Accurate, rapid and high-throughput detection of strain-specific polymorphisms in Bacillus anthracis and Yersinia pestis by next-generation sequencing.

摘要: Background: In the event of biocrimes or infectious disease outbreaks, high-resolution genetic characterization for identifying agent and attributing it to a specific source can be crucial an effective response. Until recently, in-depth required expensive time-consuming Sanger sequencing few strains, followed by genotyping small number marker loci in panel isolates at gel-based approaches such as pulsed field gel electrophoresis, which necessity ignores most genome. Next-generation, massively parallel (MPS) technology (specifically Applied Biosystems oligonucleotide ligation detection (SOLiD™) system) is powerful investigative tool rapid, cost-effective microbial whole-genome characterization. Results: To demonstrate utility MPS typing monomorphic pathogens, four Bacillus anthracis Yersinia pestis strains were sequenced parallel. Reads aligned complete reference genomes, genomic variations identified. Resequencing B. Ames ancestor strain detected no false-positive single-nucleotide polymorphisms (SNPs), mapping reads Sterne correctly identified 98% 133 SNPs that are not clustered associated with repeats. Three geographically distinct from A branch lineage found have between 352 471 each, relative genome, one harbored amplification. Sequencing Y. Orientalis 20 54 per CO92 single Bolivian isolate having approximately twice many three more closely related North American strains. Coverage plotting also revealed common deletion two amplification appear due insertion element-mediated recombination events. Most private (that is, a, variant only this set) selected validation confirmed, although rare falsepositive variable nucleotide tandem Conclusions: The high-throughput, multiplexing capability, accuracy system make suitable rapid pathogens during forensic epidemiological investigation. By interrogating nearly every base reliably discovered, thus facilitating tracking strengthening attribution.