Mol Biol Rep DOI 10.1007/s11033-014-3041-9
mtDNA ancestry of Rio de Janeiro population, Brazil Suellen Bernardo • Rose Hermida • Ma´rcia Deside´rio Dayse A. Silva • Elizeu F. de Carvalho
•
Received: 6 November 2012 / Accepted: 4 January 2014 Ó Springer Science+Business Media Dordrecht 2014
Abstract Polymorphism studies concerning HVI and HVII regions of mitochondrial DNA (mtDNA) have improved the understanding of the admixture genetic process related to the occupation of the continents by human population groups. We have analyzed the mtDNA lineages of 190 healthy and maternally unrelated individuals born in the metropolitan region of the Rio de Janeiro city, the capital of the State of Rio de Janeiro, southeastern Brazil. The data showing that 57.9, 25.3 and 16.8 % of the matrilineages found in Rio de Janeiro come from African, Amerindian and European population groups. They are, respectively, in close agreement with historical records which indicate that the admixture population of Brazil is the resulting of interethnic asymmetry crosses between individuals from those population groups. The high proportion of African mtDNA lineages in the population of Rio de Janeiro is in accordance with studies related to other Brazilian states. Keywords Mitochondrial DNA Haplogroups Rio de Janeiro Brazilian population
Electronic supplementary material The online version of this article (doi:10.1007/s11033-014-3041-9) contains supplementary material, which is available to authorized users. S. Bernardo R. Hermida M. Deside´rio D. A. Silva (&) E. F. de Carvalho Laborato´rio de Diagno´sticos por DNA, Instituto de Biologia, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, Brazil e-mail: [email protected] D. A. Silva E. F. de Carvalho Departamento de Ecologia, Instituto de Biologia, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, Brazil
Introduction As a direct reflection of the Latin American colonization process, Brazilian population has been recognized as one of the most heterogeneous in the world. Brazil is a very large country and its admixture population is the result of crosses among different ethnic groups such as Native Americans, Africans and Europeans that have been taking place for over five centuries, since Brazil was discovered and colonized by Portugal. When the Portuguese came to Brazil, the land was inhabited by several Native American tribes [1, 2]. During the XVI and XIX centuries in which only Portuguese citizens were admitted to the unique newly Portuguese American colony, in a first immigration wave, 500 thousand Portuguese citizens moved to Brazil [3]. In the first decade of the XIX century, when the Portuguese royal family moved to Rio de Janeiro in order to avoid a fight against Napolean, the first King was established down in the New World, in 1810. During the same period, approximately four million Africans were taken to Brazil as slaves [4]. From then on, an increasing number of individuals from several parts of the world immigrated to different regions of the country, resulting in a second wave of immigration, between 1910 and 1950. Portugal remained by far the most significant source of immigrants followed by Italy, Spain, and Germany. Additionally, Asian immigration took place mainly from Japan, as well as from Lebanon and Syria in the 20th century [5, 6]. Rio de Janeiro is a Southeastern Brazilian state. When the first Europeans arrived, the main Amerindian tribes that inhabited the region were Goytacazes, Paraiba, Puris, Tamoios and Goiania; smaller Amerindians groups were formed by crowned Coropo´s, Kayapo, Guarus, Tupimino´s and Temimino´s [7]. Rio de Janeiro was the major destination of slaves in Brazil until nearly half of the nineteenth
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century, receiving slaves predominately shipped from Central and West Africa through the Atlantic [8, 9]. Nowadays, Brazilian citizens are classified according to their self-denomination: white, black, brown, yellow and Amerindian. The last demographic census showed that the population in the Rio de Janeiro state is approximately composed of 16 million inhabitants, 50.1 % were classified as black, (which include black and brown skinned people) (IBGE Census 2010; http://www.ibge.gov.br) [10]. In the last decades, some genetic studies were undertaken trying to capture the footprints of demographic events behind the Brazilian populations. The first genetic studies were focused on a restricted type of marker, namely, based on protein information, or different DNA markers including autossomal, Y chromosome and mtDNA specific polymorphisms [5, 11–18]. Some other studies have start being published regarding a more complete set of different type of markers, giving more complete pictures of the history of Latin American populations [13, 14, 19–22]. Namely, mtDNA studies based on information from the hypervariable regions HVSI and HVS-II have helped to improve the resolution of the genealogy of the female lineages [23–27]. mtDNA sequence data from Rio de Janeiro Admixed population does not exist. For forensic purposes, a comprehensive and high-quality dataset is needed, which also reflects regional differences between populations. The maim purpose of this study was to analyze the maternal ancestry of admixture of Rio de Janeiro population by searching their mtDNA lineages. Investigation may, in a general way, unravel the formation of modern human populations.
