Whole-genome typing of Bacillus anthracis isolates by next-generation sequencing accurately and rapidly identifies strain-specific diagnostic polymorphisms

Cummings, Craig; Chung, Christina A. Bormann; Fang, Rixun; Barker, Melissa; Brzoska, Pius; Williamson, Phillip; Beaudry, Jodi A.; Matthews, Molly; Schupp, James M.; Wagner, David M.; Furtado, Manohar R.; Keim, Paul; Budowle, Bruce

doi:10.1016/j.fsigss.2009.08.097

Cited by 11 publications

(9 citation statements)

References 4 publications

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“…Although these draft genomes can have up to thousands of sequence gaps, the quality of the assembly is sufficient for automated annotation, and subsequent comparative genomics analyses, particularly when studying a conserved group of organisms such as the Listeria species examined here. Thus, NGS represents a feasible approach for rapid and comprehensive pathogen identification, subtyping, source tracking, and surveillance [ 62 ], and has the potential to be developed, in the long term, into routine diagnostic applications. The utility of draft genomes for identification of candidate vaccine targets has also been recently demonstrated [ 63 ].…”

Section: Discussionmentioning

confidence: 99%

Comparative genomics of the bacterial genus Listeria: Genome evolution is characterized by limited gene acquisition and limited gene loss

Cummings²,

et al. 2010

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BackgroundThe bacterial genus Listeria contains pathogenic and non-pathogenic species, including the pathogens L. monocytogenes and L. ivanovii, both of which carry homologous virulence gene clusters such as the prfA cluster and clusters of internalin genes. Initial evidence for multiple deletions of the prfA cluster during the evolution of Listeria indicates that this genus provides an interesting model for studying the evolution of virulence and also presents practical challenges with regard to definition of pathogenic strains.ResultsTo better understand genome evolution and evolution of virulence characteristics in Listeria, we used a next generation sequencing approach to generate draft genomes for seven strains representing Listeria species or clades for which genome sequences were not available. Comparative analyses of these draft genomes and six publicly available genomes, which together represent the main Listeria species, showed evidence for (i) a pangenome with 2,032 core and 2,918 accessory genes identified to date, (ii) a critical role of gene loss events in transition of Listeria species from facultative pathogen to saprotroph, even though a consistent pattern of gene loss seemed to be absent, and a number of isolates representing non-pathogenic species still carried some virulence associated genes, and (iii) divergence of modern pathogenic and non-pathogenic Listeria species and strains, most likely circa 47 million years ago, from a pathogenic common ancestor that contained key virulence genes.ConclusionsGenome evolution in Listeria involved limited gene loss and acquisition as supported by (i) a relatively high coverage of the predicted pan-genome by the observed pan-genome, (ii) conserved genome size (between 2.8 and 3.2 Mb), and (iii) a highly syntenic genome. Limited gene loss in Listeria did include loss of virulence associated genes, likely associated with multiple transitions to a saprotrophic lifestyle. The genus Listeria thus provides an example of a group of bacteria that appears to evolve through a loss of virulence rather than acquisition of virulence characteristics. While Listeria includes a number of species-like clades, many of these putative species include clades or strains with atypical virulence associated characteristics. This information will allow for the development of genetic and genomic criteria for pathogenic strains, including development of assays that specifically detect pathogenic Listeria strains.

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Section: Discussionmentioning

confidence: 99%

Comparative genomics of the bacterial genus Listeria: Genome evolution is characterized by limited gene acquisition and limited gene loss

Cummings²,

et al. 2010

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“…Null mutations in spo0F resulting in asporogenous phenotypes contribute to colony morphology variation in B. anthracis , B. thuringiensis and B. subtilis [53] , [54] , [55] . Enhanced in vitro “fitness” is also a likely driver behind the recovery of asporogenic B. anthracis mutants that were discovered during the investigation into the B. anthracis attacks of 2001 [56] . Because the process of sporulation is highly energy-intensive and irreversible once commenced, mutants that delay sporulation (or fail to sporulate altogether) to take advantage of remaining nutrients would out-compete wild-type cells during repeated passage in vitro in the absence of other selection pressures, as has been demonstrated in extended in vitro evolution studies with B. subtilis under relaxed sporulation conditions [57] .…”

