Copyright

by

Jaime Mata-Míguez

2016

The Dissertation Committee for Jaime Mata-Miguez Certifies that this is the approved version of the following dissertation:

Assessing the Demographic and Genetic Impact of State Expansion in

Pre-Hispanic and Colonial Mexico

Committee:

Deborah A. Bolnick, Supervisor

Enrique Rodriguez-Alegria

Anthony Di Fiore

Brian M. Kemp

Christopher C. Nice Assessing the Demographic and Genetic Impact of State Expansion in

Pre-Hispanic and Colonial Mexico

by

Jaime Mata-Miguez, B.S.; B.S.; M.A.

Dissertation

Presented to the Faculty of the Graduate School of

The University of Texas at Austin

in Partial Fulfillment

of the Requirements

for the Degree of

Doctor of Philosophy

The University of Texas at Austin August 2016

Assessing the Demographic and Genetic Impact of State Expansion in

Pre-Hispanic and Colonial Mexico

Jaime Mata-Miguez, PhD

The University of Texas at Austin, 2016

Supervisor: Deborah A. Bolnick

Political expansions in Mexico may have had important demographic effects.

However, these demographic effects remain unclear. In Xaltocan, a native town in central

Mexico founded by Otomi-speaking people in AD 900, archaeological and historical evidence provide somewhat contradictory accounts of such effects.

When Xaltocan expanded during the 10-13th centuries, two culturally distinct groups inhabited this town. Archaeologists hypothesize that one group comprised newcomers who settled on the periphery of Xaltocan and were unrelated to the Otomis in the interior, but this scenario remains unclear. In 1395, the Tepanec state conquered

Xaltocan, and this town was incorporated into the Aztec state in 1428. Historical documents assert that the Otomis abandoned Xaltocan following the Tepanec conquest and the Aztec state repopulated this town in 1435. However, archaeological evidence of cultural continuity through this time suggests that some Otomis remained at Xaltocan after the Tepanec and Aztec conquests, so the demographic consequences of these events

iv remain unsolved. After the Spanish conquest in 1521, the native residents adopted

Spanish material culture, but historical documents point to infrequent contact with

Spaniards. Archaeological and historical evidence also suggest that political reorganizations and trade stimulated immigration from other native populations.

However, whether this immigration increased in colonial times or not is unclear.

To clarify these demographic effects, I analyzed DNA from ancient and modern

Xaltocan residents. Mitochondrial DNA (mtDNA) indicates that the periphery residents during the 10-13th centuries were maternally unrelated to the Otomis, as archaeologists suggest. However, mtDNA, Y-chromosome, and autosomal DNA reveal genetic discontinuities associated with the Tepanec and Aztec conquests, revealing important demographic consequences unidentified in the archaeological record. Furthermore, in colonial times, mtDNA, Y-chromosome DNA, and genome-wide single nucleotide polymorphisms indicate very little (or perhaps non-existent) admixture, which is consistent with infrequent contact with Spaniards during this period. Finally, temporal changes of mtDNA diversity in colonial times were largely due to immigration from other native populations, as archaeological and historical evidence suggests.

Thus, this dissertation illustrates how anthropologists can integrate archaeological, historical, and genetic evidence to study the demographic effects of political expansions.

v Table of Contents

List of Tables ...... ix

List of Figures ...... xi

Introduction ...... 1

Chapter 1: The Genetic and Demographic Impact of Political Expansions in Pre- Hispanic Mexico: Evidence from Ancient DNA ...... 7 Introduction ...... 7 Materials and Methods ...... 14 Samples ...... 14 Dating and Temporal Classification of Samples ...... 15 DNA Extraction and Genetic Analysis ...... 16 Statistical Analyses ...... 17 Results and Discussion ...... 19 Ancient DNA Analysis ...... 19 Immigration and Population Expansion Before 1395 ...... 20 Kin Burials Within Pre-1395 Interior and Periphery Xaltocan ...... 22 Genetic and Demographic Impacts of the Tepanec Conquest ...... 23 Population Changes at Xaltocan Following the Aztec Conquest ...... 27 Kin Burials Within Tepanec and Aztec Houses ...... 28 Modeling Demographic and Genetic Changes in Pre-Hispanic Xaltocan ...... 31 Supplementary Information ...... 43 Samples ...... 43 Genetic Analyses ...... 43 Mitochondrial DNA Analyses ...... 43 Autosomal DNA Analyses and Molecular Sex Determination ..44 Y-Chromosome DNA Analyses ...... 45 Contamination Controls ...... 47 Statistical Analyses ...... 49 vi Chapter 2: A Genetic Test of Historical and Archaeological Hypotheses on the Effects of Spanish Colonialism in Mexico ...... 78 Abstract ...... 78 Introduction ...... 79 Spanish Colonialism in Mexico ...... 80 Migration from Europe and Africa: Patterns and Consequences 81 Political Reorganizations and Changing Patterns of Regional Trade ...... 85 Previous Genetic Research ...... 88 Methods...... 90 Samples ...... 90 DNA Extraction and Genotyping...... 92 Statistical Analyses ...... 93 Results and Discussion ...... 98 Gene Flow from Europe and Africa...... 99 Gene Flow Between Xaltocan and Other Native Populations in Mexico100 Conclusion ...... 102

Chapter 3: A Genetic Test of Historically-Based and Archaeological Hypotheses on the Effects of Spanish Colonialism: Genome-Wide DNA Evidence from Xaltocan, Mexico ...... 133 Abstract ...... 133 Introduction ...... 134 Previous Genetic Research ...... 135 Methods...... 138 Samples ...... 138 SNP Genotyping ...... 138 Comparative SNP Data ...... 138 Data Curation ...... 139 Data Analysis ...... 139 Results and Discussion ...... 140

vii Conclusion ...... 151

References ...... 162 Chapter 1 ...... 162 Supplementary Information ...... 166 Chapter 2 ...... 174 Chapter 3 ...... 181

viii List of Tables

Table 1.1. Summary data for the 38 individuals with validated mitochondrial DNA

data...... 34

Table 1.2. Autosomal STR genotypes for 38 individuals...... 54

Table 1.3. Y-chromosome haplogroups and haplotypes for 15 male individuals. 58 Table 1.4. Mitochondrial haplotype frequencies in Xaltocan and other Mesoamerican

populations...... 60 Table 1.5. Mitochondrial and Y-chromosome diversity statistics for ancient

Xaltocan.1 ...... 62 Table 1.6. Number of mtDNA haplotypes observed in this study and degree of mtDNA haplotype sharing within and between spatial or temporal

groups...... 63

1 Table 1.7. FST comparisons based on mitochondrial DNA haplotypes...... 65

Table 1.8. BayeSSC results...... 67 Table 1.9. Common logarithms of likelihood ratios (parent-offspring/unrelated) obtained in KINGROUP for pairwise comparisons of all of the ancient individuals from which at least seven complete autosomal STR

genotypes (both alleles) could be determined.1 ...... 68 Table 1.10. Common logarithms of likelihood ratios (full siblings/unrelated) obtained in KINGROUP for pairwise comparisons of all of the ancient individuals from which at least seven complete autosomal STR

genotypes (both alleles) could be determined.1 ...... 70 Table 1.11. Natural logarithms of likelihood ratios for trios (mother, father, and

offspring) obtained in Cervus.1 ...... 72 ix Table 2.1. Sex, recent ancestry, Y-DNA and mtDNA haplogroups, and mtDNA

haplotypes for all of the 47 present-day residents of Xaltocan...... 111 Table 2.2. mtDNA haplogroups and mtDNA haplogroup diversity in pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in

Mexico...... 118 Table 2.3. mtDNA haplotype diversity and nucleotide diversity in pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in

Mexico based on nps 16,055-16,382...... 121

Table 2.4. Y-DNA haplogroups and Y-DNA haplogroup diversity in pre-Hispanic

Xaltocan and modern Xaltocan...... 123 Table 2.5. Y-DNA haplogroups in pre-Hispanic Xaltocan, modern Xaltocan, and

other modern native populations in Mexico...... 124

Table 2.6. FST values between pre-Hispanic Xaltocan, modern Xaltocan, and other

modern native populations in Mexico...... 127

Table 2.7. Latitude and longitude of native populations in Mexico...... 129 Table 2.8. Great circle distances (in kilometers) between native populations in

Mexico...... 130

Table 2.9. BayeSSC simulations...... 132

Table 3.1. Sample sizes for all of the populations used in this study...... 144 Table 3.2. Number of SNPs present in autosomal and X-chromosome datasets at

different steps of data curation...... 145

x List of Figures

Figure 1.1. Map of Xaltocan within the Basin of Mexico in ancient times (Rodríguez-

Alegría, 2010)...... 39 Figure 1.2. Network for mtDNA haplogroup A. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding

haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

...... 40

Figure 1.3. MDS plot based on mtDNA haplotype data...... 41

Figure 1.4. MDS plot based on mtDNA haplotype data...... 42

Figure 1.5. Map of Xaltocan in ancient times and location of houses...... 73 Figure 1.6. Network for mtDNA haplogroup B. Mutations identified with reference to

the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

...... 74 Figure 1.7. Network for mtDNA haplogroup C. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding

haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

...... 75 xi Figure 1.8. Network for mtDNA haplogroup D. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

...... 76

Figure 1.9. BayeSSC demographic scenarios...... 77

Figure 2.1. Map of Xaltocan within the Basin of Mexico in ancient times (when this

town was an island in Lake Xaltocan)...... 103

Figure 2.2. BayeSSC demographic scenarios...... 104 Figure 2.3. Network for mtDNA haplogroup A. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White

circles represent haplotypes not found in the populations analyzed here.

...... 105 Figure 2.4. Network for mtDNA haplogroup B. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et

al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

...... 106

xii Figure 2.5. Network for mtDNA haplogroup C. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

...... 107 Figure 2.6. Network for mtDNA haplogroup D. Mutations identified with reference to

the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et

al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

...... 108

Figure 2.7. MDS plot based on mtDNA haplotype data...... 109 Figure 2.8. Genetic distance versus geographic distance in modern native population

in Mexico, including modern Xaltocan. Each data point represents a

pairwise comparison of populations...... 110

Figure 3.1. PCA plots based on autosomal SNPs...... 146

Figure 3.2. PCA plots based on X-chromosome SNPs...... 147

Figure 3.3. Values of K and cross-validation error for ADMIXTURE analyses based

on (a) autosomal SNPs and (b) X-chromosome SNPs...... 148 Figure 3.4. Admixture proportions based on autosomal SNPs for (a) all Native American, European, and African individuals included in this study, (b)

Xaltocan individuals with all four grandparents from Xaltocan, and (c) Xaltocan individuals with at least one grandparent not from Xaltocan.

...... 149 xiii Figure 3.5. Admixture proportions based on X-chromosome SNPs for (a) all Native American, European, and African females included in this study, (b) Xaltocan females with all four grandparents from Xaltocan, and (c)

Xaltocan females with at least one grandparent not from Xaltocan.150

xiv Introduction

The rise and fall of influential political entities in the past may have had significant demographic effects. To investigate these demographic effects, scholars have traditionally used historical and archeological evidence, and these lines of evidence have provided important insights. However, the demographic effects of such events remain unclear. For instance, did the rise and fall of influential political entities lead to significant movements of people in or out of conquered regions? Genetic evidence represents an additional tool for answering these questions. Because different demographic processes produce distinct patterns of genetic change through time, genetic analyses of a temporal sample of individuals can assess the likelihood of specific demographic scenarios. Hence, genetic evidence can play a crucial role in testing demographic hypotheses associated with the rise and fall of past political entities, especially when historical documents and archaeological finds are inconclusive.

This dissertation uses genetic evidence to clarify the demographic consequences of the expansion and collapse of ancient political entities in the Basin of Mexico. In this region, a series of influential states and empires, including the Otomi state, the Tepanec state, the Aztec empire, and the Spanish empire, emerged and declined from the 13th to

19th centuries (Brumfiel, 1983; Lockhart, 1992; Hassig, 1994; Berdan and Smith,

2003a,b; Smith and Berdan, 2003). Because of their significant socio-cultural impacts, these political entities have been the subject of much historical and archaeological

1 research. However, their demographic effects on native populations remain open to debate. To elucidate such effects, this dissertation analyzes ancient and modern DNA from Xaltocan, a town in the northern region of the Basin of Mexico.

Xaltocan was founded by a group of Otomi-speaking people around 900 C.E. At that time, Xaltocan was a small island in the middle of now drained Lake Xaltocan

(Brumfiel, 2005a). From the 11th to 13th centuries, Xaltocan grew into an influential city- state that collected tribute from neighboring communities, served as the capital of the

Otomi state, and experienced a high rate of population growth and land expansion

(Gibson, 1964; Alva Ixtlilxóchitl, 1975; Alva Ixtlilxóchitl, 1977; Sanders et al., 1979;

Carrasco-Pizana, 1987; Brumfiel, 2005a,b,c; Chimonas, 2005). However, by the mid-13th century Xaltocan and the neighboring Tepanec town of Cuauhtitlan became involved in a war against each other, significantly reducing Xaltocan’s influence in the Basin of

Mexico. With the aid of other Tepanec allies, Cuauhtitlan eventually defeated Xaltocan in

1395 (Gibson, 1964; Alva Ixtlilxóchitl, 1975; Annals of Cuauhtitlan, 1992). The emerging Aztec empire annexed Xaltocan in 1428, and Tenochtitlan (the capital of the empire) assigned rulers to govern the town (Carrasco-Pizana, 1987; Annals of

Cuauhtitlan, 1992; Hicks, 1994). Xaltocan remained under Aztec rule until 1521, when

Hernán Cortés and his troops incorporated it into the Spanish empire (Díaz del Castillo,

1965; Hernán Cortés, 1971; Hassig, 1994). In Mexico, Spanish colonialism extended until 1821 (García-Martínez, 2010).

2

Historical and archaeological evidence have left some questions about the demographic effects of these events open to debate. First, differences in house structure, burial practices, and ceramic consumption have led some archaeologists to hypothesize that two ethnically distinct groups inhabited Xaltocan as the town experienced population growth and land expansion during the Otomi period. Specifically, De Lucia and

Overholtzer (2014) have suggested that while the descendants of the original Otomies lived in the interior of the island, a group of newcomers settled on the eastern and southeastern periphery (De Lucia and Overholtzer, 2014). However, relationships of biological descend between ancient residents of the interior and the periphery houses have never been tested using genetic evidence.

Second, historical documents and archaeological finds provide contradictory accounts of the demographic effects of the Tepanec conquest of Xaltocan. According to some colonial documents, the inhabitants of Xaltocan fled when this town lost the war against the Tepanecs in 1395, leaving the town unpopulated for roughly 40 years (AGI

Justicia 123/3, 1536; Alva Ixtlilxóchitl, 1977; Carrasco-Pizana, 1987). In addition, some colonial documents state that in 1435 an Aztec ruler repopulated Xaltocan with people from other regions in the Basin of Mexico (Carrasco-Pizana, 1987; Annals of

Cuauhtitlan, 1992; Hicks, 1994). However, several lines of archaeological evidence from

Xaltocan may contradict these historical accounts. For instance, some burials date between 1395 and 1435, which is not consistent with the assertion that Xaltocan remained uninhabited during this time period (Overholtzer, 2013). In addition,

3 archaeological excavations indicate that inhabitants of Xaltocan after the Tepanec conquest continued to build houses and bury their dead in approximately the same locations as the earlier Otomi residents (Overholtzer, 2013).

Finally, historical and archaeological evidence have not fully resolved questions about the demographic effects of Spanish colonialism at Xaltocan. Some colonial documents describe this town as a pueblo de indios (i.e., ruled by native people) that

Spanish colonists rarely visited (Montúfar, 1897; Hicks, 2005), but archaeological evidence indicates that some native residents adopted Spanish material culture

(Rodríguez-Alegría, 2010; Rodríguez-Alegría et al., 2013). In addition, historical documents and archeological finds provide inconclusive evidence on how the colonial period affected migration rates between Xaltocan and other native populations (Montúfar,

1897; Gibson, 1964).

Xaltocan provides an excellent opportunity to test these historical and archaeological hypotheses using ancient and modern DNA. During 25 years of research at Xaltocan, archaeologists have recovered a collection of ancient human remains with well-preserved DNA that date to the Otomi period, the supposed abandonment period, and the Aztec period (Brumfiel, 2005a; Brumfiel and Rodríguez-Alegría, 2010;

Overholtzer, 2013). Since these samples provide a precise temporal representation of genetic diversity, they make it possible to clarify the demographic effects of political transitions during those periods. In addition, modern patterns of genetic diversity at

Xaltocan are informative of demographic processes during the colonial period.

4

This dissertation uses mitochondrial, Y-chromosome, and autosomal DNA markers to clarify the demographic effects of past political transitions. Mitochondrial

DNA is exclusively inherited maternally and present in both females and males, so these markers represent an excellent tool for determining maternal ancestry. In contrast, Y- chromosome DNA is exclusively inherited paternally and present only in males, which makes these markers useful for determining paternal ancestry in males (Pakendorf and

Stoneking, 2005; Underhill and Kivisild, 2007). Autosomal DNA is bi-parentally inherited and present in both females and males, so these markers provide a more complete picture of individual ancestry than mitochondrial and/or Y-chromosome DNA alone (Li et al., 2008). Within cells, mitochondrial DNA is much more abundant and, therefore, easier to recover from ancient human remains, than either Y-chromosome or autosomal DNA. For this reason, ancient DNA studies have traditionally focused on mitochondrial DNA (Hagelberg et al., 2015).

Chapter 1 analyzes mitochondrial haplogroups and sequences, Y-chromosome haplogroups and 23 short tandem repeat loci, and 15 autosomal short tandem repeat loci in ancient individuals to clarify the demographic effects of political transitions during the

Otomi period, the supposed abandonment period, and the Aztec period. Chapter 2 analyzes mitochondrial haplogroups and sequences in both ancient and modern individuals to shed light in the demographic effects of the colonial period. Finally,

Chapter 3 analyzes roughly 630,000 genome-wide single nucleotide polymorphisms in modern individuals to further clarify the demographic effects of the colonial period.

5

Together, these chapters illustrate how genetic evidence can be used to test historical and archaeological hypotheses on human population history.

6

Chapter 1: The Genetic and Demographic Impact of Political

Expansions in Pre-Hispanic Mexico: Evidence from Ancient DNA

INTRODUCTION

The rise and fall of states can have profound genetic and demographic consequences for human populations. During periods of expansion, political instability, and collapse, societies may face warfare, increased immigration and emigration, changes in community organization and socioeconomic structures, and shifts in the allocation of food and other resources. These events can all produce changes in the size, structure, and composition of communities, and those demographic shifts may leave signatures in the human genome. Genetic data may therefore play an important role in elucidating the consequences of major political transitions, especially when other lines of evidence are inconclusive, contradictory, or highly contested.

In the Basin of Mexico, for example, ethnographic, historical, and archaeological research have suggested that the rise and fall of the Toltec, Tepanec, and Aztec states across the Middle (1240-1350 C.E.) and Late Postclassic (1350-1521 C.E.) periods may have had important genetic and demographic effects (Mata-Míguez et al., 2012).

Indigenous histories recount a series of migrations in the Basin of Mexico, both before and during the founding of these city-states (Boone, 2000). However, the nature and extent of the demographic and genetic effects of political transitions in this region are still unclear, in part because early colonial documents and archaeological finds have yielded

7 somewhat conflicting evidence. The inferences that can be made from these lines of evidence are also limited because material culture and documents written centuries after the events in question provide only indirect evidence of genetic and demographic processes. To provide the first direct evidence of the genetic and demographic effects of the rise and fall of the successive Toltec, Tepanec, and Aztec states, I analyzed ancient

DNA (aDNA) from Xaltocan, a town in the northern part of the Basin of Mexico (Figure

1).

The Basin of Mexico went through a series of political transitions during the

Middle and Late Postclassic periods. Following the collapse of the Toltec state in 1150

C.E., city-states battled against one another for political domination and control of resources until power was gradually consolidated into a few large polities. By the end of the 14th century, the Tepanec state gained control over much of the Basin of Mexico.

However, Tepanec dominance ended in 1428, when an alliance between the city-states of

Texcoco, Tlacopan, and Tenochtitlan laid the foundations of the Aztec empire. Over the next century, the Aztec empire conquered a large number of polities and came to rule nearly the entire Basin of Mexico and vast areas in central and southern Mexico until

Spanish conquistadors took over the Aztec capital city of Tenochtitlan in 1521 (Brumfiel,

1983; Berdan and Smith, 2003a,b; Smith and Berdan, 2003).

This series of political transitions may have had important genetic and demographic consequences for communities in the Basin of Mexico like Xaltocan, a town on an artificially constructed island (about 800 x 400 m) in the middle of the now

8 drained Lake Xaltocan (Brumfiel, 2005a). Colonial documents indicate that a group of

Otomi-speaking people founded Xaltocan around 1100 C.E. (Carrasco-Pizana, 1950;

Gibson, 1964; Alva Ixtlilxóchitl, 1975; Alva Ixtlilxóchitl, 1977; Annals of Cuauhtitlan,

1992), but archaeological evidence places the foundation of the town earlier, around 900

C.E. (Brumfiel, 2005a). During the 11 to 13th centuries, Xaltocan grew into an influential city-state that collected tribute from neighboring communities, served as the capital of the

Otomi state (Gibson, 1964; Alva Ixtlilxóchitl, 1975; Alva Ixtlilxóchitl, 1977; Carrasco-

Pizana, 1987; Brumfiel, 2005a,b,c; Morehart, 2012b), and experienced a high rate of population growth (Chimonas, 2005), with the town perhaps reaching a population size of nearly 5,000 inhabitants (Sanders et al., 1979). Because the development of chinampas

(agricultural lands with plots elevated above water levels and surrounded by canals) filled the shoreline of the island with about 100 additional hectares between 1250 and 1350

(Frederick et al., 2005; Morehart and Eisenberg 2010; Morehart, 2012a; Morehart, 2014;

Morehart and Frederick, 2014), Xaltocan also experienced a substantial land expansion during this time period (Overholtzer, 2012).

The collapse of the Toltec state in the 12th century preceded this population expansion at Xaltocan. As a prosperous and influential city-state, Xaltocan may have been the focus of migration from other towns as migrants left unstable, impoverished regions to settle in more stable, affluent communities (De Lucia and Overholtzer, 2014;

Overholtzer and De Lucia, in press). Archaeological evidence is consistent with this scenario because differences in house structure, burial practices, and ceramic

9 consumption suggest that at least two groups of people, with different cultural traditions, were living in Xaltocan by 1240 C.E. In the interior of the island, multi-roomed houses with plaster floors or murals were built, infants and children were frequently buried under house floors or walls, and both Aztec I and Aztec II Black-on-Orange pottery were used alongside significant quantities of Black-on-Red and Incised Brownware serving vessels

(De Lucia 2010; De Lucia, 2014; De Lucia and Overholtzer, 2014; Overholtzer and De

Lucia, in press). Because this pattern of ceramic consumption is indicative of continuous occupation in the interior of the island since the foundation of Xaltocan, these houses likely belonged to descendants of the original Otomi residents (De Lucia and

Overholtzer, 2014; Overholtzer and De Lucia, in press). In the newly added periphery of the island, however, single-roomed houses were built with earthen floors and adobe or clay walls, and both infants and adults were buried in outside patios. These households did not use Aztec I Black-on-Orange or Incised Brownwares, but instead selectively consumed only Aztec II Black-on-Orange and significant quantities of Black-and-White on Red pottery (De Lucia and Overholtzer, 2014; Overholtzer and De Lucia, in press). In comparison to the houses in the interior of the island, periphery residents had fewer imported polychrome serving vessels and houses of poorer quality materials, and thus, likely had a lower socioeconomic status (Overholtzer and De Lucia, in press). Based on these differences, researchers have hypothesized that residents of houses in the eastern and southeastern periphery belonged to a group of newcomers who were ethnically distinct from and unrelated to the Otomi residents in the interior (De Lucia and

10

Overholtzer, 2014; Overholtzer and De Lucia, in press). While archaeological evidence supports this hypothesis, further tests are needed to directly compare the biological makeup of the people who lived in the interior versus at the periphery of Xaltocan.

The subsequent rise of the Tepanec and Aztec states in the 13th and 14th centuries, respectively, may have also had a genetic and demographic impact on communities in the

Basin of Mexico. Xaltocan and neighboring Cuauhtitlan, part of the rising Tepanec state, waged war against one another starting in 1250, with their periodic battles ending more than a century later in 1395, following a final major battle in which Cuauhtitlan emerged victorious (Gibson, 1964; Alva Ixtlilxóchitl, 1975; Annals of Cuauhtitlan, 1992). Early colonial documents assert that the inhabitants of Xaltocan fled after the Tepanec conquest, leaving the town uninhabited (AGI Justicia 123/3, 1536; Alva Ixtlilxóchitl,

1977; Carrasco-Pizana, 1987), and the emerging Aztec empire absorbed it in 1428. In addition, historical documents indicate that the Aztec ruler repopulated Xaltocan in 1435 with people who were not descendants of the original Otomis. These newcomers belonged to multiple ethnic groups, including members of the Acolhua, Colhua,

Tenochca, and Otomi, and came from a number of places in central Mexico, including

Tenochtitlan, Ixayoctonco, Totollan, Tlapallan, Tlihuacan, and Ixayoc (Carrasco-Pizana,

1987; Annals of Cuauhtitlan, 1992; Hicks, 1994). Overall, historical evidence suggests that the expansion of the Tepanec and Aztec states may have led to genetic and demographic changes at Xaltocan.

11

However, three lines of archaeological evidence from Xaltocan may contradict the historical account. First, some burials date to between 1395 and 1435, which is not consistent with the assertion that Xaltocan remained uninhabited during this time period

(Overholtzer, 2013). Second, survey evidence suggests a population decrease of only

6.5%, far less than would be expected if the site had been completely abandoned and then resettled (Chimonas, 2005; Miller, 2007). Finally, archaeological excavations indicate that inhabitants of Xaltocan after the Tepanec conquest continued to build houses and bury their dead in approximately the same locations as earlier Otomi residents

(Overholtzer, 2013). This finding may indicate that the newcomers who settled in

Xaltocan took possession of abandoned houses and shared burial practices with the earlier Otomi residents, but it is also possible that some Otomis remained in the town during the supposed abandonment period — perhaps only the Otomi elites departed after the Tepanec conquest — and even following incorporation into the Aztec empire.

Overall, archaeological evidence does not support a period without habitation, nor does it point to a complete genetic and demographic rupture across the Tepanec and Aztec imperial transitions.

To test the historical and archaeological hypotheses about the impact of past political transitions on Xaltocan, I analyzed aDNA from a collection of ancient human remains unearthed during 25 years of archaeological research at the site (Brumfiel,

2005a, 2007). These remains come from houses in both the interior and periphery of the island, and they date to before the Tepanec conquest (1240-1395), the Tepanec period

12

(1395-1428), and the Aztec period (1428-1512). Because aDNA from these individuals provides a precise temporal and spatial representation of genetic diversity at Xaltocan during the Middle and Late Postclassic periods, this analysis can clarify the genetic and demographic effects of past political transitions and help elucidate the genetic relationships among individuals buried close together at Xaltocan.

My previous analysis of mitochondrial DNA (mtDNA) from three houses on the eastern and southeastern periphery of Xaltocan included 10 individuals buried before the

Tepanec conquest and 15 residents buried after (Mata-Míguez et al., 2012). This analysis shed some light on the genetic and demographic changes associated with the Tepanec and

Aztec conquests of Xaltocan, indicating that these events were associated with a replacement of mtDNA lineages in the sampled houses. This finding is consistent with a population replacement, substantial demographic change in the absence of complete population replacement (e.g., immigration and/or population movement within Xaltocan), or the replacement of matrilines but continuity of patrilines within the community (Mata-

Míguez et al., 2012). Because my previous study analyzed only a single genetic locus and a relatively small number of individuals from three houses at Xaltocan, I could not distinguish between these possibilities. In addition, since those three houses were all located on the periphery of the town, my previous research did not compare patterns of genetic diversity between households in the interior and at the periphery of Xaltocan.

To better evaluate the genetic and demographic effects of past political transitions in the Basin of Mexico, I analyzed mtDNA sequences, Y-chromosome single nucleotide

13 polymorphisms (SNPs) and short tandem repeats (STRs), and autosomal STRs in an expanded sample of individuals from Xaltocan. These individuals were buried during the pre-Tepanec period (when two groups are hypothesized to have lived side-by-side at

Xaltocan), the supposed abandonment period, and the Aztec period, and they come from houses located in the interior and at the eastern and southeastern periphery of the town.

MATERIALS AND METHODS

Samples

Skeletal remains were recovered from six houses (Operation G, Operation Y,

Zocalo A, Zocalo C, Structure 122, and Structure 124; Figure S1) during archaeological research at Xaltocan. Elizabeth Brumfiel excavated Operation G and Operation Y

(Brumfiel, 2005; Brumfiel, 2007), Enrique Rodríguez-Alegría and Brumfiel excavated

Zocalo A and Zocalo C (Brumfiel and Rodríguez-Alegría, 2010), and Lisa Overholtzer excavated Structure 122 and Structure 124 (Overholtzer, 2012; Overholtzer, 2013; De

Lucia and Overholtzer, 2014). Operation G, Zocalo A, and Zocalo C were multi-roomed houses in the interior of the island where infants and children were buried under house floors, whereas Operation Y, Structure 122, and Structure 124 were single-roomed houses on the periphery of the island where infants and adults were buried in outside patios (Figure S1; De Lucia and Overholtzer, 2014; Overholtzer and De Lucia, in press).

Archaeological excavations at Xaltocan were carried out with support and appropriate

14 permissions from the Mexican National Institute for Anthropology and History, the town’s delegados (delegates), and local property owners.

Dating and Temporal Classification of Samples

I used Bayesian statistical modeling of radiocarbon determinations directly dating human tooth and bone from 18 individuals (Table 1; see also Overholtzer, 2013). For the remaining individuals for whom radiocarbon dates were not available, I determined approximate dates based on stratigraphic analyses and the style of pottery placed in graves.

Because deposits containing Aztec II Black-on-Orange ceramics have been radiocarbon dated to between 1240 and 1350 (Overholtzer 2013), all of the individuals that were associated with these deposits were assigned in the pre-1395 Xaltocan group

(n=26). Radiocarbon dates for individuals E14.3, E14.4, E14.5, E14.7, and E25.2 provided independent confirmation for their inclusion in pre-1395 Xaltocan. Pre-1395

Xaltocan includes individuals from all six houses, and this temporal group can be further divided into two spatial subgroups based on excavation location: pre-1395 interior (n=8) and periphery (n=14) Xaltocan.

Because Aztec III Black-on-Orange ceramic deposits are associated with radiocarbon determinations dating to between 1350 and 1521 (Overholtzer 2013), it is possible that some individuals associated with these deposits also predate 1395.

