Sediment

DNA in sediment (Figure 1a) is often from small fragments of bone and feces, with additional DNA from urine, hair and skin, plant matter, and other discarded organic material (Massilani et al. Reference Massilani, Morley, Mentzer, Aldeias, Vernot, Miller and Stahlschmidt2022; Pedersen et al. Reference Pedersen, Overballe-Petersen, Ermini, Sarkissian, Haile, Hellstrom and Spens2015). DNA from skeletal material may also diffuse into surrounding sediments (Sarhan et al. Reference Sarhan, Lehmkuhl, Straub, Tett, Wieland, Francken, Zink and Maixner2021). This results in sediment containing aDNA that likely represents the floral and faunal environment at the time the sediment was created, with traces of human aDNA depending on context. First analyzed archaeologically by Willerslev and colleagues (Reference Willerslev, Hansen, Binladen, Brand, Gilbert, Shapiro, Bunce, Wiuf, Gilichinsky and Cooper2003), sediment aDNA (sedaDNA) has been used primarily in environmental reconstruction. This includes reconstructing floral communities at a single point in time (Gugerli et al. Reference Gugerli, Alvarez and Tinner2013; Jørgensen, Haile, et al. Reference Jørgensen, Haile, Moller, Andreev, Boessenkool, Rasmussen and Kienast2012; Jørgensen, Kjaer, et al. Reference Jørgensen, Kjaer, Haile, Rasmussen, Boessenkool, Andersen and Coissac2012; Parducci et al. Reference Parducci, Bennett, Ficetola, Alsos, Suyama, Wood and Pedersen2017) and correlating floral, faunal, and environmental shifts (Anderson-Carpenter et al. Reference Anderson-Carpenter, McLachlan, Jackson, Kuch, Lumibao and Poinar2011; Andresen et al. Reference Andresen, Björck, Bennike and Bond2004; Birks and Birks Reference Birks and Birks2016; Epp et al. Reference Epp, Gussarova, Boessenkool, Olsen, Haile, Schroder-Nielsen and Ludikova2015; Seersholm et al. Reference Seersholm, Werndly, Grealy, Johnson, Early, Lundelius and Winsborough2020). Sediment has also been used to look for faunal traces to determine what kinds of animal resources a group may have used (Haile et al. Reference Haile, Holdaway, Oliver, Bunce, Gilbert, Nielsen, Munch, Ho, Shapiro and Willerslev2007; Hebsgaard et al. Reference Hebsgaard, Thomas P, Gilbert, Heyn, Allentoft, Bunce, Munch, Schweger and Willerslev2009; Seersholm et al. Reference Seersholm, Pedersen, Soe, Shokry, Mak, Ruter and Raghavan2016; Willerslev et al. Reference Willerslev, Hansen, Binladen, Brand, Gilbert, Shapiro, Bunce, Wiuf, Gilichinsky and Cooper2003) and to study extinct species and the timing and circumstances of their extinction (Graham et al. Reference Graham, Belmecheri, Choy, Culleton, Davies, Froese and Heintzman2016; Willerslev et al. Reference Willerslev, Hansen, Binladen, Brand, Gilbert, Shapiro, Bunce, Wiuf, Gilichinsky and Cooper2003). SedaDNA from burial contexts has been used to study the genetics of fossil hominins and humans, along with various fauna (Gelabert et al. Reference Gelabert, Sawyer, Bergström, Margaryan, Collin, Meshveliani and Belfer-Cohen2021; Sarhan et al. Reference Sarhan, Lehmkuhl, Straub, Tett, Wieland, Francken, Zink and Maixner2021; Slon et al. Reference Slon, Hopfe, Weiß, Mafessoni, De La Rasilla, Lalueza-Fox and Rosas2017; Vernot et al. Reference Vernot, Zavala, Gómez-Olivencia, Jacobs, Slon, Mafessoni and Romagné2021).