Materials and methods DNA samples Samples were obtained from paternity casework from the DNA Diagnostic Laboratory (LDD). All donors gave their written consent for the use of the samples in an anonymous way. Furthermore, the scientific investigation proposal, together with the informed consent, had been evaluated and approved in advance by the Science Ethical Committee of the State University of Rio de Janeiro (CAAE:0067.0.228. 000-09). We randomly selected 190 unrelated individuals born in the metropolitan region of the Rio de Janeiro city (22°540 1000 S, 43°120 2800 W), the capital of the State of Rio de Janeiro, RJ), southeastern Brazil. Genomic DNA was extracted from 100 ll of peripheral blood by the Chelex [29] or by the salting-out method [30].
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Samples genotyping Mitochondrial HVI and HVII regions were amplified using primer pairs L15997-H16391 and L048-H408, respectively [31]. Sequencing was performed in both directions and, if necessary, internal primers (H16236, L16159, H285 and L172) were used for confirmation of the quality of the sequences obtained. PCR reactions were carried out in VeritiÒ Thermal Cycler (Applied Biosystems, Foster City, CA) and the amplified products were purified by Microspin S-300 HR columns (Amersham Biosciences, Co) alternatively, 5 ll of the amplified products were purified using 2 ll of the ExoSAP 2ITÒ kit (USB Corporation). Purified products (c. 10 ng) were sequenced by cycle sequencing using an ABI PRISMÒ 310/3130/3500 Genetic Analyzer and BigDyeTM Terminator kit version 3.1 (Applied Biosystems, Foster City, CA). Sequencing was performed using the same primers set of the PCR reaction and the sequenced products were cleaned by the Sephadex G-50 resin [32]. Data analysis The sequences obtained were aligned and compared to the revised Cambridge Reference Sequence (rCRS) using SeqScapeÒ Software-version 2.5 (Applied Biosystems, Foster City, CA). Mitochondrial haplotypes found in the population examined were assigned to haplogroups, taking into account the polymorphic positions of the control region (HVI and HVII), according to the ‘‘Phylotree.org—Global Human mtDNA phylogenetic tree’’ (available at http://www.phylo tree.org/) [33]. Estimates of genetic diversity, haplotype frequencies, nucleotide diversity and genetic distances were computed using the program Arlequin version 3.5 [34]. Genetic distances were visualized in two-dimensional plots using the multidimensional scaling (MDS) method included in data analysis software STATISTICA v8.0 (StatSoft, Inc. 2007; www.statsoft.com).
Results and discussion The present study used sequence analysis of the hypervariable regions HVS-I (16024-163650) and HVS-II (73340) of the mitochondrial DNA control region from 190 unrelated individuals from Rio de Janeiro state, Brazil, and found a total of 171 polymorphic sites. The distribution of polymorphic sites across both regions clearly showed their highly polymorphic status. The mtDNA lineages yielding 180 different HVI and HVII haplotypes can be perfectly allocated to the known
Mol Biol Rep Table 1 Diversity measures estimated from the analysis of the entire mtDNA HVI and HVII regions in a population sample from Rio de Janeiro (Brazil) Parameters
HV I ? II
Number of haplotypes
180
Number of observed transitions
147
Number of observed transversions Number of substitutions
30 177
Number of polymorphic sites
171
Mean number of pairwise differences
17.352604 ± 7.745836
Nucleotide diversity (average over loci)
0.101477 ± 0.050127
Sequence diversity
0.9994 ± 0.0006
Table 2 Haplotype general frequencies in 190 Rio de Janeiro samples Haplogroups
n
Frequency
Native American A
10
20.9
B
18
37.5
C
16
33.3
D
4
8.3
48
25.3
L0 L1
9 20
8.2 18.2
L2
25
22.7
L3
54
49.1
Total African
L4
2
1.8
110
57.9
H
21
65.7
I
1
3.1
J
1
3.1
K
3
9.4
N
2
6.3
T
1
3.1
U
1
3.1
W
1
3.1
M
1
3.1
32
16.8
Total European
Total
haplogroups (Supplementary material 1). Polymorphisms of the hypervariable segments were analyzed both separately and jointly showing the nucleotide positions 16129 in the HVI and 263 in the HVII with 86.08 and 97.42 % respectively, as the one the highest frequencies of substitutions. Diversity parameters for HVI and HVII regions are presented in the Table 1.