However, of the 13 individuals who were radiocarbon dated and whose remains were associated with Aztec III Black-on-Orange ceramic deposits (E6.1, E7.1, E8.1, E8.2, 15

E8.3, E8.4, E8.5, E10.1, E10.2, E14.2, E14.6, and E34.1), ten have radiocarbon dates that fall squarely in the post-1395 period with 95% confidence (Table 1), and two others have ranges that begin in 1390, including very little of the pre-Tepanec conquest period. One

(E34.1) has a 22% likelihood of dating to 1330-1370 and a 72% likelihood of dating to

1380-1430, and thus is statistically much more likely to fall in the post-1395 period. I therefore included all individuals associated with Aztec III Black-on-Orange ceramic deposits in the post-1395 conquest group (n=16). Post-1395 Xaltocan includes individuals from two houses that were located on the southeastern periphery of the island:

Structure 122 and Structure 124. In Structure 122, eight individuals (E7.1, E8.1, E8.2,

E8.3, E8.4, E8.5, E10.1, and E10.2) are statistically more likely to date to the Tepanec period (1395-1428) than to the Aztec period (1428-1521), and three individuals (E6.1,

E14.2, and E14.6) have been confidently dated to the Aztec period. Thus, I can further divide some post-1395 individuals from Structure 122 into two temporal subgroups:

Tepanec (n=8) and Aztec (n=3) residents.

DNA Extraction and Genetic Analysis

I extracted ancient DNA (aDNA) from the skeletal samples following published protocols (Rohland and Hofreiter, 2007; Bolnick et al., 2012). For each individual, I obtained 2-5 independent extracts in order to confirm all results. I screened each extract for mutations defining the founding Native American mitochondrial DNA (mtDNA) haplogroups A, B, C, and D (Schurr et al., 1990; Torroni et al., 1993), and sequenced mtDNA nucleotide positions 16,011-16,382 of the first hypervariable region to determine 16 matrilineal haplotypes, following Bolnick et al. (2012). I used the PowerPlex® 16 System kit (Promega) to genotype 15 autosomal short tandem repeat (STR) loci in the aDNA extracts and modern samples. This kit also targets a length dimorphism in the amelogenin gene on the X and Y chromosomes, allowing us to determine the molecular sex of each individual. For some extracts, I set up additional PCRs with primers targeting only the amelogenin gene in order to confirm the molecular sex determination. For individuals genetically identified as males, I screened their aDNA extracts for mutations that define the Native American founding Y-chromosome haplogroups Q-M3 and Q-L54(xM3), and

I used the PowerPlex® 23 System kit (Promega) to genotype 23 Y-chromosome STR loci.

Because these loci are located on the non-recombining region of the paternally-inherited

Y-chromosome, they were used to define paternal haplotypes. In all of the stages of aDNA analysis, I followed strict procedures to prevent and detect contamination

(Supplementary Information).

Statistical Analyses

To investigate the genetic relationships among individuals buried at Xaltocan, I carried out three sets of analyses. First, I created median-joining networks (Bandelt et al.,

1999) to determine the phylogenetic relationships among haplotypes within each mtDNA haplogroup using Network 4.6.1.0 (www.fluxus-engineering.com). Second, I estimated genetic relatedness based on autosomal STR genotypes for all pairs of individuals using

Kingroup v.2.08+ (Konovalov et al., 2004). Third, I evaluated parentage relationships between individuals using Cervus 3.0 (Kalinowski et al., 2007). 17

To evaluate the degree of genetic differentiation between spatial or temporal groups/subgroups, I performed two sets of statistical analyses. First, I used Arlequin to compute FST values based on mtDNA haplogroup and haplotype data in order to estimate genetic distances between spatial or temporal groups/subgroups. I included comparative data from modern Mesoamerican populations (Table S3) to contextualize genetic distances between the groups and subgroups from ancient Xaltocan. The FST values were then used in multidimensional scaling (MDS) analysis in XLSTAT to generate a two- dimensional representation of genetic differentiation between the groups and subgroups from ancient Xaltocan as well as modern Mesoamerican populations. Second, I carried out exact tests of population differentiation using Arlequin. These tests evaluated the null hypothesis that haplogroup, haplotype, or allele frequencies were identical in two spatial or temporal groups/subgroups being compared. If these tests rejected the null hypothesis, then haplogroups, haplotypes, or alleles in those groups/subgroups are unlikely to have been drawn from the same population.

To assess the likelihood of the data under different demographic scenarios, I used the simulation-based program Bayesian Serial SimCoal (BayeSSC; Excoffier et al., 2000;

Anderson et al., 2005). I evaluated whether genetic drift and mutation alone could explain the number of mtDNA haplotypes shared between pre-1395 and post-1395 Xaltocan.

From historical and archaeological evidence, I gathered plausible rates of population growth, female effective population sizes (Nef), and numbers of generations between the

18 pre-1395 and post-1395 Xaltocan population samples. I then ran simulations using those rates.

RESULTS AND DISCUSSION

Ancient DNA Analysis

I sampled skeletal remains (teeth or bone) from 42 ancient individuals (Table 1) recovered from six houses (Operation G, Operation Y, Zocalo A, Zocalo C, Structure

122, and Structure 124; Figure S1). For all individuals, I attempted to (1) sequence 372 base pairs of the first hypervariable region (HVRI) of the mtDNA to determine mtDNA haplotypes, (2) identify mtDNA haplogroups based on polymorphisms in HVRI and the coding region, (3) determine molecular sex based on a length dimorphism in the amelogenin gene on the X and Y chromosomes, and (4) genotype 15 autosomal STRs. In males, I also attempted to identify Y-chromosome haplogroups and haplotypes based on

SNPs and 23 STRs, respectively, in the non-recombining region of the Y-chromosome. I followed strict procedures to avoid contamination, including the use of a restricted-access clean room for pre-PCR work, independent extractions and multiple amplifications, negative controls for both extractions and PCRs, and independent confirmation of some results in a different aDNA laboratory. In addition, I applied stringent criteria to validate all of my results (Materials and Methods, Supplementary Information).

I was able to determine the mtDNA haplogroups and haplotypes, as well as the molecular sex, of 38 individuals (Table 1). All 23 mtDNA haplotypes observed could be

19 assigned to the founding Native American haplogroups A2, B2, C1, or D1. For the period between 1395 and 1521 C.E. (after the Tepanec conquest), I could validate 13-15 autosomal STRs in 12 out of 16 individuals, and 11-23 Y-chromosome STRs in all nine males (Tables S1 and S2). However, for the period between 900 and 1395 C.E. (before the Tepanec conquest), I could validate 11-15 autosomal STRs in only two out of 22 individuals, and only one Y-chromosome STR in one out of six males (Tables S1 and

S2). Poorer nuclear DNA preservation in the individuals dated to between 900 and 1395

C.E. may be due to the fact that their remains were stored at room temperature for a couple of years rather than being transferred to a -80°C freezer shortly after excavation

(like the remains for individuals dated to between 1395 and 1521 C.E.) (Pruvost et al.,

2007). Summary statistics of mtDNA and Y-chromosome diversity are given in Table S4.

Immigration and Population Expansion Before 1395

Based on archaeological evidence, researchers have hypothesized that individuals living in houses in the interior of Xaltocan before 1395 C.E. were descendants of the original Otomi inhabitants of the town, but individuals living on the eastern and southeastern periphery were unrelated newcomers (De Lucia and Overholtzer, 2014;

Overholtzer and De Lucia, in press). To test this hypothesis, I sampled 26 individuals dating to between 900 and 1395 C.E., and divided this temporal group into two spatial subgroups based on house location: pre-1395 interior Xaltocan (n=8, from 3 houses:

Operation G, Zocalo A, and Zocalo C) and pre-1395 periphery Xaltocan (n=18, from 3 houses: Operation Y, Structure 122, and Structure 122). If the archaeological hypothesis 20 is correct, then the individuals from houses in interior Xaltocan should not be closely related genetically to individuals from houses on the periphery of the site. I determined mtDNA haplogroups and haplotypes for all individuals from interior Xaltocan and for 14 individuals from periphery Xaltocan, but many of these individuals exhibited poor Y- chromosome and/or autosomal DNA preservation. For this reason, I used only mtDNA data to investigate immigration and population expansion during the pre-Tepanec period.

My results demonstrate that individuals from the two pre-1395 spatial subgroups were not closely related maternally. First, pre-1395 interior and periphery Xaltocan exhibit substantially different mtDNA haplogroup frequencies. All eight individuals from pre-1395 interior Xaltocan (100%) belong to haplogroup A, whereas pre-1395 periphery

Xaltocan includes three individuals (21.4%) belonging to haplogroup A, seven (50%) to haplogroup B, and four (28.6%) to haplogroup D (Table S3). Second, exact tests of population differentiation based on mtDNA haplogroup and haplotype frequencies indicate that individuals from these spatial groups are unlikely to have been drawn from the same biological population (P = 0.001 and P = 0.003, respectively). Third, pre-1395 interior and periphery Xaltocan exhibit distinct mtDNA haplotypes: none are shared between individuals from different spatial groups, and median-joining haplotype networks indicate that the haplotypes present in interior Xaltocan are not closely related to the haplotypes present in periphery Xaltocan (Figures 2, S2, S3, and S4; Table S5).

Finally, the multidimensional scaling (MDS) plot based on mtDNA haplotypes suggests a high degree of genetic differentiation between pre-1395 interior and periphery Xaltocan

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(Figure 3a). Thus, the observed mtDNA differences between pre-1395 interior and periphery Xaltocan show that the two spatial groups were not closely related maternally.

These results are consistent with the hypothesis that residents living in the eastern and southeastern periphery of Xaltocan were a genetically unrelated, ethnically distinct group that moved to Xaltocan during the period of land expansion. This migration would have occurred during the period of political instability that followed the fall of the Toltec state, and may have contributed to population growth at Xaltocan and the rise of Xaltocan as an influential city-state in the Basin of Mexico.

Kin Burials Within Pre-1395 Interior and Periphery Xaltocan

While mtDNA haplotypes show significant genetic differences between pre-1395 interior and periphery Xaltocan, these data also reveal close genetic relationships between individuals within each of the two spatial subgroups. Some mtDNA haplotypes are shared by multiple individuals from the same house, such as two infant males (ZA.2 and

ZA.4) buried in the pre-1395 interior house Zocalo A. This pattern of haplotype sharing indicates the burial of maternally-related kin within the house. Similarly, the newborn female (E14.3), 7-month old male (E14.4), and 10-month old female (E14.5) buried close together in Structure 122 share the same mtDNA haplotype, indicating another kin burial in a house located at the periphery of Xaltocan.

More strikingly, I also see some mtDNA haplotypes shared by individuals from two different houses in the same area of the town (e.g., related individuals in Operations

G and Zocalo C in pre-1395 interior Xaltocan, and related individuals in Structure 122 22 and Operation Y in pre-1395 periphery Xaltocan). However, as discussed earlier, mtDNA haplotypes are not shared across neighborhood lines. The identification of matrilineal ties between houses in the same area of Xaltocan, but not across the interior-periphery boundary, further supports the notion that residents of the town’s interior and periphery were genetically distinct. Altogether, the data suggest that immigration and the geographic expansion of the town in the 13th century produced changes in the genetic composition of the Xaltocan community.

Genetic and Demographic Impacts of the Tepanec Conquest

Early colonial documents describe a population replacement at Xaltocan following the Tepanec conquest in 1395, saying that residents fled and left the town uninhabited until 1435, after it was incorporated into the Aztec empire (AGI Justicia

123/3, 1536; Carrasco-Pizana, 1950; Alva Ixtlilxóchitl, 1977). While archaeological evidence indicating that Xaltocan remained inhabited between 1395 and 1435 raises questions about the accuracy of the colonial narrative concerning a period of abandonment (Overholtzer, 2013), it is possible that a demographic shift occurred shortly after the Tepanec conquest. To investigate this possibility, I also collected genetic data from 16 individuals who date to between 1395 and 1521 C.E. and were recovered from

Structure 122 and Structure 124. I compared patterns of genetic diversity at Xaltocan before and after 1395 to measure the degree of genetic differentiation between the two temporal groups (pre- and post-1395 Xaltocan). If there was a substantial population

23 replacement at Xaltocan following the Tepanec conquest, one should see substantial genetic differences and a lack of genetic continuity between pre- and post-1395 Xaltocan.

Mitochondrial DNA, Y-chromosome, and autosomal analyses all reveal significant genetic differentiation between pre- and post-1395 Xaltocan. First, mtDNA haplotypes show clear differences between these temporal groups. Pre- and post-1395

Xaltocan do not share any mtDNA haplotypes, and haplotypes from the different temporal groups are not closely related (Figures 2, and S2-S4; Table S5). In addition, the

MDS plot based on mtDNA haplotypes suggests an important level of genetic differentiation between pre- and post-1395 Xaltocan (Figure 3b), and an exact test of population differentiation based on mtDNA haplotype frequencies indicates that pre- and post-1395 individuals are unlikely to have been drawn from the same biological population (P < 0.001).

To evaluate whether mutation and genetic drift alone could have produced the observed mitochondrial differentiation (in the absence of migration/gene flow from elsewhere), I used the simulation-based Bayesian Serial SimCoal (BayeSSC) program. I tested whether the lack of mtDNA haplotype sharing between pre- and post-1395

Xaltocan could be explained by demographic scenarios based on plausible mutation rates and parameters of population size for ancient Xaltocan that do not involve gene flow between Xaltocan and other populations. The BayeSSC simulations indicate that mutation and genetic drift alone cannot explain the lack of mtDNA haplotypes shared between pre- and post-1395 Xaltocan (Table S7), so gene flow into Xaltocan — likely

24 due to immigration — must have been an important factor in reshaping the genetic composition of this town.

Overall, these mtDNA results suggest that the Tepanec conquest may have been associated with a replacement of matrilines at Xaltocan, confirming my previous findings

(Mata-Míguez et al., 2012) with a much larger sample size and individuals sampled from both the interior and periphery of the town. Since all pre-1395 haplotypes identified to date from both interior and periphery Xaltocan appear to have been replaced, this study suggests that the change in matrilines may have affected the town as a whole. It may not have been just a localized occurrence affecting individual households. Unfortunately, like my previous research, this post-1395 sample remains limited to individuals interred in two households located on the site periphery, since those are the only ones from this later occupational phase to have been excavated to date. Thus, the nature of this available sample limits my ability to distinguish between possible migration scenarios, for example, to test whether significant reorganization within the island happened after the conquest. I can demonstrate that people dwelling in the interior of the island likely did not move to the periphery after the conquest, since none of the newly sampled interior dwellers, all of whom pre-date the 1395 conquest, are related to the post-1395 Structure

122 and 124 households. However, I cannot test the reverse pattern, wherein periphery households moved to the center, which is a more plausible scenario given that elites (who likely fled and thus vacated valuable houses and land) lived in the center of the island at the town’s height.

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Autosomal and Y-chromosome analyses were less informative because the pre-

1395 samples exhibited poorer nuclear DNA preservation, but the results are nevertheless consistent with significant population changes following the Tepanec conquest of

Xaltocan. I evaluated the degree of genetic relatedness between individuals based on their biparentally-inherited autosomal STR genotypes with the program Kingroup. This program calculates the likelihood of a particular genetic relationship for a pair of individuals (e.g., parent-offspring, siblings, half-siblings, cousins, unrelated) given their genotypes and population allele frequencies (Goodnight and Queller, 1999; Konovalov et al., 2004). Kingroup firmly ruled out a parent-offspring relationship between most pre-

1395/post-1395 pairs, and failed to identify any first-degree relationships (e.g., parent- offspring or full siblings) between any pre-1395 individual and any post-1395 individual with statistical significance (Tables S8 and S9).

Similarly, my analyses of Xaltocan Y-chromosomes indicate substantial differences in Y-chromosome haplogroup frequencies between the two temporal populations (Table 1). Haplogroup Q-L54(xM3) was common in pre-1395 Xaltocan

(66.7%) but much rarer in post-1395 Xaltocan (11.1%), while haplogroup Q-M3 showed the opposite pattern (33.3% in pre-1395 Xaltocan versus 88.9% in post-1395 Xaltocan).

An exact test of population differentiation based on Y-chromosome haplogroup frequencies failed to reject the null hypothesis that pre- and post-1395 Xaltocan individuals could have been drawn from the same population (P = 0.090), probably due to the small sample sizes (n=6 and n=9, respectively). Altogether, though, the mtDNA,

26 autosomal, and Y-chromosome evidence indicates that the pre-1395 residents were not closely related to post-1395 residents. Thus, this study suggests that the Tepanec conquest of Xaltocan was associated with significant demographic and genetic changes, and it seems likely that there was a substantial movement of people in and out of

Xaltocan at this time.

Population Changes at Xaltocan Following the Aztec Conquest

Early colonial documents state that an Aztec ruler populated Xaltocan in 1435 with people who were not descendants of the earlier residents of the town (Carrasco-

Pizana, 1950; Annals of Cuauhtitlan, 1992; Hicks, 1994). As an initial test of this assertion, I examined genetic data from 11 post-1395 individuals from a single house

(Structure 122) who could be divided into two temporal subgroups: Tepanec residents

(1395-1428 C.E.; n=8) and Aztec residents (1428-1521 C.E.; n=3). By analyzing genetic relatedness among the Tepanec and Aztec period residents of this house, I was able to evaluate fine-scale genetic changes across the Aztec imperial transition in this household.

Specifically, I tested the hypothesis that the Aztec period residents of Structure 122 were not descended from the Tepanec residents, as would be the case if colonial assertions about Aztec immigration to Xaltocan were accurate.

My analyses indicate that the Aztec residents of Structure 122 were not descendants of the earlier Tepanec residents. The Aztec residents exhibited distinct mtDNA haplotypes from the Tepanec residents, indicating that none of the Aztec individuals were descended from any of the Tepanec females. Kingroup analyses of 27 autosomal STR genotypes also firmly ruled out a parent-offspring relationship between two of the three Aztec period individuals and any of the eight Tepanec individuals (Table

S8). More generally, Kingroup did not identify any first-degree kinship relationships (i.e., parent-offspring or full siblings) between any Aztec individual and any Tepanec individual with statistical significance (Table S8 and Table S9). Analyses of Y- chromosome haplotypes also suggest that the one Aztec male in this household did not descend from any of the earlier Tepanec male residents (Table S2). Overall, the mtDNA, autosomal, and Y-chromosome evidence show that the Aztec period residents of this house were not closely related to the earlier Tepanec residents. This finding is consistent with the suggestion in early colonial documents that an Aztec governor populated

Xaltocan in 1435 with people who were not descendants of the earlier residents of the town. However, because I have only examined the Tepanec-Aztec transition in a single household, further research is needed to determine if the observed genetic discontinuity was limited to this house or if it represents a broader shift in the town’s population after

Xaltocan was incorporated into the Aztec empire.

Kin Burials Within Tepanec and Aztec Houses

The extensive excavations, large number of individuals with preserved aDNA interred, and the fine-grained chronological framework of Structure 122 provided an exceptional opportunity to examine kinship, household descent, and demographic shifts within the context of a single household. Even though there are no close genetic relationships between the Tepanec and Aztec individuals from Structure 122, I do see 28 close genetic relationships between individuals within each of the two temporal groups.

In addition to Kingroup, I used the program Cervus 3.0 (Kalinowski et al., 2007) to assess potential parentage relationships between 17 individuals recovered in undisturbed, primary burials in the Structure 122 house mound. This program applies a maximum- likelihood approach to evaluate how likely a female and a male are to be the parents of a given individual (e.g., how likely those individuals are to be a trio) (Kalinowski et al.,

2007). Cervus results are indicative of a nuclear family during the Tepanec and likely early Aztec imperial period: an adult female (E8.2) and an adult male (E8.1) were the parents of a male newborn (E8.4) and two adult females (E8.5 and E10.2) (Tables S8-

S10). Kingroup results are consistent with a father-son relationship between E8.1 and

E8.4 (P < 0.050) as well as with full sibling relationships between E8.5 and E10.2 (P <

0.050), between E8.5 and E8.4 (P > 0.050), and between E10.2 and E8.4 (P > 0.050). In addition, Y-chromosome haplotype data confirm that E8.1 and E8.4 were closely related paternally (Table S2), consistent with a father-son relationship between these individuals, and E8.2, E8.4, E8.5, and E10.2 share the same mtDNA haplotype, indicating that they are all matrilineally related. Because a 5-year old male (E7.1) and an adult female (E8.3) buried nearby also exhibit the same mtDNA haplotype, this study points to the presence of an extended family and provides evidence that six individuals who belonged to the same matriline lived, died, and were buried in a single structure during the 14th and 15th centuries, probably over multiple generations.

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Ancient DNA evidence also suggests that the Structure 122 household received some immigrants during the mid-15th century. Specifically, Structure 122 exhibits a cluster of three individuals interred on the eastern side of the patio during the Aztec period who were not closely related to the residents of the western side during the

Tepanec period. Kingroup indicates that two of the individuals on the eastern side of the patio, an adult female (E14.6) and a 14-year old male (E6.1), were likely mother and son

(Table S8). In accordance with this inference, E14.6 and E6.1 shared the same mtDNA haplotype (Table 1). Because E6.1 and E14.6 were buried close together on the eastern side of the Structure 122 patio, this study provides strong evidence for the presence of another family living in this house in Aztec times. E14.6 and E6.1 may have briefly lived in the same house as a joint household, a very common household structure in ancient central Mexico (Carrasco 1976a), before solely inheriting the house. This family’s appearance in the Structure 122 household sometime during the early to mid 15th century coincides chronologically with historical accounts of an imperial Aztec effort in 1435 to repopulate the town with taxpayers from elsewhere with people who were not descendants of the earlier residents of the town (Carrasco-Pizana, 1950; Annals of

Cuauhtitlan, 1992; Hicks, 1994). Thus, by using mtDNA, autosomal, and Y-chromosome markers to identify the close genetic relationships between individuals buried near one another during the Tepanec or Aztec period at Xaltocan, I have been able, for the first time, to verify the presence of kin burials in pre-Hispanic Mexico.

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Modeling Demographic and Genetic Changes in Pre-Hispanic Xaltocan

By synthesizing the historical and archaeological evidence from Xaltocan with the new genetic data reported here, I can develop an integrated model of the genetic and demographic changes associated with political transitions in the Basin of Mexico.

Between the 11th and the 13th centuries, the Otomi town of Xaltocan experienced substantial land expansion and population growth (Chimonas, 2005; Frederick et al.,

2005; Morehart, 2010; Morehart and Frederick, 2014). My genetic data show that the newcomers who settled in the southeastern periphery of Xaltocan during this time period were not closely related to the original Otomi residents of the town. Perhaps attracted by its growing prosperity between the 11th and the 13th centuries, the newcomers who were genetically unrelated to the original Otomis built new habitable land and a new lakeshore on the periphery of this Xaltocan, settled there during this time period, and exhibited distinct patterns of household organization and burial practices compared to the Otomi residents living in more interior areas of Xaltocan (De Lucia and Overholtzer, 2014;

Overholtzer and De Lucia, in press).

During the 13th and 14th centuries, the Tepanec state became a dominant political power in the Basin of Mexico, and conflict with Xaltocan ensued. After the Tepanec city of Cuauhtitlan defeated Xaltocan in 1395, it appears that many (especially elite) residents of Xaltocan abandoned the town, as reported in early colonial accounts of the town’s history. However, in conflict with those accounts, immigration into Xaltocan must have taken place shortly after the Tepanec conquest, since newcomers occupied some of the

31 abandoned houses that were described as vacant in historical records, and the site remained inhabited during the period of Tepanec rule (1395-1428 C.E.). The genetic data presented here indicate a substantial population replacement at this time, as the residents of Xaltocan during the Tepanec period do not appear to be descended from the earlier

Otomi inhabitants of the town.

In 1428, the Aztec empire emerged as a result of the conquest of the Tepanecs and the Triple Alliance between the city-states of Texcoco, Tlacopan, and Tenochtitlan.

Shortly after its foundation, the Aztec empire conquered Xaltocan, and in 1435 an Aztec governor sent taxpayers to complement the existing population and increase the taxpayer base (Annals of Cuauhtitlan, 1992; Hicks, 1994). My results suggest that these newcomers may not have been closely related to the original Otomis or the residents of

Xaltocan during the Tepanec period, yet they continued to live in houses and perform burials in the same places as earlier residents (Overholtzer, 2013).

In addition to migration at the population level, I have used ancient DNA to elucidate kinship patterns and migration at the micro-level of the household. I have reconstructed the genetic kinship of a nuclear and extended family who lived in one house during the period of Tepanec rule, and I have detected the arrival of a new, unrelated matriline to that house in the early- to mid-15th century. This matriline arrived precisely when historical documents report that an Aztec governor sent taxpayers who were not descendants of the original Otomis to complement the existing population and increase the taxpayer base (Annals of Cuauhtitlan, 1992; Hicks, 1994).

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This study shows how a multidisciplinary approach can help anthropologists to detect large-scale population mobility, clarify the genetic impacts of past periods of political upheaval, and understand migration at both macro and micro scales. By integrating historical, archaeological, and genetic evidence from ancient human remains recovered at Xaltocan, my study sheds light into the genetic and demographic effects of a gradual process of consolidation of political power in the Basin of Mexico during the

Middle and Late Postclassic periods. It demonstrates that population discontinuities due to both local and long-distance migration may have been the norm during the volatile and dynamic Postclassic period in the Basin of Mexico; in fact, long-term stability within sites and within individual households may have been the exception. My study helps clarify questions that Mesoamerican historians and archaeologists have long discussed and serves as a reference for future investigations addressing similar questions in other regions of the world.

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Table 1.1. Summary data for the 38 individuals with validated mitochondrial DNA data.

Individual House Age at death Radiocarbon Molecular Y- (Mitochondrial haplogroup)

ID date (C.E.)3 Sex4 chromosome Mitochondrial haplotype5

haplogroup

Pre-1395

Interior

G.1 Operation G NA NA Female (A2) 111T, 223T, 274A, 290T, 319A, 362C

G4.4 Operation G NA NA Female (A2) 111T, 223T, 274A, 290T, 319A, 362C

G7.4 Operation G NA NA Female (A2) 111T, 223T, 274A, 290T, 319A, 362C

ZA.1 Zocalo A NA NA Male Q-L54(xM3) (A2) 111T, 189C, 223T, 258G, 290T, 295T, 319A,

362C

ZA.2 Zocalo A NA NA Male Q-L54(xM3) (A2) 223T, 240G, 290T, 319A, 362C

ZA.4 Zocalo A NA NA Male Q-M3 (A2) 223T, 240G, 290T, 319A, 362C

ZC.1 Zocalo C NA NA Female (A2) 111T, 223T, 287G, 290T, 319A, 362C

ZC.2 Zocalo C NA NA Female (A2) 111T, 223T, 274A, 290T, 319A, 362C

Periphery

E14.3 Structure 122 (east) Birth-3 months 1310-1350 Female (D1) 223T, 292T, 325C, 362C

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Table 1.1 (continued)

E14.4 Structure 122 (east) 7 months ± 3 months 1300-1350 Male Q-L54(xM3) (D1) 223T, 292T, 325C, 362C

E14.5 Structure 122 (east) 10 months – 1 year 1290-1350 Female (D1) 223T, 292T, 325C, 362C

E14.7 Structure 122 (east) 9 months – 1 year 1300-1350 Female (B2) 182C, 183C, 189C, 217C, 295T

E25.2 Structure 124 9 months – 1 year 1300-1360 Male Q-M3 (A2) 111T, 223T, 290T, 319A, 335G

Y2.2 Operation Y NA NA Female (B2) 183C, 189C, 217C, 278T

Y2.3 Operation Y NA NA Female (A2) 111T, 223T, 290T, 319A, 344T, 362C

Y2.7 Operation Y NA NA Female (B2) 182C, 183C, 189C, 217C

Y3.4 Operation Y NA NA Female (B2) 182C, 183C, 189C, 217C, 295T

Y3.6 Operation Y NA NA Female (B2) 183C, 189C, 194C, 195C, 217C, 362C

Y3.6I Operation Y NA NA Male Q-L54(xM3) (B2) 183C, 189C, 217C, 274A

Y3.8 Operation Y NA NA Female (A2) 95T, 111T, 223T, 290T, 319A, 326G, 362C

Y3.10A Operation Y NA NA Female (D1) 223T, 292T, 325C, 362C

Y3.10C Operation Y NA NA Female (B2) 182C, 183C, 189C, 217C, 234T

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Table 1.1 (continued)

Post-1395

Tepanec

E7.1 Structure 122 5 years ± 1.5 years 1400-1450 Male Q-M3 (A2) 93C, 111T, 136C, 223T, 290T, 311C, 319A,

(west) 362C

E8.1 Structure 122 30 – 40 years 1390-1460 Male Q-M3 (A2) 111T, 183C, 189C, 223T, 290T, 291T, 319A,

(west) 362C

E8.2 Structure 122 More than 50 years 1400-1470 Female (A2) 93C, 111T, 136C, 223T, 290T, 311C, 319A,

(west) 362C

E8.3 Structure 122 30 – 40 years 1410-1490 Female (A2) 93C, 111T, 136C, 223T, 290T, 311C, 319A,

(west) 362C

E8.4 Structure 122 1 – 3 months 1390-1440 Male Q-M3 (A2) 93C, 111T, 136C, 223T, 290T, 311C, 319A,

(west) 362C

E8.5 Structure 122 30 – 40 years 1390-1440 Female (A2) 93C, 111T, 136C, 223T, 290T, 311C, 319A,

(west) 362C

E10.1 Structure 122 20 – 35 years 1400-1470 Male Q-M3 (A2) 172C, 223T, 290T, 319A, 362C

(west)

E10.2 Structure 122 More than 50 years 1400-1450 Female (A2) 93C, 111T, 136C, 223T, 290T, 311C, 319A,

(west) 362C

36

Table 1.1 (continued)

Aztec

E6.1 Structure 122 (east) 14 years ± 2 years 1410-1450 Male Q-M3 (B2) 182C, 183C, 189C, 217C, 357C

E14.2 Structure 122 (east) 1 month 1430-1520 Female (A2) 111T, 192T, 223T, 290T, 319A, 362C

E14.6 Structure 122 (east) 30 – 35 years 1430-1500 Female (B2) 182C, 183C, 189C, 217C, 357C

Undefined2

E5.1 Structure 122 Fetus-7 months in NA Male Q-M3 (A2) 93C, 111T, 136C, 223T, 290T, 311C, 319A,

utero 362C

E14.1 Structure 122 (east) 8 – 9 years 1390-1470 Female (B2) 183C, 189C, 217C, 258C, 260.1T6

E30.A Structure 124 Fetus-birth NA Male Q-L54(xM3) (D1) 223T, 325C, 362C

E30.3 Structure 124 30 – 35 years NA Male Q-M3 (C1) 172C, 223T, 298C, 325C, 327T

E34.1 Structure 124 Adult 1330-1430 Male Q-M3 (D1) 223T, 325C, 362C

1 Mitochondrial data from four pre-1395 individuals (Y2.5, Y3.1, Y3.9, and Y3.12) could not be validated. NA: not available.

