Browse
Submit Data
Databases
API
Help

Dataset Information

29 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Single-nucleotide repeat analysis for subtyping Bacillus anthracis isolates.

ABSTRACT: Single-nucleotide repeats (SNRs) are variable-number tandem repeats that display very high mutation rates. In an outbreak situation, the use of a marker system that exploits regions with very high mutation rates, such as SNRs, allows the differentiation of isolates with extremely low levels of genetic diversity. This report describes the identification and analysis of SNR loci of Bacillus anthracis. SNR loci were selected in silico, and the loci with the highest diversity were used to design and test locus-specific primers against a number of B. anthracis strains with the same multilocus variable-number tandem repeat analysis (MLVA) genotype. SNR markers that allowed strains with the same MLVA genotype to be differentiated from each other were identified. The resulting SNR marker system can be used as a molecular epidemiological tool in a natural outbreak or bioterrorism event, offering the best chance of distinguishing very closely related isolates.

SUBMITTER: Stratilo CW

PROVIDER: S-EPMC1393151 | biostudies-literature | 2006 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Single-nucleotide repeat analysis for subtyping Bacillus anthracis isolates.

Stratilo Chad W CW Lewis Christopher T CT Bryden Louis L Mulvey Michael R MR Bader Doug D

Journal of clinical microbiology 20060301 3

Single-nucleotide repeats (SNRs) are variable-number tandem repeats that display very high mutation rates. In an outbreak situation, the use of a marker system that exploits regions with very high mutation rates, such as SNRs, allows the differentiation of isolates with extremely low levels of genetic diversity. This report describes the identification and analysis of SNR loci of Bacillus anthracis. SNR loci were selected in silico, and the loci with the highest diversity were used to design and ...[more]

PMID: 16517854

Similar Datasets

Strain-specific single-nucleotide polymorphism assays for the Bacillus anthracis Ames strain.

Project description:Highly precise diagnostics and forensic assays can be developed through a combination of evolutionary analysis and the exhaustive examination of genomic sequences. In Bacillus anthracis, whole-genome sequencing efforts revealed ca. 3,500 single-nucleotide polymorphisms (SNPs) among eight different strains and evolutionary analysis provides the identification of canonical SNPs. We have previously shown that SNPs are highly evolutionarily stable, and the clonal nature of B. anthracis makes them ideal signatures for subtyping this pathogen. Here we identified SNPs that define the lineage of B. anthracis that contains the Ames strain, the strain used in the 2001 bioterrorist attacks in the United States. Sequencing and real-time PCR were used to validate these SNPs across B. anthracis strains, including (i) 88 globally and genetically diverse isolates; (ii) isolates that were shown to be genetic relatives of the Ames strain by multiple-locus variable number tandem repeat analysis (MLVA); and (iii) several different lab stocks of the Ames strain, including a clinical isolate from the 2001 letter attack. Six SNPs were found to be highly specific for the Ames strain; four on the chromosome, one on the pX01 plasmid, and one on the pX02 plasmid. All six SNPs differentiated the B. anthracis Ames strain from the 88 unique B. anthracis strains, while five of the six separated Ames from its close genetic relatives. The use of these SNPs coupled with real-time PCR allows specific and sensitive (<100 fg of template DNA) identification of the Ames strain. This evolutionary and genomics-based approach provides an effective means for the discovery of strain-specific SNPs in B. anthracis.

| S-EPMC1828967 | biostudies-literature

Pathogenomic sequence analysis of Bacillus cereus and Bacillus thuringiensis isolates closely related to Bacillus anthracis.

Project description:Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis are closely related gram-positive, spore-forming bacteria of the B. cereus sensu lato group. While independently derived strains of B. anthracis reveal conspicuous sequence homogeneity, environmental isolates of B. cereus and B. thuringiensis exhibit extensive genetic diversity. Here we report the sequencing and comparative analysis of the genomes of two members of the B. cereus group, B. thuringiensis 97-27 subsp. konkukian serotype H34, isolated from a necrotic human wound, and B. cereus E33L, which was isolated from a swab of a zebra carcass in Namibia. These two strains, when analyzed by amplified fragment length polymorphism within a collection of over 300 of B. cereus, B. thuringiensis, and B. anthracis isolates, appear closely related to B. anthracis. The B. cereus E33L isolate appears to be the nearest relative to B. anthracis identified thus far. Whole-genome sequencing of B. thuringiensis 97-27and B. cereus E33L was undertaken to identify shared and unique genes among these isolates in comparison to the genomes of pathogenic strains B. anthracis Ames and B. cereus G9241 and nonpathogenic strains B. cereus ATCC 10987 and B. cereus ATCC 14579. Comparison of these genomes revealed differences in terms of virulence, metabolic competence, structural components, and regulatory mechanisms.

| S-EPMC1447445 | biostudies-literature

Multilocus genotyping assays for single nucleotide polymorphism-based subtyping of Listeria monocytogenes isolates.

