The proposed REU supplement would support research by Meagan Rubel, an undergraduate anthropology major at the Arizona State University. In addition to learning laboratory techniques such as ancient DNA extraction and quantitative PCR (qPCR), Meagan will learn to design qPCR assays in order to assess the type(s) of mycobacterial DNA in ancient skeletal materials from the medieval English Spitalfields cemetery, a skeletal collection with known tuberculosis status for many of the individuals. The primary goal of this project is to integrate information from the ancient and historic past into an understanding of the evolution of M. tuberculosis and how it relates to human migration and other population events. We have collected 237 skeletal samples with evidence of TB from a wide temporal and geographic range. Analyses to date have focused on New World pre-Age of Exploration samples and are still underway (we plan to request a no-cost extension shortly). The Spitalfields sample, however, dates to 1200 -1539 A.D. It is a known TB cemetery and spans time before and during the Age of Exploration. After assessing DNA preservation in the samples, we are selecting a subset for further analysis using high-throughput sequencing (454 sequencing) to address several hypotheses about the evolutionary history and biogeography of tuberculosis. Specifically, DNA fragments that are phylogenetically informative for modern strains within M. tuberculosis and between members of the Mycobacterium tuberculosis complex (MTBC), which includes M. tuberculosis, M. bovis, M. microti, M. africanum and M. canettii, and other mycobacterial species will be used to address the following hypotheses: Prehistoric TB in the Americas was caused by M. tuberculosis strains that are most closely related to modern strains found in Asia. New strains of M. tuberculosis were introduced into the Americas at European contact. These strains were most closely related to those prevalent today in Europe. The increase in prevalence of tuberculosis in Europe during the 17th and 18th centuries was due to new strains that were the same as those introduced into the Americas. The proposed REU supplement will contribute to the project in three ways. First, we will be able to assess the level of M. tuberculosis DNA present in the Spitalfields skeletal remains. This will allow us to determine which samples are candidates for 454 sequencing analyses. A large amount of DNA (1-5 micrograms) is necessary for massively parallel sequencing, and this typically can only be obtained through multiple extractions of a large amount of bone. Many of our samples are too small for massively parallel sequencing, but still indicate preservation of ancient MTBC DNA. Further, massively parallel sequencing is sufficiently expensive that we can only process a relatively small number of samples. Thus, a second important contribution of the proposed REU supplement would be the development of qPCR assays capable of determining the lineage of ancient M. tuberculosis with a much smaller amount of DNA. In this way, we maximize the genetic information obtained from all of our ancient samples, which will be phylogenetically informative, albeit to a somewhat lesser degree than the strain-level information from full sequences. Finally, strain and/or lineage information obtained from the Spitalfields cemetery will provide information that allows us to test our hypotheses that TB strains prevalent in NW Europe during the Age of Exploration were different than those prevalent in the New World at this time.
|Effective start/end date||8/1/06 → 7/31/11|
- National Science Foundation (NSF): $271,355.00
High-Throughput Nucleotide Sequencing
Polymerase Chain Reaction
Costs and Cost Analysis
Bone and Bones