2 Post-1395 individuals who could not be assigned to either Tepanec or Aztec Xaltocan.

3 Ranges indicate 95 percent certainty in Bayesian statistical model. When classifying samples for which radiocarbon dates are available into temporal groups, their archaeological context was also taken into account. Based on Mata-Miguez et al. (2012),

Overholtzer (2013), and ?.

4 Based on amelogenin analysis. 37

5 Sequences from nucleotide positions (np) 16,011-16,382. Mutations specified by the last three digits (e.g., 111 refers to np

16,111) and identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999).

6 Insertion between nucleotide positions 16,260 and 16,261.

38

Figure 1.1. Map of Xaltocan within the Basin of Mexico in ancient times (Rodríguez-

Alegría, 2010).

39

Figure 1.2. Network for mtDNA haplogroup A. Mutations identified with reference to the

Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al.,

1999). Circle with a thick black line represents the founding haplotype.

Circle size is proportional to the number of samples. White circles represent

haplotypes not found in the populations analyzed here.

40

Figure 1.3. MDS plot based on mtDNA haplotype data.

41

Figure 1.4. MDS plot based on mtDNA haplotype data.

42

SUPPLEMENTARY INFORMATION

Samples

To help evaluate whether any results could be due to contamination during archaeological or genetic research (see section “Contamination Controls”), I also collected saliva samples from 11 archaeologists and buccal swabs from four researchers who worked in the Ancient DNA Laboratory at the University of Texas at Austin. I extracted DNA from these saliva and buccal swabs using the prepIT•L2P kit (DNA

Genotek) and the DNeasy Blood & Tissue kit (Qiagen), respectively.

Genetic Analyses

Mitochondrial DNA Analyses

For the researchers’ samples, I sequenced nucleotide positions (np) 15,960-16,566 of the mitochondrial genome following Kemp et al. (2010). In the ancient extracts, some mitochondrial amplicons exhibited C > T or G > A misincorporations that were not present in other amplifications of the same DNA fragment from the same individual.

Because the post-mortem deamination of cytosine into uracil could produce these misincorporations, I included AmpErase® Uracil N-Glycosylase (UNG; Life

Technologies), as indicated by the manufacturer, in subsequent PCRs from the same

DNA extracts. The inclusion of AmpErase® UNG eliminated those previously observed misincorporations, indicating that they were probably due to cytosine deamination. To

43 validate the mitochondrial DNA (mtDNA) haplogroup and haplotype for each ancient individual, I replicated all results in at least two independent extracts as well as through at least two independent amplifications from one of those extracts.

Autosomal DNA Analyses and Molecular Sex Determination

I set up PowerPlex® 16 HS PCRs using the proportions of reagents recommended by the manufacturer, but I reduced the total reaction volume to 3-10 µL. In PCRs with the researchers’ samples, I used the original number of cycles suggested by the manufacturer

(10 cycles in the first set of cycles and 22 cycles in the second set of cycles). In PCRs with aDNA extracts, I increased the number of cycles in the second set of cycles (32 instead of 22) in order to overcome potential problems that are common in aDNA research, such as a low number of initial target molecules and reduced amplification efficiency due to inhibitors (Hummel, 2003). PCR products were submitted to the DNA

Sequencing Facility of the University of Texas at Austin for fragment analysis, and

GeneMapper® Software v4.0 (Applied Biosystems) was used to visualize chromatograms and call alleles.

I used three stringent criteria to validate autosomal STR genotypes for each ancient individual. First, I considered an allele to be validated if it was identified in at least two different extracts from the same individual. Second, because allelic dropout is a common problem in aDNA studies, I also accepted any allele that was amplified in the same PCR as an allele that had been validated following my first criterion. Finally, because allelic dropout also makes it more difficult to determine whether an individual is 44 homozygous, I accepted a homozygous genotype as validated as long as the same allele

(and only that allele) was present in chromatograms from a minimum of four PCRs in total and at least one PCR from each extract.

For some extracts, I set up additional PCRs with a forward primer and a fluorescently-labeled reverse primer that target the amelogenin gene (Amelogenin-

Forward: 5’-CCC TGG GCT CTG TAA AGA ATA GTG-3’; Amelogenin-Reverse: 5’-

ATC AGA GCT TAA ACT GGG AAG CTG-3’) to clarify their molecular sex. PCR conditions were as follows: 1) initial 4-minute denaturing hold at 94˚C; 2) 60 cycles of

30-second successive holds at 94˚C (denaturation), 60˚C (annealing), and 72˚C

(extension); and 3) final 7-minute extension hold at 72˚C. PCR products were submitted to the DNA Sequencing Facility of the University of Texas at Austin for fragment analysis, and GeneMapper® Software v4.0 (Applied Biosystems) was used to visualize chromatograms and call alleles. To validate these amelogenin alleles, I employed the same criteria used to validate autosomal STR alleles.

Y-Chromosome DNA Analyses

To determine if the mutation that defines haplogroup Q-M3 was present, I performed restriction fragment length polymorphism analyses following Kemp et al.

(2007). To determine if the mutation that defines haplogroup Q-L54(xM3) was present, I designed a pair of primers (L54-Forward: 5’-CTC AGA CAA GGG TGA CAG TT-3’;

L54-Reverse: 5’-ACA GGG TCT CAT GGT CTT-3’) targeting a 134-base-pair fragment. Y-chromosome haplogroup Q-L54(xM3) is characterized by a G to A 45 transition at np 85 of this fragment (Battaglia et al., 2013). In PCRs with these primers, reagent volumes followed Bolnick et al. (2012), and PCR conditions were as follows: 1) initial 3-minute denaturing hold at 94˚C; 2) 60 cycles of 15-second successive holds at

94˚C (denaturation), 50˚C (annealing), and 72˚C (extension); and 3) final 3-minute extension hold at 72˚C. To obtain sequences, I submitted PCR products to the DNA

Sequencing Facility of the University of Texas at Austin. C > T and G > A misincorporations were observed in a few amplicons, likely due to post-mortem deamination, so I included AmpErase® Uracil N-Glycosylase (UNG; Life Technologies), as indicated by the manufacturer, in subsequent PCRs for those extracts. The resulting sequences did not include the previously observed misincorporations, which indicates that such changes were due to cytosine deamination. To validate the Y-chromosome haplogroup for each male, I required replication from at least two independent extracts as well as from at least two independent amplifications in one of those extracts.

To amplify Y-chromosome STR loci using the PowerPlex® 23 System kit

(Promega), I set up PCRs using the proportion of reagents recommended by the manufacturer, but I reduced the total reaction volume to 3-10 µL. In PCRs with modern samples from male archaeologists and geneticists, I used the original number of cycles suggested by the manufacturer (26 cycles). However, I increased the number of cycles in

PCRs with aDNA extracts to 60 cycles to ensure adequate amplification from samples with a low number of initial target molecules and reduced efficiency due to inhibitors

(Hummel, 2003). For some extracts from male individual E6.1, I set up additional PCRs

46 with a forward primer and a fluorescently-labeled reverse primer that target the Y- chromosome STR locus DYS458 (DYS458-Forward: 5’-AGC AAC AGG AAT GAA

ACT CCA AT-3’; DYS458-Reverse: 5’-CCA CCA CGC CCA CCC TCC-3’; Redd,

2002) to clarify his allele at this locus. PCR conditions were as follows: 1) initial 2- minute denaturing hold at 96˚C; 2) 60 cycles of a 10-second denaturing hold at 94˚C, a 1- minute annealing hold at 61˚C, and a 30-second extension hold at 72˚C (extension); and

3) final 7-minute extension hold at 72˚C. PCR volumes followed Bolnick et al. (2012).

To perform fragment analysis, visualize chromatograms and call alleles, I used the procedures described above, in section “Autosomal DNA Analyses and Molecular Sex

Determination.” For each individual, I considered an allele to be validated if it was present in chromatograms from at least two extracts.

Contamination Controls

Ancient DNA studies are methodologically challenging because aDNA is usually degraded, present in small amounts, and may be obscured by contamination from exogenous sources (O’Rourke et al., 2000; Kaestle and Horsburgh, 2002; Pääbo et al.,

2004; Orlando et al., 2015). To prevent contamination, I followed standard precautions in aDNA research at all stages of the analysis. During archeological work, I wore gloves and facemasks while excavating the burials at Structure 122 and Structure 124. As soon as the burials were documented, I removed samples for genetic analysis and wrapped them in aluminum foil. All pre-PCR work was then performed in the Ancient DNA

Laboratory of the University of Texas at Austin. This laboratory consists of a restricted- 47 access clean room that is exclusively dedicated to pre-PCR aDNA research. This laboratory is equipped with dedicated equipment, overhead UV lights, positive air pressure, and HEPA-filtered ventilation. While working in the aDNA laboratory, additional precautions included wearing disposable coveralls, hair covers, facemasks, sleeve covers, shoe covers, and gloves at all times, regularly decontaminating workspaces and equipment with bleach, decontaminating the entire laboratory weekly with 3% sodium hypochlorite (50% v/v household bleach), UV irradiating the entire laboratory for at least two hours after each use, irradiating both reagents and tubes with 254 nm UV light prior to use whenever possible, using reagents that were certified DNA-free and/or molecular grade whenever possible, performing DNA extractions and PCR set-up in a laminar flow hood, using aerosol resistant filter tips in all pre-PCR analyses, and treating samples with both bleach and 254 nm UV light to eliminate any surface contamination.

Following PCR set-up in the aDNA laboratory, the geneticists performed all of the post-

PCR work in the Anthropological Genetics Laboratory of the University of Texas at

Austin, which is located on a separate building. To avoid carry-over of PCR products into the aDNA laboratory, personnel movement between laboratories was unidirectional (from pre-PCR aDNA lab to post-PCR lab) each day.

I also followed standard precautions in aDNA research to detect contamination.

During extractions with ancient human remains and PCR set-up with aDNA extracts, I included negative controls (blanks) to detect any contamination that may have occurred.

To evaluate whether any of my results could be due to contamination during

48 archaeological or genetic research, I analyzed mitochondrial and nuclear DNA in archaeologists who participated in the Xaltocan excavations and in all researchers who work in the Ancient DNA Laboratory of the University of Texas at Austin. To further ensure the validity of my results, I performed at least two independent extractions for each ancient individual (different bones and/or teeth) and confirmed my results through multiple amplifications. In addition, mitochondrial and nuclear DNA results for ancient individuals were validated using the stringent criteria described above. Finally, I sent skeletal samples from two ancient individuals (E8.4 and E14.7) to the Ancient DNA

Laboratory at Washington State University for independent extraction and confirmation of my results (Mata-Míguez et al. 2012).

Statistical Analyses

I performed statistical analyses to characterize patterns of genetic variation in different spatial or temporal groups and to assess how likely different demographic scenarios are to explain my data. All analyses of mtDNA haplotype data did not include np 16,183 (because mutations at this site are strictly dependent on the presence of a C at np 16,189; Pfeiffer et al., 1999) or insertions in poly-C stretches (due to uncertainty in the exact position of such mutations).

To characterize patterns of genetic variation in spatial or temporal groups and subgroups, I estimated standard indexes of genetic variation using the program Arlequin

3.5.1.2 (Excoffier and Lischer, 2010). For mitochondrial and Y-chromosome haplogroups and haplotypes, I estimated haplogroup and haplotype diversity (h). For mitochondrial 49 haplotypes, I also estimated nucleotide diversity (π). When calculating nucleotide diversity, I used the Kimura 2-parameters model, which allows multiple substitutions at each nucleotide position as well as different substitution rates between transitions and transversions, with the transition-transversion ratio estimated from the data (Excoffier and Lischer, 2010).

Networks were used to illustrate the extent of haplotype sharing within and between temporal groups. Whenever possible, network analyses also included comparative data from modern Mesoamerican populations (Table S3) to contextualize the haplotype variation in spatial or temporal groups at ancient Xaltocan. To maximize the number of individuals from modern Mesoamerican populations that could be included in these networks, I restricted my analysis based on mtDNA haplotypes to np 16,055-16,382 of the first hypervariable region (HVRI). Because preliminary networks based on mtDNA haplotypes showed high levels of reticulation, I followed Kemp et al. (2010) in applying a default weight of 10 to all sites and down-weighting polymorphic sites with higher relative mutation rates as estimated by Meyer et al. (1999). Sites with more than 4-fold higher rates were down-weighted to 4, sites with 3-fold higher rates were down-weighted to 5, and sites with 2-fold higher rates were down-weighted to 6.

I used Kingroup v.2.08+ to calculate the likelihood of a close genetic kin relationship for pairs of individuals (Goodnight and Queller, 1999; Konovalov et al.,

2004). For this analysis, I used autosomal STR genotypes from ancient Xaltocan to estimate population allele frequencies. To account for the fact that population allele

50 frequencies were obtained from the same dataset as the pair of individuals whose genetic relatedness was being calculated, I applied a bias correction that excludes individuals from population allele frequency calculations if they may have been closely related to the pair of individuals whose genetic relatedness is being evaluated (Queller and Goodnight,

1989). To apply this bias correction, I created groups of individuals that may have been closely related by assembling individuals who belonged to the same time period and were recovered from the same house together. To account for the fact that Kingroup calculates the likelihood of multiple hypothetical genetic relationships, I adjusted P-values using a standard Bonferroni correction (Rice, 1989).

In Cervus 3.0, I analyzed parentage relationships using the option “parent pair

(sexes known)” because molecular sex for each individual was analyzed as part of this study. Cervus simulations were run using a pool of 5 candidate mothers and 5 candidate fathers for each offspring (which I presumed to represent 75% of total candidate mothers and 75% of total candidate mothers), 53% of all loci typed (based on my data), and a genotyping error rate of 1%. To obtain confidence levels associated with paternity assignments, I simulated parentage for 100,000 offspring based on allele frequencies derived from ancient Xaltocan.

To evaluate the degree of genetic differentiation between pre-1395 and post-1395

Xaltocan, I performed two sets of statistical analyses. First, I carried out exact tests of population differentiation using Arlequin. These tests evaluated the null hypothesis that haplogroup, haplotype, or allele frequencies were identical in pre-1395 and post-1395

51

Xaltocan. If these tests reject the null hypothesis, then haplogroups, haplotypes, or alleles in pre-1395 and post-1395 Xaltocan are unlikely to have been drawn from the same population. Second, I used Arlequin to compute FST values based on haplogroup and mtDNA haplotype data, as well as RST values based on the STR data, in order to estimate genetic distances between pre-1395 and post-1395 Xaltocan. FST values based on the mtDNA haplotype data were estimated in Arlequin from pairwise differences with a gamma value of 0. To maximize the number of individuals from modern Mesoamerican populations that could be included in this analysis, I restricted my FST analysis based on mtDNA haplotypes to np 16,055-16,382 of HVRI.

To evaluate whether mutation and genetic drift alone could have produced the observed mtDNA differences, I used BayeSSC, which implements a coalescence-based approach to simulate phylogenetic trees given different parameters (e.g., population size, rate of population growth, mutation rate). Summary statistics are generated from the simulated trees that can be compared to summary statistics from the observed data

(Excoffier et al., 2000; Anderson et al., 2005). Using this program, I tested three possible rates of population growth: 0% per generation, -6.5% per generation (Chimonas, 2005), and -17% per generation (Brumfiel, 1991), with the last two values drawn from previous archaeological investigations (Figure S5; Brumfiel, 1991; Chimonas, 2005). For the number of generations between pre-1395 and post-1395 Xaltocan, I tested two values: 1 and 11. The first value represents the minimum possible, in the unlikely situation that the pre-1395 and post-1395 subpopulations were separated by only a single generation. The

52 second value was estimated using an average generation length of 27 years (Weiss, 1973;

Fenner, 2005) and the dates AD 1240 and 1520 for the two Xaltocan groups (the maximum possible range of dates based on radiocarbon analyses), which suggests that

10.4 generations may have separated some individuals. For Nef, I tested two values: 750 and 50. The first value corresponds to a population with a census size of 5,000 individuals (Cabana et al., 2008), which represents my maximum estimate for population size at Xaltocan before it was conquered in 1395 (Sanders et al., 1979). The second value corresponds to a census size of ~330, my minimum estimate. In all simulations, I assumed a mutation rate of 1.642733 x 10-7 substitutions per nucleotide per year (Soares et al., 2009), a transition:transversion ratio of 0.9841 (Kimura, 1980), and a gamma distribution of rates with shapes parameters of 0.205 (theta) and 10 (kappa) (Ho and

Endicott, 2008). Because I tested three values for population growth, two values for the number of generations, and two values for Nef, I performed a total of 12 sets of simulations (Figure S6). For each set, I ran 1,000 simulations. I used the number of mtDNA haplotypes shared between temporal groups as a summary statistic. To determine

P-values, I calculated the fraction of simulations that yielded equal to or fewer mtDNA haplotypes shared between temporal groups than what I observed between pre-1395 and post-1395 Xaltocan.

53

Table 1.2. Autosomal STR genotypes for 38 individuals.

Individual Autosomal STR Loci2

ID

FGA

vWA

TH01

TPOX

Penta E

Penta D

D18S51 D21S11 D5S818 D7S820

CSF1PO

D13S317 D16S519 D3S1358 D8S1179

Pre-1395

Interior

G.1 -/- 12/- -/- -/- 30/31 15/16 11/- 11-/ 10/- -/- -/- -/- 6/7 -/- 16/17

G4.4 -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- 19/-

G7.4 -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- 7/- -/- -/- -/-

ZA.1 -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- -/- -/-

ZA.2 -/- 9/- -/- -/- -/- 15/16 11/11 -/- -/- -/- -/- -/- -/- -/- 16/17

ZA.4 -/- 8/9 -/- -/- -/- 15/16 -/- 10/- 10/- -/- -/- -/- 7/9.3 -/- 16/19

ZC.1 -/- 9/- -/- 14/- 30/- 15/16 9/13 12/12 10/14 -/- -/- -/- 6/7 -/- 15/16

ZC.2 -/- -/- -/- -/- -/- -/- 10/11 -/- -/- -/- -/- -/- -/- -/- 16/17

54

Table 1.2 (continued)

Periphery

E14.3 -/- -/- -/- -/- -/- 15/17 10/11 10/12 10/12 -/- -/- -/- 6/7 8/- 16/17

E14.4 -/- 14/- -/- -/- -/- 15/16 9/11 11/- -/- -/- -/- -/- 9.3/- -/- 16/16

E14.5 -/- 9/11 -/- -/- -/- 15/16 12/12 11/12 13/15 -/- -/- -/- 6/7 -/- 16/18

E14.7 -/- -/- -/- -/- -/- 15/15 -/- -/- -/- -/- -/- -/- -/- -/- -/-

E25.2 -/- -/- -/- -/- -/- 15/- -/- -/- -/- -/- -/- -/- -/- -/- -/-

Y2.2 -/- -/- -/- -/- -/- -/- 11/12 -/- -/- -/- -/- -/- -/- -/- 16/19

Y2.3 -/- 9/11 10/12 -/- -/- 15/15 7/12 11/- -/- -/- -/- -/- 6/6 12/- 16/17

Y2.7 -/- 8/9 -/- -/- -/- 15/15 11/12 11/12 12/- 24/25 -/- -/- 6/7 -/- 17/18

Y3.4 -/- 11/13 -/- -/- -/- 15/15 7/11 11/11 13/- -/- -/- -/- 6/7 8/12 16/18

Y3.6 -/- -/- -/- -/- 29/30 15/15 9/11 -/- -/- -/- -/- -/- -/- -/- 17/18

Y3.6I -/- -/- -/- -/- -/- 15/15 9/- -/- -/- -/- -/- -/- -/- -/- -/-

Y3.8 11/13 12/13 10/10 15/18 30/31 16/16 11/12 10/13 10/14 24/27 10/12 12/15 6/9.3 8/11 16/18

Y3.10A -/- 9/11 10/12 -/- 30/33.2 15/16 11/12 10/11 13/14 22/24 9/13 -/- 6/9.3 8/- 15/16

Y3.10C -/- -/- -/- -/- -/- -/- 11/12 -/- -/- -/- -/- -/- -/- -/- -/-

55

Table 1.2 (continued)

Post-1395

Tepanec

E7.1 12/12 9/10 10/12 15/22 29/30 16/16 11/11 11/12 14/14 20/24 9/11 12/14 6/7 8/8 16/19

E8.1 11/13 13/14 10/10 17/22 30/33.2 16/16 9/11 11/12 13/14 20/25 11/11 15/19 7/7 8/13 16/18

E8.2 12/12 9/10 10/12 13/17 29/30.2 16/16 11/11 11/12 13/14 20/27 11/11 14/16 6/6 8/13 17/18

E8.3 12/12 10/13 10/10 21/22 29/30 15/16 9/11 11/11 14/15 20/24 10/11 -/- 6/7 8/13 17/17

E8.4 12/13 10/14 10/12 17/22 30/30.2 16/16 9/11 11/12 13/14 20/25 11/11 14/15 6/7 8/8 17/18

E8.5 12/13 9/13 10/10 17/22 29/30 16/16 9/11 11/12 14/14 20/- 11/11 14/15 6/7 8/13 16/17

E10.1 12/12 9/9 11/11 14/17 29/29 15/16 9/12 12/12 10/14 21/25 9/13 15/22 6/7 8/11 16/16

E10.2 -/- 9/13 10/10 13/22 30/30.2 16/16 9/11 11/12 13/14 20/27 11/11 14/15 6/7 8/13 16/17

Aztec

E6.1 11/12 12/13 9/12 14/17 30.2/32.2 15/16 11/13 10/12 14/14 24/27 11/- -/- 6/7 -/- 16/18

E14.2 -/- 8/9 12/- -/- -/- 16/16 9/11 -/- 13/- -/- -/- -/- 6/7 -/- 16/17

E14.6 10/11 11/12 9/11 14/17 30/32.2 15/17 11/13 10/10 13/14 21/24 11/13 15/20 6/7 8/11 16/16

Undefined1

E5.1 10/- 9/13 10/- -/- 30/31.2 15/16 11/11 11/11 13/14 -/- -/- -/- 6/9.3 8/12 16/18

56

Table 1.2 (continued)

E14.1 -/- 12/13 10/11 16/16 -/- 15/16 9/11 10/11 13/13 -/- -/- -/- 7/9.3 8/- 16/17

E30.A 12/13 10/13 12/12 -/- 33.2/33.2 15/15 9/11 11/12 11/14 -/- -/- -/- 6/6 8/- 16/20

E30.3 11/12 9/9 11/12 13/19 30/32.2 15/17 11/11 8/12 12/13 23/24 9/12 17/19 7/9.3 8/12 16/17

E34.1 11/12 9/11 11/12 17/17 30/31.2 16/16 11/13 10/10 12/14 19/23 11/13 18/19 6/7 12/12 16/17

1 Post-1395 Undefined Xaltocan includes all of the post-1395 individuals who could not be assigned to either Tepanec or

Aztec Xaltocan.

2 (-): allele could not be amplified and/or validated.

57

Table 1.3. Y-chromosome haplogroups and haplotypes for 15 male individuals.

Indivi Y-chromosome haplotypes

dual (based on genotypes at STR loci)2

ID

H4

-

DYS19

GATA

DYS390 DYS391 DYS392 DYS393 DYS437 DYS438 DYS439 DYS448 DYS456 DYS458 DYS481 DYS533 DYS549 DYS570 DYS576 DYS635 DYS643

DYS385a -

DYS385b

DYS389 I

DYS389 II

Y

Pre-

1395

Periph ery

E14.4 ------13 ------

58

Table 1.3 (continued)

Post-1395

Tepanec

E7.1 13 14 17 13 30 24 10 14 13 14 11 12 21 16 17 24 11 13 20 17 22 10 11

E8.1 13 14 18 14 32 24 10 14 13 14 12 12 20 15 14 23 11 13 18 21 22 10 12

E8.4 13 14 18 14 32 24 10 14 13 - 12 12 - 15 14 23 11 13 18 21 22 - 12

E10.1 13 14 19 13 30 26 10 14 13 14 11 12 19 16 17 24 11 12 17 19 22 - 12

Aztec

E6.1 - 14 - 14 32 24 - - 13 - 12 12 - 15 15 - - - 18 - 22 - -

Undefined1

E5.1 13 15 18 12 - 24 9 - 13 - - - - - 18 25 11 12 18 - 22 - -

E30.A - 15 18 14 - 23 10 14 13 - 11 - 21 - 15 24 12 13 16 20 22 10 -

E30.3 13 15 20 12 28 23 9 13 13 14 11 13 19 15 19 25 11 13 18 18 23 10 11

E34.1 16 14 18 13 30 23 10 14 13 14 11 12 20 16 18 23 11 12 17 - 22 10 12

1 Post-1395 Undefined Xaltocan includes all of the post-1395 individuals who could not be assigned to either Tepanec or

Aztec Xaltocan.

2 (-): allele could not be amplified and/or validated.

59

Table 1.4. Mitochondrial haplotype frequencies in Xaltocan and other Mesoamerican populations.

Population Haplotype A B C D Reference

N2

Ancient Xaltocan (All) 38 0.553 0.263 0.026 0.158 This study

Pre-1395 Xaltocan (All) 22 0.500 0.318 0.000 0.182 This study

Pre-1395 interior Xaltocan 8 1.000 0.000 0.000 0.000 This study

Pre-1395 periphery Xaltocan 14 0.214 0.500 0.000 0.286 This study

Post-1395 Xaltocan (All) 16 0.625 0.188 0.062 0.125 This study

Tepanec Structure 122 8 1.000 0.000 0.000 0.000 This study

Aztec Structure 122 3 0.333 0.667 0.000 0.000 This study

Post-1395 undefined Xaltocan1 5 0.200 0.200 0.200 0.400 This study

60

Table 1.4 (continued)

Otomi 68 0.397 0.250 0.294 0.059 Sandoval et al., 2009

Zapotec 72 0.431 0.236 0.292 0.041 Kemp et al., 2010

Mixtec 84 0.714 0.167 0.071 0.048 Sandoval et al., 2009; Kemp et al., 2010

Triqui 107 0.720 0.280 0.000 0.000 Sandoval et al., 2009

Nahua-Cuetzalan 34 0.676 0.088 0.206 0.030 Malhi et al., 2003; Kemp et al., 2010

Nahua-Atocpan 44 0.386 0.409 0.182 0.023 Kemp et al., 2010

Nahua-Xochimilco 35 0.771 0.143 0.086 0.000 Sandoval et al., 2009

Nahua-Zitlala 14 1.000 0.000 0.000 0.000 Sandoval et al., 2009

Nahua-Ixhuatlancillo 10 0.400 0.100 0.300 0.200 Sandoval et al., 2009

Nahua-Necoxtla 25 0.480 0.520 0.000 0.000 Sandoval et al., 2009

1 Post-1395 undefined Xaltocan includes all of the post-1395 individuals who could not be assigned to either Tepanec or Aztec Xaltocan. 2 Haplotypes belonging to haplogroups others than A, B, C, D, or X were not included. 61

Table 1.5. Mitochondrial and Y-chromosome diversity statistics for ancient Xaltocan.1

Mitochondrial Mitochondrial Mitochondrial Y-chromosome

haplogroup haplotype nucleotide haplogroup

diversity (h) diversity (h) diversity () diversity (h)

Ancient Xaltocan (All) 0.680 0.947 0.018 0.476

Pre-1395 Xaltocan (All) 0.645 0.939 0.017 0.533

Pre-1395 interior Xaltocan 0.000 0.750 0.006 0.500

Pre-1395 periphery Xaltocan 0.670 0.923 0.017 1.000

Post-1395 Xaltocan (All) 0.592 0.808 0.017 0.222

Tepanec Structure 122 1.000 0.464 0.006 NA

Aztec Structure 122 0.667 0.667 0.184 0.222

1 NA: not available.

62

Table 1.6. Number of mtDNA haplotypes observed in this study and degree of mtDNA haplotype sharing within and between

spatial or temporal groups.

Haplogr N1 n2 Between Between Between Between Within Within Between Between Between Within Within Within Singleton

oup pre-1395 pre-1395 pre-1395 pre-1395 pre-1395 pre-1395 post- post- post- post- post- post- 4

and interior interior peripher interior peripher 1395 1395 1395 1395 1395 1395

post- and Xaltocan y Xaltocan y Tepanec Tepanec Aztec Tepanec Aztec undefine

1395 peripher houses Xaltocan houses Xaltocan and and and Xaltocan Xaltocan d

Xaltocan y houses houses Aztec undefine undefine Xaltocan

Xaltocan Xaltocan d3 d3 3

Xaltocan Xaltocan A 21 12 0.000 0.000 0.083 0.000 0.083 0.000 0.000 0.000 0.000 0.083 0.000 0.000 0.750

(0) (0) (1) (0) (1) (0) (0) (0) (0) (1) (0) (0) (9)

B 10 8 0.000 0.000 0.000 0.083 0.000 0.000 0.000 0.000 0.000 0.000 0.125 0.000 0.750

(0) (0) (0) (1) (0) (0) (0) (0) (0) (0) (1) (0) (6)

C 1 1 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 1.000

(0) (0) (0) (0) (0) (0) (0) (0) (0) (0) (0) (0) (1)

D 6 2 0.000 0.000 0.000 0.500 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.500 0.000

(0) (0) (0) (1) (0) (0) (0) (0) (0) (0) (0) (1) (0)

63

Table 1.6 (continued)

Total 38 23 0.000 0.000 0.043 0.086 0.043 0.000 0.000 0.000 0.000 0.043 0.043 0.043 0.696

(0) (0) (1) (2) (1) (0) (0) (0) (0) (1) (1) (1) (16)

1 Number of individuals.

2 Number of mtDNA haplotypes.

3 Post-1395 undefined Xaltocan includes all of the post-1395 individuals who could not be assigned to either Tepanec or Aztec

Xaltocan.

