Advances in the molecular detection of tuberculosis in pre-contact Andean South America

Abstract

Andean paleopathological research has significantly enhanced knowledge about the geographical distribution and evolution of tuberculosis (TB) in pre-Columbian South America. In this paper, we review the history and progress of research on ancient tuberculosis (TB) in the Andean region, focusing on the strengths and limitations of current approaches for the molecular detection of ancient pathogens, with special attention to TB. As a case study, we describe a molecular screening approach for the detection of ancient Mycobacterium tuberculosis in individuals from Late Intermediate Period (1000–1400 CE) contexts at the site of Huari, Peru. We evaluate 34 commingled human vertebrae and combine morphological assessments of pathology with high throughput sequencing and a non-selective approach to ancient pathogen DNA screening. Our method enabled the simultaneous detection of ancient M. tuberculosis DNA and an evaluation of the environmental microbial composition of each sample. Our results show that despite the dominance of environmental DNA, molecular signatures of M. tuberculosis were identified in eight vertebrae, six of which had no observable skeletal pathology classically associated tuberculosis infection. This screening approach will assist in the identification of candidate samples for downstream genomic analyses. The method permits higher resolution disease identification in cases where pathology may be absent, or where the archaeological context may necessitate a broad differential diagnosis based on morphology alone.

Introduction

Archaeological collections of Andean South America have been an important influence on methodological advances in the field of paleopathology. The environmental conditions of certain regions naturally promote the preservation of human remains, yielding an abundance of biological material including bone, teeth, hair, soft tissue and coprolites, all of which are rich resources for the study of past disease (Bos et al., 2014; Harkins et al., 2015; Salo et al., 1994; Verano, 1997). Pioneers of the field in the 19th and 20th centuries, such as Hrdlička and Moodie, took special interest in the anatomy, health, and disease of ancient Peruvians (Hrdlička, 1914; Moodie, 1923; Tello, 1909; Weiss et al., 1984), though their focus centered heavily on individual case studies. This early attention set the stage for the many historical and ethnographic descriptions of disease in the region (Hoyle et al., 1945; de Ayala, 1980; Tello and Williams, 1930; Urteaga-Ballon, 1991), which have yielded a wealth of information concerning the health of its past peoples.

In 1997, John Verano published Advances in the Paleopathology of Andean South America. In this work, he discussed the foundations of paleopathology in the Andean region and highlighted the advances research on Andean populations stimulated in this increasingly multidisciplinary field of investigation. In the 20 years following its publication, Andean collections have continued to play a central role in anthropological theory (Lozada and Tantaleán, 2019) and bioarchaeological methods (Blom et al., 2005; Klaus, 2014) as related to paleopathology, while promoting interdisciplinary collaboration through the incorporation of new technologies to better understand population health and disease in the past (Aufderheide et al., 2004; Bos et al., 2014; Wilbur and Buikstra, 2006).

Notable progress in paleopathology has come from the considerable contributions of genetic methods as applied to infectious disease detection and characterization in archaeological specimens, which now extend as far back as the Neolithic (Bos et al., 2014; Mühlemann et al., 2018; Salo et al., 1994; Schuenemann et al., 2018; Spyrou et al., 2018; Vågene et al., 2018). The recent introduction of high throughput sequencing greatly assists our quest to address medically and historically relevant questions. Genome-level analyses permit a glimpse at complex evolutionary history, thus yielding a better understanding of disease presence, geographic range, and ecology. The addition of genetic analyses also provide a means to move beyond morphologically detectable diseases to those that leave only molecular remnants in archaeological tissues such as infections of Salmonella enterica and hepatitis B virus (Mühlemann et al., 2018; Vågene et al., 2018). In the case of Andean populations, genome-level investigations of tuberculosis (TB) have altered our perceptions of ancient disease ecology, where the potential of zoonotic transmissions from marine mammals are now entering into discussions on disease origin in the region (Bos et al., 2014).

Here we explore how molecular analyses of archaeological material from the Andes are contributing to new and much needed standards in the detection of Mycobacterium tuberculosis DNA in ancient remains. We begin by describing the disease itself in a social and biological framework and then examine its known distribution in the Andes of South America by drawing upon previously published paleopathological studies. Finally, we explore the benefits of incorporating molecular data into morphology-based research on skeletal tuberculosis, along with a consideration of archaeological context to establish an interpretive framework. This is demonstrated via the presentation of TB screening data for individuals from Late Intermediate Period (LIP) contexts at the site of Huari, the former capital city of the Wari Empire, located in the Central Andes of Peru.