Sediment is abundant; it is ubiquitous to archaeological sites, resampling is possible, and very small amounts are needed for genetic analysis. Sediment is commonly collected during excavation and survey, and sediment can easily be subsampled for use in both destructive and nondestructive analyses. Sediment's ability to act as a proxy is particularly useful, because it allows for the study of human DNA without necessitating the destruction of Ancestral remains. SedaDNA research can be placed into two broad contexts: (1) environmental studies focusing on flora and nonhuman fauna and (2) using sediment as a proxy for Ancestors and other culturally significant organisms. When used for the former, sediment collected from areas with no known human occupation, cultural activity, or cultural significance at any time point is unlikely to contain endogenous human DNA. An example is the sampling and genetic analysis of lake sediments to reconstruct past environments (Anderson-Carpenter et al. Reference Anderson-Carpenter, McLachlan, Jackson, Kuch, Lumibao and Poinar2011; Birks and Birks Reference Birks and Birks2016). Although there is little cause for concern with these kinds of studies given that there is no human association, researchers should be cognizant of whether any of the flora and fauna they detect have cultural significance to the traditional custodians of the land. If so, additional considerations akin to those when working with human DNA may be needed. If sediment is collected from sites with known human occupations, human DNA may be recovered. This is especially true of sites with burials, and human DNA may also be present in layers devoid of visible Ancestral remains. Although sediment can be an excellent alternative to sampling Ancestral remains due to its abundance, human DNA yields from sediment will likely be lower than those gained from sampling Ancestral remains (Sarhan et al. Reference Sarhan, Lehmkuhl, Straub, Tett, Wieland, Francken, Zink and Maixner2021). SedaDNA context can be further complicated by DNA leeching from higher stratigraphic layers (Haile et al. Reference Haile, Holdaway, Oliver, Bunce, Gilbert, Nielsen, Munch, Ho, Shapiro and Willerslev2007). Recent work has shown that genetically screening sediment from throughout a profile to identify specific taxa and then targeting those taxa from microfeatures could mitigate the effects of leeching (Massilani et al. Reference Massilani, Morley, Mentzer, Aldeias, Vernot, Miller and Stahlschmidt2022). Alternatively, knowledge of a site's formation and assemblages can be used in conjunction with sedaDNA to reconstruct chronologies.

Coprolites

DNA in coprolites (Figure 1b) primarily comes from gut microbes, dietary elements, and the defecator (Rose et al. Reference Rose, Parker, Jefferson and Cartmell2015). The aDNA in coprolites provides the identity of the depositing organism and its gut microbiome, along with what it consumed on a given day. Coprolites, which were first genetically analyzed by Poinar and colleagues (Reference Poinar, Hofreiter, Geoffrey Spaulding, Martin, Artur Stankiewicz, Bland, Evershed, Possnert and Paabo1998), are often used to discuss the defecator and its diet. The floral and faunal components of coprolites are useful for reconstructing past diets in both human and nonhuman animals (Boast et al. Reference Boast, Weyrich, Wood, Metcalf, Knight and Cooper2018; Gilbert et al. Reference Gilbert, Jenkins, Götherstrom, Naveran, Sanchez, Hofreiter and Thomsen2008; Poinar et al. Reference Poinar, Kuch, Sobolik, Barnes, Stankiewicz, Kuder, Geofferey Spaulding, Bryant, Cooper and Paabo2001; Wood et al. Reference Wood, Rawlence, Rogers, Austin, Worthy and Cooper2008; Wood, Wilmshurst, Wagstaff et al. Reference Wood, Wilmshurst, Wagstaff, Worthy, Rawlence and Cooper2012; Wood, Wilmshurst, Worthy et al. Reference Wood, Wilmshurst, Worthy and Cooper2012; Wood et al. Reference Wood, Wilmshurst, Richardson, Rawlence, Wagstaff, Worthy and Cooper2013), and the microbial DNA has been used to study the gut microbiome (Lugli et al. Reference Lugli, Milani, Mancabelli, Turroni, Ferrario, Duranti, van Sinderen and Ventura2017; Santiago-Rodriguez et al. Reference Santiago-Rodriguez, Fornaciari, Luciani, Toranzos, Marota, Giuffra and Cano2017; Tito et al. Reference Tito, Knights, Metcalf, Obregon-Tito, Cleeland, Najar and Roe2012; Wibowo et al. Reference Wibowo, Yang, Borry, Hübner, Huang, Tierney and Zimmerman2021). Several researchers have studied defecator phylogenies and movements across a landscape (Botella et al. Reference Botella, Afonso Vargas, de la Rosa, Leles, Reimers, Vicente and Iniguez2010; Gilbert et al. Reference Gilbert, Jenkins, Götherstrom, Naveran, Sanchez, Hofreiter and Thomsen2008; Karpinski et al. Reference Karpinski, Mead and Poinar2017; Poinar et al. Reference Poinar, Kuch, McDonald, Martin and Paabo2003).