4 Unique haplotype.

64

1 Table 1.7. FST comparisons based on mitochondrial DNA haplotypes.

Atocpan Cuetzalan Xochimilco Zitlala Ixhuatlancillo

- - - - -

1395 Tepanec 1395 Tepanec 1395 Aztec 1395 Xaltocan

- - -

1395 interior 1395 periphery 1395 (All) Xaltocan - - -

Pre Xaltocan Pre Xaltocan Post Xaltocan Post Xaltocan Pre Post (All) Mixtec Zapotec Triqui Otomi Nahua Nahua Nahua Nahua Nahua

Pre-1395 periphery 0.342

Post-Xaltocan1395 Tepanec 0.433 0.464 -0.050 0.050 0.150 0.250 0.350 0.450 0.550

Xaltocan 0.049 0.149 0.249 0.349 0.449 0.549 0.649 Post-1395 Aztec 0.493 -0.028 0.595

Pre-1395Xaltocan Xaltocan

Post-1395(All) Xaltocan 0.080

Mixtec(All) 0.083 0.145 0.244 0.170 0.031 0.054

Zapotec 0.174 0.053 0.282 0.057 0.032 0.073 0.053

Triqui 0.121 0.187 0.329 0.214 0.050 0.118 0.020 0.098

Otomi 0.210 0.045 0.333 0.079 0.031 0.097 0.068 0.000 0.119

65

Table 1.7 (continued)

Nahua-Atocpan 0.217 0.010 0.304 -0.032 0.025 0.078 0.077 0.009 0.112 0.012

Nahua-Cuetzalan 0.095 0.149 0.214 0.178 0.050 0.032 0.010 0.030 0.063 0.044 0.063

Nahua-Xochimilco 0.074 0.200 0.256 0.233 0.062 0.064 0.004 0.064 0.027 0.088 0.099 0.004

Nahua-Zitlala 0.126 0.429 0.470 0.634 0.217 0.200 0.089 0.194 0.113 0.236 0.249 0.102 0.059

Nahua-Ixhuatlancillo 0.210 0.075 0.329 0.088 0.037 0.051 0.049 -0.023 0.138 -0.017 0.004 -0.011 0.061 0.272

Nahua-Necoxtla 0.275 0.037 0.402 -0.050 0.040 0.145 0.105 0.086 0.114 0.097 0.035 0.144 0.154 0.336 0.128

1 Haplotypes defined here by nucleotide positions 16,055-16,382 to facilitate comparisons with modern Mesoamerican populations. Underlined values indicate statistical significance (P < 0.050). Cells in grey indicate either non-reported or non- applicable FST values.

66

Table 1.8. BayeSSC results.

Simulation Population growth Number of generations Pre-1395 Xaltocan Post-1395 Xaltocan P-value

(%/generation) Nef Nef

1 0 1 750 750 0.000

2 0 1 50 50 0.000

3 0 11 750 750 0.000

4 0 11 50 50 0.000

5 -6.5 1 750 750 x e (-0.065 x 1) 0.000

6 -6.5 1 50 50 x e (-0.065 x 1) 0.000

7 -6.5 11 750 750 x e (-0.065 x 11) 0.000

8 -6.5 11 50 50 x e (-0.065 x 11) 0.000

9 -17 1 750 750 x e (-0.17 x 1) 0.000

10 -17 1 50 50 x e (-0.17 x 1) 0.000

11 -17 11 750 750 x e (-0.17 x 11) 0.000

12 -17 11 50 50 x e (-0.17 x 11) 0.005

67

Table 1.9. Common logarithms of likelihood ratios (parent-offspring/unrelated) obtained in KINGROUP for pairwise comparisons of all of the ancient individuals from which at least seven complete autosomal STR genotypes (both alleles) could be determined.1

Y2.7 Y3.4 Y3.8 Y310 E5.1 E14.1 E30.A E30.3 E34.1 E7.1 E8.1 E8.2 E8.3 E8.4 E8.5 E10.1 E10.2 E6.1

Y3.4 X A

Y3.8 X X

Y3.10A X -0.428 X

E5.1 -0.142 0.800 X 0.009

E14.1 X -0.595 X X 0.442

E30.A X -0.048 X X X X

E30.3 -0.263 X X X X X X

E34.1 X X X X X X X X

E7.1 X X X -0.728 -0.501 X X X X

E8.1 X X X X X X X X X X

E8.2 X X X X X X X X X X X

E8.3 X X X X X X X X X X X X

E8.4 X X X X X X X X X X 2.540 2.067 0.615

E8.5 X X X X -0.656 X X X X 0.765 1.690 1.006 1.778 X

E10.1 X X X X X X X X X X X X X X X

68

Table 1.9 (continued)

E10.2 X X X X -0.566 X X X X -0.003 1.444 2.287 1.011 X 2.316 X

E6.1 X X X X X X X X X X X X X X X X X

E14.6 X X X X X X X X X X X X X X X X X 2.878

1 “X” indicates that parent-offspring relatedness is ruled out. Cells shaded in gray correspond to comparisons between pre-1395 and post-1395 Xaltocan individuals. Logarithms in bold are statistically significant (P < 0.050) after Bonferroni correction

(Rice, 1989).

69

Table 1.10. Common logarithms of likelihood ratios (full siblings/unrelated) obtained in KINGROUP for pairwise

comparisons of all of the ancient individuals from which at least seven complete autosomal STR genotypes (both

alleles) could be determined.1

Y2.7 Y3.4 Y3.8 Y310 E5.1 E14.1 E30.A E30.3 E34.1 E7.1 E8.1 E8.2 E8.3 E8.4 E8.5 E10.1 E10.2 E6.1

Y3.4 -0.411 A

Y3.8 -1.747 -1.705

Y3.10A -1.153 -0.945 -1.413

E5.1 -0.807 0.725 -0.719 0.012

E14.1 -1.681 -1.084 -1.077 -0.813 -0.291

E30.A -0.972 -0.441 -3.456 -1.491 -1.623 -2.166

E30.3 -1.028 -1.615 -5.086 -1.964 -0.830 -1.294 -3.420

E34.1 -2.312 -1.300 -5.153 -1.428 -0.756 -2.019 -3.644 -1.047

E7.1 -1.281 -1.683 -2.957 -1.484 -1.130 -3.136 -1.467 -3.689 -4.065

E8.1 -1.235 -0.953 -2.513 -2.516 -0.777 -1.087 -1.423 -5.070 -3.954 -2.584

E8.2 -0.758 -1.826 -4.693 -2.684 -0.788 -2.698 -1.475 -4.303 -3.947 0.333 -1.865

E8.3 -1.001 -0.775 -3.495 -3.123 -1.341 -1.151 -1.214 -5.093 -4.762 -0.031 -1.189 -0.688

E8.4 -0.369 -1.558 -4.220 -3.069 -1.302 -1.866 -0.962 -5.425 -4.048 -0.118 2.396 1.525 -0.673

E8.5 -0.846 -1.479 -2.857 -2.507 -1.069 -1.345 -1.319 -4.823 -2.855 0.353 1.654 0.121 1.197 1.472

70

Table 1.10 (continued)

E10.1 -0.456 -2.175 -4.263 -2.029 -2.686 -2.470 -2.591 -3.889 -3.768 -2.746 -4.265 -4.101 -4.057 -3.898 -2.061

E10.2 -1.448 -1.479 -2.948 -2.732 -0.693 -0.695 -1.900 -3.914 -3.962 -0.861 1.375 1.906 0.322 1.680 3.119 -3.931

E6.1 -1.189 -0.426 -0.651 -2.453 -1.152 -1.676 -2.023 -2.722 -0.976 -2.381 -2.412 -1.503 -3.443 -2.239 -2.036 -2.374 -1.858

E14.6 -2.205 -1.168 -3.088 -1.378 -2.295 -1.170 -3.895 -2.531 -0.638 -5.245 -4.387 -6.024 -5.360 -4.997 -4.436 -1.308 -4.477 1.895

1 Cells shaded in gray correspond to comparisons between pre-1395 and post-1395 Xaltocan individuals. Logarithms in bold are statistically significant

(P < 0.050) after Bonferroni correction (Rice, 1989).

71

Table 1.11. Natural logarithms of likelihood ratios for trios (mother, father, and offspring) obtained in Cervus.1

Candidate Candidate Candidate mothers

offspring fathers (post-1395 females)

(post-1395 E8.2 E8.3 E8.5 E10.2 E14.6 E8.4 E8.1males) 4.023 3.100 -1.050 3.500 -8.155

E8.4 E10.1 -5.600 -6.279 -5.328 -4.159 -11.988

E8.4 E30.3 -7.502 -8.780 -7.720 -6.904 -13.646

E8.4 E34.1 -6.959 -8.345 -7.176 -5.110 -13.184

E8.5 E8.1 9.840 4.490 NA 4.490 -7.475

E8.5 E10.1 -4.675 -4.790 NA -22.426 -10.030

E8.5 E30.3 -8.862 -6.279 NA -6.497 -13.129

E8.5 E34.1 -7.312 -3.914 NA -3.778 -10.737

E10.2 E8.1 3.398 -8.210 1.420 NA -8.563

E10.2 E10.1 -4.648 -5.110 -4.920 NA -11.933

E10.2 E30.3 -7.203 -4.920 -4.621 NA -12.015

E10.2 E34.1 -7.068 -7.584 -7.068 NA -11.199 1 Likelihood ratios with 95% confidence or higher are in bold. NA: not applicable (candidate offspring and candidate mother are the same individual).

72

Figure 1.5. Map of Xaltocan in ancient times and location of houses.

73

Figure 1.6. Network for mtDNA haplogroup B. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

74

Figure 1.7. Network for mtDNA haplogroup C. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

75

Figure 1.8. Network for mtDNA haplogroup D. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

76

Figure 1.9. BayeSSC demographic scenarios.

77

Chapter 2: A Genetic Test of Historical and Archaeological Hypotheses on the Effects of Spanish Colonialism in Mexico

ABSTRACT

Since the Spanish conquest in 1521, native populations in Mexico have experienced important demographic changes. Spanish colonialism, for instance, facilitated new patterns of migration, reshaping how individuals from diverse geographic, cultural, and linguistic backgrounds interacted with one another. Historical, archaeological, and genetic studies have shed light on these interactions, demonstrating that major population centers experienced substantial migration from Europe and Africa, but the extent of migration and gene flow into other native communities is not as well understood. Furthermore, while archaeological and historical evidence suggests that political reorganizations and changing patterns of regional trade may have increased gene flow among native populations, it remains unclear if native populations outside major population centers experienced significant genetic changes during the colonial period. To help clarify the genetic history of native populations in Mexico since the onset of Spanish colonialism, I analyzed mitochondrial and Y-chromosome DNA from the pre-Hispanic and modern residents of Xaltocan, a small town in central Mexico. I found no evidence of female- or male-mediated gene flow from Europe or Africa during the colonial period.

However, temporal changes in genetic diversity from pre-Hispanic to present-day

Xaltocan were observed. Overall, these results are consistent with very low, or perhaps non-existent, levels of genetic change due to migration from Europe and Africa after the

78 onset of Spanish colonialism, but substantial genetic change due to migration and female- mediated gene flow from other native populations into Xaltocan.

INTRODUCTION

Colonial expansions have greatly affected patterns of migration between human populations. Often, colonialism has resulted in migration between distant populations that had previously remained isolated from each other. In addition, changes of the political, economic, and socio-cultural context may have altered movements of people within regions where colonial expansions took place. Because changing patterns of long- distance and local migration associated with colonialism affected gene flow between human populations, colonial expansions mat have had significant genetic effects.

However, those genetic effects of these changing patterns of migration remain unclear.

To better understand such changes, I analyzed mitochondrial DNA (mtDNA) and

Y-chromosome DNA (Y-DNA) from pre-Hispanic and modern Xaltocan, a native population in central Mexico. In Xaltocan, historical and archaeological evidence provide contradictory accounts about the extent of genetic change due to immigration from

Europe and Africa after the Spanish conquest. Therefore, Xaltocan constitutes a perfect case study for testing historical and archaeological hypotheses about the genetic effects of changing patterns of migration in native communities in Mexico since 1521.

79

Spanish Colonialism in Mexico

Spanish colonialism in the Americas began when Christopher Columbus and his

Spanish fleet first reached several Caribbean islands in AD 1492, including Cuba and

Hispaniola (Hassig, 1994). As Columbus led three additional transatlantic voyages from

Spain to the Caribbean over the next 11 years, Spanish colonialism gradually extended throughout the region (Bergreen, 2011). The first contact between native Mesoamericans and Spanish colonizers probably took place when Spanish conquistador Francisco

Hernández de Córdoba led a naval expedition from Cuba in 1517 that reached the

Yucatán Peninsula (Hassig, 1994; García-Martínez, 2010).

In 1519, Spanish conquistador Hernán Cortés led another naval expedition from

Cuba with an army of about 600 soldiers. This expedition, which landed near the present- day Mexican city of Veracruz (García-Martínez, 2010), eventually resulted in a military campaign against the Aztec empire. On his way to the Aztec capital city of Tenochtitlan,

Cortés formed an alliance with native residents from the state of Tlaxcala, which had long been in conflict with the Aztec empire (Restall and Schwaller, 2011). Cortés, his troops, and his Tlaxcalan allies fought against native residents and Aztec soldiers at

Xaltocan in 1521, leading to a temporary abandonment of the town (Díaz del Castillo,

1965), shortly before they conquered Tenochtitlan and laid the foundations of Spanish colonialism in the territory where present-day Mexico is located.

Tenochtitlan became known as Mexico City, capital of the Viceroyalty of New

Spain, and served as an important focus of migration from Spain (Hassig 1994; Evans,

80

2008; Restall and Schwaller, 2011). Because Mexico achieved independence from Spain in 1821, the colonial period in this region ended up extending for 300 years (García-

Martínez, 2010). During this period, the arrival of newcomers from Europe and Africa, political reorganizations, and changing patterns of regional trade substantially affected movements of people into and between native populations, leading to new types of interaction.

Migration from Europe and Africa: Patterns and Consequences

Spanish colonialism led to a significant influx of European colonists and enslaved

Africans into Mexico (Boyd-Bowman, 1976; García-Martínez, 2010). Most European newcomers were from Spain, but a small percentage migrated from other nations, including Portugal and Italy (Boyd-Bowman, 1963; Boyd-Bowman, 1973). African slaves were primarily from Cabo Verde and the Senegambian region of West Africa

(Landers, 2006; Restall and Schwaller, 2011). Early migrants from both Europe and

Africa were predominantly male: only about 6 percent of the Spanish migrants who reached colonial Mexico shortly after the conquest were women (Boyd-Bowman, 1963;

García-Martínez, 2010). The earliest European newcomers were predominantly middle- class men, but by late 16th century, Spanish settlers included a wider diversity of people, including nobles, friars, clergy, administrators, and merchants (Altman, 2001; Restall and

Schwaller, 2011). Women and children also represented a higher percentage of migrants from Spain by this time (Altman, 2001; Boyd-Bowman, 1973; Powers, 2005; Restall and

Schwaller, 2011). 81

Despite this influx of migration from Europe and Africa, native people remained a large majority. About 20,000 individuals of Spanish ancestry and 15,000 of African ancestry lived in Mexico by the mid-16th century, but they may have accounted for only

1.5 percent of the total population (García-Martínez, 2010). Because the main destinations for these newcomers were large cities, especially Mexico City (Gibson,

1964; Charlton et al., 2005; Restall and Schwaller, 2011), contact between Spanish colonists, enslaved Africans, and native individuals was probably most frequent in these locations (Socolow, 1996), and urban areas experienced high levels of gene flow from

Europe and Africa (Moreno-Estrada et al., 2014). However, scholars have long debated the extent that newcomers and native individuals interacted elsewhere in Mexico (Hassig,

1985; Lockhart, 1992; Rodríguez-Alegría et al., 2013), and the amount of gene flow from

Europe and Africa into these native communities remains unclear.

Several lines of evidence suggest that interactions between newcomers and native individuals in rural communities may have been quite rare. Notably, throughout the colonial period, most people in Mexico lived in relatively small native communities scattered across the countryside (Socolow, 1996). Spanish conquistadors did not exert much influence on these communities, in part because of their comparatively small numbers (Hamnett, 2006). Cortés created a system of encomiendas shortly after the conquest, in which native communities were subject to a Spanish landlord, or encomendero, who was tasked with collecting tribute from native people and supposedly defending and Christianizing them, but many communities remained largely independent,

82 with native leaders retaining substantial political control (Gibson, 1964; Ramírez, 1986;

Hamnett, 2006; Endfield, 2008; García-Martínez, 2010; Restall and Schwaller, 2011;

Hernández-Sánchez, 2012). Spanish colonists referred to these communities as pueblos de indios (literally, “Indians’ towns”; Restall and Schwaller, 2011), and interactions between Spanish authorities and the native peoples in such communities may have been very limited.

Other political and socio-cultural factors may have also restricted gene flow from

Europe and Africa into communities outside major population centers. Some laws barred

Spaniards and Africans from living in pueblos de indios (Restall and Schwaller, 2011) or trading with native individuals (Gibson, 1964), and marriages between Spaniards and native individuals were not customarily endorsed (Curcio-Nagy, 2011; Deed, 2011).

Thus, interactions between Spanish colonizers and native individuals in the countryside may have been largely restricted to limited contexts such as the church and the court of law (Gibson, 1964; Schwartz, 2000; Hicks, 2005). Archeological evidence is also consistent with limited acquisition of Spanish material culture in some communities during the 16th and early 17th century (Charlton, 1968). Furthermore, because Nahua languages incorporated few Spanish terms between 1521 and 1650 (Lockhart, 1992), linguistic evidence is also consistent with limited contact between newcomers and native people during this time period.

In Xaltocan, historical evidence suggests low levels of gene flow from Europe and Africa during the colonial period. Even though Spanish authorities occasionally

83 visited Xaltocan, the town remained a pueblo de indios (Begines Juárez, 1999; Hicks,

2005). In a 1569 letter addressed to the archbishop of Mexico, a Spanish priest called

Pedro Infante affirmed that there were about 9,000 native individuals but only two married Spaniards who owned land in his parish, which included Xaltocan (Montúfar,

1897:91-96). Hence, this letter suggests little genetic impact of Spanish colonists and

African slaves in Xaltocan.

However, the extent of gene flow from Europe and Africa into Xaltocan is not entirely clear, as some scholars have suggested that the veracity of Pedro Infante’s letter should be taken with caution (Rodríguez-Alegría et al., 2013). In addition, archaeological evidence from Xaltocan indicates that some native residents adopted Spanish material culture (Rodríguez-Alegría, 2010; Rodríguez-Alegría et al., 2013). While the presence of

Spanish material culture is not necessarily indicative of direct contact between newcomers and native people, this finding raises the possibility that interactions between them at Xaltocan may not have been as rare as historical documents suggest.

Similarly, some scholars have proposed that interactions between newcomers and native individuals elsewhere in the countryside were more frequent than others have assumed. For example, even though many native leaders continued to rule the pueblos de indios, some Spanish conquistadors married the daughters of those leaders to gain political influence (Simpson, 1982), and individuals of Spanish and native ancestry

(mestizos) became important political figures in native communities (Restall and

Schwaller, 2011). Furthermore, some encomenderos and their agents moved to the

84 countryside with their families (Espejo-Ponce Hunt, 1976; Ramírez, 1986), leading to increased opportunities for contact, and Spanish friars were sent to the encomiendas to help Christianize the native people (Montúfar, 1897; Simpson, 1982). Contact between

Spanish and native peoples likely became more frequent over time during the colonial era, and linguistic evidence suggests that interactions were common enough by 1650 that

Nahua-Spanish bilingualism developed in many native communities (Lockhart, 1992).

Lastly, it is possible that enslaved Africans also interacted with native communities in significant ways. For instance, even though most of the ~200,000 African slaves that were brought to New Spain during the colonial period worked in urban areas, some were used as intermediaries between encomenderos and native communities

(Curcio-Nagy, 2011; Restall and Schwaller, 2011). Accordingly, Lockhart (1992: 4) describes encomiendas as “a whole staff of Europeans, Africans, and Indians in permanent Spanish employ.”

Political Reorganizations and Changing Patterns of Regional Trade

Other changes in the political organization of native communities and regional trade patterns also affected the composition of native populations during the colonial era.

Political reorganizations of native communities stemmed in part from the Spanish decimation of those communities through war, mistreatment, slavery, and increased mortality due to the introduction of diseases from Europe and Africa, including smallpox, measles, typhus, bubonic plague, yellow fever, and malaria (Gibson, 1964; Burkhart and

Gasco, 2007). As large numbers of native communities became depopulated during the 85 late 16th and early 17th centuries, Spanish authorities relocated native peoples to congregaciones in order to concentrate depopulated, scattered communities and facilitate their continued efforts to impose tribute, Spanish laws, and Christianity on the native peoples of Mexico (Gibson, 1964; Lockhart, 1992; Endfield, 2008; Hernández-Sánchez,

2012).

The establishment of the congregación system almost certainly altered migration patterns and the genetic composition of native communities, but the specific effects of this system remain unclear. Many congregaciones were created through Spanish coercion, with native individuals being forced to relocate from their natal communities to new locations (Gibson, 1964). The capture, relocation, and trade of native slaves to distant communities was also common in some regions, especially at the beginning of the colonial period (Altman, 1976; Lockhart, 1992; English Martin, 1996). Some native individuals also migrated without direct Spanish coercion, usually to locations where the congregaciones had become important regional capitals that attracted migrants from many distant communities (Lockhart, 1992; García-Martínez, 2010).

By the end of the 16th century, Xaltocan became part of a congregación, and historical documents indicate that the town received a substantial number of newcomers.

While these documents do not specify their exact origin, the migrants were probably native individuals from nearby communities (Gibson, 1964). Hence, historical evidence suggests that political reorganizations may have increased gene flow from other native communities into Xaltocan.

86

Spanish colonialism also altered patterns of regional trade. In pre-Hispanic times, native individuals from many different communities in Mexico were involved in long- distance trade of a wide variety of goods (Lockhart, 1992). Spanish colonialism initially had little effect on trade patterns, but over time, some communities lost their status as major trade centers because demographic collapse weakened their commercial influence

(Gudmundson, 1996), demand for their certain goods decreased (Gibson, 1964), and/or access became more difficult or simply unfeasible as Spanish colonists drained major lakes and disrupted canoe-based transportation (Hassig, 1985). Trade patterns also shifted as demand for other goods increased and new communities gained economic influence

(Gibson, 1964; English Martin, 1996). When silver mines were discovered in central and northern Mexico after 1550 AD, for example, locations that were scarcely populated in pre-Hispanic times became key economic targets, attracting large numbers of workers and growing into important regional trade centers (Stuart, 1996). The creation of new roads (Gibson, 1964; Gudmundson, 1996) and the introduction of draft animals (e.g., horses and mules), wheeled carts, and wagons improved land transportation and trade among regions across Mexico (Hassig, 1985 Gudmundson, 1996; Rodríguez-Alegría et al., 2013). These changes affected patterns of movement and interaction between native communities, and facilitated the growth of trade relationships between Spanish colonizers and native individuals. Archaeological evidence shows, for instance, that some native communities used Spanish material culture from Mexico City (Rodríguez-Alegría, 2010;

87

Curcio-Nagy, 2011; Rodríguez-Alegría et al., 2013) and Spanish people in Mexico City used native pottery (Rodríguez-Alegría, 2005).

In Xaltocan, historical and archaeological evidence suggests that changing patterns of regional trade promoted prosperity (Gibson, 1964) and increased gene flow from other native populations. Although Pablo Infante described Xaltocan as “very poor” by the mid-16th century (Montúfar, 1897: 96), the town emerged as an important center of trade and became more prosperous during the colonial period (Gibson, 1964; Rodríguez-

Alegría, 2008). In addition, archaeological research has shown that Xaltocan obtained obsidian from a larger number of sources during the colonial period than in pre-Hispanic times, and ceramics from the colonial period came from a wide variety of locations, including Cuauhtitlan, Otumba, and Texcoco (Rodríguez-Alegría et al., 2013). Thus, trade from and into Xaltocan likely resulted in frequent interactions with individuals from other native communities.

Previous Genetic Research

Previous studies have examined the genetic effects on native populations in

Mexico following the Spanish conquest. Most of such studies have analyzed mtDNA.

Because the mitochondrial genome, present in both females and males, is maternally inherited and does not undergo recombination, mtDNA is an excellent tool for studying maternal ancestry. Previous genetic research has shown that the vast majority of individuals in native populations in Mexico belong to one of four major founding Native

American mtDNA haplogroups (A2, B2, C1, and D1) (Torroni et al., 1994; Peñaloza- 88

Espinosa et al., 2007; Sandoval et al., 2009; Kemp et al., 2010; Perego et al., 2010;

Gorostiza et al., 2012; González-Martín et al., 2015). However, a small percentage of individuals in some native populations belong to mtDNA haplogroups that were present in Africa and/or Europe but not in the Americas in pre-Hispanic times (Peñaloza-

Espinosa et al., 2007).

Other studies have analyzed Y-DNA. Because the Y chromosome, only present in males, is paternally inherited and possesses a region that does not recombine, Y-DNA is a useful tool for studying paternal ancestry. Even though these studies have also shown that most males in native Mexican populations belong to the founding Native American Y-

DNA haplogroup Q, a considerable percentage of males in some populations belong to

Y-DNA haplogroups that were present in Africa and/or Europe but not in the Americas in pre-Hispanic times (Torroni et al., 1994; Kemp et al., 2010; O’Rourke and Raff, 2010;

Sandoval et al., 2012). Therefore, mtDNA and Y-DNA evidence is consistent with sex- biased gene flow from Europe and Africa into native populations in Mexico, where male newcomers have contributed more than female newcomers.

Two studies analyzed large numbers of genome-wide markers in individuals from native populations in Mexico (Reich et al., 2012; Moreno-Estrada et al., 2014). Because genome-wide markers provide a much more complete representation of the human genome and are bi-parentally inherited, they represent an extremely useful tool for studying overall individual ancestry and population history. These studies have shown that native populations generally have low levels of European and African admixture

89

(Reich et al., 2012; Moreno-Estrada et al., 2014). Overall, mtDNA, Y-DNA, and genome-wide SNP studies have clarified the genetic effects of Spanish colonialism in certain regions in Mexico. However, the genetic impact of Spanish colonialism may have varied substantially across different regions (Green et al., 2000; Sandoval et al., 2012).

Finally, because patterns of genetic diversity before and after the colonial period in a single native population in Mexico have never been compared, no study has directly examined temporal changes in genetic diversity across the colonial period. Here, I present data from both pre-Hispanic and modern residents of Xaltocan, allowing this study to overcome an important methodological limitation of previous research on the genetic effects after the Spanish conquest.

METHODS

Samples

To characterize patterns of genetic diversity in pre-Hispanic Xaltocan, I sampled skeletal remains (teeth or bone) from 16 ancient individuals. This group of pre-Hispanic individuals includes all of the post-1395 AD individuals that I previously analyzed

(Mata-Míguez et al., in prep.). Ancient DNA data from pre-1395 AD individuals from

Xaltocan (Mata-Míguez et al., in prep.) were excluded here because my previous research suggests that the pre-1395 and post-1395 individuals are unlikely to have belonged to the same biological population due to demographic changes in Xaltocan across the Aztec imperial transition (Mata-Míguez et al., 2012; Mata-Míguez et al., in prep.). Because

90 post-1395 individuals are more immediately anterior to the Hispanic period, I considered them alone to characterize patterns of genetic diversity in pre-Hispanic Xaltocan just before the onset of Spanish colonialism.

To characterize patterns of genetic diversity in modern Xaltocan, I analyzed DNA from the present-day residents of this town. In July 2013, I obtained permits from the

Ayuntamiento Constitucional (Town Council), the Consejo de Participación Ciudadana

(literally, Board of Citizen Participation), and the town’s delegados (delegates), and then collected saliva samples with individual informed consent from 47 residents in Xaltocan.

The Institutional Review Board of the University of Texas at Austin also approved the collection and analysis of these samples (protocol #2012-05-0105). To collect and preserve saliva samples, we used Oragene•DISCOVER (OGR-500) kits (DNA Genotek

Inc.). For all study participants, I documented their sex, place of birth, and language, and their parent’s and grandparents’ place of birth and language. I preferentially collected samples from individuals whose maternal grandmother and paternal grandfather were born in Xaltocan. By taking this approach, I minimized the effects of mtDNA and/or Y-

DNA gene flow into Xaltocan over the last two generations. Of those 47 residents, 42 reported that her or his maternal grandmother was born in Xaltocan (Table 1). Of the 27 male study participants, 23 reported that his paternal grandfather was born in Xaltocan

(Table 1). In addition, I preferentially collected samples from individuals who are not closely related (i.e., parents and their children, or siblings) because sampling close relatives can skew population allele frequencies. Among the 47 individuals sampled,

91 there are only three pairs who are parent-offspring or siblings. As comparative data for some analyses, I used samples from other native populations in Mexico that have been previously published (Torroni et al., 1994; Malhi et al., 2003; Peñaloza-Espinosa et al.,

2007; Sandoval et al., 2009; Kemp et al., 2010; Sandoval et al., 2012).

DNA Extraction and Genotyping

DNA from the ancient samples was extracted as described elsewhere (Bolnick et al., 2012; Mata-Míguez et al., 2012). To extract DNA from the saliva samples, I used the prepIT•L2P kit (DNA Genotek Inc.) and followed the manufacturer’s protocol. For the ancient DNA extracts, I determined mtDNA haplotypes (specific sequences) and haplogroups (monophyletic group of haplotypes that are closely related) following

Bolnick et al. (2012). For the modern DNA extracts, I amplified nucleotide positions

(nps) 15,960-16,566 of the control region of the mitochondrial genome following Kemp et al. (2010). I determined Y-DNA haplogroups Q-L54(xM3) and Q-M3 in the ancient extracts as described elsewhere (Mata-Míguez et al., in prep.). For the modern extracts, I determined Y-DNA haplogroup Q-L54(xM3) following Battaglia et al. (2013), Y-DNA haplogroup Q-M3 following Bolnick et al. (2006), and Y-DNA haplogroups C, P, and R following Underhill et al. (2001). When sequencing was necessary for Y-DNA analysis, I submitted amplified fragments to the DNA Sequencing Facility at the University of

Texas at Austin for direct sequencing, with sequencing performed in both directions. I visualized and edited sequence data using Sequencher 5.0.1.

92

In all analyses of mtDNA haplotype data, I excluded nucleotide position (np)

16,183 because mutations at this site are strictly dependent on the presence of a C at np

16,189 (Pfeiffer et al., 1999). I also excluded insertions in poly-C stretches due to uncertainty in the exact position of such mutations. In analyses that only included mtDNA haplotype data from pre-Hispanic and modern Xaltocan, I analyzed nps 16,035-

16,382 in order to maximize the number of individuals and nucleotides that could be used. In analyses that included mtDNA haplotype data from pre-Hispanic Xaltocan, modern Xaltocan, and other native populations in Mexico, I analyzed nps 16,055-16,382.

Statistical Analyses

I performed statistical analyses to (1) characterize patterns of genetic variation in pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in

Mexico, (2) ascertain genetic relationships between individuals from pre-Hispanic

Xaltocan, modern Xaltocan, and other modern native populations in Mexico, (3) estimate genetic distances between pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in Mexico, (4) evaluate whether genetic distances between populations are correlated with geographic and/or temporal distances or not, and (5) assess how likely different demographic scenarios are to explain mtDNA data from pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in Mexico.

To characterize patterns of genetic variation in pre-Hispanic Xaltocan, modern

Xaltocan, and other modern native populations in Mexico, I estimated standard indexes of genetic variation using Arlequin 3.5.1.2 (Excoffier and Lischer, 2010). I estimated 93 haplogroup and haplotype diversity (h), which are defined as the probability that two randomly chosen haplogroups or haplotypes, respectively, are different in the population sampled. I also estimated nucleotide diversity (π), which indicates the probability that two randomly chosen homologous nucleotide positions are different (Excoffier and

Lischer, 2010).

