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Haplogroup X Confirmed in Prehistoric North America

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Haplogroup X Confirmed in Prehistoric North America AMERICAN JOURNAL OF PHYSICAL ANTHROPOLOGY 119:84–86 (2002) Brief Communication: Haplogroup X Confirmed in Prehistoric North America Ripan S. Malhi1* and David Glenn Smith2,3 1Department of Human Genetics, University of Michigan, Ann Arbor, Michigan 48109 2Department of Anthropology, University of California, Davis, California 95616 3California Regional Primate Research Center, University of California, Davis, California 95616 KEY WORDS Native American; mtDNA; mitochondrial; ancient DNA; migration ABSTRACT Haplogroup X represents approximately in Vantage, Washington. The presence of haplogroup X in 3% of all modern Native North American mitochondrial prehistoric North America, along with recent findings of lineages. Using RFLP and hypervariable segment I haplogroup X in southern Siberians, confirms the hypoth- (HVSI) sequence analyses, we identified a prehistoric in- esis that haplogroup X is a founding lineage. Am J Phys dividual radiocarbon dated to 1,340 Ϯ 40 years BP that is Anthropol 119:84–86, 2002. © 2002 Wiley-Liss, Inc. a member of haplogroup X, found near the Columbia River Using RFLP and hypervariable segment I (HVSI) (Kaestle, 2000; Merriwether and Cabana, 2000; sequence analyses, a mitochondrial DNA haplo- Smith et al., 2000b), other early Holocene skeletons group that represents approximately 3% of all mod- in North America with cranial features unlike mod- ern Native North American samples (Forster et al., ern Native Americans (e.g., from Wizard’s Beach 1996; Brown et al., 1998; Smith et al., 1999) and and Hourglass Cave) exhibit haplogroups that are whose known worldwide distribution also includes found in Native Americans but not in Europeans West Asia and Europe (Torroni et al., 1996) has been (Stone and Stoneking, 1996; Kaestle and Smith, identified. Due to the absence of haplogroup X in 2001). These lines of evidence, together with recent East Asia, presumed to be the homeland of the criticism of similarities between the Clovis and So- founders of Native America, diverse hypotheses lutrean cultures (Straus, 2000) that were cited by were proposed to explain the presence of this haplo- Stanford (1997), strongly suggest that haplogroup X group in North America. Brown et al. (1998) sug- did not reach the Americas via an ancient European gested the possibility of an ancient European migra- migration. tion to North America. Stanford (1997) suggested The wide geographic and linguistic distribution of that early Holocene Europeans used a trans-Atlan- haplogroup X in modern Native North American tic route to colonize the Americas and that Clovis groups (Forster et al., 1996; Brown et al., 1998; technology was derived from the Solutrean tradition Smith et al., 1999) and the predominance among of the Iberian Peninsula. The presence of haplo- those of a characteristic mutation in HVSI of the group X in North America might also be due to control region not found in European or Asian mem- recent European admixture, since this haplogroup is bers of haplogroup X (the G 3 A transition at np found in high frequency in the Ojibwa (Brown et al., 16,213), imply that it is a founding Native American 1998; Malhi et al., 2001), among whom nuclear lineage. In addition, the presence of three (allegedly markers exhibit evidence of extensive European ad- full-blood) Algonquian-speaking Native Americans mixture (Szathmary and Auger, 1983). (Smith et al., 2000a; Malhi et al., 2001) who ex- Most of these hypotheses have been refuted by hibit both the haplogroup X with the transition more recent research. Derenko et al. (2001) reported the presence of haplogroup X in Altaian populations Grant sponsor: NSF. from southern Siberia, where the other four Native American founding haplogroups are also present. *Correspondence to: Ripan S. Malhi, Department of Human Genet- Therefore, the Altai are the only known modern ics, 3726 Medical Science II, 1301 E. Catherine St., University of ethnic group whose membership represents all five Michigan, Ann Arbor, MI 48109-0618. E-mail: [email protected] Native American haplogroups and, assuming the New World was colonized by a single migration, Received 3 August 2001; accepted 5 February 2002. constitute a possible origin of the founders of Native DOI 10.1002/ajpa.10106 America. Despite unsuccessful attempts to extract Published online in Wiley InterScience (www.interscience.wiley. ancient mtDNA from the Kennewick specimen com). © 2002 WILEY-LISS, INC. PREHISTORIC HAPLOGROUP X 85 at np 16,213 and the rare Albumin marker, dover site (Hauswirth et al., 1994), and the Amazon Albumin*Naskapi, also only found in North Amer- Basin (Ribeiro-Dos-Santos et al., 1996) exhibit the ica, suggests that haplogroup X is not found in North characteristic HVSI control region markers found in America due to recent European admixture. individuals assigned to haplogroup X, but they could The