Unlike sediment, coprolites are not common to all archaeological sites and, when found, are present in varying amounts and differing states of preservation. Potential human coprolites are often found in middens or cesspits (Shillito et al. Reference Shillito, Blong, Green and Van Asperen2020) and may not be collected given that they can be difficult to distinguish from the surrounding sediment. Additionally, human coprolites can be similar in size, shape, and color to the coprolites of nonhuman animals such as canines, making identifications difficult without additional analysis. Although coprolites are primarily a source of dietary and environmental information, they can sometimes act as proxies for their depositing organism. Two broad categories of coprolite analysis are (1) analyses of nonhuman coprolites and (2) analyses of human coprolites. Regardless of their source, coprolites are finite. As with any archaeological or natural resource, sampling should only be done when there are either enough coprolites to leave a portion of the assemblage unanalyzed or when the coprolites are large enough that any sampling would not result in the destruction of the complete coprolite. An example of a large coprolite assemblage is that of Hind's Cave, Texas, where hundreds of coprolites were recovered and only a fraction analyzed (Dean Reference Dean2006), leaving most coprolites intact. As for coprolite size, fecal material weighing as little as 5 g can be used in a variety of macro, micro, and molecular analyses while still maintaining a voucher sample (Wood and Wilmshurst Reference Wood and Wilmshurst2016). Having one to a few coprolites at a site does not mean research cannot be conducted on them, but more care is needed to ensure that multiproxy analyses can take place.

An important consideration with coprolite analysis is that identifying the defecator may require some form of analysis, especially when distinguishing human from canine coprolites at North American sites. If coprolites come from environmental sites or are clearly nonhuman, such as the occasional mislabeled owl pellet, they can be considered as environmental traces. However, as a default, unknown coprolites recovered from archaeological sites should be considered human until additional analyses prove otherwise. In coprolite analysis, this requires either engaging in traditional identification methods by looking at the contents and rehydration liquid (Fry Reference Fry, Gilbert and Mielke1985; Reinhard and Bryant Reference Reinhard and Bryant1992) or doing genetic analyses (Borry et al. Reference Borry, Cordova, Perri, Wibowo, Honap, Ko and Yu2020; Knights et al. Reference Knights, Kuczynski, Charlson, Zaneveld, Mozer, Collman, Bushman, Knight and Kelley2011; Poinar et al. Reference Poinar, Kuch, McDonald, Martin and Paabo2003). As with sediment, preservation of human and other DNA may not be as good as in skeletal material. Even coprolites confirmed as human are not guaranteed to contain analyzable human DNA.

Birch Pitch and Other Chewed Materials

Birch pitch (for a similar material, see Figure 1c), an adhesive substance used for tasks including hafting, waterproofing, and mending vessels, is a more recent subject of DNA analysis. Small amounts of birch pitch can be common at European archaeological sites (Jensen et al. Reference Jensen, Niemann, Iversen, Fotakis, Gopalakrishnan, Vågene and Pedersen2019; Kashuba et al. Reference Kashuba, Kırdök, Damlien, Manninen, Nordqvist, Persson and Götherström2019; Mazza et al. Reference Mazza, Martini, Sala, Magi, Colombini, Giachi, Landucci, Lemorini, Modugno and Ribechini2006; Ottoni et al. Reference Ottoni, Borić, Cheronet, Sparacello, Dori, Coppa and Antonović2021; Rageot et al. Reference Rageot, Lepère, Henry, Binder, Davtian, Filippi and Fernandez2021; Sykes Reference Sykes, Coward, Hosfield, Pope and Wenban-Smith2015). Although birch pitch is not found in North American archaeological sites, other plant pitches, adhesives, and gums are found that could contain similar materials and genetic traces as birch pitch (Fox et al. Reference Fox, Heron and Sutton1995; Langejans et al. Reference Langejans, Aleo, Fajardo, Kozowyk and Aldenderfer2022). Pitches can be found with tooth and tool marks and, in some cases, fingerprints (Aveling and Heron Reference Aveling and Heron1999; Kashuba et al. Reference Kashuba, Kırdök, Damlien, Manninen, Nordqvist, Persson and Götherström2019; Sykes Reference Sykes, Coward, Hosfield, Pope and Wenban-Smith2015). Plant pitches that were chewed before use can contain DNA from humans, their diet, and their oral microbiome (Jensen et al. Reference Jensen, Niemann, Iversen, Fotakis, Gopalakrishnan, Vågene and Pedersen2019; Kashuba et al. Reference Kashuba, Kırdök, Damlien, Manninen, Nordqvist, Persson and Götherström2019; Lawton Reference Lawton2021; Ottoni et al. Reference Ottoni, Borić, Cheronet, Sparacello, Dori, Coppa and Antonović2021). Kashuba and colleagues (Reference Kashuba, Kırdök, Damlien, Manninen, Nordqvist, Persson and Götherström2019), the first to genetically analyze birch pitch, and Jensen and colleagues (Reference Jensen, Niemann, Iversen, Fotakis, Gopalakrishnan, Vågene and Pedersen2019) have shown that birch pitch can provide information about individuals, including their genetic affinities, their oral microbiomes, and their environmental and ecological contexts (Jensen et al. Reference Jensen, Niemann, Iversen, Fotakis, Gopalakrishnan, Vågene and Pedersen2019; Kashuba et al. Reference Kashuba, Kırdök, Damlien, Manninen, Nordqvist, Persson and Götherström2019; Rageot et al. Reference Rageot, Lepère, Henry, Binder, Davtian, Filippi and Fernandez2021; Stacey et al. Reference Stacey, Dunne, Brunning, Devièse, Mortimer, Ladd, Parfitt, Evershed and Bull2020). Quids—chewed wads of plant matter—are more common to North American archaeological sites than chewed pitches. Quids may display unique morphologies and can contain similar genetic information to pitches. LeBlanc and colleagues (Reference LeBlanc, Cobb Kreisman, Kemp, Smiley, Carlyle, Dhody and Benjamin2013) successfully extracted mitochondrial DNA from quids from the southwestern United States, identifying the haplogroups of the chewers.