To ascertain genetic relationships between individuals from pre-Hispanic

Xaltocan, modern Xaltocan, and other modern native populations in Mexico, I created median-joining networks (Bandelt et al., 1999) among haplotypes within each mtDNA haplogroup using Network 4.6.1.0 (www.fluxus-engineering.com). These networks were used to determine phylogenetic relationships between haplotypes and to illustrate the extent of haplotype sharing within and between populations. Because preliminary networks showed high levels of reticulation, I followed Kemp et al. (2010) in applying a default weight of 10 to all sites and down-weighting polymorphic sites with higher relative mutation rates as estimated by Meyer et al. (1999). Sites with more than 4-fold higher rates were down-weighted to 4, sites with 3-fold higher rates were down-weighted to 5, and sites with 2-fold higher rates were down-weighted to 6. When constructing these networks, I excluded individuals who did not belong to founding Native American mtDNA haplogroups.

To estimate genetic distances between pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in Mexico, I performed two sets of statistical analyses. First, I computed FST values based on haplogroup and haplotype data to

94 estimate genetic distances between pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in Mexico using Arlequin. When estimating FST values based on haplotypes, I computed a distance matrix using pairwise differences with a gamma value equal to 0. I then used the FST values as variables in multidimensional scaling

(MDS) analysis to generate a two-dimensional representation of genetic distances between populations using XLSTAT (Addinsoft SARL). Second, I carried out exact tests of population differentiation between pre-Hispanic and modern Xaltocan using Arlequin.

These tests evaluate the null hypothesis that haplogroup or haplotype frequencies were identical in pre-Hispanic and modern Xaltocan. If the null hypothesis is rejected, mtDNA haplogroups or haplotypes in the two population samples are unlikely to have been drawn from the same population. For these analyses, I excluded individuals who did not belong to founding Native American mtDNA haplogroups.

To evaluate whether genetic distances between populations are correlated with geographic and/or temporal distances, I performed Mantel tests (Mantel, 1967) using

Arlequin. Mantel tests calculate the correlation between two distance matrices while taking into account that pairwise geographic distances between populations are not independent from each other and temporal distances between populations are not independent from each other. To assess these correlations, Mantel tests evaluate the null hypothesis that the correlation coefficient (rXY) equals 0 (i.e., that genetic distances are not correlated with geographic and/or temporal distances) using a permutational approach. For these tests, I used FST values based on mtDNA haplotypes as genetic

95 distances between populations. To estimate geographic distances between populations, I determined their approximate geographic location using information from their original publication, and assigned geographic coordinates (latitude and longitude) to those locations using Google Earth (Table 7). Based on these geographic coordinates, I then calculated great circle distances between populations (Table 8) at www8.nau.edu/cvm/latlongdist.html (Chris Michels, Northern Arizona University). To estimate temporal distances between populations, I assigned pre-Hispanic Xaltocan the date AD 1475 (average based on radiocarbon analyses) and all of the modern populations, including Xaltocan, the date AD 2013 (when the samples from modern

Xaltocan were collected). For these analyses, I excluded individuals who did not belong to founding Native American mtDNA haplogroups. When evaluating whether genetic distances are correlated with geographic distances, I only included modern populations (I excluded pre-Hispanic Xaltocan).

To assess how likely different demographic scenarios are to explain mtDNA data from pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in

Mexico, I used BayeSSC (Excoffier et al., 2000; Anderson et al., 2005). This program runs simulations using a coalescence-based approach with different parameters (e.g., population size, rate of population growth, mutation rate) to generate trees. Summary statistics are then calculated from the simulated trees and compared to summary statistics from observed data. I used this approach to evaluate whether genetic drift and mutation alone could explain the observed genetic differences between pre-Hispanic and modern

96

Xaltocan. To this end, I ran simulations testing different numbers of generations between pre-Hispanic and modern Xaltocan as well as different female effective population sizes

(Nef) for pre-Hispanic Xaltocan (Figure 2).

For the number of generations between pre-Hispanic and modern Xaltocan, I used two values: 20 and 25. Assuming a generation time of 25 years (Reynolds et al., 2015), the first value (20) situates pre-Hispanic Xaltocan in AD 1521, when Hernán Cortés conquered this town. Thus, this value of 20 generations indicates the minimal temporal distance between pre-Hispanic and modern Xaltocan individuals. The second value (25) situates pre-Hispanic in AD 1395. I used this value to account for the fact that some pre-

Hispanic individuals may date back to 1395.

The values considered for female effective population sizes were also based on archaeological and historical evidence. The Instituto Nacional de Estadística y Geografía

(Statistics and Geography National Institute) in Mexico has estimated that around 8,500 people live in Xaltocan today (INEGI, 2008), so I used this census size (which corresponds to a female effective population size of 1,275; Cabana et al., 2008) to characterize the effective population size of modern Xaltocan in all simulations. For the female effective population sizes in pre-Hispanic Xaltocan, I used three values: 50, 750, and 1,275. The first value (50), which corresponds to a census size of 333 individuals

(Cabana et al., 2008), represents my minimum estimate of population size for pre-

Hispanic Xaltocan (Mata-Míguez et al., 2012). This value corresponds to a scenario in which Xaltocan experienced a strong population expansion (from Nef = 50 to Nef = 1,275)

97 during the colonial period. The second value (750), which corresponds to a census size of

5,000 individuals (Cabana et al., 2008), represents my maximum estimate of population size based on Sanders et al. (1979). This value corresponds to a scenario in which

Xaltocan experienced a mild population expansion (from Nef = 750 to Nef = 1,275) during the colonial period. The third value (1,275) is unlikely to characterize population size in pre-Hispanic times, but I used this value to create a scenario in which population size has remained constant and the effects of genetic drift are smaller than those present in simulations modeling population expansion.

Following Bramanti et al. (2009), Fehren-Schmitz et al. (2014), and Reynolds et al. (2015), I used a generation time of 25 years, a fixed mutation rate of 1.642733 x 10-7

(Soares et al., 2009), a transition:transversion ratio of 0.9841 (Kimura, 1980), and a gamma distribution of mutation rates with shape parameters of 0.205 (theta) and 10

(kappa) (Ho and Endicott, 2008). Each simulation was run 10,000 times. To evaluate how likely the observed mtDNA haplotype data are given each combination of parameters, I used the number of shared haplotypes as the summary statistic. P-values indicate what fraction of simulations yielded a value for this summary statistic that was equal to or smaller than what I observed between pre-Hispanic and modern Xaltocan.

RESULTS AND DISCUSSION

In post-1395 Xaltocan, we were able to determine mtDNA haplogroups and haplotypes in all of the 16 individuals (Mata-Míguez et al., in prep.). In this temporal group, ten individuals (62.5%) belonged to haplogroup A, three individuals (18.8%) 98 belonged to haplogroup B, one individual (6.2%) belonged to haplogroup C, and two individuals (12.5%) belonged to haplogroup D. These mtDNA haplogroup frequencies are also consistent with those found in previous studies of ancient and modern native populations in Mexico (Torroni et al., 1994; González-Oliver et al., 2001; Malhi et al.,

2003; Kemp et al., 2005; Peñaloza-Espinosa et al., 2007; Sandoval et al., 2009; Kemp et al., 2010).

In modern Xaltocan, we were able to determine mtDNA haplogroups and haplotypes in all of the 47 individuals (Table 1). In this temporal group, 26 individuals

(55.3%) belong to haplogroup A, 12 individuals (25.5%) belong to haplogroup B, and 9 individuals (19.1%) belong to haplogroup C. Modern Xaltocan is more diverse than pre-

Hispanic Xaltocan in terms of mtDNA haplogroup and haplotype diversity (Tables 2 and

3). However, pre-Hispanic Xaltocan is more diverse than modern Xaltocan in terms of mtDNA nucleotide diversity (Table 3).

Gene Flow from Europe and Africa

Early colonial documents indicate that Xaltocan remained a pueblo de indios that

Spanish colonists rarely visited (Montúfar, 1897; Hicks, 2005), but questions remain about up to what extent native communities remained isolated from Spanish colonists.

This study showed that all of the modern residents belong to founding Native American mtDNA haplogroups, and all of the modern male residents with a parental grandfather from Xaltocan belong to the founding Native American Y-DNA haplogroup Q (Table 1).

Thus, these results are consistent with low, or perhaps non-existent, levels of both 99 female- and male-mediated gene flow from Europe or Africa into Xaltocan during the colonial period. This finding is in agreement with early colonial documents, which suggest that newcomers from Europe and Africa rarely visited Xaltocan (Montúfar,

1897). Furthermore, these results are consistent with previous genetic evidence showing that most individuals from modern native populations in Mexico belong to one of four major founding Native American mtDNA haplogroups (A, B, C, or D; Table 2), and most males from those populations belong to founding Native American Y-DNA haplogroup

Q (Table 5).

In addition, this study showed that two out of three males with a paternal grandfather who was not from Xaltocan belong to non-founding Native American Y-

DNA haplogroups (Table 2). This observation provides evidence for a significant introduction of non-founding Native American Y-DNA haplogroups into Xaltocan over the last two generations.

Gene Flow Between Xaltocan and Other Native Populations in Mexico

Historical and archaeological evidence suggest that political reorganizations and changing patterns of regional trade increased gene flow from other native communities into Xaltocan. Consistent with this evidence, this study found significant levels of genetic differentiation between pre-Hispanic and modern Xaltocan. Networks show that pre-

Hispanic and modern Xaltocan do not share any haplotypes (Figures 3, 4, 5, and 6).

However, modern Xaltocan shares some derived Native American mtDNA haplotypes with modern native populations (Figures 3 and 5), which could be due to recent gene 100 flow between these populations. In addition, the MDS plot based on FST values shows that there is a substantial degree of genetic differentiation between pre-Hispanic and modern Xaltocan (Figure 7), and the test of population differentiation indicates that the pre-Hispanic and modern Xaltocan mtDNA haplotypes are unlikely to represent the same biological population (P < 0.001).

This kind of genetic change over time can result from a variety of evolutionary processes, including gene flow, genetic drift, and mutation. Coalescent-based simulations designed to identify the relevant processes indicated that demographic scenarios characterized by plausible parameter values (e.g., for population size and population growth) for ancient Xaltocan, realistic mutation rates, and the absence of migration are unlikely to explain the lack of shared mtDNA haplotypes between pre-Hispanic and modern Xaltocan (Table 9). Because genetic drift and mutation alone do not explain the observed genetic differences between these temporal populations, this result suggests that gene flow played an important role in shaping such genetic differences.

Finally, this study shows that geographic distances do not explain mtDNA similarities between modern native populations in Mexico (Figure 8). Specifically, a

Mantel test failed to reject the null hypothesis that genetic distances between modern native populations, including modern Xaltocan, are uncorrelated with geographic distances (P=0.541). This finding suggests that factors other than isolation by distance played an important role in shaping the pattern of genetic similarities and differences among populations in Mexico today. Movements of people due to political

101 reorganizations and/or trade during colonial times, for instance, may have had a greater influence than geographic distances on determining similarities between Xaltocan and other native communities.

CONCLUSION

By comparing pre-Hispanic and modern patterns of mtDNA and Y-DNA diversity, this study shows that Xaltocan, a native community in central Mexico, has experienced little gene flow from Europe and Africa, but important levels of gene flow from other native populations since the onset of Spanish colonialism. These results are consistent with historical and archeological evidence, which suggest that Xaltocan received a low number of Spanish and African newcomers yet a high number of native migrants due to political reorganizations and changing patterns of regional trade. This study illustrates how anthropologists can use genetic evidence to test historical and archaeological hypothesis on the effects of past colonial expansions.

102

Figure 2.1. Map of Xaltocan within the Basin of Mexico in ancient times (when this town was an island in Lake Xaltocan).

103

Figure 2.2. BayeSSC demographic scenarios.

104

Figure 2.3. Network for mtDNA haplogroup A. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

105

Figure 2.4. Network for mtDNA haplogroup B. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

106

Figure 2.5. Network for mtDNA haplogroup C. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

107

Figure 2.6. Network for mtDNA haplogroup D. Mutations identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). Circle with a thick black line represents the founding haplotype. Circle size is proportional to the number of samples. White circles represent haplotypes not found in the populations analyzed here.

108

Figure 2.7. MDS plot based on mtDNA haplotype data.

109

Figure 2.8. Genetic distance versus geographic distance in modern native population in Mexico, including modern Xaltocan. Each data point represents a pairwise comparison of populations.

110

Table 2.1. Sex, recent ancestry, Y-DNA and mtDNA haplogroups, and mtDNA haplotypes for all of the 47 present-day residents of Xaltocan.

Individual Maternal Paternal mtDNA Sex grandmother grandfather Y-DNA (mtDNA Haplogroup) Haplotype ID born in born in Haplogroup mtDNA Haplotype1 Range Xaltocan Xaltocan

Female No 15,986- XalMod1 Yes (B2) 182C, 183C, 189C, 217C, 519C 16,528

Male 15,960- XalMod2 Yes Yes Q-M3 (A2) 111T, 223T, 290T, 319A, 362C 16,566

Female 15,960- XalMod3 Yes Yes (C1) 51G, 223T, 298C, 311C, 325C, 327T, 519C 16,566

Female 15,960- XalMod4 Yes Yes (A2) 111T, 223T, 290T, 319A, 362C 16,566

Male 15,960- XalMod5 Yes Yes Q-M3 (A2) 111T, 223T, 278T, 290T, 319A, 519C, 524G 16,566

111 Table 2.1 (continued)

Female 15,960- XalMod6 Yes Yes (C1) 223T, 227G, 298C, 325C, 327T 16,566

Male 15,995- XalMod7 Yes Yes Q-M3 (B2) 182C, 183C, 189C, 217C, 362C, 519C 16,404

Female 15,996- XalMod8 Yes Yes (B2) 183C, 189C, 217C, 519C 16,544

Male 15,960- XalMod9 Yes Yes Q-M3 (A2) 111T, 223T, 290T, 319A, 362C 16,566

Female 15,974- (B2) 182C, 183C, 189C, 217C, 239T, 311Y2, 353T, XalMod10 Yes Yes 16,542 519C

Female 16,035- XalMod11 Yes Yes (B2) 182C, 183C, 189C, 271C, 519C 16,541

Male Unknown 15,987- (A2) 111T, 189C, 209C, 223T, 290T, 319A, 362C, XalMod12 Yes Q-M3 16,542 519C

Male 15,960- (C1) 51G, 223T, 298C, 311C, 325C, 327T, 488Y2, XalMod13 Yes Yes Q-L54(xM3) 16,566 519C

112 Table 2.1 (continued) Male 15,960- (A2) 111T, 223T, 290T, 319A, 362C, 390A, 519C, XalMod14 Yes Yes Q-M3 16,566 525G

Male 15,960- (A2) 111T, 223T, 290T, 319A, 362C, 390A, 519C, XalMod15 Yes Yes Q-L54(xM3) 16,566 525G

Female 15,960- XalMod16 Yes No (A2) 75C, 111T, 223T, 239T, 290T, 319A, 362C 16,566

Female 15,960- XalMod17 Yes Yes (C1) 223T, 227G, 298C, 325C, 327T 16,566

Male 15,960- XalMod18 Yes Yes Q-M3 (A2) 111T, 129A, 217C, 223T, 290T, 319A, 362C 16,566

Female 15,995- XalMod19 Yes Yes (B2) 182C, 183C, 189C, 217C, 519C 16,528

Male 15,960- XalMod20 Yes Yes Q-L54(xM3) (A2) 111T, 175G, 223T, 290T, 300G, 319A, 362C 16,566

Male 15,960- (C1) 51G, 223T, 298C, 311C, 325C, 327T, 488Y2, XalMod21 Yes Yes Q-L54(xM3) 16,566 519C

113 Table 2.1 (continued)

Female 15,960- XalMod22 No Yes (A2) 111T, 223T, 290T, 311C, 319A, 362C, 526A 16,566

Female 15,978- XalMod23 Yes Yes (B2) 182C, 183C, 189C, 217C, 362C, 519C 16,544

Male Non-P3, 15,960- (A2) 111T, 223T, 290T, 319A, 362C, 390A, 519C, XalMod24 Yes No Non-C 16,566 525G

Male 15,960- XalMod25 Yes Yes Q-M3 (C1) 223T, 227G, 298C, 325C, 327T 16,566

Female No 15,960- XalMod26 No (A2) 111T, 223T, 290T, 319A, 325T, 362C 16,566

Male 15,960- XalMod27 Yes Yes Q-L54(xM3) (A2) 75C, 111T, 223T, 239T, 290T, 319A, 362C 16,542

Female 15,983- XalMod28 Yes Yes (B2) 182C, 183C, 189C, 217C, 519C 16,528

Male 15,960- XalMod29 No No Q-M3 (C1) 153A, 223T, 298C, 325C, 327T 16,566

114 Table 2.1 (continued)

Female 15,960- (C1) 51G, 223T, 298C, 311C, 325C, 327T, 488Y2, XalMod30 Yes Yes 16,566 519C

Male 15,982- XalMod31 Yes Yes Q-M3 (B2) 182C, 183C, 189C, 217C, 362C, 519C 16,544

Female 15,960- XalMod32 Yes Yes (A2) 75C, 111T, 223T, 239T, 290T, 319A, 362C 16,566

Male 15,974- XalMod33 Yes Yes Q-M3 (A2) 111T, 189C, 209C, 223T, 290T, 319A, 362C 16,541

Male 15,960- XalMod34 No No R (A2) 111T, 223T, 290T, 319A, 362C 16,566

Female 15,960- XalMod35 Yes Yes (A2) 111T, 223T, 290T, 291T, 319A, 362C 16,566

Female 15,960- XalMod36 Yes Yes (A2) 75C, 111T, 223T, 239T, 290T, 319A, 362C 16,566

Male 15,995- XalMod37 Yes Yes Q-L54(xM3) (B2) 182C, 183C, 189C, 217C, 519C 16,542

115 Table 2.1 (continued)

Male 15,960- XalMod38 Yes Yes Q-L54(xM3) (C1) 51G, 223T, 298C, 311C, 325C, 327T, 519C 16,566

Male 15,960- XalMod39 Yes Yes Q-M3 (A2) 111T, 223T, 290T, 319A, 362C 16,566

Female 15,960- XalMod40 Yes Yes (A2) 111T, 223T, 290T, 299G, 319A, 362C 16,494

Male 15,960- XalMod41 Yes Yes Q-L54(xM3) (A2) 111T, 175G, 223T, 290T, 300G, 319A, 362C 16,566

Male 15,960- XalMod42 Yes Yes Q-M3 (A2) 111T, 223T, 290T, 319A, 362C 16,566

Male 15,960- XalMod43 Yes Yes Q-M3 (A2) 111T, 223T, 278T, 290T, 319A, 519C, 524G 16,566

Male 15,960- XalMod44 No Yes Q-M3 (A2) 111T, 223T, 290T, 319A, 362C, 387G, 519C 16,566

Male 15,983- XalMod45 Yes Yes Q-L54(xM3) (B2) 182C, 183C, 189C, 217C, 362C, 519C 16,542

116 Table 2.1 (continued) Male 15,960- (A2) 111T, 223T, 278T, 290T, 319A, 362C, 519C, XalMod46 Yes Yes Q-M3 16,566 524G

Female 15,974- XalMod47 Yes Yes (B2) 182C, 183C, 189C, 217C, 362C, 519C 16,542

1 Mutations are specified with the last three digits within range 16,051-16,566 and identified with reference to the Cambridge Reference Sequence (Anderson et al., 1981; Andrews et al., 1999). 2 Y : C-T heteroplasmy. 3 Y-DNA haplogroup P includes haplogroups Q and R.

117 Table 2.2. mtDNA haplogroups and mtDNA haplogroup diversity in pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in Mexico.

Population Reference n1 Founding Non-Founding mtDNA Native American Native American Haplogroup mtDNA Haplogroups mtDNA Diversity2 (%) Haplogroups (h) (%) A B C D X J L NS3

Pre-Hispanic This study 16 10 3 1 2 0 0 0 0 0.592 Xaltocan (62.5) (18.8) (6.2) (12.5) (0.0) (0.0) (0.0) (0.0)

Modern Xaltocan This study 47 26 12 9 0 0 0 0 0 0.605 (55.3) (25.5) (19.2) (0.0) (0.0) (0.0) (0.0) (0.0)

Maternal This study 42 22 12 8 0 0 0 0 0 0.622 grandmother (52.4) (28.6) (19.0) (0.0) (0.0) (0.0) (0.0) (0.0) from Xaltocan Maternal This study 5 4 0 1 0 0 0 0 0 0.400 grandmother not (80.0) (0) (20.0) (0.0) (0.0) (0.0) (0.0) (0.0) from Xaltocan Cora Kemp et al., 72 22 37 10 3 0 0 0 0 0.630 2010 (30.5) (51.4) (13.9) (4.2) (0.0) (0.0) (0.0) (0.0)

Huichol Kemp et al., 62 19 33 10 0 0 0 0 0 0.607 2010 (30.7) (53.2) (16.1) (0.0) (0.0) (0.0) (0.0) (0.0)

Maya Sandoval et 52 32 9 8 3 0 0 0 0 0.575 118 Table 2.2 (continued)

al., 2009 (61.5) (17.3) (15.4) (5.8) (0.0) (0.0) (0.0) (0.0)

Mixe Kemp et al., 52 16 15 15 6 0 0 0 0 0.740 2010 (30.8) (28.8) (28.8) (11.5) (0.0) (0.0) (0.0) (0.0)

Mixtec Kemp et al., 67 45 14 5 3 0 0 0 0 0.505 2010 (67.1) (20.9) (7.5) (4.5) (0.0) (0.0) (0.0) (0.0)

Mixtec Alta Peñaloza- 16 6 9 1 0 0 0 0 0 0.575 Espinosa et (37.5) (56.2) (6.2) (0.0) (0.0) (0.0) (0.0) (0.0) al., 2007 Mixtec Baja Peñaloza- 11 7 2 1 1 0 0 0 0 0.600 Espinosa et (63.6) (18.2) (9.1) (9.1) (0.0) (0.0) (0.0) (0.0) al., 2007 Nahua Atocpan Kemp et al., 50 19 20 9 2 0 0 0 0 0.675 2010 (38.0) (40.0) (18.0) (4.0) (0.0) (0.0) (0.0) (0.0)

Nahua Peñaloza- 41 19 14 3 5 0 0 0 0 0.665 Chilacachapa Espinosa et (46.3) (34.2) (7.3) (12.2) (0.0) (0.0) (0.0) (0.0) al., 2007 Nahua Coyolillo Peñaloza- 38 26 3 0 6 0 0 3 0 0.424 Espinosa et (68.4) (7.9) (0.0) (15.8) (0.0) (0.0) (7.9) (0.0) al., 2007 Nahua Cuetzalan Kemp et al., 46 29 9 7 1 0 0 0 0 0.553 2010 (63.0) (19.6) (15.2) (2.2) (0.0) (0.0) (0.0) (0.0)

Nahua Sandoval et 10 4 1 3 2 0 0 0 0 0.778 Ixhuatlancillo al., 2009 (40.0) (10.0) (30.0) (20.0) (0.0) (0.0) (0.0) (0.0)

Nahua Necoxtla Sandoval et 25 12 13 0 0 0 0 0 0 0.520 al., 2009 (48.0) (52.0) (0.0) (0.0) (0.0) (0.0) (0.0) (0.0)

119 Table 2.2 (continued)

Nahua Xochimilco Sandoval et 35 27 5 3 0 0 0 0 0 0.388 al., 2009 (77.1) (14.3) (8.6) (0.0) (0.0) (0.0) (0.0) (0.0)

Nahua Zitlala Sandoval et 14 14 0 0 0 0 0 0 0 0.000 al., 2009 (100.0) (0.0) (0.0) (0.0) (0.0) (0.0) (0.0) (0.0)

Otomi Sandoval et 68 27 17 20 4 0 0 0 0 0.700 al., 2009 (39.7) (25.0) (29.4) (5.9) (0.0) (0.0) (0.0) (0.0)

Pima Sandoval et 98 11 3 82 1 0 0 0 1 0.274 al., 2009 (11.2) (3.0) (83.7) (1.0) (0.0) (0.0) (0.0) (1.0)

Purepecha Sandoval et 34 20 3 8 3 0 0 0 0 0.600 al., 2009 (58.8) (8.8) (23.5) (8.8) (0.0) (0.0) (0.0) (0.0)

Tarahumara Kemp et al., 73 25 21 23 4 0 0 0 0 0.707 2010 (34.2) (28.8) (31.5) (5.5) (0.0) (0.0) (0.0) (0.0)

Triqui Sandoval et 10 77 30 0 0 0 0 0 0 0.407 al., 2009 7 (72.0) (28.0) (0.0) (0.0) (0.0) (0.0) (0.0) (0.0)

Tzeltal Peñaloza- 35 21 5 5 4 0 0 0 0 0.603 Espinosa et (60.0) (14.3) (14.3) (11.4) (0.0) (0.0) (0.0) (0.0) al., 2007 Zapotec Kemp et al., 88 36 19 25 5 0 1 2 0 0.689 2010 (40.9) (21.6) (28.4) (5.7) (0.0) (1.1) (2.3) (0.0)

1 n: total sample size. 2 mtDNA haplogroup diversity was calculated taking into account only founding Native American mtDNA haplogroups. 3 NS: the authors reported the presence of non-founding Native American mtDNA haplogroups, but they did not list specific haplogroups.

120 Table 2.3. mtDNA haplotype diversity and nucleotide diversity in pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in Mexico based on nps 16,055-16,382.

Population Reference n1 Founding mtDNA mtDNA Native American Haplotype Nucleotide mtDNA Haplogroups Diversity (h) Diversity (π) (%) A B C D

Pre-Hispanic Xaltocan This study 16 10 3 1 2 0.808 0.019 (62.5) (18.8) (6.2) (12.5)

Modern Xaltocan This study 47 26 12 9 0 0.916 0.018 (55.3) (25.5) (19.2) (0.0)

Maternal This study 42 22 12 8 0 0.914 0.018 grandmother from (52.4) (28.6) (19.0) (0.0) Xaltocan Maternal This study 5 4 0 1 0 0.900 0.011 grandmother not (80.0) (0) (20.0) (0.0) from Xaltocan Mixtec Sandoval et al., 2009; 84 60 14 6 4 0.945 0.016 Kemp et al., 2010 (71.4) (16.7) (7.1) (4.8)

Nahua Atocpan Kemp et al., 2010 44 17 18 8 1 0.957 0.023 (38.6) (40.9) (18.2) (2.3)

Nahua Cuetzalan Malhi et al., 2003; 34 23 3 7 1 0.973 0.019 Kemp et al., 2010 (67.6) (8.8) (20.6) (3.0) 121 Table 2.3 (continued)

Nahua Ixhuatlancillo Sandoval et al., 2009 10 4 1 3 2 0.956 0.023 (40.0) (10.0) (30.0) (20.0)

Nahua Necoxtla Sandoval et al., 2009 25 12 13 0 0 0.910 0.018 (48.0) (52.0) (0.0) (0.0)

Nahua Xochimilco Sandoval et al., 2009 35 27 5 3 0 0.928 0.015 (77.1) (14.3) (8.6) (0.0)

Nahua Zitlala Sandoval et al., 2009 14 14 0 0 0 0.593 0.004 (100.0) (0.0) (0.0) (0.0)

Otomi Sandoval et al., 2009 68 27 17 20 4 0.960 0.020 (39.7) (25.0) (29.4) (5.9)

Triqui Sandoval et al., 2009 107 77 30 0 0 0.548 0.013 (72.0) (28.0) (0.0) (0.0)

Zapotec Kemp et al., 2010 72 31 17 21 3 0.950 0.022 (43.1) (23.6) (29.2) (4.1)

1 n: total sample size. Individuals who do not belong to founding Native American haplogroups were not included in these analyses.

122 Table 2.4. Y-DNA haplogroups and Y-DNA haplogroup diversity in pre-Hispanic Xaltocan and modern Xaltocan.

Population Reference n1 Founding Non-Founding Y-DNA Native American Native American Haplogroup Y-DNA Haplogroups Y-DNA Haplogroups Diversity2 (h) (%) (%) Q C R Other

L54(xM3) M3

Pre-Hispanic Xaltocan Mata-Míguez et al., 9 1 8 0 0 0 0.222 in prep. (11.1) (88.9) (0.0) (0.0) (0.0)

Modern Xaltocan This study 27 9 16 0 1 1 0.480 (33.3) (59.2) (0.0) (3.7) (3.7)

Parental grandfather This study 23 9 14 0 0 0 0.498 from Xaltocan (39.1) (60.9) (0.0) (0.0) (0.0)

Parental grandfather This study 3 0 1 0 1 1 1.000 not from Xaltocan (0.0) (33.3) (0.0) (33.3) (33.3)

Unknown whether This study 1 0 1 0 0 0 1.000 parental grandparent (0.0) (100.0) (0.0) (0.0) (0.0) from Xaltocan or not 1 n: total sample size. 2 Y-DNA haplogroup diversity was calculated taking into account only founding Native American Y-DNA haplogroups.

123 Table 2.5. Y-DNA haplogroups in pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in Mexico.

Population Reference n1 Founding Other Native American (%) Y-DNA Haplogroups (%) Q C

Pre-Hispanic Xaltocan Mata-Míguez et al., in prep. 9 9 0 0 (100.0) (0.0) (0.0)

Modern Xaltocan This study 27 25 0 2 (92.6) (0.0) (7.4)

Parental grandfather from Xaltocan This study 23 23 0 0 (100.0) (0.0) (0.0)

Parental grandfather not from This study 3 1 0 2 Xaltocan (33.3) (0.0) (66.7)

Unknown whether parental This study 1 1 0 0 grandparent from Xaltocan or not (100.0) (0.0) (0.0)

Cora Kemp et al. (2010) 43 43 0 0 (100.0) (0.0) (0.0)

Huichol Kemp et al. (2010) 12 12 0 0 (100.0) (0.0) (0.0)

Mixe Kemp et al. (2010) 7 7 0 0 124 Table 2.5 (continued) (100.0) (0.0) (0.0)

Mixtec Kemp et al. (2010) 22 20 0 2 (90.9) (0.0) (9.1)

Mixtec Sandoval et al. (2012) 2 2 0 0 (100.0) (0.0) (0.0)

Nahua Atocpan Kemp et al. (2010) 7 6 0 1 (85.7) (0.0) (14.3)

Nahua Cuetzalan Kemp et al. (2010) 10 10 0 0 (100.0) (0.0) (0.0)

Nahua San Pedro Sandoval et al. (2012) 9 9 0 0 (100.0) (0.0) (0.0)

Nahua Santo Domingo Sandoval et al. (2012) 17 15 0 2 (88.2) (0.0) (11.8)

Nahua Xochimilco Sandoval et al. (2012) 22 15 0 7 (68.2) (0.0) (31.8)

Nahua Zitlala Sandoval et al. (2012) 22 19 0 3 (86.4) (0.0) (13.6)

Otomi Sandoval et al. (2012) 7 4 0 3 (57.1) (0.0) (42.9)

Pima Sandoval et al. (2012) 51 49 0 2 (96.1) (0.0) (3.9)

125 Table 2.5 (continued)

Purepecha Sandoval et al. (2012) 8 6 0 2 (75.0) (0.0) (25.0)

Tarahumara Kemp et al. (2010) 20 19 0 1 (95.0) (0.0) (5.0)

Tarahumara Sandoval et al. (2012) 18 13 0 5 (72.2) (0.0) (27.8)

Triqui Sandoval et al. (2012) 22 22 0 0 (100.0) (0.0) (0.0)

Yucatec Maya Sandoval et al. (2012) 19 19 0 0 (100.0) (0.0) (0.0)

Zapotec Kemp et al. (2010) 11 6 0 5 (54.5) (0.0) (45.4)

1 n: total sample size.