It was observed that 110 maternal lineages belong to various African haplogroups, 48 mtDNA lineages fall into the Native American, whereas 32 belong to European haplogroups (Table 2). As expected of an admixed population, high HVI and HVII diversities were observed in Rio de Janeiro (haplotype diversity: 0.9994 ± 0.0006). These results clearly indicate the absence of founder effects related to the European colonization of Rio de Janeiro state, Brazil. Still, an expressive number of maternal lineages linked to African and Native American ancestries with 57.9 and 25.3 %, respectively was observed, while only 16.8 % of the Rio de Janeiro individuals harbored European maternal inheritance. According to the historical records, it is expected that the maternal and paternal proportion of African, European and Amerindian ancestry are variable in different Brazilian regions due to their specific processes of colonization, as noted in the southern States of the country, such as Santa Catarina and Rio Grande do Sul. In Santa Catarina, the European proportion accounts for 63.7 % of maternal heritage of this population, followed by the African contribution (21.3 %) and Native American maternal ancestry (15 %) [26]. In Rio Grande do Sul, such proportions are at the order of 48, 36 and 16 % [27], for the same ethnic groups. The greatest contribution of European mitochondrial lineages in the Southern region is justified by the predominantly European colonization in this region, with a minor use of African slave labor. In different regions of the North Brazil, a prevalence of Amerindian mtDNA Haplogroups [14, 24] has been observed. The prevalence of Amerindian mitochondrial lineages in the Northern region is justified by the fact that the dense Amazon rainforest was a natural obstacle to the process of Portuguese colonization, thus, making possible a greater resistance and isolation of native American peoples in the region [15, 25, 35–37]. Our results are consistent with those found for the mixed population of states from the Southeast (Sa˜o Paulo) and northeast (Alagoas) Brazil. In Sa˜o Paulo, the African proportion account for 46.6 % of maternal heritage of this population, followed by the European contribution (27.3 %) and Native American maternal ancestry (26.1 %) and Alagoas has a distribution of African, Native American and European maternal lineage of 44, 33 and 21 %, respectively [25]. The predominance of ancestral African maternal lineages in the Southeast and Northeast of Brazil is justified by the similarity of the processes of occupation and settlement, which had a strong influence on the slave trade by more than 250 years, exactly as expected from known historical and anthropological studies of Brazilians [38].
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With respect to the paternal ancestry, by analyzing SNPs markers on the non-recombining portion of the Y-chromosome, a greater European contribution for the male component of the Rio de Janeiro population was shown. Paternal inheritance is somewhat different from the maternal, in Rio de Janeiro they were estimated to be European (88.1 %) and, to a lesser extent, sub-Saharan African (8.7 %). Native Americans, even though the first inhabitants of this region, contributed less (1.6 %) to the present gene pool [39] as a result of the economic and cultural aspects that have governed the rules for crosses between male and females from each one of the three major components admixed during the process of the country colonization [3]. Therefore, as have been established in different studies [14, 15, 23, 38], our results also showed that the level crossing among Europeans, Africans and Amerindians was extensive and that the weight of the asymmetric mating in the colonial era, predominantly European men with Africans and Indigenous women, was determinant for noonday’s genetic pool of the admixed population of Rio de Janeiro state. Among African origin samples, the most common groups L-type are characteristic of Sub Saharan Africa [40]. In our cumulative data, the most common were L3 (49.1 %), followed by L2 (22.7 %), L1 (18.2 %), L0 (8.2 %) and L4 (1.8 %). These haplogroups are characteristic of specific regions of Africa, the traditional ports of the exportation of slaves to colonial America. By using only HVI information and additional coding region polymorphisms that can increase the resolution of the mtDNA haplogroup estimations, Alves-Silva et al. [23] suggested that the majority of mtDNA lineages of ancestry in Rio de Janeiro sample would have a West-Central Africa origin. Our results are consistent with those population studies in