Section: Discussionmentioning

confidence: 99%

“…Whole-genome approaches are becoming critical components of microbial forensics. The SNPs and indels identified in the analysis of evidentiary materials currently become the basis for higher-throughput assays to screen large numbers of samples [56] , [72] . Decreasing costs of whole-genome sequencing, and the comprehensive nature of the analysis, may make this the preferred method of forensic analysis of microbial samples in the future.…”

Section: Discussionmentioning

confidence: 99%

Genomic Signatures of Strain Selection and Enhancement in Bacillus atrophaeus var. globigii, a Historical Biowarfare Simulant

Gibbons¹,

Broomall²,

McNew

et al. 2011

PLoS ONE

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BackgroundDespite the decades-long use of Bacillus atrophaeus var. globigii (BG) as a simulant for biological warfare (BW) agents, knowledge of its genome composition is limited. Furthermore, the ability to differentiate signatures of deliberate adaptation and selection from natural variation is lacking for most bacterial agents. We characterized a lineage of BGwith a long history of use as a simulant for BW operations, focusing on classical bacteriological markers, metabolic profiling and whole-genome shotgun sequencing (WGS).ResultsArchival strains and two “present day” type strains were compared to simulant strains on different laboratory media. Several of the samples produced multiple colony morphotypes that differed from that of an archival isolate. To trace the microevolutionary history of these isolates, we obtained WGS data for several archival and present-day strains and morphotypes. Bacillus-wide phylogenetic analysis identified B. subtilis as the nearest neighbor to B. atrophaeus. The genome of B. atrophaeus is, on average, 86% identical to B. subtilis on the nucleotide level. WGS of variants revealed that several strains were mixed but highly related populations and uncovered a progressive accumulation of mutations among the “military” isolates. Metabolic profiling and microscopic examination of bacterial cultures revealed enhanced growth of “military” isolates on lactate-containing media, and showed that the “military” strains exhibited a hypersporulating phenotype.ConclusionsOur analysis revealed the genomic and phenotypic signatures of strain adaptation and deliberate selection for traits that were desirable in a simulant organism. Together, these results demonstrate the power of whole-genome and modern systems-level approaches to characterize microbial lineages to develop and validate forensic markers for strain discrimination and reveal signatures of deliberate adaptation.

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“…Next generation and third generation sequencing technologies can be expected to be more widely used in epidemiological investigations as costs decrease and speed, coverage and accuracy increase [28] , [31] – [33] . However, other molecular methods are needed to bridge the gap between vast amounts of data gathered from a small number of strains and fewer data from a large number of isolates.…”

Section: Discussionmentioning

confidence: 99%

Molecular Investigations of a Locally Acquired Case of Melioidosis in Southern AZ, USA

et al. 2011

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Melioidosis is caused by Burkholderia pseudomallei, a Gram-negative bacillus, primarily found in soils in Southeast Asia and northern Australia. A recent case of melioidosis in non-endemic Arizona was determined to be the result of locally acquired infection, as the patient had no travel history to endemic regions and no previous history of disease. Diagnosis of the case was confirmed through multiple microbiologic and molecular techniques. To enhance the epidemiological analysis, we conducted several molecular genotyping procedures, including multi-locus sequence typing, SNP-profiling, and whole genome sequence typing. Each technique has different molecular epidemiologic advantages, all of which provided evidence that the infecting strain was most similar to those found in Southeast Asia, possibly originating in, or around, Malaysia. Advancements in new typing technologies provide genotyping resolution not previously available to public health investigators, allowing for more accurate source identification.

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Whole-genome typing of Bacillus anthracis isolates by next-generation sequencing accurately and rapidly identifies strain-specific diagnostic polymorphisms

Cited by 11 publications

References 4 publications

Comparative genomics of the bacterial genus Listeria: Genome evolution is characterized by limited gene acquisition and limited gene loss

Comparative genomics of the bacterial genus Listeria: Genome evolution is characterized by limited gene acquisition and limited gene loss

Genomic Signatures of Strain Selection and Enhancement in Bacillus atrophaeus var. globigii, a Historical Biowarfare Simulant

Molecular Investigations of a Locally Acquired Case of Melioidosis in Southern AZ, USA

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