126 Table 2.6. FST values between pre-Hispanic Xaltocan, modern Xaltocan, and other modern native populations in Mexico.

Cuetzalan

Hispanic Hispanic - Pre Xaltocan Modern Xaltocan Mixtec Atocpan Nahua Nahua Nahua Ixhuatlancillo Necoxtla Nahua Xochimilco Nahua Zitlala Nahua Otomi Triqui

Modern Xaltocan 0.062

Mixtec 0.054 0.020

Nahua Atocpan 0.078 0.018 0.077

Nahua Cuetzalan 0.032 0.018 0.010 0.063

Nahua 0.051 0.007 0.049 0.004 -0.011 Ixhuatlancillo Nahua Necoxtla 0.145 0.065 0.105 0.035 0.144 0.128

Nahua 0.064 0.030 0.004 0.100 0.003 0.061 0.154 Xochimilco Nahua Zitlala 0.200 0.165 0.089 0.249 0.102 0.272 0.336 0.059

127 Table 2.6 (continued)

Otomi 0.097 0.020 0.068 0.012 0.044 -0.017 0.097 0.088 0.236

Triqui 0.118 0.044 0.020 0.112 0.063 0.138 0.114 0.027 0.113 0.119

Zapotec 0.073 0.008 0.053 0.009 0.030 -0.023 0.086 0.064 0.194 0.000 0.099

128 Table 2.7. Latitude and longitude of native populations in Mexico.

Latitude Longitude Reference

Xaltocan 19° 43' 0.26'' N 99° 2' 24.54'' W This study

Mixtec 16° 59' 48.19'' N 96° 46' 53.36'' W Sandoval et al., 2009

Zapotec 16° 47' 12.03'' N 96° 40' 8.97'' W Kemp et al., 2010

Triqui 17° 16' 3.22'' N 97° 40' 48.85'' W Sandoval et al., 2009

Otomi 20° 5' 51.41'' N 98° 42' 34.48'' W Sandoval et al., 2009

Nahua Atocpan 19° 12' 12.54'' N 99° 2' 55.32'' W Kemp et al., 2010

Nahua Cuetzalan 20° 1' 3.49'' N 97° 31' 24.13'' W Malhi et al., 2003; Kemp et al., 2010

Nahua Xochimilco 19° 15' 26.03'' N 99° 6' 10.68'' W Sandoval et al., 2009

Nahua Zitlala 17° 41' 30.65'' N 99° 11' 11.01'' W Sandoval et al., 2009

Nahua Ixhuatlancillo 18° 53' 44.96'' N 97° 8' 56.76'' W Sandoval et al., 2009

Nahua Necoxtla 18° 48' 15.43'' N 97° 11' 4.66'' W Sandoval et al., 2009

129 Table 2.8. Great circle distances (in kilometers) between native populations in Mexico.

Xaltocan Mixtec Atocpan Nahua Cuetzalan Nahua Nahua Ixhuatlancillo Necoxtla Nahua Nahua Xochimilco Zitlala Nahua Otomi Triqui

Mixtec 385.5

Nahua Atocpan 57.1 343.3

Nahua Cuetzalan 162.2 345.3 183.8

Nahua 218.7 214.9 202.8 130.9 Ixhuatlancillo Nahua Necoxtla 219.9 205.7 201.2 139.7 10.8

Nahua 51.6 351.6 8.0 186.0 209.5 208.1 Xochimilco Nahua Zitlala 225.9 267.0 168.9 312.6 253.6 245.2 174.5

Otomi 54.7 400.7 105.7 124.3 211.5 215.3 102.2 272.5

Triqui 308.1 100.2 259.6 306.6 189.8 178.9 166.4 166.8 333.2

130 Table 2.8 (continued)

Zapotec 411.3 26.3 368.6 370.8 240.2 231.1 376.8 286.0 426.0 120.2

131 Table 2.9. BayeSSC simulations.

Pre-Hispanic Pre-Hispanic Modern Modern Number of P-value * ** * ** Nc Nef Nc Nef generations 333 50 8,500 1,275 20 0.000

5000 750 8,500 1,275 20 0.000

8500 1275 8,500 1,275 20 0.000

333 50 8,500 1,275 25 0.000

5000 750 8,500 1,275 25 0.000

8500 1275 8,500 1,275 25 0.000

* Nc: census size. ** Nef: female effective population size. Nef = (Nc/2) x 0.3 (Cabana et al., 2008).

132 Chapter 3: A Genetic Test of Historically-Based and Archaeological

Hypotheses on the Effects of Spanish Colonialism: Genome-Wide DNA

Evidence from Xaltocan, Mexico

ABSTRACT

Since the Spanish conquest in 1521, Mexico has received a substantial number of immigrants from Europe and Africa. This immigration resulted in extensive admixture in locations where the newcomers mostly settled, such as Mexico City, but the extent of admixture in other communities in Mexico remains unclear. In Xaltocan, a native community in central Mexico, archaeological evidence indicates that some native residents adopted Spanish material culture, but colonial documents suggest that Spanish colonists and enslaved Africans rarely visited this town. To clarify the extent of admixture in Xaltocan, I analyzed genome-wide single nucleotide polymorphisms (SNPs) from present-day residents of this community. I found that residents with all four grandparents from Xaltocan have an average proportion of Native American ancestry of

99.4% based on autosomal SNPs and 100% based on X-chromosome SNPs. These results indicate that Xaltocan experienced very low, or perhaps non-existent, levels of admixture after the Spanish conquest. However, I found that residents with at least one grandparent who was not from Xaltocan exhibit an average proportion of Native American ancestry of 82.8% based on autosomal SNPs and 89.3% based on X-chromosome SNPs. Thus, this

133 study also suggests moderate levels of admixture in the present-day community due to immigration into Xaltocan over the last two generations.

INTRODUCTION

In 1521, Hernán Cortés conquered the Aztec capital city of Tenochtitlan and laid the foundations of Spanish colonialism in the territory comprising present-day Mexico.

Since the onset of the colonial period, Mexico has received a substantial number of immigrants from Europe and Africa. For instance, about 20,000 individuals of Spanish ancestry and 15,000 of African ancestry lived in Mexico by the mid-16th century (García-

Martínez, 2010). This immigration led to admixture in locations where the newcomers became established, such as Mexico City. However, the extent of admixture in other communities during the colonial period remains unclear. Spanish colonialism came to an end when Mexico gained independence in 1821. Since then, further immigration into

Mexico (or even between communities within Mexico) may have increased the extent of admixture in certain communities.

In Xaltocan, a native community in central Mexico, historical and archaeological evidence provide inconclusive accounts on the extent of admixture during the colonial period. Archaeological findings show that the native residents of Xaltocan adopted

Spanish material culture. While upper elites relied on Spanish dresses and weaponry to consolidate their power, lower elites and commoners used Spanish ceramics for display to improve their social status (Rodríguez-Alegría, 2010). However, colonial documents suggest little interaction between Spanish or African newcomers and the native residents 134 of Xaltocan. In a 1569 letter addressed to the archbishop of Mexico, for instance, a

Spanish priest called Pedro Infante affirmed that there were about 9,000 native individuals but only two married Spaniards who owned land in his parish, which included

Xaltocan (Montúfar, 1897). Therefore, Xaltocan provides an excellent opportunity for testing historically-based and archaeological hypotheses about the extent of admixture during the colonial period. To evaluate the genetic impact of Spanish colonialism in

Xaltocan, this study analyzed genome-wide DNA markers from present-day residents of this town.

PREVIOUS GENETIC RESEARCH

To evaluate the extent of admixture in Xaltocan, I previously analyzed mitochondrial DNA (mtDNA) and Y-chromosome DNA (Y-DNA) haplogroups from 47 present-day residents of this town (Mata-Míguez et al., in prep.). I found that all of the residents belong to founding Native American mtDNA haplogroups, and all of the 23 males whose parental grandfather was from Xaltocan belong to a founding Native

American Y-DNA haplogroup. However, out of the three males with a paternal grandfather who was not from Xaltocan, two do not belong to a founding Native

American Y-DNA haplogroup. Therefore, my previous research suggested very low, or perhaps non-existent, levels of mtDNA and Y-DNA admixture during the colonial period in Xaltocan, but some introduction of non-founding Native American Y-DNA haplogroups due to immigration into this town over the last two generations.

135

This work helped elucidate the extent and timing of admixture since the onset of

Spanish colonialism, but the conclusions that could be drawn were limited for two reasons. First, my previous research provided little information on overall genomic variation because both mtDNA and Y-DNA (especially mtDNA) represent a small proportion of the human genome. Second, mtDNA and Y-DNA are exclusively inherited maternally and paternally, respectively, so they provide a very skewed picture of individual ancestry. To overcome these limitations, this study analyzed genome-wide single nucleotide polymorphisms (SNPs) from those 47 present-day residents of

Xaltocan.

To date, only two published studies have analyzed a large number of genome- wide SNPs in individuals from native populations in Mexico (Reich et al., 2012; Moreno-

Estrada et al., 2014). These studies have shown that native populations generally have low levels of European and African admixture. However, such levels vary considerably across different regions. For instance, Seri individuals from northwestern Mexico present very low levels of European ancestry, but Triqui individuals from southern Mexico exhibit notably higher levels of European admixture (Moreno-Estrada et al., 2014). In addition, SNP data showed that urban populations such as Veracruz (where the main harbor of arrival of transoceanic migration was located) have substantially higher levels of European and African admixture than native populations (Moreno-Estrada et al.,

2014).

136

While these two studies have helped elucidate patterns of admixture in native communities in Mexico, their sampling strategies made it difficult to differentiate admixture during the colonial period (1521-1821) from post-colonial admixture. Namely, because both studies included individuals from a native community even if those individuals’ immediate ancestors (e.g., parents and grandparents) had not lived in such a community, their results could partly reflect post-colonial admixture. To help differentiate colonial from post-colonial admixture, I distinguished residents with four grandparents who inhabited Xaltocan from residents with at least one grandparent who did not inhabit Xaltocan. Although this distinction does not perfectly tell apart colonial and post-colonial admixture, at least it makes it possible to differentiate admixture that took place more than two generations ago from admixture that may have happened over the last two generations.

Finally, at least three additional studies have analyzed a large number of genome- wide SNPs in Mexican American individuals (Bryc et al., 2010; Wall et al., 2011; Bryc et al., 2015). However, patterns of genetic diversity in Mexican Americans are problematic for studying population history during the colonial period in Mexico, as such patterns may have been strongly influenced by recent historical events.

137

METHODS

Samples

Samples were collected and extracted as described in Mata-Míguez et al. (in prep.).

SNP Genotyping

A total of 627,151 SNPs were genotyped using the Axiom® Genome-Wide

Human Origins 1 Array (Affymetrix, Inc.) at the Center for Applied Genomics at the

Children’s Hospital of Philadelphia. This array has low ascertainment bias because it was developed by identifying heterozygous SNP loci in individuals of know ancestry who were sampled from 11 worldwide populations (San, Yoruba, French, Han, Papuan,

Cambodian, Bougainville, Sardinian, Mbuti, Mongolian, and Karitiana) (Patterson et al.,

2012). Therefore, this collection of SNPs represents an excellent tool to characterize genome-wide diversity and reconstruct population history in human populations.

Comparative SNP Data

I merged genome-wide SNP data from Xaltocan with equivalent data from other native populations in North America (Mixe), Central America (Cabecar), and South

America (Piapoco, Surui, Apalai, Arara, Urubu Kaapor, Xavante, Karitiana) and likely sources of admixture from Europe (Spain and Italy), northern Africa (Algeria, Egypt, and

Tunisia), and west-central Africa (Biaka, Mandenka, and Yoruba) (Cann et al., 2012;

138

Reich et al., 2012; Lazaridis et al., 2014; Skoglund et al., 2015; Reich et al., n.d.). This merged dataset contained a total of 402 individuals (Table 1). I divided the merged dataset into two subsets of SNPs: autosomal (620,731 SNPs) and X-chromosome (4,331

SNPs). Because males are haploid for most X-chromosome loci, I only included females

(n=166; Table 1) in further analyses using the X-chromosome SNP dataset.

Data Curation

Using PLINK (Purcell et al., 2007), I removed all individuals and loci whose rate of genotyping success was lower than 90 percent (Moreno-Estrada et al., 2014), I verified that no individual showed an excess or deficiency of heterozygosity (Reich et al., 2012), and pruned SNPs using a window size of 50 SNPs advanced by 10 SNPs and an R2 threshold of 0.1 in order to minimize linkage disequilibrium. Following data curation, the autosomal dataset consisted of 402 individuals and 101,463 SNPs, and the X- chromosome dataset consisted of 166 females and 825 SNPs (Table 2).

Data Analysis

To visually explore the autosomal and X-chromosome SNP datasets, I obtained two-dimensional representations of genetic distances between individuals by creating principal component analysis (PCA) plots in EIGENSOFT (Patterson et al., 2006; Price et al., 2006). I created these PCA plots using default parameters for outlier removal

(numoutlieriter=5, numoutlierevec=10, outliersigmathresh=6). To assess European and

African admixture in Xaltocan, I estimated admixture proportions in each individual

139 using the program ADMIXTURE (Alexander et al., 2009) with the autosomal and X- chromosome SNP datasets. For each dataset, I performed 19 runs in ADMIXTURE with values of K (i.e., number of genetic clusters) that ranged from 2 to 20, following

Lazaridis et al. (2014) and Moreno-Estrada et al. (2014).

RESULTS AND DISCUSSION

Early colonial documents indicate that Xaltocan remained a pueblo de indios that

Spanish colonists and enslaved Africans rarely visited (Montúfar, 1897). Thus, historical evidence suggests little admixture between native residents and newcomers from Europe and Africa during the colonial period in Xaltocan. To test this historically-based hypothesis, I analyzed autosomal and X-chromosome SNP data in modern residents of

Xaltocan in conjunction with equivalent data from individuals in other native populations in North America, Central America, and South America as well as likely source populations of admixture in Europe and Africa.

The PCA plots based on autosomal and X-chromosome SNPs clustered the vast majority of Xaltocan residents with all four grandparents from Xaltocan with Native

American individuals (Figures 1a and 1b). In the PCA plot based on autosomal SNPs, only one out of 38 resident with all four grandparents from Xaltocan (2.6%) fell outside the Native American cluster, being located closer to the European and northern African clusters. In addition, the PCA plot based on autosomal SNPs showed that, out of nine residents with at least one grandparent not from Xaltocan, four (44.4%) Xaltocan were

140 clustered with Native American individuals, but five (55.6%) fell outside the Native

American cluster and were closer to the European and northern African clusters.

ADMIXTURE analyses indicated that the most suitable modeling choice corresponded to K=4 in both the autosomal and X-chromosome datasets (Figures 2a and

2b). For K=4, European and west-central African individuals’ ancestry corresponds almost entirely with the blue and red genetic components, respectively (Figures 3a and

4a). Thus, the blue component can be used as a proxy for European ancestry, whereas the red component can be used as a proxy for west-central African ancestry. Since Native

American individuals’ ancestry is almost entirely represented by green and/or yellow, the green and yellow components can be used as a proxy for Native American ancestry.

ADMIXTURE analyses showed that the Xaltocan residents with all four grandparents from Xaltocan have an average proportion of Native American ancestry of

99.4% based on autosomal SNPs (Figure 3b) and 100% based on X-chromosome SNPs

(Figure 4b). More specifically, only two out of 38 Xaltocan individuals with all four grandparents from Xaltocan exhibit evidence of European admixture in their autosomal chromosomes. However, the Xaltocan residents with at least one grandparent not from

Xaltocan have average proportions of Native American ancestry of about 82.8% based on autosomal SNPs (Figure 3c) and 89.3% based on X-chromosome SNPs (Figure 4c). More specifically, out of nine Xaltocan residents with at least one grandparent not from

Xaltocan, two exhibited evidence of both European and west-central African ancestry and six showed evidence of European ancestry on their autosomal chromosomes. X-

141 chromosome analysis indicated that out of four females, one exhibited both European and west-central African ancestry, one exhibited European ancestry, and one exhibited some

African ancestry on their X-chromosomes.

Overall, this study is consistent with historical evidence suggesting little admixture between native residents and newcomers from Europe and Africa during the colonial period in Xaltocan (Montúfar, 1897). Namely, ADMIXTURE analyses showed no evidence of European or west-central African ancestry in the vast majority of

Xaltocan residents with all four grandparents from Xaltocan. Because two individuals with all four grandparents from Xaltocan showed evidence of some European ancestry on their autosomal chromosomes, this study cannot entirely rule out the possibility that some admixture took place during the colonial period in Xaltocan. Namely, this study cannot determine whether European ancestry in those two individuals is due to admixture during the colonial period or perhaps the early post-colonial period. However, even if admixture in those individuals did date to colonial times, the extent of admixture in Xaltocan during that time period was clearly still very low.

Interestingly, this study also found that, on average, levels of European and west- central African ancestry in residents with at least one grandparent not from Xaltocan were considerably higher than in residents with all four grandparents from Xaltocan.

Even though it is possible that European or west-central African ancestry in some residents with at least one grandparent not from Xaltocan was due to admixture in

Xaltocan during the colonial period or perhaps during the early post-colonial period, this

142 possibility is unlikely given the patterns observed in the residents with all four grandparents from Xaltocan. Instead, this study primarily suggests that European and west-central African ancestry in the residents with at least one grandparent not from

Xaltocan was introduced into this town through immigration over the last two generations. Further research analyzing and modeling local ancestry across the chromosomes of all of the Xaltocan individuals showing evidence of admixture will clarify the time and provenance of European and west-central African ancestry.

143

Table 3.1. Sample sizes for all of the populations used in this study.

Population Reference n Female Male Total Americas Xaltocan This study Four grandparents from Xaltocan 16 22 38 At least one grandparent not from Xaltocan 4 5 9 Mixe Reich et al. (2012) 6 4 10 Cabecar Reich et al. (2012) 3 3 6 Piapoco Cann et al. (2002)1 4 0 4 Surui Cann et al. (2002)1 5 7 12 Apalai Skoglund et al. (2015) 2 2 4 Arara Reich et al. (2012) 1 3 4 Urubu Kaapor Skoglund et al. (2015) 2 1 3 Xavante Skoglund et al. (2015) 9 2 11 Karitiana Cann et al. (2002)1 8 8 16

Europe Spain Spanish Lazaridis et al. (2014) 27 26 53 Spanish North Lazaridis et al. (2014) 2 3 5 Basque Cann et al. (2002) 11 18 29 Italy Italian North Cann et al. (2002)1 6 14 20 Italian South Lazaridis et al. (2014) 0 1 1 Sardinian Cann et al. (2002)1 12 15 27 Sicilian Lazaridis et al. (2014) 0 11 11

Africa Northern Africa Algerian Lazaridis et al. (2014) 5 2 7 Egyptian Lazaridis et al. (2014) 0 17 17 Tunisian Lazaridis et al. (2014) 5 3 8 West-central Africa Biaka Cann et al. (2002) 0 20 20 Mandenka Cann et al. (2002) 4 13 17 Yoruba Cann et al. (2002) 34 36 70

Total sample size 166 236 402 1 References where the samples were first published. Human Origins 1 Array data obtained by Reich et al. (n.d.).

144

Table 3.2. Number of SNPs present in autosomal and X-chromosome datasets at different steps of data curation.

Step Number of SNPs Autosomal X-chromosome

Initial (before data curation) 620,731 4,331 After removing SNPs with low success rate (<90%) 610,899 4,264 After minimizing linkage disequilibrium 101,463 825

145

Figure 3.1. PCA plots based on autosomal SNPs.

146

Figure 3.2. PCA plots based on X-chromosome SNPs.

147

Figure 3.3. Values of K and cross-validation error for ADMIXTURE analyses based on (a) autosomal SNPs and (b) X-chromosome SNPs.

148

Figure 3.4. Admixture proportions based on autosomal SNPs for (a) all Native American, European, and African individuals included in this study, (b) Xaltocan individuals with all four grandparents from Xaltocan, and (c) Xaltocan individuals with at least one grandparent not from Xaltocan.

149

Figure 3.5. Admixture proportions based on X-chromosome SNPs for (a) all Native American, European, and African females included in this study, (b) Xaltocan females with all four grandparents from Xaltocan, and (c) Xaltocan females with at least one grandparent not from Xaltocan.

150

Conclusion

As anthropologists, we are sometimes interested in elucidating how individuals are or have been spatially and temporally distributed relative to genetic relatedness. For instance, to what extent is the spatial location of an individual at a given point in time similar to the spatial location of a closely related individual at either the same time or at a previous point in time? In addition, we are interested in clarifying why these spatial and temporal distributions of individuals relative to their genetic relatedness exist. For instance, what factors cause the spatial location of an individual at a given point in time to be very different from the spatial location of another closely related individual, living either at the same time or at a previous point in time?

In this dissertation, I have attempted to shed light on these two questions. To determine how some individuals are or have been spatially and temporally distributed relative to genetic relatedness, I used archaeological evidence, radiocarbon dates, and fieldwork observations for 99 individuals who at some point lived in Xaltocan, a town in central Mexico, and I analyzed DNA markers in these individuals. To clarify why the spatial and temporal distributions of these individuals exist, relative to their genetic relatedness, I considered hypotheses based on both historical documents and archaeological finds. However, none of these lines of evidence is without limitations. In this conclusion, I summarize what this dissertation has and has not been able to answer, and discuss some issues that should be considered as one thinks about and tries to answer those two overarching questions. 151

Chapter 1

To elucidate how 42 ancient individuals who died between 900 and 1521 were spatially and temporally distributed relative to genetic relatedness, I used archaeological evidence, radiocarbon dates, and analyses of a variety of DNA markers in these individuals, as described in Chapter 1. Out the 42 ancient individuals, 22 died between

900 and 1395 (“900-1395 individuals”) and were recovered from interior and periphery houses. In all of these 22 individuals, I could validate mtDNA haplogroups and 372 base- pair mtDNA haplotypes. Based on differences in burial practices, house structure, and pottery styles found during archaeological work, De Lucia and Overholtzer (2014) hypothesized that none of the 900-1395 individuals from interior houses were closely related to any of the 900-1395 individuals from periphery houses. I found that no 900-

1395 individual from an interior house shared their mtDNA haplotype with any 900-1395 individual from a periphery house, which is consistent with De Lucia and Overholtzer’s

(2014) hypothesis.

Out of the 42 ancient individuals, 16 others died between 1395 and 1521 (“1395-

1521 individuals”) and were recovered from Structure 122 and Structure 124. In all of these 16 individuals, I could validate mtDNA haplogroups and 372 base-pair mtDNA haplotypes. Colonial documents state that the residents of Xaltocan fled after the Tepanec conquest in 1395 and this town remained uninhabited until 1435. However, radiocarbon analyses indicate that some burials date to between 1395 and 1435. In addition, archaeological evidence indicates that houses were built and burials were interred during

152 this period of time in the same locations as before 1395. This cultural continuity might indicate that at least some of the 900-1395 individuals were closely related genetically to some 1395-1521 individuals. However, I found that no 900-1395 individual shared the same mtDNA haplotype as any 1395-1521 individual, a finding which does not provide support for that scenario.

Out of the 16 1395-1521 individuals, 8 individuals from Structure 122 died between 1395 and 1435 (“1395-1435 individuals”) and 3 individuals from Structure 122 died between 1435 and 1521 (“1435-1521 individuals”). In these 1395-1435 and 1435-

1521 individuals, I could validate the alleles present at some or all of 15 autosomal STR loci. When an individual exhibited a Y-chromosome, I could also validate the Y-DNA haplogroup and some or all of the alleles present at 23 STR loci in the non-recombining region of the Y-chromosome. Historical documents state that in 1435, an Aztec governor repopulated Xaltocan with people who were not related to the earlier residents. However, archaeological evidence indicates that, after 1435, houses were built and burials were interred in the same locations as those used prior to 1435 and even 1395. I found that no

1395-1435 individual shared a mtDNA haplotype with any 1435-1521 individual, and I failed to find any evidence for first-degree kinship relationships (e.g., parent-offspring or full siblings) between any 1395-1435 individual and any 1435-1521 individual. In addition, I found that the three 1395-1435 males did not share a Y-DNA haplotype with the one 1435-1521 male. Overall, I failed to find close genetic relationships between any

153

1395-1435 individual and any 1435-1521 individual, which is consistent with the scenario suggested in historical documents.

Chapter 2

To further elucidate patterns of genetic relatedness among individuals who either are living or have lived in Xaltocan, Chapter 2 included data from 47 individuals who donated a saliva sample while I was doing fieldwork in Xaltocan in July 2013 (extant individuals). For all of these individuals, I could validate mtDNA haplogroups and 613 base-pair mtDNA haplotypes. When a Y-chromosome was present, I could also validate the Y-DNA haplogroup. I then integrated these genetic data with archaeological evidence, radiocarbon dates, and historical records to evaluate how colonial and post- colonial history has shaped genetic relatedness among the contemporary and ancient residents of Xaltocan. Archaeological evidence from Xaltocan indicates that some individuals adopted Spanish material culture during the colonial period. However, some historical documents suggest that individuals of Spanish ancestry rarely visited Xaltocan.

For instance, in a 1569 letter, a Spanish priest who was assigned to Xaltocan wrote that there were about 9,000 native people but only two Spaniards in his parish. I found that all of the extant individuals belong to one of the following haplogroups: A2, B2, or C1.

Since genetic research suggests that these haplogroups were present in the Americas, but absent or rare in Europe and Asia, when European contact took place in 1492, this result indicates that the mitochondrial ancestors of these 47 individuals lived in the Americas

154 before European contact. I also found that 24 of the 26 individuals with a Y-chromosome belong to the Y-DNA haplogroup Q-L54. Since genetic research suggests that this haplogroup was present in the Americas, but absent or rare in Europe and Africa, in 1492, this result indicates that the Y-chromosome ancestors of these 24 males lived in the

Americas before European contact. The two other males belong to a Y-DNA haplogroup other than haplogroups Q-M242 or C-M207. Since genetic research indicates that the vast majority of Y-chromosome haplotypes present in the Americas in 1492 belonged to either haplogroup Q-M242 or haplogroup C-M207, this result indicates that those two individuals had Y-chromosome ancestors who did not live in the Americas then. These findings are consistent with historical records indicating little contact between the residents of Xaltocan and Spaniards during colonial times. I also found that 46 of the 47 extant residents of Xaltocan did not share their 372 base-pair mtDNA haplotypes with any of the 900-1521 individuals. Thus, these mitochondrial data provide no evidence of a direct matrilineal connection between the ancient and contemporary residents of

Xaltocan, despite their spatial proximity.

Chapter 3

To elucidate how the 47 contemporary residents of Xaltocan are related to individuals living in other locations in the Americas, Spain, Italy, northern Africa, and west-central Africa, Chapter 3 presented analyses of genome-wide SNPs in all of these extant individuals. I found that, across their entire genome, 37 of the contemporary

155

Xaltocan residents share more recent common ancestry with individuals identified as

Native Americans in my dataset than any other individuals. A few of the contemporary

Xaltocan residents do share some recent ancestry with individuals living elsewhere in the world. Altogether, though, the genome-wide data indicate that contemporary residents of

Xaltocan have little genetic ancestry from non-Native American populations, consistent with other lines of evidence suggesting that the inhabitants of Xaltocan may have had comparatively little contact with peoples who moved to Mesoamerica after European contact.

Reflections and Considerations for Future Research

This dissertation has shed light on the spatial and temporal genetic patterns at

Xaltocan, as well as on why these spatial and temporal distributions of the sampled individuals exist, relative to their genetic relatedness. In addition, this dissertation illustrates some key limitations of the approach taken here, and it helps draw attention to some considerations for future research.

Limitations of mtDNA Haplogroup and Haplotype Data. Because mtDNA is more abundant than nuclear DNA in each cell, it is often easier to analyze mtDNA in DNA samples from ancient human remains. This dissertation therefore used mtDNA haplogroup and haplotype data to help characterize the genetic relatedness between individuals. However, these data have important limitations. First, mtDNA may provide very skewed information about genetic relatedness because mtDNA represents a very

156 small percentage of the human genome and is uniparentally inherited. Specifically, mtDNA is exclusively maternally inherited, so one’s mtDNA is derived from an increasingly small proportion of one’s ancestors in each generation as one moves back further through the generations. For this reason, a lack of close mitochondrial relatedness between two individuals does not necessarily imply a lack of overall genetic relatedness.

Second, mtDNA haplogroup or haplotype data may provide only limited information about mitochondrial relatedness. In particular, mtDNA haplogroups are broad indicators of common mitochondrial ancestry1. While mtDNA haplogroups can show that two individuals are very distantly related mitochondrially2, these data alone do not provide evidence for close mitochondrial relationships. mtDNA haplotypes can be more specific indicators of common mitochondrial ancestry because these data may contain shared mutations that originated in a relatively recent mitochondrial ancestor. However, it is important to remember that such shared mutations do not necessarily indicate very close mitochondrial relationships3.

1 For instance, let us consider two present-day individuals whose mtDNA share the mutations that define mtDNA haplogroup A2 due to common ancestry (i.e., homoplasy is disregarded). If one estimates that these mutation occurred around 17,000 years ago (Kumar et al., 2011), all one could say based on this information is that these two individuals’ most recent mitochondrial ancestor lived at some point over the last 17,000 years. 2 For instance, consider two present-day individuals where one belongs to mtDNA haplogroup A2, but the other one does not. If estimates of the mitochondrial mutation rate indicates that these two mtDNA haplogroups diverged more than 17,000 years ago (Kumar et al., 2011), then one can say based on this information that these two individuals’ last shared a mitochondrial ancestor more than 17,000 years ago. 3 For instance, consider two present-day individuals whose mtDNA belong to haplogroup A2 and share a more derived mutation (relative to the most recent common ancestor of haplogroup A2) due to common ancestry (i.e., homoplasy is disregarded). If this mutation is estimated to have occurred 500 years ago, this information would tell us that the most recent mitochondrial ancestor of these two individuals lived at some point over the last 500 years, but we would not be able to pinpoint exactly how long ago the two individuals shared a direct maternal ancestor without additional information. 157

Limitations of Autosomal STR Data. To help evaluate whether the patterns of genetic relatedness observed in the mtDNA data are representative of those seen elsewhere in the genome, I also analyzed 15 autosomal STR loci. However, this portion of my dataset also has several limitations. First, even thought most 1395-1435 and 1435-

1521 individuals showed remarkable levels of aDNA preservation4, I often could not determine whether two individuals were more likely to be first-degree relatives (e.g., parent-offspring or full siblings) or unrelated with statistical significance. For instance, out of the 24 pairwise analyses investigating kinship relationships between a 1395-1435 individual and a 1435-1521 individual, 22 analyses (~91.7%) yielded a non-significant result. Blouin (2003) suggests that 15-20 unlinked autosomal STR loci often provide high power (0.9) to determine whether two individuals are more likely to be full siblings or unrelated with statistical significance, whereas 10 loci provide equally high power to assess whether they were parent and offspring or unrelated5. However, it is possible that my current dataset has limited statistical power for distinguishing first-degree kinship relationships because some autosomal STR alleles could not be genotyped, some loci exhibit low genetic variation in the population studied, and additional loci are needed

(Blouin, 2003).