most convincing evidence that haplogroup X not be confidently assigned that haplogroup because is not the result of Viking or even more recent Eu- they were not tested for the AccI restriction site at ropean admixture would be its presence in ancient np 14,465. We confirmed the presence of haplogroup Native Americans. Ancient samples from the Norris X in one prehistoric sample excavated at a site on Farms site (Stone and Stoneking, 1998), the Win- the Columbia River near Vantage, Washington and radiocarbon dated to 1,340 Ϯ 40 years BP. Extensive TABLE 1. precautions were taken to limit contamination of Nucleotide positions of the mutations for the prehistoric sample and lab researcher this sample and to detect any such contamination when it did occur. To eliminate surface contamina- tion, the tooth was soaked in 10% bleach for 10 min, followed by ultraviolet light (254 nm) irradiation. Extraction and amplification setups were performed 16189 16213 16223 16234 16240 16242 16278 16311 14465 AccI in a dedicated ancient DNA laboratory that is rou- CRS T G C C A C C T Ϫ tinely bleach-sterilized. Negative controls were in- Vantage C A T . T . ϩ cluded at various stages of the extraction and am- RSM. .TCG.CϪ plification setup processes. Multiple extractions were performed on the tooth to confirm haplogroup CRS is the Cambridge Reference (Anderson et al., 1981). A dot (.) denotes identity with the Cambridge Reference sequence; a plus assignment. For a complete review of the extraction sign (ϩ) denotes a site gain; minus sign (Ϫ) denotes a site loss. method and precautions used to prevent contamina- Fig. 1. Electropherogram of mutational sites for the prehistoric sample. a: Reverse complement electropherogram, displaying the Poly C region created by a T 3 C transition at np 16,189. b: Reverse complement electropherogram, displaying the G 3 A transition and C 3 T transition at np 16,213 and np 16,223, respectively. c: Electropherogram displaying the C 3 T transition at np 16,278. 86 R.S. MALHI AND D.G. SMITH tion of the sample, see Smith et al. (2000b). Extrac- Hauswirth WW, Dickel CD, Rowold DJ, Hauswirth MA. 1994. tion and most amplification setup negative controls Inter- and intrapopulation studies of ancient humans. Experi- entia 50:585–591. exhibited no evidence of contamination. When neg- Kaestle FA. 2000. Report on DNA analysis of the remains of ative controls did exhibit contamination, the data “Kennewick Man” from Columbia Park, Washington. Washing- was excluded from the analysis. Samples from the ton, DC: National Park Service. Vantage site (500–2,000 years BP) were well-pre- Kaestle FA, Smith DG. 2001. Ancient mitochondrial DNA evi- served, and a high percentage (88%) could confi- dence for prehistoric population movement: the Numic expan- sion. Am J Phys Anthropol 115:1–12. dently be assigned to a haplogroup (Malhi, 2001). Malhi RS. 2001. Investigating prehistoric population movements This sample exhibited the HVSI control region in North America with ancient and modern mtDNA. Ph.D. markers (transitions at np 16,189, 16,223, and dissertation, Department of Anthropology, University of Cali- 16,278) and the AccI restriction site at np 14,465 fornia at Davis. found in individuals assigned to haplogroup X as Malhi RS, Schultz BA, Smith DG. 2001. Distribution of mitochon- 3 drial DNA lineages among Native American tribes of north- well as the G A transition at np 16,213 that is eastern North America. Hum Biol 73:17–55. specific to the Native American subclade of haplo- Merriwether DA, Cabana GS. 2000. Kennewick Man ancient group X (Table 1; Fig. 1). To the best of our knowl- DNA analysis: final report, submitted to the Department of the edge, this is the first evidence of haplogroup X in Interior. Washington, DC: National Park Service. prehistoric America to be confirmed using both con- Ribeiro-Dos-Santos AKC, Santos SEB, Machado AL, Guapindaia V, Zago MA. 1996. Heterogeneity of mitochondrial DNA haplo- trol region markers and the diagnostic restriction types in pre-Columbian natives of the Amazon region. Am J site gain in the coding region. This verifies a prehis- Phys Anthropol 101:29–37. toric presence of haplogroup X in North America Smith DG, Malhi RS, Eshleman J, Lorenz JG, Kaestle FA. 1999. that is probably derived from a Siberian source pop- Distribution of mtDNA haplogroup X among Native North ulation. Americans. Am J Phys Anthropol 110:271–284. Smith DG, Lorenz J, Rolfs BK, Bettinger RL, Green B, Eshleman ACKNOWLEDGMENTS J, Schultz B, Malhi R. 2000a. Implications of the distribution of Albumin Naskapi and Albumin Mexico for New World prehis- We thank Steven Hackenberger and James Chat- tory. Am J Phys Anthropol 111:557–572. ters for providing information and the sample used Smith DG, Malhi R, Eshleman JA, Kaestle FA. 2000b. Report on in this study. This research was funded by an NSF DNA analysis of the remains of “Kennewick Man” from Colum- bia Park, Washington. Washington, DC: National Park Service. Dissertation Improvement Grant to R.S.M.
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