Within-Africa Ancestry Figure 1. Pan-continental group ancestry of African-descended Americans. (a) mitochondrial DNA, (b) Y-chromosome (reflecting maternal and paternal admixture, respectively), (c) Ancestry Informative Markers (AIMs, reflecting the autosomal genome), showing the relative ratio of the three world populations that significantly contribute to the admixed populations of the United States (represented here as the entire US or by our sample from Philadelphia), the Caribbean Islands, and Brazil. The diverse socio-cultural histories of South and North America are reflected in sex- specific admixture and in overall admixture levels that differ between North and South America. The autosomal genome admixture proportions (c) approximately reflect the combination of the maternal and paternal contributions (a and b). AIMs-based estimates for Senegalese and Nigerian samples (far right) were added for comparison of African ancestral vs. American admixed populations. (Note: * designates samples typed and analyzed by the authors, while # designates previously published estimates. The remaining sample sets were collected from the literature as raw sequence and/or haplotype data and analyzed by the authors.) doi:10.1371/journal.pone.0014495.g001 further divided into Bantu, Pygmy, and ‘‘other’’ populations. We used SAMOVA [43] to first identify several genetically distinct groups (Figure 2a): a) West Pygmy from Cameroon, C.A.R. and Gabon, b) Khoisan speakers from South Africa and Botswana, c) Individuals from D.R.C. that consisted mainly of East Pygmy Mbuti, and d) Moroccans (mainly Berbers) from North Africa. After excluding the outliers from the calculations (West and East Pygmy, Khoisan speakers and North Africans, outside of lighter insert in Figure 2a), the remaining countries were split by SAMOVA into 4 groups (Figure 2b): 1) West Central/South West (WC/SW) Bantu from Angola, Cameroon, Gabon and Equatorial Guinea, 2) East/Southeast (E/SE) African individuals from Kenya and Mozambique Bantu speakers, 3) Northeast/East (NE/E) African individuals from Egypt, Sudan, Eritrea, Ethiopia, and Somalia, and finally 4) West/West Central (W/WC) countries after excluding Bantu speakers and Pygmy hunter-gatherers that cluster closely when divided by countries (details in Table S5 in File S3). These relationships are parallel to the published genetic structure based on the autosomal polymorphic markers [44]. mtDNA variation in West/West-Central Africa. We were interested to investigate the W/WC African population in more depth. We have assembled a large amount of data that includes over fifty ethnic groups sampled from 9 W/WC countries, Chad (C) and Mauritania/Western Sahara (NW), yet this region seemed to be relatively homogeneous when dissecting mtDNA pool within the whole of Africa. Our goal was to define clusters within W/WC African populations composed of data from Burkina Faso, Cameroon, Chad, Guinea-Bissau, Mali, Mauritania, Niger, Nigeria, Senegal, Sierra Leone, and Western Sahara that would group genetically similar units based on the information in our database: language, geography or ethnic affiliation. We evaluated clustering using multi-dimensional scaling and AMOVA methods [43], maximizing the between-group variation (vA) and minimizing the within-group variation (vB). First, five geographically defined clusters were identified: 1) Mauritania and Western Sahara, 2) Burkina Faso, 3) Niger, Nigeria, and Cameroon, 4) Guinea-Bissau, Mali, Senegal, and Sierra Leone, and 5) Chad (where vA = 2.29%, vB = 0.45%, Figure 2b and Figure S2 in File S3). Second, three language-defined clusters were identified: 1) Mande and Atlantic North/South speakers of the Niger-Congo family, 2) Berber and Semitic speakers of the Afro- Asiatic family, and 3) heterogeneous cluster grouping speakers of Nilo-Saharan, Chadic of the Afro-Asiatic family and non-Bantu Volta-Congo of the Niger-Congo family (vA = 2.26%, vB = 0.31%, Figure S3 in File S3). Finally, we grouped W/WC Africa by ethnicity (see Figure S4, Figure S5, and Text S1 in File S3, and File S5). While we have evaluated the within-Africa data in a variety of ways, each grouping provides additional information while none proved to be superior. Therefore, we used all three clustering approaches (by geography, language, and ethnicity) in our admixture analysis but for simplicity, we refer mainly to clustering by geography in the main text. Within-Africa ancestry of admixed populations of African descent We first established which of the previously identified eight African clusters depicted in Figure 2a contributed significantly to the admixed American populations using ADMIX software. Then, we dissected these regions further to obtain high within-Africa resolution when estimating the contribution of specific African regions to the admixed populations. We tested our approach on admixed populations from archipelagos off the African coast. Admixed Populations in Africa. To confirm that we can correctly assess the African contribution to admixture in American populations, data from two geographical regions off the W/WC coast of Africa, Cabo Verde and Sa˜o Tome´ e Pr´ıncipe, were evaluated (Figure S6 and Table S6 in File S3). These two archipelagos were former Portuguese hubs of the Atlantic slave trade and historical records of contributing African populations are available [45]. Since these archipelagos have relatively small populations with well-described histories, they can serve as a kind of natural control analysis for subsequent analysis of the larger and more diverse populations of the Americas. Our admixture analysis indicated that the current population of Cabo Verde derives solely from West Africa (,100% from W/WC, not including Bantu speakers or Pygmy), namely from West Niger-Congo speakers of Guinea-Bissau, Senegal, and Sierra Leone (,90%) and Semitic/ Berber speakers of Mauritania, Mali and Western Sahara (,10%). PLoS ONE | www.plosone.org 3 January 2011 | Volume 6 | Issue 1 | e14495