4 For instance, I could genotype both alleles for all 15 autosomal STR loci in five 1395-1435 individuals and one 1435-1521 individual, and I could genotype both alleles for at least 13 autosomal STR loci in the remaining 1395-1435 individuals and another 1435-1521 individual. 5 Blouin (2003) also suggests that a given number of autosomal STR loci generally provide as much statistical power for assessing kinship as three to four times that number of autosomal SNP loci. For instance, 10 autosomal STR loci would provide as much statistical power as 30-40 autosomal SNP loci. 158

To counteract these limitations, future research could further attempt to genotype the autosomal STR loci that I could not genotype in some individuals for this dissertation.

Moreover, future research will benefit from genotyping additional polymorphic6 diploid loci. Additional loci could consist of any loci that are independent (i.e., located far enough apart on a chromosome that they are inherited separately due to recombination) of the 15 autosomal STR loci that I studied. Ongoing analyses are targeting more than a million genome-wide SNPs in some 900-1521 individuals, and these data will increase statistical power to assess kinship relationships between individuals. .

Considerations When Formulating Hypotheses.When formulating hypotheses and establishing one’s expectations for what data will be consistent with a study’s hypotheses, it is important to enunciate hypotheses that are as specific as possible. For example, because historical documents state that an Aztec governor repopulated Xaltocan in 1435 with people who were not descendents of the earlier residents, I evaluated the hypothesis that none of the 1395-1435 individuals was closely related to any of the 1435-

1521 individuals. However, this account and its corresponding hypothesis are somewhat vague. Namely, what is considered related versus unrelated was not specified. For instance, should we consider that grandparents and grandchildren, half siblings, and/or cousins are closely related or not? This decision will always be arbitrary, but this arbitrary threshold should be specified. In this project, I tested for first-degree kinship relationships using the autosomal STR loci, but I also used statistical tests that are not

6 Polymorphism is necessary because monomorphic loci provide identical likelihood values for any hypothesized kinship relationship. As a result, monomorphic loci do not help assess whether a hypothesized kinship relationship is more or less likely than any other (Goodnight and Queller, 1999). 159 directly tied to specific kin relationships (such as exact tests of population differentiation).

It is important to note that the type of dataset needed in any study depends on which specific kinship relationships are going to be treated as close relatedness, and therefore tested. More nuclear loci are required to detect both second degree relationships

(grandparent-grandchildren, half siblings, and avuncular) and third degree relationships

(great grandparent-great grandoffspring, first cousins), and even more are needed for more distant relationships.

In this project, I could validate a very limited number of nuclear STR alleles in most of the 900-1395 individuals. Out of these 22 individuals, I could validate Y- chromosome DNA haplogroups in all of the six individuals in which I detected a Y- chromosome. However, I could not validate any Y-chromosome STRs in five of these individuals. I could also validate only a very limited number of autosomal STR genotypes in the pre-1395 samples. Thus, in some cases, I could only determine maternal relationships through mtDNA comparisons. Ongoing analyses of genome-wide SNPs will further clarify the degree of relatedness among these individuals.

Finally, I want to note that genetic relatedness between two individuals is a very complex concept. Genetic relatedness can be defined as a measure of the fraction of identical-by- descent alleles that are shared between two individuals (Blouin, 2003). This measure is relatively straightforward when two individuals are parent and offspring. Because an individual inherits nearly 50% of her autosomal DNA from her mother and nearly and

160

50% from her father, parent-offspring relatedness equals 0.5. However, genetic relatedness becomes less straightforward for more distant kinship relationships because independent assortment and recombination tend to skew the relative genetic contribution of each ancestor. For instance, individuals inherit on average 25% of their autosomal

DNA from each of their grandparents. However, it is more likely that an individual actually inherits slightly more or less than 25% from each grandparent. This deviation around the average percentage may therefore increase for each further generation of ancestors. In addition, the average contribution per ancestor for each generation of ancestors decreases exponentially. The relative difference between contributions from each ancestor therefore increases as one moves backwards through the generations. Thus, one should remember that genetic relatedness itself can be complicated. Altogether, attention to these considerations when undertaking future research will improve projects.

161

References

CHAPTER 1 AGI Justicia Justicia 123/2. 1536. Gil González de Benavides y el Governador y

principales de Xaltocan, con los del pueblo de Tlatelolco, sobre ciertas estancias,

Vol. Ramo justicia, Leg. 123. Seville, Spain: Archivo General de Indias.

Alva Cortés Ixtlilxóchitl F de. 1975. Obras históricas, tomo 1, translated by O’Gorman E.

Mexico City, Mexico: Universidad Autónoma de México.

Alva Cortés Ixtlilxóchitl F de. 1977. Obras históricas, tomo 2, translated by O’Gorman E.

Mexico City, Mexico: Universidad Autónoma de México.Berdan FF, Smith ME.

2003a. The Aztec empire. In: Smith ME, Berdan FF, editors. The Postclassic

Mesoamerican world. Salt Lake City, UT: The University of Utah Press. p 67-72.

Annals of Cuauhtitlan. 1992. Annals of Cuauhtitlan. In: Bierhorst J, translator. History

and mythology of the Aztecs: The Codex Chimalpopoca. Tucson, AZ: The

University of Arizona Press. p 17-138.

Berdan FF, Smith ME. 2003b. The evolution of a core zone: the Basin of Mexico. In:

Smith ME, Berdan FF, editors. The Postclassic Mesoamerican world. Salt Lake

City, UT: The University of Utah Press. p 238-248.

Brumfiel EM. 1983. Aztec state making: ecology, structure, and the origin of the state.

American Anthropologist 85:261-284.

Brumfiel EM. 2005a. Introduction: production and power at Postclassic Xaltocan. In:

Brumfiel EM, editor. Production and power at Postclassic Xaltocan. Mexico City,

Mexico: Instituto Nacional de Antropología e Historia. p 27-41. 162

Brumfiel EM. 2005b. Conclusions: production and power at Xaltocan. In: Brumfiel EM,

editor. Production and power at Postclassic Xaltocan. Mexico City, Mexico:

Instituto Nacional de Antropología e Historia. p 349-367.

Brumfiel EM. 2005c. Opting in and opting out: Tula, Cholula, and Xaltocan. In: Blanton

RE, editor. Settlement, subsistence, and social complexity: essays honoring the

legacy of Jeffrey R. Parsons. Los Angeles, CA: Cotsen Institute of Archaeology at

UCLA. p 63-88.

Brumfiel EM, Rodríguez-Alegría E. 2010. Estrategias de las élites y cambios políticos en

Xaltocan, México. Field report on file at the Consejo de Arqueología, Instituto

Nacional de Antropología e Historia, Mexico.

Carrasco-Pizana P. 1987. Los otomíes: cultura e historia prehispánica de los pueblos

mesoamericanos de habla otomiana. Toluca, Mexico: Ediciones del Gobierno del

Estado de México.

Chimonas S. 2005. Occupational history of Prehispanic Xaltocan. In: Brumfiel EM,

editor. Production and power at Postclassic Xaltocan. Mexico City, Mexico:

Instituto Nacional de Antropología e Historia. p 169-193.

De Lucia K, Overholtzer L. 2014. Everyday action and the rise and decline of ancient

polities: household strategy and political change in Postclassic Xaltocan, Mexico.

Ancient Mesoamerica 25:441-458.

Díaz del Castillo B. 1965. The discovery and conquest of Mexico, 1517-1521. New York,

NY: Noonday Press.

163

García-Martínez B. 2010. Los años de la conquista. In: Velásquez-García E, Nalda E,

Escalante-Gonzalbo P, García-Martínez B, Hausberger B, Mazín O, Tanck de

Estrada D, Marichal C, Ávila A, Jáuregui L, Serrano-Ortega JA, Zoraida-Vázquez

J, Lira A, Staples A, Kuntz-Ficker S, Speckman-Guerra E, Garciadiego J, Aboites

L, Loyo E, loaeza S, Rodríguez-Kuri A, González-Mello R, Márquez G, Meyer L,

editors. Nueva historia general de México. Mexico City, Mexico: El Colegio de

México. p 169-215.

Gibson C. 1964. The Aztecs under Spanish rule: a history of the Indians of the Valley of

Mexico 1519-1810. Stanford, CA: Stanford University Press.

Hagelberg E, Hofreiter M, Keyser C. 2015. Ancient DNA: the first three decades.

Philosophical Transactions of the Royal Society B 370:20130371.

Hassig R. 1994. Mexico and the Spanish conquest. New York, NY: Longman.

Hernán Cortés. 1971. Cartas de relación. Mexico DF, Mexico: Editorial Porrúa, S.A.

Hicks F. 1994. Xaltocan under Mexican domination, 1435-1520. In: Marcus J, Zeitlin JF,

editors. Caciques and their people. Ann Arbor, MI: Museum of Anthropology,

University of Michigan. p 67-85.

Hicks F. 2005. Mexico, Acolhuacan, and the rulership of Late Postclassic Xaltocan. In:

Brumfiel EM, editor. Production and power at Postclassic Xaltocan. Mexico City,

Mexico: Instituto Nacional de Antropología e Historia. p 195-206.

Li JZ, Absher DM, Tang H, Southwick AM, Casto AM, Ramachandran S, Cann HM,

Barsh GS, Feldman M, Cavalli-Sforza LL, Myers RM. 2008. Worldwide human

164

relationships inferred from genome-wide patterns of variation. Science 319:1100-

1104.

Lockhart J. 1992. The Nahuas after the conquest. Stanford, CA: Stanford University

Press.

Montúfar A. 1897. Descripción del Arzobispado de México hecha en 1570. Mexico City,

Mexico: José Joaquín Terrazas e Hijas, Impresores.

Overholtzer L. 2013. Archaeological interpretation and the rewriting of history:

deimperializing and decolonizing the past at Xaltocan, Mexico. American

Anthropologist 115:481-495.

Pakendorf B, Stoneking M. 2005. Mitochondrial DNA and human evolution. Annual

Review of Genomics and Human Genetics 6:165-183.

Rodríguez-Alegría E. 2010. Incumbents and challengers: indigenous politics and the

adoption of Spanish material culture in colonial Xaltocan, Mexico. Historical

Archaeology 44:51-71.

Rodríguez-Alegría E, Millhauser JK, Stoner WD. 2013. Trade, tribute, and neutron

activation: the colonial political economy of Xaltocan, Mexico. Journal of

Anthropological Archaeology 32:397-414.

Sanders WT, Parsons JR, Santley RS. 1979. Basin of Mexico: ecological processes in the

evolution of a civilization. New York, NY: Academic Press.

165

Smith ME, Berdan FF. 2003. Postclassic Mesoamerica. In: Smith ME, Berdan FF,

editors. The Postclassic Mesoamerican world. Salt Lake City, UT: The University

of Utah Press. p 3-13.

Underhill PA, Kivisild T. 2007. Use of Y chromosome and mitochondrial DNA population structure in tracing human migrations. Annual Review of Genetics 41:539-

564.

Supplementary Information AGI Justicia Justicia 123/2. 1536. Gil González de Benavides y el Governador y

principales de Xaltocan, con los del pueblo de Tlatelolco, sobre ciertas estancias,

Vol. Ramo justicia, Leg. 123. Seville, Spain: Archivo General de Indias.

Alva Cortés Ixtlilxóchitl F de. 1975. Obras históricas, tomo 1, translated by O’Gorman E.

Mexico City, Mexico: Universidad Autónoma de México.

Alva Cortés Ixtlilxóchitl F de. 1977. Obras históricas, tomo 2, translated by O’Gorman E.

Mexico City, Mexico: Universidad Autónoma de México.

Annals of Cuauhtitlan. 1992. Annals of Cuauhtitlan. In: Bierhorst J, translator. History

and mythology of the Aztecs: The Codex Chimalpopoca. Tucson, AZ: The

University of Arizona Press. p 17-138.

Berdan FF, Smith ME. 2003a. The Aztec empire. In: Smith ME, Berdan FF, editors. The

Postclassic Mesoamerican world. Salt Lake City, UT: The University of Utah

Press. p 67-72.

166

Berdan FF, Smith ME. 2003b. The evolution of a core zone: the Basin of Mexico. In:

Smith ME, Berdan FF, editors. The Postclassic Mesoamerican world. Salt Lake

City, UT: The University of Utah Press. p 238-248.

Bolnick DA, Smith DG. 2007. Migration and social structure among the Hopewell:

evidence from ancient DNA. American Antiquity 72:627-644.

Brumfiel EM. 1983. Aztec state making: ecology, structure, and the origin of the state.

American Anthropologist 85:261-284.

Brumfiel EM. 2005a. Introduction: production and power at Postclassic Xaltocan. In:

Brumfiel EM, editor. Production and power at Postclassic Xaltocan. Mexico City,

Mexico: Instituto Nacional de Antropología e Historia. p 27-41.

Brumfiel EM. 2005b. Conclusions: production and power at Xaltocan. In: Brumfiel EM,

editor. Production and power at Postclassic Xaltocan. Mexico City, Mexico:

Instituto Nacional de Antropología e Historia. p 349-367.

Brumfiel EM. 2005c. Opting in and opting out: Tula, Cholula, and Xaltocan. In: Blanton

RE, editor. Settlement, subsistence, and social complexity: essays honoring the

legacy of Jeffrey R. Parsons. Los Angeles, CA: Cotsen Institute of Archaeology at

UCLA. p 63-88.

Brumfiel EM. 2007. Otras investigaciones en Xaltocan, 1997-2005. In: Brumfiel EM,

editor. Estrategias de las unidades domésticas en Xaltocan postclásico, México:

informe final al Instituto Nacional de Antropología e Historia. Mexico City,

Mexico: Instituto Nacional de Antropología e Historia. p 71-109.

167

Brumfiel EM, Rodríguez-Alegría E. 2010. Estrategias de las élites y cambios políticos en

Xaltocan, México. Field report on file at the Consejo de Arqueología, Instituto

Nacional de Antropología e Historia, Mexico.

Cabana GS, Hunley K, Kaestle FA. 2008. Population continuity or replacement? A novel

computer simulation approach and its application to the Numic expansion

(Western Great Basin, USA). American Journal of Physical Anthropology

135:438-447.

Carrasco-Pizana P. 1987. Los otomíes: cultura e historia prehispánica de los pueblos

mesoamericanos de habla otomiana. Toluca, Mexico: Ediciones del Gobierno del

Estado de México.

Chimonas S. 2005. Occupational history of Prehispanic Xaltocan. In: Brumfiel EM,

editor. Production and power at Postclassic Xaltocan. Mexico City, Mexico:

Instituto Nacional de Antropología e Historia. p 169-193.

De Lucia K. 2010. A child’s house: social memory, identity, and the construction of

childhood in Early Postclassic Mexican households. American Anthropologist

112:607-624.

De Lucia K. 2014. Everyday practice and ritual space: the organization of domestic ritual

in pre-Aztec Xaltocan, Mexico. Cambridge Archaeological Journal 24:379-403.

De Lucia K, Overholtzer L. 2014. Everyday action and the rise and decline of ancient

polities: household strategy and political change in Postclassic Xaltocan, Mexico.

Ancient Mesoamerica 25:441-458.

168

Fehren-Schmitz L, Haak W, Mächtle B, Masch F, Llamas B, Cagigao ET, Sossna V,

Schittek K, Cuadrado JI, Eitel B, Reindel M. 2014. Climate change underlies

global demographic genetic, and cultural transitions in pre-Columbian southern

Peru. Proceedings of the National Academy of Sciences USA 111:9443-9448.

Frederick CD, Winsborough B, Popper VS. 2005. Geoarchaeological investigations in the

northern Basin of Mexico. In: Brumfiel EM, editor. Production and power at

Postclassic Xaltocan. Mexico City, Mexico: Instituto Nacional de Antropología e

Historia. p 71-115.

Gibson C. 1964. The Aztecs under Spanish rule: a history of the Indians of the Valley of

Mexico 1519-1810. Stanford, CA: Stanford University Press.

González-Oliver A, Márquez-Morfín L, Jiménez JC, Torre-Blanco A. 2001. Founding

Amerindian mitochondrial DNA lineages in ancient Maya from Xcaret, Quintana

Roo. American Journal of Physical Anthropology 116:230-235.

Hicks F. 1994. Xaltocan under Mexican domination, 1435-1520. In: Marcus J, Zeitlin JF,

editors. Caciques and their people. Ann Arbor, MI: Museum of Anthropology,

University of Michigan. p 67-85.

Hodge MG, Neff H. 2005. Xaltocan in the economy of the Basin of Mexico: a view from

ceramic tradewares. In: Brumfiel EM, editor. Production and power at Postclassic

Xaltocan. Mexico City, Mexico: Instituto Nacional de Antropología e Historia. p

319-347.

169

Kaestle FA, Smith DG. 2001. Ancient mitochondrial DNA evidence for prehistoric

population movement: the Numic expansion. American Journal of Physical

Anthropology 115:1-12.

Kemp BM, Reséndez A, Román-Berrelleza JA, Malhi RS, Smith DG. 2005. An analysis

of ancient Aztec mtDNA from Tlatelolco: pre-Columbian relations and the spread

of Uto-Aztecan. In: Reed DM, editor. Biomolecular archaeology: genetic

approaches to the past. Carbondale, IL: Center for Archaeological Investigations

of Southern Illinois University at Carbondale. p 22-46.

Kemp BM, Tung TA, Summar ML. 2009. Genetic continuity after the collapse of the

Wari Empire: mitochondrial DNA profiles from Wari and post-Wari populations

in the ancient Andes. American Journal of Physical Anthropology 140:80-91.

Kemp BM, González-Oliver A, Malhi RS, Monroe C, Schroeder KB, McDonough J,

Rhett G, Resendez A, Peñaloza-Espinosa RI, Buentello-Malo L, Gorodesky C,

Smith DG. 2010. Evaluating the farming/language dispersal hypothesis with

genetic variation exhibited by populations in the Southwest and Mesoamerica.

Proceedings of the National Academy of Sciences USA 107:6759-6764.

Lewis CM, Buikstra JE, Stone AC. 2007. Ancient DNA and genetic continuity in the

South Central Andes. Latin American Antiquity 18:145-160.

Malhi RS, Mortensen HM, Eshleman JA, Kemp BM, Lorenz JG, Kaestle FA, Jonhson

JR, Gorodezky C, Smith DG. 2003. Native American mtDNA prehistory in the

American Southwest. American Journal of Physical Anthropology 120:108-124.

170

Malhi RS, Breece KE, Shook BAS, Kaestle FA, Chatters JC, Hackenberger S, Smith DG.

2004. Patterns of mtDNA diversity in Northwestern North America. Human

Biology 76:33-54.

Mata-Míguez J, Overholtzer L, Rodríguez-Alegría E, Kemp BM, Bolnick DA. 2012. The

genetic impact of Aztec imperialism: ancient mitochondrial DNA evidence from

Xaltocan, Mexico. American Journal of Physical Anthropology 149:504-516.

Miller A. 2007. Water mountain: a GIS analysis of Xaltocan’s integration into the Aztec

empire. BA Dissertation, Northwestern University.

Moraga M, Santoro CM, Standen VG, Carvallo P, Rothhammer F. 2005. Microevolution

in prehistoric Andean populations: chronologic mtDNA variation in the desert

valleys of northern Chile. American Journal of Physical Anthropology 127:170-

181.

Morehart CT. 2010. The archaeology of farmscapes: production, place, and the

materiality of landscape at Xaltocan, Mexico. PhD Dissertation, Northwestern

University, Chicago, IL.

Morehart CT. 2012a. Mapping ancient chinampas landscapes in the Basin of Mexico: a

remote sensing and GIS approach. Journal of Archaeological Science 39:2541-

2551.

Morehart CT. 2012b. What if the Aztec empire never existed? The prerequisites of

empire and the politics of plausible alternative histories. American Anthropologist

114:267-281.

171

Morehart CT. 2014. The potentiality and the consequences of surplus: agricultural

production and institutional transformation in the northern Basin of Mexico.

Economic Anthropology 1:154-166.

Morehart CT, Eisenberg DTA. 2010. Prosperity, power, and change: modeling maize at

Postclassic Xaltocan, Mexico. Journal of Anthropological Archaeology 29:94-

112.

Morehart CT, Frederick CD. 2014. The chronology and collapse of pre-Aztec raised field

(chinampa) agriculture in the northern Basin of Mexico. Antiquity 88:531-548.

O’Fallon BD, Fehren-Schmitz L. 2011. Native Americans experienced a strong

population bottleneck coincident with European contact. Proceedings of the

National Academy of Sciences USA 108:20444-20448.

Overholtzer L. 2012. Empire and everyday material practices: a household archaeology

of Aztec and Spanish imperialism at Xaltocan, Mexico. PhD Dissertation,

Northwestern University, Chicago, IL.

Overholtzer L. 2013. Archaeological interpretation and the rewriting of history:

deimperializing and decolonizing the past at Xaltocan, Mexico. American

Anthropologist 115:481-495.

Peñaloza-Espinosa RI, Arenas-Aranda D, Cerda-Flores RM, Buentello-Malo L,

González-Valencia G, Torres J, Álvarez B, Mendoza I, Flores M, Sandoval L,

Loeza F, Ramos I, Muñoz L, Salamanca F. 2007. Characterization of mtDNA

haplogroups in 14 Mexican indigenous populations. Human Biology 79:313-320.

172

Pruvost M, Schwarz R, Bessa Correia V, Champlot S, Braguier S, Morel N, Fernandez-

Jalvo Y, Grange T, Geigi E-M. 2007. 2007. Freshly excavated fossil bones are

best for amplification of ancient DNA. Proceedings of the National Academy of

Sciences USA 104:739-744.

Reynolds AW, Raff JA, Bolnick DA, Cook DC, Kaestle FA. 2015. Ancient DNA from

the Schild site in Illinois: implications for the Mississippian transition in the

Lower Illinois River Valley. American Journal of Physical Anthropology

156:434-448.

Sanders WT, Parsons JR, Santley RS. 1979. Basin of Mexico: ecological processes in the

evolution of a civilization. New York, NY: Academic Press.

Sandoval K, Buentello-Mallo L, Peñaloza-Espinosa R, Avelino H, Salas A, Calafell F,

Comas D. 2009. Linguistic and maternal genetic diversity are not correlated in

Native Americans. Human Genetics 126:521-531.

Shook BA, Smith DG. 2008. Using ancient mtDNA to reconstruct the population history

of northeastern North America. American Journal of Physical Anthropology

137:14-29.

Skoglund P, Sjödin P, Skoglund T, Lascoux M, Jakobsson M. 2014. Investigating

population history using temporal genetic differentiation. Molecular Biology and

Evolution 31:2516-2527.

173

Smith ME, Berdan FF. 2003. Postclassic Mesoamerica. In: Smith ME, Berdan FF,

editors. The Postclassic Mesoamerican world. Salt Lake City, UT: The University

of Utah Press. p 3-13.

Snow MH, Durand KR, Smith DG. 2010. Ancestral Puebloan mtDNA in context of

greater southwest. Journal of Archaeological Science 37:1635-1645.

Torroni A, Chen Y-S, Semino O, Santachiara-Beneceretti SS, Scott CR, Lott MT,

Winter M, Wallace DC. 1994. mtDNA and Y-chromosome polymorphisms in four

Native American populations from southern Mexico. American Journal of Human

Genetics 54:303-318.

CHAPTER 2 Anderson S, Bankier AT, Barrell BG, Brujin MHL, Coulson AR, Drouin J, Eperon IC,

Nierlich DP, Roe BA, Sanger F, Schreir PH, Smith AJH, Staden R, Young IG.

1981. Sequence and organization of the human mitochondrial genome. Nature

290:457-465.

Anderson CNK, Ramakrishan U, Chan YL, Hadly EA. 2005. Serial SimCoal: a

population genetic model for data from multiple populations and points in time.

Bioinformatics 21:1733-1734.

Andrews RM, Kubacka I, Chinnery PF, Lightowlers RN, Turnbull DM, Howell N. 1999.

Reanalysis and revision of the Cambridge reference sequence for human

mitochondrial DNA. Nature Genetics 23:147.

174

Bandelt H-J, Forster P, Röhl A. 1999. Median-joining networks for inferring intraspecific

phylogenies. Molecular Biology and Evolution 16:37-48.

Battaglia V, Grugni V, Perego UA, Angerhofer N, Gomez-Palmieri JE, Woodward SR,

Achilli A, Myres N, Torroni A, Semino O. 2013. The first peopling of South

America: new evidence from Y-chromosome haplogroup Q. PLOS ONE DOI:

10.1371/journal.pone.0071390

Bolnick DA, Bonine H, Mata-Míguez J, Kemp B, Meradeth S, LeBlanc S. 2012. Non-

destructive sampling of human skeletal remains yields ancient nuclear and

mitochondrial DNA. American Journal of Physical Anthropology 147:293-300.

Brumfiel EM. 1991. Tribute and commerce in imperial cities: the case of Xaltocan,

Mexico. In: Claesen HJM, van de Velde P, editors. Early state economics. New

Brunswick, Canada: Transaction. p 177-198.

Brumfiel EM. 2007. Otras investigaciones en Xaltocan, 1997-2005. In: Brumfiel EM,

editor. Estrategias de las unidades domésticas en Xaltocan postclásico, México:

informe final al Instituto Nacional de Antropología e Historia. Mexico City,

Mexico: Instituto Nacional de Antropología e Historia. p 71-109.

Cabana GS, Hunley K, Kaestle FA. 2008. Population continuity or replacement? A novel

computer simulation approach and its application to the Numic expansion

(Western Great Basin, USA). American Journal of Physical Anthropology

135:438-447.

175

Chimonas S. 2005. Occupational history of Prehispanic Xaltocan. In: Brumfiel EM,

editor. Production and power at Postclassic Xaltocan. Mexico City, Mexico:

Instituto Nacional de Antropología e Historia. p 169-193.

Cooper G, Amos W, Hoffman D, Rubinsztein DC. 1996. Network analysis of human Y

microsatellite haplotypes. Human Molecular Genetics 5:1759-1766.

De Lucia K, Overholtzer L. 2014. Everyday action and the rise and decline of ancient

polities: household strategy and political change in Postclassic Xaltocan, Mexico.

Ancient Mesoamerica 25:441-458.

Excoffier L, Novembre J, Schneider S. 2000. SIMCOAL: a general coalescent program

for simulation of molecular data in interconnected populations with arbitrary

demography. Journal of Heredity 91:506-509.

Excoffier L, Lischer HEL. 2010. Arlequin suite ver 3.5: a new series of programs to

perform population genetic analyses under Linux and Windows. Molecular

Ecology Resources 10:564-567.

Fenner, JN. 2005. Cross-cultural estimation of the human generation interval for use in

genetics-based population divergence studies. American Journal of Physical

Anthropology 128:415-423.

Goodnight KF, Queller DC. 1999. Computer software for performing likelihood tests of

pedigree relationship using genetic markers. Molecular Ecology 8:1231-1234.

176

Ho SY, Endicott P. 2008. The crucial role of calibration in molecular date estimates for

the peopling of the Americas. American Journal of Human Genetics 831:142-146,

author reply 146-147.

Hummel S. 2003. Ancient DNA typing: methods, strategies, and applications. Berlin,

NY: Springer.

Jobling MA, Tyler-Smith C. 1995. Fathers and sons: the Y chromosome and human

evolution. Trends in Genetics 11:449-456.

Kaestle FA, Horsburgh KA. 2002. Ancient DNA in Anthropology: methods, applications,

and ethics. Yearbook of Physical Anthropology 45:92-130.

Kalinowski ST, Taper ML, Marshall TC. 2007. Revising how the computer program

CERVUS accommodates genotyping error increases success in paternity

assignment. Molecular Ecology 16:1099-1106.

Kemp BM, Smith DG. 2005. Use of bleach to eliminate contaminating DNA from the

surface of bones and teeth. Forensic Science International 154:53-61.

Kemp BM, Malhi RS, McDonough J, Bolnick DA, Eshleman JA, Rickards O, Martinez-

Labarga C, Johnson JR, Lorenz JG, Dixon EJ, Fifield TE, Heaton TH, Worl R,

Smith DG. 2007. Genetic analysis of early Holocene skeletal remains from

Alaska and its implications for the settlement of the Americas. American Journal

of Physical Anthropology 132:605-621.

Kemp BM, González-Oliver A, Malhi RS, Monroe C, Schroeder KB, McDonough J,

Rhett G, Resendez A, Peñaloza-Espinosa RI, Buentello-Malo L, Gorodesky C,

177

Smith DG. 2010. Evaluating the farming/language dispersal hypothesis with

genetic variation exhibited by populations in the Southwest and Mesoamerica.

Proceedings of the Natural Academy of Sciences 107:6759-6764.

Kimura M. 1980. A simple method for estimating evolutionary rates of base substitutions

through comparative studies of nucleotide sequences. Journal of Molecular

Evolution 162:111-120.

Konovalov DA, Manning C, Henshaw MT. 2004. KINGROUP: a program for pedigree

relationship reconstruction and kin group assignments using genetic markers.

Molecular Ecology Notes 4:779-782.

Malhi RS, Mortensen HM, Eshleman JA, Kemp BM, Lorenz JG, Kaestle FA, Jonhson

JR, Gorodezky C, Smith DG. 2003. Native American mtDNA prehistory in the

American Southwest. American Journal of Physical Anthropology 120:108-124.

Meyer S, Weiss G, von Haeseler A. 1999. Pattern of nucleotide substitution and rate

heterogeneity in the hypervariable regions I and II of human mtDNA. Genetics

152:1103-1110.

Orlando L, Gilbert MT, Willerslev E. 2015. Reconstructing ancient genomes and

epigenomes. Nature Reviews Genetics 16:395-408.

O’Rourke DH, Hayes MG, Carlyle SW. 2000. Ancient DNA studies in Physical

Anthropology. Annual Review of Anthropology 29:217-242.

178

Overholtzer L. 2012. Empire and everyday material practices: a household archaeology

of Aztec and Spanish imperialism at Xaltocan, Mexico. PhD Dissertation,

Northwestern University.