Even if pitch and chewed materials are plentiful at a site or in a region, the amount that contains clear bite marks, fingerprints, and other distinct morphologies is smaller. Chewed materials can be divided into (1) fragments with no evidence of chewing and (2) fragments with clear evidence of chewing, such as bite marks. Fragments from the former may contain environmental DNA, but they should not be considered as a potential proxy for humans. Chewed materials may contain better preserved human DNA than coprolites and sediment, but due to their rarity, they are not good proxies.

Dental Calculus

Dental calculus (Figure 1d) is a mineralized biofilm commonly found on teeth that may contain DNA from oral microbes and the individual, along with food remains and environmental particles (Dagli et al. Reference Dagli, Dagli, Baroudi and Tarakji2015; Preus et al. Reference Preus, Marvik, Selvig and Bennike2011; Weyrich et al. Reference Weyrich, Dobney and Cooper2015). De La Fuente and colleagues (Reference De La Fuente, Flores and Moraga2013) were the first to successfully extract and sequence oral microbe DNA from dental calculus. The first high-throughput study of dental calculus DNA was done by Adler and colleagues (Reference Adler, Dobney, Weyrich, Kaidonis, Walker, Haak and Bradshaw2013), who studied questions of diet, pathology, and health. Dental calculus is well preserved, has relatively high DNA yields, and can be less susceptible to contamination compared to composites such as sediment and coprolites (Dagli et al. Reference Dagli, Dagli, Baroudi and Tarakji2015).

Because dental calculus is found on teeth, it is the only composite discussed here that can be explicitly associated with an individual. Dental calculus can be considered in one of two ways: (1) dental calculus is a biofilm that is separate from the individual, or (2) dental calculus, as it is formed in the body and is on teeth, is part of the individual. Although neither is inherently incorrect, they are two opposing viewpoints that would result in different concerns around studying dental calculus. If dental calculus is viewed as a biofilm, its sampling is not under the same ethical considerations as if the tooth were sampled. Our view is that dental calculus, as it is found on teeth and is directly linked to an individual, should be treated in the same way as the tooth.

Working with Composites: An Overview

A benefit to working with composites—beyond the information they can provide about diets, environments, microbiomes, and change over time—is that composites sampled from certain contexts can be used to study human genetics while avoiding the destruction of Ancestral remains. The potentially lower genetic yields from extracting DNA from composites and the possibility that no human DNA is found are acceptable trade-offs to avoid sampling Ancestral remains. However, the sampling and analysis of human DNA from composites cannot be seen as a workaround for collaboration and communication with descendants and other stakeholders (Tsosie et al. Reference Tsosie, Begay, Fox and Nanibaa'A.2020).