South America and other regions of Brazil, regarding the haplogroup distribution [23, 37, 38, 41]. The haplogroup B (37.5 %) is the most frequent haplogroup within the Native American fraction, closely followed by the haplogroup C (33.3 %), with A (20.9 %) coming next and D (8.3 %) coming last. Similar proportion of frequencies are observed in the southern and southeastern regions of Brazil [23]. Therefore, the major founder haplotypes of the Native American haplogroups A–D [42] are present in Brazil and shared with many other American populations [25, 43]. In relation to European maternal ancestry, the haplogroup H is the most frequent found (65.6 %) in the population. A strong correlation was also observed between those haplogroups found in the population of Rio de Janeiro and the Iberian Peninsula ones [23, 44, 45]. Pairwise genetic distance analysis was performed between the study sample and previously published data concerning five Brazilian populations [24–27, 38] and those representatives of continental population groups: Europe [46], Africa [47], Asia [48] and Native American (Silva DA, personal communication). In comparison with populations from other Brazilian regions, significant differences were observed when comparing Rio de Janeiro (RJ) to Santa Catarina (SC) and Rio Grande do Sul (RS). For the other Brazilian populations: Alagoas (AL), Amazonas (AM) and Sa˜o Paulo (SP), a significant genetic difference was not identified. Genetic distance among Rio de Janeiro state and other population groups is shown in Table 3. In relation to the ancestral groups that potentially contribute to the formation of the Brazilian population, the MDS plot of Fig. 1, indicates the sample from Rio de Janeiro plotted closer to the African (AF) and it has shown no significant genetic differentiation between them. This is
Table 3 FST genetic distances between the studied population (RJ) and other populations RJ
AF
TE
PT
CH
AL
AM
SC
RS
RJ
–
AF
0.03159
–
TE PT
0.14945* 0.22524*
0.26905* 0.3172*
– 0.34375
–
CH
0.14945*
0.22406*
0.14021*
0.20979*
–
AL
0.01048
0.06162*
0.10817*
0.17225*
0.10594*
–
AM
0.02378
0.10851*
0.07073*
0.25653*
0.10403*
0.02276*
–
SC
0.08377*
0.16654*
0.17058*
0.05694*
0.10514*
0.04528*
0.09485*
RS
0.06399*
0.14865*
0.124*
0.13116*
0.08633*
0.02948*
0.05193*
0.01141
–
SP
0.00773
0.06146*
0.13073*
0.14578*
0.10745*
0.00311
0.02428*
0.03183*
0.02628*
SP
– –
RJ Rio de Janeiro, AF Africa, TE Terena, PT Portugal, CH China, AL Alagoas, AM Amazonas, SC Santa Catarina, RS Rio Grande do Sul, SP Sa˜o Paulo * Significant genetic distance values after applying Bonferroni correction in FST P value
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References
Fig. 1 MDS plot of the pairwise genetic distances among six Brazilian samples [Rio de Janeiro population of present study (RJ); Alagoas (AL); Sa˜o Paulo (SP); Amazonas (AM), Santa Catarina (SC); Rio Grande do Sul (RS)] and four reference samples from Africa (Angola-AF), Europe (Portugal-PT), Asia (China-CH) and Native American (Terena-TE)
explained by the prevalence of African haplogroups in the study sample. Significant genetic differentiation was observed among Rio de Janeiro state, Native American (Terena-TE), European (Portugal-PT) and Asian (ChinaCH) populations.
Conclusions In conclusion, we have shown that 57.9 % of the mtDNA lineages from the admixed population of Rio de Janeiro state are associated to African ancestry. The results confirm that the population derived from crosses among Native Americans interethnic Africans and Europeans for over 500 years but there was an asymmetry favouring crosses among male Europeans and Africans or Amerindian women as evidenced by historical, anthropological and genetic studies already carried out with mitochondrial DNA and the Y-chromosome lineage markers. The study demonstrates, therefore, that admixed population cannot be adequately classified taking into consideration just arbitrary parameters, as skin color, because of the heterogeneity of our population. Currently, there are sociological discussions happening in the Brazilian political life concerning the introduction of public policies favoring the non-white population in a population demonstrably mixed. Moreover, not only the application of phylogenetic methods to our data, but also the comparison with closely related sequences will provide valuable information for forensic and population genetic analysis in the admixed population of Rio de Janeiro state.