Project description:Listeria monocytogenes is responsible for serious invasive illness associated with consumption of contaminated food and places a significant burden on public health and the agricultural economy. We recently developed a multilocus genotyping (MLGT) assay for high-throughput subtype determination of L. monocytogenes lineage I isolates based on interrogation of single nucleotide polymorphisms (SNPs) via multiplexed primer extension reactions. Here we report the development and validation of two additional MLGT assays that address the need for comprehensive DNA sequence-based subtyping of L. monocytogenes. The first of these novel MLGT assays targeted variation segregating within lineage II, while the second assay combined probes for lineage III strains with probes for strains representing a recently characterized fourth evolutionary lineage (IV) of L. monocytogenes. These assays were based on nucleotide variation identified in >3.8 Mb of comparative DNA sequence and consisted of 115 total probes that differentiated 93% of the 100 haplotypes defined by the multilocus sequence data. MLGT reproducibly typed the 173 isolates used in SNP discovery, and the 10,448 genotypes derived from MLGT analysis of these isolates were consistent with DNA sequence data. Application of the MLGT assays to assess subtype prevalence among isolates from ready-to-eat foods and food-processing facilities indicated a low frequency (6.3%) of epidemic clone subtypes and a substantial population of isolates (>30%) harboring mutations in inlA associated with attenuated virulence in cell culture and animal models. These mutations were restricted to serogroup 1/2 isolates, which may explain the overrepresentation of serotype 4b isolates in human listeriosis cases.

| S-EPMC2607178 | biostudies-literature

Phylogenetic discovery bias in Bacillus anthracis using single-nucleotide polymorphisms from whole-genome sequencing.

Project description:Phylogenetic reconstruction using molecular data is often subject to homoplasy, leading to inaccurate conclusions about phylogenetic relationships among operational taxonomic units. Compared with other molecular markers, single-nucleotide polymorphisms (SNPs) exhibit extremely low mutation rates, making them rare in recently emerged pathogens, but they are less prone to homoplasy and thus extremely valuable for phylogenetic analyses. Despite their phylogenetic potential, ascertainment bias occurs when SNP characters are discovered through biased taxonomic sampling; by using whole-genome comparisons of five diverse strains of Bacillus anthracis to facilitate SNP discovery, we show that only polymorphisms lying along the evolutionary pathway between reference strains will be observed. We illustrate this in theoretical and simulated data sets in which complex phylogenetic topologies are reduced to linear evolutionary models. Using a set of 990 SNP markers, we also show how divergent branches in our topologies collapse to single points but provide accurate information on internodal distances and points of origin for ancestral clades. These data allowed us to determine the ancestral root of B. anthracis, showing that it lies closer to a newly described "C" branch than to either of two previously described "A" or "B" branches. In addition, subclade rooting of the C branch revealed unequal evolutionary rates that seem to be correlated with ecological parameters and strain attributes. Our use of nonhomoplastic whole-genome SNP characters allows branch points and clade membership to be estimated with great precision, providing greater insight into epidemiological, ecological, and forensic questions.

| S-EPMC518758 | biostudies-literature

Microarray-based resequencing of multiple Bacillus anthracis isolates.

Project description:We used custom-designed resequencing arrays to generate 3.1 Mb of genomic sequence from a panel of 56 Bacillus anthracis strains. Sequence quality was shown to be very high by replication (discrepancy rate of 7.4 x 10(-7)) and by comparison to independently generated shotgun sequence (discrepancy rate < 2.5 x 10(-6)). Population genomics studies of microbial pathogens using rapid resequencing technologies such as resequencing arrays are critical for recognizing newly emerging or genetically engineered strains.

| S-EPMC549062 | biostudies-literature

Use of single nucleotide polymorphisms in the plcR gene for specific identification of Bacillus anthracis.

Project description:A TaqMan-minor groove binding assay designed around a nonsense mutation in the plcR gene was used to genotype Bacillus anthracis, B. cereus, and B. thuringiensis isolates. The assay differentiated B. anthracis from these genetic near-neighbors and determined that the nonsense mutation is ubiquitous across 89 globally and genetically diverse B. anthracis strains.

| S-EPMC1081367 | biostudies-literature

A single-nucleotide-polymorphism-based multilocus genotyping assay for subtyping lineage I isolates of Listeria monocytogenes.

Project description:Listeria monocytogenes is a facultative intracellular pathogen responsible for food-borne disease with high mortality rates in humans and is the leading microbiological cause of food recalls. Lineage I isolates of L. monocytogenes are a particular public health concern because they are responsible for most sporadic cases of listeriosis and the vast majority of epidemic outbreaks. Rapid, reproducible, and sensitive methods for differentiating pathogens below the species level are required for effective pathogen control programs, and the CDC PulseNet Task Force has called for the development and validation of DNA sequence-based methods for subtyping food-borne pathogens. Therefore, we developed a multilocus genotyping (MLGT) assay for L. monocytogenes lineage I isolates based on nucleotide variation identified by sequencing 23,251 bp of DNA from 22 genes distributed across seven genomic regions in 65 L. monocytogenes isolates. This single-well assay of 60 allele-specific probes captured 100% of the haplotype information contained in approximately 1.5 Mb of comparative DNA sequence and was used to reproducibly type a total of 241 lineage I isolates. The MLGT assay provided high discriminatory power (Simpson's index value, 0.91), uniquely identified isolates from the eight listeriosis outbreaks examined, and differentiated serotypes 1/2b and 4b as well as epidemic clone I (ECI), ECIa, and ECII. In addition, the assay included probes for a previously characterized truncation mutation in inlA, providing for the identification of a specific virulence-attenuated subtype. These results demonstrate that MLGT represents a significant new tool for use in pathogen surveillance, outbreak detection, risk assessment, population analyses, and epidemiological investigations. DNA sequences were deposited in the GenBank database under accession numbers DQ 812146 to DQ 812517, DQ 843664 to DQ 844598, and AY 512391 to AY 512502.