Historic Mistrust

Studying Ancestor remains and belongings, including composites, is integral to archaeological research in the United States, but historically, it has been done with little discussion with or involvement of descendant communities. Through much of its history, archaeology was the purview of Western archaeologists who took upon themselves primary authority to analyze, interpret, and represent cultures with which they were not affiliated (Atalay Reference Atalay2006; Colwell Reference Colwell2016; Colwell-Chanthaphonh et al. Reference Colwell-Chanthaphonh, Ferguson, Lippert, McGuire, Nicholas, Watkins and Zimmerman2010; Nassaney Reference Nassaney2021; Prendergast and Sawchuk Reference Prendergast and Sawchuk2018; Tsosie et al. Reference Tsosie, Begay, Fox and Nanibaa'A.2020; Van Dyke Reference Van Dyke2020; Wilcox Reference Wilcox2010). These research practices were centered on Western viewpoints and exploited the pasts of descendant groups without taking care to ensure that the knowledge gained benefited them. The lack of recognized descendant autonomy over cultural and physical remains and the failure of archaeologists to include descendants in the research process have contributed to a culture of mistrust between descendant communities and archaeologists. This same mistrust and historic exclusion is seen in genetic research when Indigenous populations faced a lack of control over Ancestor remains, cultural artifacts, and data (Claw et al. Reference Claw, Lippert, Bardill, Cordova, Fox, Yracheta and Bader2017; Colwell Reference Colwell2018; Garrison et al. Reference Garrison, Hudson, Ballantyne, Garba, Martinez, Taualii, Arbour, Caron and Rainie2019; Handsley-Davis et al. Reference Handsley-Davis, Kowal, Russell and Weyrich2021; Malhi and Bader Reference Malhi and Bader2015; Tackney and Raff Reference Tackney and Raff2019; Tsosie et al. Reference Tsosie, Begay, Fox and Nanibaa'A.2020; Van Dyke Reference Van Dyke2020; Wilcox Reference Wilcox2010). Indigenous communities are increasingly involved in the planning and interpretation of archaeogenetic research, but this development is unfolding against the backdrop of hundreds of years of exploitation and marginalization of Indigenous peoples in North America (Nassaney Reference Nassaney2021; Van Dyke Reference Van Dyke2020).

Context and Interpretation

Studies focused on tracing the genetic histories of multiple diverse people groups can be extensive. Although these analyses are useful and necessary for broad characterizations, the detection of trends, and evolutionary studies, researchers do not always engage with the cultural context of or the Indigenous knowledge about the Ancestors they study. This lack of engagement and potential disregard for traditional ways of knowing and cultural data can be common in paleogenetic studies (Crellin and Harris Reference Crellin and Harris2020; Fox Reference Fox2019; Gokcumen and Frachetti Reference Gokcumen and Frachetti2020; Tackney and Raff Reference Tackney and Raff2019). Genetic data that are separated from their cultural context or used in ways not consented to by descendant communities run the risk of being interpreted in ways that contradict the oral and historic traditions of descendant groups. This disjunct can harm a group's or an individual's sense of self and may remove nuance from data interpretation in favor of simpler, straightforward narratives (Austin et al. Reference Austin, Sholts, Williams, Kistler and Hofman2019; Crellin and Harris Reference Crellin and Harris2020; Hakenbeck Reference Hakenbeck2019). Additionally, genetic data have the potential to cause lasting harm to Indigenous peoples by weakening land claims and political standings, stigmatizing groups, and playing into racist ideologies (Garrison et al. Reference Garrison, Hudson, Ballantyne, Garba, Martinez, Taualii, Arbour, Caron and Rainie2019; Handsley-Davis et al. Reference Handsley-Davis, Kowal, Russell and Weyrich2021; Nassaney Reference Nassaney2021).

Access

Access relates to data stewardship and to the ability to participate in aDNA work. From a research perspective, it is standard practice to publicly share paleogenomic datasets, and in fact, such sharing is often a requirement for publication (Alpaslan-Roodenberg et al. Reference Alpaslan-Roodenberg, Anthony, Babiker, Bánffy, Booth, Capone and Deshpande-Mukherjee2021; Anagnostou et al. Reference Anagnostou, Capocasa, Milia, Sanna, Battaggia, Luzi and Bisol2015; Sedig Reference Sedig2019). Few standards exist regarding data format and content. The result is a variety of databases in different formats that have different rules for access and differing amounts of associated metadata (Fox Reference Fox2019; Fox and Hawks Reference Fox and Hawks2019; Powell Reference Powell2021). However, a greater concern is the frequent lack of access or control that descendant communities have over data generated from their Ancestors (Fleskes et al. Reference Fleskes, Bader, Tsosie, Wagner, Claw and Garrison2022; Mackey et al. Reference Mackey, Calac, Chenna Keshava, Yracheta, Tsosie and Fox2022; Tsosie et al. Reference Tsosie, Begay, Fox and Nanibaa'A.2020). Publicly sharing genetic datasets without full collaboration with Indigenous communities largely benefits researchers while potentially harming descendant communities, and not all cultural knowledge or genetic information is meant for public consumption (Carney et al. Reference Carney, Diedrich, Blong, Guedes, Fulkerson, Kite, Leonard-Doll, LeCompte-Mastenbrook, Tushingham and Zimmermann2022; Fleskes et al. Reference Fleskes, Bader, Tsosie, Wagner, Claw and Garrison2022; Kowal et al. Reference Kowal, Weyrich, Argüelles, Bader, Colwell, Cortez and Davis2023; Nassaney Reference Nassaney2021; Van Dyke Reference Van Dyke2020).