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mtDNA ancestry of Rio de Janeiro population, Brazil Suellen Bernardo • Rose Hermida • Ma´rcia Deside´rio Dayse A. Silva • Elizeu F. de Carvalho
•
Received: 6 November 2012 / Accepted: 4 January 2014 Ó Springer Science+Business Media Dordrecht 2014
Abstract Polymorphism studies concerning HVI and HVII regions of mitochondrial DNA (mtDNA) have improved the understanding of the admixture genetic process related to the occupation of the continents by human population groups. We have analyzed the mtDNA lineages of 190 healthy and maternally unrelated individuals born in the metropolitan region of the Rio de Janeiro city, the capital of the State of Rio de Janeiro, southeastern Brazil. The data showing that 57.9, 25.3 and 16.8 % of the matrilineages found in Rio de Janeiro come from African, Amerindian and European population groups. They are, respectively, in close agreement with historical records which indicate that the admixture population of Brazil is the resulting of interethnic asymmetry crosses between individuals from those population groups. The high proportion of African mtDNA lineages in the population of Rio de Janeiro is in accordance with studies related to other Brazilian states. Keywords Mitochondrial DNA Haplogroups Rio de Janeiro Brazilian population
Electronic supplementary material The online version of this article (doi:10.1007/s11033-014-3041-9) contains supplementary material, which is available to authorized users. S. Bernardo R. Hermida M. Deside´rio D. A. Silva (&) E. F. de Carvalho Laborato´rio de Diagno´sticos por DNA, Instituto de Biologia, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, Brazil e-mail: [email protected] D. A. Silva E. F. de Carvalho Departamento de Ecologia, Instituto de Biologia, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, Brazil
Introduction As a direct reflection of the Latin American colonization process, Brazilian population has been recognized as one of the most heterogeneous in the world. Brazil is a very large country and its admixture population is the result of crosses among different ethnic groups such as Native Americans, Africans and Europeans that have been taking place for over five centuries, since Brazil was discovered and colonized by Portugal. When the Portuguese came to Brazil, the land was inhabited by several Native American tribes [1, 2]. During the XVI and XIX centuries in which only Portuguese citizens were admitted to the unique newly Portuguese American colony, in a first immigration wave, 500 thousand Portuguese citizens moved to Brazil [3]. In the first decade of the XIX century, when the Portuguese royal family moved to Rio de Janeiro in order to avoid a fight against Napolean, the first King was established down in the New World, in 1810. During the same period, approximately four million Africans were taken to Brazil as slaves [4]. From then on, an increasing number of individuals from several parts of the world immigrated to different regions of the country, resulting in a second wave of immigration, between 1910 and 1950. Portugal remained by far the most significant source of immigrants followed by Italy, Spain, and Germany. Additionally, Asian immigration took place mainly from Japan, as well as from Lebanon and Syria in the 20th century [5, 6]. Rio de Janeiro is a Southeastern Brazilian state. When the first Europeans arrived, the main Amerindian tribes that inhabited the region were Goytacazes, Paraiba, Puris, Tamoios and Goiania; smaller Amerindians groups were formed by crowned Coropo´s, Kayapo, Guarus, Tupimino´s and Temimino´s [7]. Rio de Janeiro was the major destination of slaves in Brazil until nearly half of the nineteenth
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century, receiving slaves predominately shipped from Central and West Africa through the Atlantic [8, 9]. Nowadays, Brazilian citizens are classified according to their self-denomination: white, black, brown, yellow and Amerindian. The last demographic census showed that the population in the Rio de Janeiro state is approximately composed of 16 million inhabitants, 50.1 % were classified as black, (which include black and brown skinned people) (IBGE Census 2010; http://www.ibge.gov.br) [10]. In the last decades, some genetic studies were undertaken trying to capture the footprints of demographic events behind the Brazilian populations. The first genetic studies were focused on a restricted type of marker, namely, based on protein information, or different DNA markers including autossomal, Y chromosome and mtDNA specific polymorphisms [5, 11–18]. Some other studies have start being published regarding a more complete set of different type of markers, giving more complete pictures of the history of Latin American populations [13, 14, 19–22]. Namely, mtDNA studies based on information from the hypervariable regions HVSI and HVS-II have helped to improve the resolution of the genealogy of the female lineages [23–27]. mtDNA sequence data from Rio de Janeiro Admixed population does not exist. For forensic purposes, a comprehensive and high-quality dataset is needed, which also reflects regional differences between populations. The maim purpose of this study was to analyze the maternal ancestry of admixture of Rio de Janeiro population by searching their mtDNA lineages. Investigation may, in a general way, unravel the formation of modern human populations.
