title: Screening ancient tuberculosis with qPCR: challenges and opportunities.
abstract: The field of ancient DNA (aDNA) has rapidly accelerated in recent years as a result of new methods in next-generation sequencing, library preparation and targeted enrichment. Such research is restricted, however, by the highly variable DNA preservation within different tissues, especially when isolating ancient pathogens from human remains. Identifying positive candidate samples via quantitative PCR (qPCR) for downstream procedures can reduce reagent costs, increase capture efficiency and maximize the number of sequencing reads of the target. This study uses four qPCR assays designed to target regions within the Mycobacterium tuberculosis complex (MTBC) to examine 133 human skeletal samples from a wide geographical and temporal range, identified by the presence of skeletal lesions typical of chronic disseminated tuberculosis. Given the inherent challenges working with ancient mycobacteria, strict criteria must be used and primer/probe design continually re-evaluated as new data from bacteria become available. Seven samples tested positive for multiple MTBC loci, supporting them as strong candidates for downstream analyses. Using strict and conservative criteria, qPCR remains a fast and effective screening tool when compared with screening by more expensive sequencing and enrichment technologies.
authors.affiliation: School of Human Evolution and Social Change, Arizona State University, Tempe, AZ, USA.
authors.forename: Kelly M
authors.identifier: http://orcid.org/0000-0002-0756-4250
authors.initials: KM
authors.lastname: Harkins
authors.affiliation: School of Human Evolution and Social Change, Arizona State University, Tempe, AZ, USA.
authors.forename: Jane E
authors.initials: JE
authors.lastname: Buikstra
authors.affiliation: Department of Archaeology, University of Cape Town, Cape Town, South Africa.
authors.forename: Tessa
authors.initials: T
authors.lastname: Campbell
authors.affiliation: Institute for Archaeological Sciences, University of Tübingen, Tübingen, Germany.
authors.forename: Kirsten I
authors.initials: KI
authors.lastname: Bos
authors.affiliation: School of Life Sciences, Arizona State University, Tempe, AZ, USA.
authors.forename: Eric D
authors.initials: ED
authors.lastname: Johnson
authors.affiliation: Institute for Archaeological Sciences, University of Tübingen, Tübingen, Germany.
authors.forename: Johannes
authors.initials: J
authors.lastname: Krause
authors.affiliation: School of Human Evolution and Social Change, Arizona State University, Tempe, AZ, USA acstone@asu.edu.
authors.forename: Anne C
authors.initials: AC
authors.lastname: Stone
mesh_terms: D001842:Bone and Bones / Q000737:chemistry*; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics* / Q000266:history; D005580:Fossils; D049690:History, Ancient; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D010164:Paleopathology / Q000379:methods* / Q000639:trends; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial
keywords: ancient DNA; bioarchaeology; qPCR; tuberculosis