Archaeological materials, including composites, are finite resources. Researchers must rely on either existing collections or new excavations. DNA from newly excavated material is generally better preserved, but additional DNA degradation can occur when samples are stored in less than ideal conditions. This additional degradation can make destructive aDNA analyses (Figure 2) less feasible and less justifiable (Brunson Reference Brunson2019; Brunson and Reich Reference Brunson and Reich2019; Fleskes et al. Reference Fleskes, Bader, Tsosie, Wagner, Claw and Garrison2022; Fox and Hawks Reference Fox and Hawks2019; Pálsdóttir et al. Reference Pálsdóttir, Bläuer, Rannamäe, Boessenkool and Hallsson2019; Pruvost et al. Reference Pruvost, Schwarz, Correia, Champlot, Braguier, Morel, Fernandez-Jalvo, Grange and Geigl2007; Sirak and Sedig Reference Sirak and Sedig2019). Ancestor remains are irreplaceable, and once they are destroyed for genetic analysis, they cannot be reconstructed, studied in nondestructive ways, or returned to descendants. The result of small amounts of available materials, the high costs of genetic research, the need for dedicated facilities, and the specialized knowledge required for genomics is that most studies are performed by a few prominent labs (Austin et al. Reference Austin, Sholts, Williams, Kistler and Hofman2019; Callaway Reference Callaway2017; Fleskes et al. Reference Fleskes, Bader, Tsosie, Wagner, Claw and Garrison2022; Fox Reference Fox2019; Fox and Hawks Reference Fox and Hawks2019; Lewis-Kraus Reference Lewis-Kraus2019; Makarewicz et al. Reference Makarewicz, Marom and Bar-Oz2017; Mulligan Reference Mulligan2006; Pálsdóttir et al. Reference Pálsdóttir, Bläuer, Rannamäe, Boessenkool and Hallsson2019; Prendergast and Sawchuk Reference Prendergast and Sawchuk2018; Sedig Reference Sedig2019). This has led to a research landscape that too often prevents descendant communities, along with smaller labs and research groups, from accessing materials or engaging in archaeogenetic research without collaborating with or giving research control to one of the larger groups (Lewis-Kraus Reference Lewis-Kraus2019; Somel et al. Reference Somel, Altınışık, Özer and Ávila-Arcos2021). Furthermore, cultural material and genetic samples are often stored in academic institutions or museums that may be far removed from descendants (Colwell-Chanthaphonh et al. Reference Colwell-Chanthaphonh, Ferguson, Lippert, McGuire, Nicholas, Watkins and Zimmerman2010; Lippert Reference Lippert2006; Nilsson Stutz Reference Nilsson Stutz2018; Wilcox Reference Wilcox2010). When descendants are not involved in research, they can lack access to their physical past and to the interpretation of that past (Atalay Reference Atalay2006; Handsley-Davis et al. Reference Handsley-Davis, Kowal, Russell and Weyrich2021).

FIGURE 2. Representation of the destructive nature of composite research: (a) whole paleofecal sample before subsampling; (b) exterior was removed and the sample cut in half. Subsamples for aDNA analysis were collected from the center and homogenized; (c) half the remaining material was disaggregated for macroremains and microremains analysis. (Photos provided by Taryn Johnson.)