Materials and methods DNA samples Samples were obtained from paternity casework from the DNA Diagnostic Laboratory (LDD). All donors gave their written consent for the use of the samples in an anonymous way. Furthermore, the scientific investigation proposal, together with the informed consent, had been evaluated and approved in advance by the Science Ethical Committee of the State University of Rio de Janeiro (CAAE:0067.0.228. 000-09). We randomly selected 190 unrelated individuals born in the metropolitan region of the Rio de Janeiro city (22°540 1000 S, 43°120 2800 W), the capital of the State of Rio de Janeiro, RJ), southeastern Brazil. Genomic DNA was extracted from 100 ll of peripheral blood by the Chelex [29] or by the salting-out method [30].
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Samples genotyping Mitochondrial HVI and HVII regions were amplified using primer pairs L15997-H16391 and L048-H408, respectively [31]. Sequencing was performed in both directions and, if necessary, internal primers (H16236, L16159, H285 and L172) were used for confirmation of the quality of the sequences obtained. PCR reactions were carried out in VeritiÒ Thermal Cycler (Applied Biosystems, Foster City, CA) and the amplified products were purified by Microspin S-300 HR columns (Amersham Biosciences, Co) alternatively, 5 ll of the amplified products were purified using 2 ll of the ExoSAP 2ITÒ kit (USB Corporation). Purified products (c. 10 ng) were sequenced by cycle sequencing using an ABI PRISMÒ 310/3130/3500 Genetic Analyzer and BigDyeTM Terminator kit version 3.1 (Applied Biosystems, Foster City, CA). Sequencing was performed using the same primers set of the PCR reaction and the sequenced products were cleaned by the Sephadex G-50 resin [32]. Data analysis The sequences obtained were aligned and compared to the revised Cambridge Reference Sequence (rCRS) using SeqScapeÒ Software-version 2.5 (Applied Biosystems, Foster City, CA). Mitochondrial haplotypes found in the population examined were assigned to haplogroups, taking into account the polymorphic positions of the control region (HVI and HVII), according to the ‘‘Phylotree.org—Global Human mtDNA phylogenetic tree’’ (available at http://www.phylo tree.org/) [33]. Estimates of genetic diversity, haplotype frequencies, nucleotide diversity and genetic distances were computed using the program Arlequin version 3.5 [34]. Genetic distances were visualized in two-dimensional plots using the multidimensional scaling (MDS) method included in data analysis software STATISTICA v8.0 (StatSoft, Inc. 2007; www.statsoft.com).
Results and discussion The present study used sequence analysis of the hypervariable regions HVS-I (16024-163650) and HVS-II (73340) of the mitochondrial DNA control region from 190 unrelated individuals from Rio de Janeiro state, Brazil, and found a total of 171 polymorphic sites. The distribution of polymorphic sites across both regions clearly showed their highly polymorphic status. The mtDNA lineages yielding 180 different HVI and HVII haplotypes can be perfectly allocated to the known
Mol Biol Rep Table 1 Diversity measures estimated from the analysis of the entire mtDNA HVI and HVII regions in a population sample from Rio de Janeiro (Brazil) Parameters
HV I ? II
Number of haplotypes
180
Number of observed transitions
147
Number of observed transversions Number of substitutions
30 177
Number of polymorphic sites
171
Mean number of pairwise differences
17.352604 ± 7.745836
Nucleotide diversity (average over loci)
0.101477 ± 0.050127
Sequence diversity
0.9994 ± 0.0006
Table 2 Haplotype general frequencies in 190 Rio de Janeiro samples Haplogroups
n
Frequency
Native American A
10
20.9
B
18
37.5
C
16
33.3
D
4
8.3
48
25.3
L0 L1
9 20
8.2 18.2
L2
25
22.7
L3
54
49.1
Total African
L4
2
1.8
110
57.9
H
21
65.7
I
1
3.1
J
1
3.1
K
3
9.4
N
2
6.3
T
1
3.1
U
1
3.1
W
1
3.1
M
1
3.1
32
16.8
Total European
Total
haplogroups (Supplementary material 1). Polymorphisms of the hypervariable segments were analyzed both separately and jointly showing the nucleotide positions 16129 in the HVI and 263 in the HVII with 86.08 and 97.42 % respectively, as the one the highest frequencies of substitutions. Diversity parameters for HVI and HVII regions are presented in the Table 1.