Overholtzer L. 2013. Archaeological interpretation and the rewriting of history:

deimperializing and decolonizing the past at Xaltocan, Mexico. American

Anthropologist 115:481-495.

Overholtzer L. 2015. Agency, practice, and chronological context: a Bayesian approach

to household chronologies. Journal of Anthropological Archaeology 37:37-47.

Pääbo S, Poinar H, Serre D, Jaenike-Després V, Hebler J, Rohland N, Kuch M, Krause J,

Vigilant L, Hofreiter M. 2004. Genetic analyses from ancient DNA. Annual

Review of Genetics 38:645-679.

Peñaloza-Espinosa RI, Arenas-Aranda D, Cerda-Flores RM, Buentello-Malo L,

González-Valencia G, Torres J, Álvarez B, Mendoza I, Flores M, Sandoval L,

Loeza F, Ramos I, Muñoz L, Salamanca F. 2007. Characterization of mtDNA

haplogroups in 14 Mexican indigenous populations. Human Biology 79:313-320.

Pfeiffer H, Brinkmann B, Hühne J, Rolf B, Morris AA, Steighner R, Holland MM,

Forster P. 1999. Expanding the forensic German mitochondrial DNA control

region database: genetic diversity as a function of sample size and

microgeography. International Journal of Legal Medicine 112:291-298.

Queller DC, Goodnight KF. 1989. Estimating relatedness using genetic markers.

Evolution 43:258-275.

179

Redd AJ, Agellon AB, Kearney VA, Contreras VA, Karafet T, Park H, Knijff P, Butler

JM, Hammer MF. 2002. Forensic value of 14 novel STRs on the human Y

chromosome. Forensic Science International 130:97-111.

Reynolds AW, Raff JA, Bolnick DA, Cook DC, Kaestle FA. 2015. Ancient DNA from

the Schild site in Illinois: implications for the Mississippian Transition in the

Lower Illinois River Valley. American Journal of Physical Anthropology

156:434-448.

Rice WR. 1989. Analyzing tables of statistical tests. Evolution 43:223-225.

Roland N, Hofreiter M. 2007. Ancient DNA extraction from bones and teeth. Nature

Protocols 2:1756-1762.

Sanders WT, Parsons JR, Santley RS. 1979. Basin of Mexico: ecological processes in the

evolution of a civilization. New York, NY: Academic Press.

Sandoval K, Buentello-Mallo L, Peñaloza-Espinosa R, Avelino H, Salas A, Calafell F,

Comas D. 2009. Linguistic and maternal genetic diversity are not correlated in

Native Americans. Human Genetics 126:521-531.

Schurr TG, Ballinger SW, Gan Y-Y, Hodge JA, Merriwether DA, Lawrence DN,

Knowler WC, Weiss KM, Wallace DC. 1990. Amerindian mitochondrial DNAs

have rare Asian mutations at high frequencies, suggesting they derived from four

primary maternal lineages. American Journal of Human Genetics 46:613-623.

Soares P, Ermini L, Thomson N, Mormina M, Rito T, Röhl A, Salas A, Oppenheimer S,

Macaulay V, Richards MB. 2009. Correcting for purifying selection: an improved

180

human mitochondrial molecular clock. American Journal of Human Genetics

84:740-759.

Torroni A, Schurr TG, Cabell MF, Brown MD, Neel JV, Larsen M, Smith DG, Vullo

CM, Wallace DC. 1993. Asian affinities and continental radiation of the four

founding Native American mtDNAs. American Journal of Human Genetics

53:563-590.

Torroni A, Chen Y-S, Semino O, Santachiara-Beneceretti SS, Scott CR, Lott MT, Winter

M, Wallace DC. 1994. mtDNA and Y-chromosome polymorphisms in four

Native American populations from southern Mexico. American Journal of Human

Genetics 54:303-318.

Weiss KM. 1973. Demographic models for anthropology. In: Wobst HM, editor.

Memoirs of the Society for American Archaeology, number 27. Washington, DC: Society for American Archaeology. p 1–186.

CHAPTER 3 Alexander DH, Novembre J, Lange K. 2009. Fast model-based estimation of ancestry in

unrelated individuals. Genome Research 19:1655-1664.

Altman I. 1976. A family and region in the northern fringe lands: the marqueses de

Aguayo of Nuevo León and Coahuila. In: Altman I, Lockhart J, editors. Provinces

of early Mexico: variants of Spanish American regional evolution. Los Angeles,

CA: UCLA Latin American Center Publications. p 253-272.

181

Bauer AJ. 1996. The colonial economy. In: Hoberman LS, Socolow SM, editors. The

countryside in colonial Latin America. Albuquerque, NM: University of New

Mexico Press. p 19-48.

Begines Juárez JM. 1999. Nextlalpan de Felipe Sánchez Solís, monografía municipal.

Toluca, Mexico: Instituto Mexiquense de Cultura.

Bergreen L. 2011. Columbus: the four voyages. New York, NY: Viking Penguin.

Bolnick DA, Bolnick DI, Smith DG. 2006. Asymmetric male and female genetic histories

among Native Americans from eastern North America. Molecular Biology and

Evolution 23:2161-2174.

Borah W, Cook SF. 1963. The aboriginal population of central Mexico on the eve of the

Spanish conquest. In: Rowe JH, Borah W, King JF, editors. Ibero-Americana: 45.

Berkeley, CA: University of California Press.

Boyd-Bowman P. 1963. La emigración peninsular a América: 1520 a 1539. Historia

Mexicana 13:165-192.

Boyd-Bowman P. 1973. Patterns of Spanish emigration to the New World (1493-1580).

Buffalo, NY: Council on International Studies, State University of New York at

Buffalo.

Boyd-Bowman P. 1976. Patterns of Spanish emigration to the Indies until 1600. The

Hispanic American Historical Review 56:580-604.

182

Burkhart LM, Gasco J. 2007. Mesoamerica and Spain: the conquest. In: Carmack RM,

Gasco J, Gossen GH, editors. The legacy of Mesoamerica: history and culture of a

Native American civilization. Upper Saddle River, NJ: Prentice Hall. p 151-181.

Charlton TH. 1968. Post-conquest Aztec ceramics: implications for archaeological

interpretation. The Florida Anthropologist 21:96-101.

Charlton TH, Charlton CLO, Fournier-García P. 2005. The Basin of Mexico A.D. 1450-

1620: archaeological dimensions. In: Kepecs S, Alexander RT. The Postclassic to

Spanish-era transition in Mesoamerica. Albuquerque, NM: University of New

Mexico Press. p 49-64.

Cook SF, Simpson LB. 1948. The population of central Mexico in the sixteenth century.

In: Sauer CO, Kinnaird L, Torres-Rioseco A, editors. Ibero-Americana: 31.

Berkeley, CA: University of California Press.

Cortés H. 1971. Cartas de relación. Mexico City, Mexico: Editorial Porrúa.

Curcio-Nagy LA. 2011. The kingdom of New Spain in the seventeenth century. In:

Beezley WH, editor. A companion to Mexican history and culture. Malden, MA:

Wiley-Blackwell. p 209-229.

Deeds SM. 2011. The enilghtened colony. In: Beezley WH, editor. A companion to

Mexican history and culture. Malden, MA: Wiley-Blackwell. p 230-248.

Díaz del Castillo B. 1965. The discovery and conquest of Mexico, 1517-1521. New York,

NY: Noonday Press.

183

Endfield GH. 2008. Climate and society in colonial Mexico. Malden, MA: Blackwell

Publishing.

Espejo-Ponce Hunt M. 1976. The process of the development of Yucatan, 1600-1700. In:

Altman I, Lockhart J, editors. Provinces of early Mexico: variants of Spanish

American regional evolution. Los Angeles, CA: UCLA Latin American Center

Publications. p 33-62.

Evans ST. 2008. Ancient Mexico and Central America: archaeology and culture history.

London, UK: Thames & Hudson.

García-Martínez B. 2010. Los años de la conquista. In: Velásquez-García E, Nalda E,

Escalante-Gonzalbo P, García-Martínez B, Hausberger B, Mazín O, Tanck de

Estrada D, Marichal C, Ávila A, Jáuregui L, Serrano-Ortega JA, Zoraida-Vázquez

J, Lira A, Staples A, Kuntz-Ficker S, Speckman-Guerra E, Garciadiego J, Aboites

L, Loyo E, Loaeza S, Rodríguez-Kuri A, González-Mello R, Márquez G, Meyer

L, editors. Nueva historia general de México. Mexico City, Mexico: El Colegio de

México. p 169-215.

Gibson C. 1964. The Aztecs under Spanish rule: a history of the Indians of the Valley of

Mexico 1519-1810. Stanford, CA: Stanford University Press.

González-Martín A, Gorostiza A, Regalado-Liu L, Arroyo-Peña S, Tirado S, Nuño-Arana

I, Rubi-Castellanos R, Sandoval K, Coble MD, Rangel Villalobos H. 2015.

Demographic history of indigenous populations in Mesoamerica based on

mtDNA sequence data. PLoS ONE 10(8):e0131791.

184

Gorostiza A, Acunha-Alonzo V, Regalado-Liu L, Tirado S, Granados J, Sámano D,

Rangel-Villalobos H, González-Martín A. 2012. Reconstructing the history of

Mesoamerican populations through the study of the mitochondrial DNA control

region. PLoS ONE 7(9):e44666.

Green LD, Derr JN, Knight A. 2000. mtDNA affinities of the peoples of North-Central

Mexico. American Journal of Human Genetics 66:989-998.

Haak W, Lazaridis I, Patterson N, Rohland N, Mallick S, Llamas B, Brandt G, Nordenfelt

S, Harney E, Stewardson K, Fu Q, Mittnik A, Bánffy E, Economou C, Francken

M, Friederich S, Garrido Pena R, Hallgren F, Khartanovich V, Khokhlov A,

Kunst M, Kuznetsov P, Meller H, Mochalov O, Moiseyev V, Nicklisch N, Pichler

SL, Risch R, Rojo Guerra MA, Roth C, Szécsényi-Nagy A, Wahl J, Meyer M,

Krause J, Brown D, Anthony D, Cooper A, Werner K, Reich D. 2015. Massive

migration from the steppe was a source for Indo-European languages in Europe.

Nature 522:207-211.

Hamnett BR. 2006. A concise history of Mexico. Cambridge, UK: Cambridge University

Press.

Hassig R. 1985. Trade, tribute, and transportation: the sixteenth century political

economy of the Valley of Mexico. Norman, OK: University of Oklahoma Press.

Hassig R. 1994. Mexico and the Spanish conquest. New York, NY: Longman.

Hernández-Sánchez G. 2012. Ceramics and the Spanish conquest. Boston, MA: Brill.

185

Hicks F. 2005. Mexico, Acolhuacan, and the rulership of Late Postclassic Xaltocan. In:

Brumfiel EM, editor. Production and power at Postclassic Xaltocan. Mexico City,

Mexico: Instituto Nacional de Antropología e Historia. p 195-206.

Kehdy FSG, Gouveia MH, Machado M, Wagner CSM, Horimoto AR, Horta BL, Moreira

RG, Leal TP, Scliar MO, Soares-Souza GB, Rodrigues-Soares F, Araújo GS,

Zamudio R, Sant Anna HP, Santos HC, Duarte NE, Fiaccone RL, Figueirido CA,

Silva TM, Costa GNO, Beleza S, Berg DE, Cabrera L, Debortoli G, Duarte D,

Ghirotto S, Gilman RH, Gonçalves VF, Marrero AR, Muniz YC, Weissensteiner

H, Yeager M, Rodrigues LC, Barreto ML, Lima-Costa MF, Pereira AC,

Rodrigues MR, Tarazona-Santos E, The Brazilian EPIGEN Project Consortium.

2015. Origin and dynamics of admixture in Brazilians and its effect on the pattern

of deleterious mutations. Proceedings of the Natural Academy of Sciences USA

112:8696-8701.

Kemp BM, González-Oliver A, Malhi RS, Monroe C, Schroeder KB, McDonough J,

Rhett G, Resendez A, Peñaloza-Espinosa RI, Buentello-Malo L, Gorodesky C,

Smith DG. 2010. Evaluating the farming/language dispersal hypothesis with

genetic variation exhibited by populations in the Southwest and Mesoamerica.

Proceedings of the National Academy of Sciences USA 107:6759-6764.

Landers JG. 2006. Cimarrón and citizen: African ethnicity, corporate identity, and the

evolution of free black towns in the Spanish Circum-Caribbean. In: Landers JG,

186

Robinson BM, editors. Slaves, subjects, and subversives: blacks in colonial Latin

America. Albuqurque, NM: University of New Mexico Press. p 111-145.

Lazaridis I, Patterson N, Mittnik A, Renaud G, Mallick S, Kirsanow K, Sudmant PH,

Schraiber JG, Castellano S, Lipson M, Berger B, Economou C, Bollongino R, Fu

Q, Bos KI, Nordenfelt S, Li H, de Filippo, Prüfer K, Sawyer S, Posth C, Haak W,

Hallgren F, Fornander E, Rohland N, Delsate D, Francken M, Guinet J-M, Wahl

J, Ayodo G, Babiker HA, Bailliet G, Balanovska E, Balanovsky O, Barrantes R,

Bedoya G, Ben-Ami H, Bene J, Berrada F, Bravi CM, Brisighelli F, Busby GBJ,

Cali F, Churnosov M, Cole DEC, Corach D, Damba L, van Driem G, Dryomov S,

Dugoujon J-M, Fedorova SA, Gallego Romero I, Gubina M, Hammer M, Henn

BM, Hervig T, Hodoglugil U, Jha AR, Karachanak-Yankova S, Khusainova R,

Khusnutdinova E, Kittles R, Kisivild T, Klitz W, Kucinskas V, Kushniarevich A,

Laredj L, Litvinov S, Loukidis T, Mahley RW, Melegh B, Metspalu E, Molina J,

Mountain J, Näkkäläjärvi K, Nesheva D, Nyambo T, Osipova L, Parik J, Platonov

F, Posukh O, Romano V, Rothhammer F, Rudan I, Ruizbakiev R, Sahakyan H,

Sajantila A, Salas A, Starikovskaya EB, Tarekegn A, Toncheva D, Turdikulova S,

Uktveryte I, Utevska O, Vasquez R, Villena M, Voevoda M, Winkler CA,

Yepiskoposyan L, Zalloua P, Zemunik T, Cooper A, Capelli C, Thomas MG,

Ruiz-Linares A, Tishkoff SA, Singh L, Thangaraj K, Villems R, Comas D,

Sukernik R, Metspalu M, Meyer M, Eichler EE, Burger J, Slatkin M, Pääbo S,

187

Kelso J, Reich, Krause J. 2014. Ancient human genomes suggest three ancestral

populations for present-day Europeans. Nature 513:409-413.

Lewontin RC. 1972. The apportionment of human diversity. Evolutionary Biology 6:381-

398.

Lockhart J. 1992. The Nahuas after the conquest. Stanford, CA: Stanford University

Press.

Malhi RS, Mortensen HM, Eshleman JA, Kemp BM, Lorenz JG, Kaestle FA, Jonhson

JR, Gorodezky C, Smith DG. 2003. Native American mtDNA prehistory in the

American Southwest. American Journal of Physical Anthropology 120:108-124.

Montúfar A. 1897. Descripción del Arzobispado de México hecha en 1570. Mexico City,

Mexico: José Joaquín Terrazas e Hijas, Impresores.

Moreno-Estrada A, Gravel S, Zakharia F, McCauley, Byrnes JK, Gignoux CR, Ortiz-

Tello PA, Martínez RJ, Hedges DJ, Morris RW, Eng C, Sandoval K, Acevedo-

Acevedo S, Norman PJ, Layrisse Z, Parham P, Martínez-Cruzado JC, González

Burchard E, Cuccaro ML, Martin ER, Bustamante CD. 2013. PloS Genetics

9(11):e1003925

Moreno-Estrada A, Gignoux CR, Fernández-López JC, Zakharia F, Sikora M, Contreras

AV, Acuña-Alonzo V, Sandoval K, Eng C, Romero-Hidalgo, Ortiz-Tello P,

Robles V, Kenny EE, Nuño-Arana I, Barquera-Lozano R, Macín-Pérez G,

Granados-Arriola J, Huntsman S, Galanter JM, Via M, Ford JG, Chapela R,

Rodriguez-Cintron W, Rodríguez-Santana JR, Romieu I, Sienra-Monge JJ, del

188

Rio Navarro B, London SJ, Ruiz-Linares A, Garcia-Herrera R, Estrada K,

Hidalgo-Miranda A, Jimenez-Sanchez G, Carnevale A, Soberón X, Canizales-

Quinteros S, Rangel-Villalobos H, Silva-Zolezzi I, Gonzalez Burchard E,

Bustamante CD. 2014. The genetics of Mexico recapitulates Native American

substructure and affects biomedical traits. Science 344:1280-1285.

O’Rourke DH, Raff JA. 2010. The human genetic history of the Americas: the final

frontier. Current Biology 20:R202-207.

Patterson N, Price AL, Reich D. 2006. Population structure and eigenanalysis. PLoS

Genetics 2:e190.

Peñaloza-Espinosa RI, Arenas-Aranda D, Cerda-Flores RM, Buentello-Malo L,

González-Valencia G, Torres J, Álvarez B, Mendoza I, Flores M, Sandoval L,

Loeza F, Ramos I, Muñoz L, Salamanca F. 2007. Characterization of mtDNA

haplogroups in 14 Mexican indigenous populations. Human Biology 79:313-320.

Perego UA, Angerhofer N, Pala M, Olivieri A, Lancioni H, Hooshiar Kashani B, Carossa

V, Ekins JE, Gómez-Carballa A, Huber G, Zimmermann B, Corach D, Babudri N,

Panara F, Myres NM, Parson W, Semino O, Salas A, Woodward SR, Achilli A,

Torroni A. 2010. The initial peopling of the Americas: a growing number of

founding mitochondrial genomes from Beringia. Genome Research 20:1174-

1179.

Price AL, Patterson NJ, Plenge RM, Weinblatt ME, Shadick NA, Reich D. 2006. Nature

Genetics 38:904-909.

189

Purcell S, Neale B, Todd-Brown K, Thomas L, Ferreira MAR, Bender D, Maller J, Sklar

P, de Bakker PIW, Daly MJ, Sham PC. 2007. PLINK: a toolset for whole-genome

association and population-based linkage analysis. American Journal of Human

Genetics 81:559-575.

Ramírez SE. 1986. Large landowners. In: Hoberman LS, Socolow SM, editors. Cities &

society in colonial Latin America. Albuquerque, NM: University of New Mexico

Press. p 19-45.

Reich D, Patterson N, Campbell D, Tandon A, Mazieres S, Ray N, Parra MV, Rojas W,

Duque C, Mesa N, García LF, Triana O, Blair S, Maestre A, Dib JC, Bravi CM,

Bailliet G, Corach D, Hünemeier T, Bortolini MC, Salzano FM, Petzl-Erler ML,

Acuña-Alonzo V, Aguilar-Salinas C, Canizales-Quinteros S, Tusié-Luna T, Riba

L, Rodríguez-Cruz M, Lopez-Alarcón M, Coral-Vazquez R, Canto-Cetina T,

Silva-Zolezzi I, Fernandez-Lopez JC, Contreras AV, Jimenez-Sanchez G, Gómez-

Vázquez MJ, Molina J, Carracedo A, Salas A, Gallo C, Poletti G, Witonsky DB,

Alkorta-Aranburu G, Sukernik RI, Osipova L, Fedorova SA, Vasquez R, Villena

M, Moreau C, Barrantes R, Pauls D, Excoffier L, Bedoya G, Rothhammer F,

Dugujon J-M, Larrouy G, Klitz W, Labuda D, Kidd J, Kidd K, Di Rienzo A,

Freimer NB, Price AL, Ruiz-Linares A. 2012. Reconstructing Native American

population history. Nature 488:370-374.

190

Restall M, Schwaller R. 2011. The gods return: conquest and conquest society (1502-

1610). In: Beezley WH, editor. A companion to Mexican history and culture.

Malden, MA: Wiley-Blackwell. p 195-208.

Rodríguez-Alegría E. 2005. Consumption and the varied ideologies of domination in

colonial Mexico City. In: Kepecs S, Alexander RT, editors. The Postclassic to

Spanish-Era transition in Mesoamerica. Albuquerque, NM: University of New

Mexico Press. p 35-48.

Rodríguez-Alegría E. 2008. Narratives of conquest, colonialism, and cutting-edge

technology. American Anthropologist 110:33-43.

Rodríguez-Alegría E. 2010. Incumbents and challengers: indigenous politics and the

adoption of Spanish material culture in colonial Xaltocan, Mexico. Historical

Archaeology 44:51-71.

Rodríguez-Alegría E, Millhauser JK, Stoner WD. 2013. Trade, tribute, and neutron

activation: the colonial political economy of Xaltocan, Mexico. Journal of

Anthropological Archaeology 32:397-414.

Sanders WT, Parsons JR, Santley RS. 1979. Basin of Mexico: ecological processes in the

evolution of a civilization. New York, NY: Academic Press.

Sandoval K, Buentello-Mallo L, Peñaloza-Espinosa R, Avelino H, Salas A, Calafell F,

Comas D. 2009. Linguistic and maternal genetic diversity are not correlated in

Native Americans. Human Genetics 126:521-531.

191

Sandoval K, Moreno-Estrada A, Mendizabal I, Underhill PA, Lopez-Valenzuela M,

Peñaloza-Espinosa R, Lopez-Lopez M, Buentello-Malo L, Avelino H, Calafell F,

Comas D. 2012. Y-chromosome diversity in native Mexicans reveals continental

transition of genetic structure in the Americas. American Journal of Physical

Anthropology 148:395-405.

Schwartz SB. 2000. Victors and vanquished. New York, NY: Bedford/St. Martin’s.

Simpson LB. 1982. The encomienda in New Spain: the beginning of Spanish Mexico.

Berkeley, CA: University of California Press.

Socolow SM. 1996. Introduction to the rural past. In: Hoberman LS, Socolow SM,

editors. The countryside in colonial Latin America. Albuquerque, NM: University

of New Mexico Press. p 3-18.

Super JC. 1976. The agricultural near north: Querétaro in the seventeenth century. In:

Altman I, Lockhart J, editors. Provinces of early Mexico: variants of Spanish

American regional evolution. Los Angeles, CA: UCLA Latin American Center

Publications. p 231-252.

Thomas H. 2003. Rivers of gold: the rise of the Spanish empire. London, UK:

Weidenfeld & Nicolson.

Torroni A, Chen YS, Semino O, Santachiara-Beneceretti AS, Scott CR, Lott MT, Winter

M, Wallace DC. 1994. mtDNA and Y-chromosome polymorphisms in four

Native American populations from southern Mexico. American Journal of Human

Genetics 54:303-318.

192

Witherspoon DJ, Wooding S, Rogers AR, Marchani EE, Watkins WS, Batzer MA, Jorde

LB. 2007. Genetic similarities within and between human populations. Genetics

176:351-359.

Zegura SL, Karafet TM, Zhivotovsky LA, Hammer MF. 2004. High-resolution SNPs and

microsatellite haplotypes point to a single, recent entry of Native American Y

chromosomes into the Americas. Molecular Biology and Evolution 21:164-175.

Alexander DH, Novembre J, Lange K. 2009. Fast model-based estimation of ancestry in

unrelated individuals. Genome Research 19:1655-1664.

Bryc K, Velez C, Karafet T, Moreno-Estrada A, Reynolds A, Auton A, Hammer M,

Bustamante CD, Ostrer H. 2010. Genome-wide patterns of population structure

and admixture among Hispanic/Latino populations. Proceedings of the National

Academy of Sciences USA 107:8954-8961.

Bryc K, Durand EY, Macpherson JM, Reich D, Mountain JL. 2015. The genetic ancestry

of African Americans, Latinos, and European Americans across the United States.

American Journal of Human Genetics 96:37-53.

Cann HM, de Toma C, Cazes L, Legrand M-F, Morel V, Piouffre L, Bodmer J, Bodmer

WF, Bonne-Tamir B, Cambon-Thomsen A, Chen Z, Chu J, Carcassi C, Contu L,

Du R, Excoffier L, Ferrara GB, Friedlaender JS, Groot H, Gurwitz D, Jenkins T,

Herrera RJ, Huang X, Kidd J, Kidd KK, Langaney A, Lin AA, Qasim Mehdi S,

Parham P, Piazza A, Pia Pistillo M, Qian Y, Shu Q, Xu, J, Zhu S, Weber JL,

193

Greely HT, Feldman MW, Thomas G, Dausset J, Cavalli-Sforza LL. 2002. A

human genome diversity cell line panel. Science 5566:261-262.

Lazaridis I, Patterson N, Mittnik A, Renaud G, Mallick S, Kirsanow K, Sudmant PH,

Schraiber JG, Castellano S, Lipson M, Berger B, Economou C, Bollongino R, Fu

Q, Bos KI, Nordenfelt S, Li H, de Filippo, Prüfer K, Sawyer S, Posth C, Haak W,

Hallgren F, Fornander E, Rohland N, Delsate D, Francken M, Guinet J-M, Wahl

J, Ayodo G, Babiker HA, Bailliet G, Balanovska E, Balanovsky O, Barrantes R,

Bedoya G, Ben-Ami H, Bene J, Berrada F, Bravi CM, Brisighelli F, Busby GBJ,

Cali F, Churnosov M, Cole DEC, Corach D, Damba L, van Driem G, Dryomov S,

Dugoujon J-M, Fedorova SA, Gallego Romero I, Gubina M, Hammer M, Henn

BM, Hervig T, Hodoglugil U, Jha AR, Karachanak-Yankova S, Khusainova R,

Khusnutdinova E, Kittles R, Kisivild T, Klitz W, Kucinskas V, Kushniarevich A,

Laredj L, Litvinov S, Loukidis T, Mahley RW, Melegh B, Metspalu E, Molina J,

Mountain J, Näkkäläjärvi K, Nesheva D, Nyambo T, Osipova L, Parik J, Platonov

F, Posukh O, Romano V, Rothhammer F, Rudan I, Ruizbakiev R, Sahakyan H,

Sajantila A, Salas A, Starikovskaya EB, Tarekegn A, Toncheva D, Turdikulova S,

Uktveryte I, Utevska O, Vasquez R, Villena M, Voevoda M, Winkler CA,

Yepiskoposyan L, Zalloua P, Zemunik T, Cooper A, Capelli C, Thomas MG,

Ruiz-Linares A, Tishkoff SA, Singh L, Thangaraj K, Villems R, Comas D,

Sukernik R, Metspalu M, Meyer M, Eichler EE, Burger J, Slatkin M, Pääbo S,

194

Kelso J, Reich, Krause J. 2014. Ancient human genomes suggest three ancestral

populations for present-day Europeans. Nature 513:409-413.

Mata-Míguez J, Overholtzer L, Rodríguez-Alegría E, Bolnick DA. In preparation.

Ancient and modern DNA informs on the genetics of native populations in

Mexico since the Spanish conquest: mitochondrial and Y-chromosome DNA

evidence from Xaltocan, Mexico.

Montúfar A. 1897. Descripción del Arzobispado de México hecha en 1570. Mexico City,

Mexico: José Joaquín Terrazas e Hijas, Impresores.

Moreno-Estrada A, Gignoux CR, Fernández-López JC, Zakharia F, Sikora M, Contreras

AV, Acuña-Alonzo V, Sandoval K, Eng C, Romero-Hidalgo, Ortiz-Tello P,

Robles V, Kenny EE, Nuño-Arana I, Barquera-Lozano R, Macín-Pérez G,

Granados-Arriola J, Huntsman S, Galanter JM, Via M, Ford JG, Chapela R,

Rodriguez-Cintron W, Rodríguez-Santana JR, Romieu I, Sienra-Monge JJ, del

Rio Navarro B, London SJ, Ruiz-Linares A, Garcia-Herrera R, Estrada K,

Hidalgo-Miranda A, Jimenez-Sanchez G, Carnevale A, Soberón X, Canizales-

Quinteros S, Rangel-Villalobos H, Silva-Zolezzi I, Gonzalez Burchard E,

Bustamante CD. 2014. The genetics of Mexico recapitulates Native American

substructure and affects biomedical traits. Science 344:1280-1285.

Patterson N, Price AL, Reich D. 2006. Population structure and eigenanalysis. PLoS

Genetics 2:e190.

195

Patterson N, Moorjani P, Luo Y, Mallick S, Rohland N, Zhan Y, Genschoreck T,

Webster T, Reich D. 2012. Ancient admixture in human history. Genetics

192:1065-1093.

Price AL, Patterson NJ, Plenge RM, Weinblatt ME, Shadick NA, Reich D. 2006.

Principal components analysis corrects for stratification in genome-wide

association studies. Nature Genetics 38:904-909.

Purcell S, Neale B, Todd-Brown K, Thomas L, Ferreira MAR, Bender D, Maller J, Sklar

P, de Bakker PIW, Daly MJ, Sham PC. 2007. PLINK: a toolset for whole-genome

association and population-based linkage analysis. American Journal of Human

Genetics 81:559-575.

Reich D, Patterson N, Campbell D, Tandon A, Mazieres S, Ray N, Parra MV, Rojas W,

Duque C, Mesa N, García LF, Triana O, Blair S, Maestre A, Dib JC, Bravi CM,

Bailliet G, Corach D, Hünemeier T, Bortolini MC, Salzano FM, Petzl-Erler ML,

Acuña-Alonzo V, Aguilar-Salinas C, Canizales-Quinteros S, Tusié-Luna T, Riba

L, Rodríguez-Cruz M, Lopez-Alarcón M, Coral-Vazquez R, Canto-Cetina T,

Silva-Zolezzi I, Fernandez-Lopez JC, Contreras AV, Jimenez-Sanchez G, Gómez-

Vázquez MJ, Molina J, Carracedo A, Salas A, Gallo C, Poletti G, Witonsky DB,

Alkorta-Aranburu G, Sukernik RI, Osipova L, Fedorova SA, Vasquez R, Villena

M, Moreau C, Barrantes R, Pauls D, Excoffier L, Bedoya G, Rothhammer F,

Dugujon J-M, Larrouy G, Klitz W, Labuda D, Kidd J, Kidd K, Di Rienzo A,

196

Freimer NB, Price AL, Ruiz-Linares A. 2012. Reconstructing Native American

population history. Nature 488:370-374.

Rodríguez-Alegría E. 2010. Incumbents and challengers: indigenous politics and the

adoption of Spanish material culture in colonial Xaltocan, Mexico. Historical

Archaeology 44:51-71.

Skoglund P, Mallick S, Bortollini MC, Chennagiri N, Hünemeier T, Petzl-Erler ML,

Salzano FM, Patterson N, Reich D. 2015. Genetic evidence for two founding

populations of the Americas. Nature 525:104-108.

Wall JD, Jiang R, Gignoux C, Chen GK, Eng C, Huntsman S, Marjoram P. 2011.

Molecular Biology and Evolution 28:2231-2237.

197