| S-EPMC1797101 | biostudies-literature

Molecular subtyping of Bacillus anthracis and the 2001 bioterrorism-associated anthrax outbreak, United States.

Project description:Molecular subtyping of Bacillus anthracis played an important role in differentiating and identifying strains during the 2001 bioterrorism-associated outbreak. Because B. anthracis has a low level of genetic variability, only a few subtyping methods, with varying reliability, exist. We initially used multiple-locus variable-number tandem repeat analysis (MLVA) to subtype 135 B. anthracis isolates associated with the outbreak. All isolates were determined to be of genotype 62, the same as the Ames strain used in laboratories. We sequenced the protective antigen gene (pagA) from 42 representative outbreak isolates and determined they all had a pagA sequence indistinguishable from the Ames strain (PA genotype I). MLVA and pagA sequencing were also used on DNA from clinical specimens, making subtyping B. anthracis possible without an isolate. Use of high-resolution molecular subtyping determined that all outbreak isolates were indistinguishable by the methods used and probably originated from a single source. In addition, subtyping rapidly identified laboratory contaminants and nonoutbreak-related isolates.

| S-EPMC2730295 | biostudies-literature

Rapid Identification of Bacillus anthracis <i>In Silico</i> and On-Site Using Novel Single-Nucleotide Polymorphisms.

Project description:Bacillus anthracis is a spore-forming bacterium that causes life-threatening infections in animals and humans and has been used as a bioterror agent. Rapid and reliable detection and identification of B. anthracis are of primary interest for both medical and biological threat-surveillance purposes. Few chromosomal sequences provide enough polymorphisms to clearly distinguish B. anthracis from closely related species. We analyzed 18 loci of the chromosome of B. anthracis and discovered eight novel single-nucleotide polymorphism (SNP) sites that can be used for the specific identification of B. anthracis. Using these SNP sites, we developed software-named AGILE V1.1 (anthracis genome-based identification with high-fidelity E-probe)-for easy, user-friendly identification of B. anthracis from whole-genome sequences. We also developed a recombinase polymerase amplification-Cas12a-based method that uses nucleic acid extracts for the specific, rapid, in-the-field identification of B. anthracis based on these SNPs. Via this method and B. anthracis-specific CRISPR RNAs for the target CR5_2, CR5_1, and Ba813 SNPs, we clearly detected 5 aM genomic DNA. This study provides two simple and reliable methods suitable for use in local hospitals and public health programs for the detection of B. anthracis. IMPORTANCE Bacillus anthracis is the etiologic agent of anthrax, a fatal disease and a potential biothreat. A specific, accurate, and rapid method is urgently required for the identification of B. anthracis. We demonstrate the potential of using eight novel SNPs for the rapid and accurate detection of B. anthracis via in silico and laboratory-based testing methods. Our findings have important implications for public health responses to disease outbreaks and bioterrorism threats.

| S-EPMC9241702 | biostudies-literature

Characterization of Bacillus cereus isolates associated with fatal pneumonias: strains are closely related to Bacillus anthracis and harbor B. anthracis virulence genes.

Project description:Bacillus cereus is ubiquitous in nature, and while most isolates appear to be harmless, some are associated with food-borne illnesses, periodontal diseases, and other more serious infections. In one such infection, B. cereus G9241 was identified as the causative agent of a severe pneumonia in a Louisiana welder in 1994. This isolate was found to harbor most of the B. anthracis virulence plasmid pXO1 (13). Here we report the characterization of two clinical and one environmental B. cereus isolate collected during an investigation of two fatal pneumonia cases in Texas metal workers. Molecular subtyping revealed that the two cases were not caused by the same strain. However, one of the three isolates was indistinguishable from B. cereus G9241. PCR analysis demonstrated that both clinical isolates contained B. anthracis pXO1 toxin genes. One clinical isolate and the environmental isolate collected from that victim's worksite contained the cap A, B, and C genes required for capsule biosynthesis in B. anthracis. Both clinical isolates expressed a capsule; however, neither was composed of poly-D-glutamic acid. Although most B. cereus isolates are not opportunistic pathogens and only a limited number cause food-borne illnesses, these results demonstrate that some B. cereus strains can cause severe and even fatal infections in patients who appear to be otherwise healthy.

| S-EPMC1594744 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data