A Cautionary Tale

Researchers extracted DNA from nine ancestors interred with funerary objects from Pueblo Bonito in Chaco Canyon with the goal of adding genetic data to archaeological debates about the role of kinship in the development of complex societies. They found that the individuals shared mitochondrial genomes and represented an elite matriline. The researchers further determined the relatedness of six of the Ancestors by genotyping single-nucleotide polymorphisms (SNPs) from their nuclear DNA (Kennet et al. Reference Kennett, Plog, George, Culleton, Watson, Skoglund and Rohland2017). Consultation of local tribes was not legally required because the Ancestors were considered culturally unaffiliated by their housing institution. This study now serves as an example of how a lack of descendant collaboration in research can cause harm. Claw and colleagues (Reference Claw, Lippert, Bardill, Cordova, Fox, Yracheta and Bader2017) brought up three main concerns with the study: (1) tribal groups were not consulted, (2) some data descriptions were culturally insensitive, and (3) the researchers did not consider how the study might affect descendants. Tribal knowledge also includes descriptions of matrilineal structures, which would have aided interpretation and put the findings in a broader cultural context. The study had the added effect of degrading long-term collaborative relationships between descendant communities and regional archaeologists (Claw et al. Reference Claw, Lippert, Bardill, Cordova, Fox, Yracheta and Bader2017; Cortez et al. Reference Cortez, Bolnick, Nicholas, Bardill and Colwell2021; Van Dyke Reference Van Dyke2020).

Legal and Biomedical Frameworks: NAGPRA and the Belmont Report

In the United States, the Native American Graves Protection and Repatriation Act of 1990, or NAGPRA, provides legal protections for human remains, funerary objects, sacred objects, and objects of cultural patrimony. This legislation rarely applies to composites and does not prohibit DNA analysis (Fleskes et al. Reference Fleskes, Bader, Tsosie, Wagner, Claw and Garrison2022; Van Dyke Reference Van Dyke2020), but the consultation process that NAGPRA inspires (but, significantly, does not require) can serve as a guide during the planning phase of a research project. The concept of stakeholder engagement, which involves early and frequent communication with stakeholders, is central to the NAGPRA document (United Nations Evaluation Group 2017). The basic consultation steps involve identifying stakeholders, sharing research information, discussing how the consultation process should proceed, and disseminating data at the conclusion of the project (Bureau of Land Management 2016; Monette et al. Reference Monette, Greenwood, Gonzales-Rogers, Durham, Johnson and Aikin2018).

A related biomedical framework is the standardized treatment of humans by biomedical researchers. The Belmont Report, a response to the infamous “Tuskegee Study of Untreated Syphilis in the Negro Male in the United States,” emphasizes that researchers need to treat people with respect and to avoid causing harm. The report provides a framework for the treatment of living subjects centered on three ethical principles: (1) respect for persons, (2) beneficence, and (3) justice, which in practice calls researchers to respect personal autonomy, minimize harms, and do research in ways that benefit affected communities (National Commission for the Protection of Human Subjects of Biomedical and Behavioral Research 1979). Another principle outlined in the Belmont Report is voluntary participation. The deceased cannot give informed consent, so for ancient genetic analyses, it falls to descendant communities to consent to research. If the letter of the law is followed, composite research is rarely subject to consultation. Therefore, although NAGPRA and the Belmont Report can provide some guidance, their specific focus on living humans and Ancestor remains necessitates the development of an ethical framework specifically for composites.

Collaborative Frameworks

Stakeholders are the individuals, communities, museums, and other institutions that have cultural or professional connections to a study (Alpaslan-Roodenberg et al. Reference Alpaslan-Roodenberg, Anthony, Babiker, Bánffy, Booth, Capone and Deshpande-Mukherjee2021; Pálsdóttir et al. Reference Pálsdóttir, Bläuer, Rannamäe, Boessenkool and Hallsson2019). We recognize that descendant communities often have strong, vested interests in aDNA research extending beyond those of generic “stakeholders.” However, for ease of conversation, we are including them within the broader category of stakeholders in this discussion. Stakeholders may include descendant populations and local, state, national, and tribal entities, in addition to archaeologists, other researchers, and curating institutions. When researchers work with Indigenous North American materials, the stakeholders include the modern tribal or Indigenous groups, including both lineal and cultural descendant communities. Collections managers and other museum professionals who are often involved in curating composites, who engage in conversations with the descendant communities whose objects they house, and who approve research proposals involving destructive analysis should also be included in conversations about the potential benefits and limitations of proposed research.

