specific PCR targeting single-copy genes. Here, we validated a previously recognized multi-copy target, a species-specific single nucleotide polymorphism (SNP) present in 2–5 copies in every B. anthracis genome analyzed. For this, a hydrolysis probe-based real-time PCR assay was developed and rigorously tested. The assay was specific as only B. anthracis DNA yielded positive results, was linear over 9 log10 units, and was sensitive with a limit of detection (LoD) of 2.9 copies/reaction. Though not exhibiting a lower LoD than established single-copy PCR targets (dhp61 or PL3), the higher copy number of the B. anthracis–specific 16S rRNA gene alleles afforded ≤2 unit lower threshold (Ct) values. To push the detection limit even further, the assay was adapted for reverse transcription PCR on 16S rRNA transcripts. This RT-PCR assay was also linear over 9 log10 units and was sensitive with an LoD of 6.3 copies/reaction. In a dilution series of experiments, the 16S RT-PCR assay achieved a thousand-fold higher sensitivity than the DNA-targeting assays. For molecular diagnostics, we recommend a real-time RT-PCR assay variant in which both DNA and RNA serve as templates (thus, no requirement for DNase treatment). This can at least provide results equaling the DNA-based implementation if no RNA is present but is superior even at the lowest residual rRNA concentrations.

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“…The A clade is the most important, globally dispersed causative form of anthrax (>90% of all cases) [ 9 ]. Modern whole genome sequence single nucleotide polymorphism- (SNP-)analyses and the definition of derived canonical SNPs (canSNPs) are used to further subdivide these main clades into phylogeographical sub-branches [ 10–14 ].…”

Section: Introductionmentioning

confidence: 99%

Review: The risk of contracting anthrax from spore-contaminated soil – A military medical perspective

Finke¹,

Beyer

Loderstädt

et al. 2020

European Journal of Microbiology and Immunology

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AbstractAnthrax is an infectious disease of relevance for military forces. Although spores of Bacillus anthracis obiquitously occur in soil, reports on soil-borne transmission to humans are scarce. In this narrative review, the potential of soil-borne transmission of anthrax to humans is discussed based on pathogen-specific characteristics and reports on anthrax in the course of several centuries of warfare. In theory, anthrax foci can pose a potential risk of infection to animals and humans if sufficient amounts of virulent spores are present in the soil even after an extended period of time. In praxis, however, transmissions are usually due to contacts with animal products and reported events of soil-based transmissions are scarce. In the history of warfare, even in the trenches of World War I, reported anthrax cases due to soil-contaminated wounds are virtually absent. Both the perspectives and the experience of the Western hemisphere and of former Soviet Republics are presented. Based on the accessible data as provided in the review, the transmission risk of anthrax by infections of wounds due to spore-contaminated soil is considered as very low under the most circumstance. Active historic anthrax foci may, however, still pose a risk to the health of deployed soldiers.

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Phylogenetic Placement of Isolates Within the Trans-Eurasian Clade A.Br.008/009 of Bacillus anthracis

Cited by 14 publications

References 29 publications

A Whole-Genome-Based Gene-by-Gene Typing System for Standardized High-Resolution Strain Typing of Bacillus anthracis

A Whole-Genome-Based Gene-by-Gene Typing System for Standardized High-Resolution Strain Typing of Bacillus anthracis

Ultrasensitive Detection of Bacillus anthracis by Real-Time PCR Targeting a Polymorphism in Multi-Copy 16S rRNA Genes and Their Transcripts

Review: The risk of contracting anthrax from spore-contaminated soil – A military medical perspective

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