Protocol

Diagnostic Bacteriology

Volume 1616 of the series Methods in Molecular Biology pp 1-22

Date:

Whole-Genome Enrichment Using RNA Probes and Sequencing of Chlamydia trachomatis Directly from Clinical Samples

  • Amanda Claire BrownAffiliated withOxford Gene TechnologyDepartment of Microbiology and Immunology, Cornell University Email author 
  • , Mette T. ChristiansenAffiliated withUniversity College London

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Abstract

Whole-genome sequencing is a powerful, high-resolution tool that can be used to generate accurate data on bacterial population structure, phylogeography, and mutations associated with antimicrobial resistance. The ability to sequence pathogen genomes directly from clinical specimens, without the requirement for in vitro culturing, is attractive in terms of time- and labor-saving, especially in the case of slow-growing, or obligate intracellular pathogens, such as Chlamydia trachomatis. However clinical samples typically contain too low levels of pathogen nucleic acid, plus relatively high levels of human and natural microbiota DNA/RNA, to make this a viable option. Using a combination of whole-genome enrichment and deep sequencing, which has been proven to be a non-mutagenic approach, we can capture all known variations found within C. trachomatis genomes. The method is a consistent and sensitive tool that enables rapid whole-genome sequencing of C. trachomatis directly from clinical samples and has the potential to be adapted to other pathogens with a similar clonal nature.

Key words

Whole-genome enrichment Whole-genome sequencing Chlamydia trachomatis Clinical samples SureSelectXT