Composite research—and more broadly, archaeology—is inherently collaborative work that integrates multiple perspectives and methods. Working with stakeholders on an egalitarian basis creates space for combining multiple viewpoints that go beyond the Western framework and leads to better science (Colwell-Chanthaphonh et al. Reference Colwell-Chanthaphonh, Ferguson, Lippert, McGuire, Nicholas, Watkins and Zimmerman2010; Nilsson Stutz Reference Nilsson Stutz2018). Studies need to be considered within their own contexts, and one viewpoint or a single approach cannot give a complete picture of the past. Opening the work to differing viewpoints, ways of knowing, and cultural knowledge leads to a more nuanced, fuller view of the past that shifts the focus of research back onto human questions (Colwell Reference Colwell2016; Colwell-Chanthaphonh et al. Reference Colwell-Chanthaphonh, Ferguson, Lippert, McGuire, Nicholas, Watkins and Zimmerman2010). Inclusivity further allows archaeology to both contend with and build on its past and inform contemporary debates and solutions (McAnany and Rowe Reference McAnany and Rowe2015; Nilsson Stutz Reference Nilsson Stutz2018).

Involved communities should include those who are culturally affiliated and those with links to the Ancestors, and communication should occur throughout a research project. This means that communities and local research groups are equal partners in all parts of the research process. In cases where descendant communities cannot be identified, where there are conflicts, or where community consent is withdrawn, research may need to stop (Fleskes et al. Reference Fleskes, Bader, Tsosie, Wagner, Claw and Garrison2022; Kowal et al. Reference Kowal, Weyrich, Argüelles, Bader, Colwell, Cortez and Davis2023). Researchers who decide to move forward with research need to explicitly state why moving forward is justifiable. The goals of engagement should be focused on collaboration and building competency (Claw et al. Reference Claw, Anderson, Begay, Tsosie, Fox and Nanibaa'A.2018; Tackney and Raff Reference Tackney and Raff2019; Wagner et al. Reference Wagner, Colwell, Claw, Stone, Bolnick, Hawks, Brothers and Nanibaa'A.2020). Researchers should assess how composites are viewed by the relevant communities and ensure that all project terminology, documentation, and goals align with their belief structures. All aspects of research should be discussed before and throughout a project (Alpaslan-Roodenberg et al. Reference Alpaslan-Roodenberg, Anthony, Babiker, Bánffy, Booth, Capone and Deshpande-Mukherjee2021; Garrison et al. Reference Garrison, Hudson, Ballantyne, Garba, Martinez, Taualii, Arbour, Caron and Rainie2019; Matisoo-Smith Reference Matisoo-Smith2019; Wagner et al. Reference Wagner, Colwell, Claw, Stone, Bolnick, Hawks, Brothers and Nanibaa'A.2020), and discussion should occur before any sampling is done (Tackney and Raff Reference Tackney and Raff2019). Communicating openly and on an equal basis with communities and individuals who have cultural and historical connections to research opens the door for true collaboration.

Including community members enables them to educate professional researchers about their past and their concerns about research plans (Atalay Reference Atalay2006; Colwell Reference Colwell2016; Colwell-Chanthaphonh et al. Reference Colwell-Chanthaphonh, Ferguson, Lippert, McGuire, Nicholas, Watkins and Zimmerman2010; McAnany and Rowe Reference McAnany and Rowe2015). This leads to a shift from archaeological gatekeeping to a more collaborative framework that has modern relevance, that benefits descendant communities, and that has a wider audience. Giving equal consideration to other views can lead to new research avenues and reveal theoretical and personal biases in data interpretation while putting materials in context and bringing broader understanding (Brunson and Reich Reference Brunson and Reich2019; Colwell Reference Colwell2016; Colwell-Chanthaphonh et al. Reference Colwell-Chanthaphonh, Ferguson, Lippert, McGuire, Nicholas, Watkins and Zimmerman2010; Fox and Hawks Reference Fox and Hawks2019; Kiddey Reference Kiddey2020; McAnany and Rowe Reference McAnany and Rowe2015; Nassaney Reference Nassaney2021).

A Quick Guide to Genetic Sequencing Methods

Different sequencing methods will yield different types and amounts of information, allowing collaborators to choose methods that are best suited to the goals of the project. For any aDNA project using composites, the goal should be to choose methods that result in the least amount of destruction with the greatest yield of the target DNA. Once DNA has been extracted from a composite, it may be amplified and sequenced. Amplification involves copying extracted DNA to create millions of new copies; sequencing refers to identifying the order of bases (adenine, guanine, thymine, cytosine) in each strand of DNA. Ancient DNA methods are summarized in detail elsewhere (Liu et al. Reference Liu, Andrew Bennett and Fu2022; Orlando et al. Reference Orlando, Allaby, Skoglund, Sarkissian, Stockhammer, Ávila-Arcos and Fu2021). Here, we briefly describe Sanger sequencing, metabarcoding, and shotgun sequencing to provide a short introduction for individuals unfamiliar with genetic analyses.