It was observed that 110 maternal lineages belong to various African haplogroups, 48 mtDNA lineages fall into the Native American, whereas 32 belong to European haplogroups (Table 2). As expected of an admixed population, high HVI and HVII diversities were observed in Rio de Janeiro (haplotype diversity: 0.9994 ± 0.0006). These results clearly indicate the absence of founder effects related to the European colonization of Rio de Janeiro state, Brazil. Still, an expressive number of maternal lineages linked to African and Native American ancestries with 57.9 and 25.3 %, respectively was observed, while only 16.8 % of the Rio de Janeiro individuals harbored European maternal inheritance. According to the historical records, it is expected that the maternal and paternal proportion of African, European and Amerindian ancestry are variable in different Brazilian regions due to their specific processes of colonization, as noted in the southern States of the country, such as Santa Catarina and Rio Grande do Sul. In Santa Catarina, the European proportion accounts for 63.7 % of maternal heritage of this population, followed by the African contribution (21.3 %) and Native American maternal ancestry (15 %) [26]. In Rio Grande do Sul, such proportions are at the order of 48, 36 and 16 % [27], for the same ethnic groups. The greatest contribution of European mitochondrial lineages in the Southern region is justified by the predominantly European colonization in this region, with a minor use of African slave labor. In different regions of the North Brazil, a prevalence of Amerindian mtDNA Haplogroups [14, 24] has been observed. The prevalence of Amerindian mitochondrial lineages in the Northern region is justified by the fact that the dense Amazon rainforest was a natural obstacle to the process of Portuguese colonization, thus, making possible a greater resistance and isolation of native American peoples in the region [15, 25, 35–37]. Our results are consistent with those found for the mixed population of states from the Southeast (Sa˜o Paulo) and northeast (Alagoas) Brazil. In Sa˜o Paulo, the African proportion account for 46.6 % of maternal heritage of this population, followed by the European contribution (27.3 %) and Native American maternal ancestry (26.1 %) and Alagoas has a distribution of African, Native American and European maternal lineage of 44, 33 and 21 %, respectively [25]. The predominance of ancestral African maternal lineages in the Southeast and Northeast of Brazil is justified by the similarity of the processes of occupation and settlement, which had a strong influence on the slave trade by more than 250 years, exactly as expected from known historical and anthropological studies of Brazilians [38].
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With respect to the paternal ancestry, by analyzing SNPs markers on the non-recombining portion of the Y-chromosome, a greater European contribution for the male component of the Rio de Janeiro population was shown. Paternal inheritance is somewhat different from the maternal, in Rio de Janeiro they were estimated to be European (88.1 %) and, to a lesser extent, sub-Saharan African (8.7 %). Native Americans, even though the first inhabitants of this region, contributed less (1.6 %) to the present gene pool [39] as a result of the economic and cultural aspects that have governed the rules for crosses between male and females from each one of the three major components admixed during the process of the country colonization [3]. Therefore, as have been established in different studies [14, 15, 23, 38], our results also showed that the level crossing among Europeans, Africans and Amerindians was extensive and that the weight of the asymmetric mating in the colonial era, predominantly European men with Africans and Indigenous women, was determinant for noonday’s genetic pool of the admixed population of Rio de Janeiro state. Among African origin samples, the most common groups L-type are characteristic of Sub Saharan Africa [40]. In our cumulative data, the most common were L3 (49.1 %), followed by L2 (22.7 %), L1 (18.2 %), L0 (8.2 %) and L4 (1.8 %). These haplogroups are characteristic of specific regions of Africa, the traditional ports of the exportation of slaves to colonial America. By using only HVI information and additional coding region polymorphisms that can increase the resolution of the mtDNA haplogroup estimations, Alves-Silva et al. [23] suggested that the majority of mtDNA lineages of ancestry in Rio de Janeiro sample would have a West-Central Africa origin. Our results are consistent with those population studies in
South America and other regions of Brazil, regarding the haplogroup distribution [23, 37, 38, 41]. The haplogroup B (37.5 %) is the most frequent haplogroup within the Native American fraction, closely followed by the haplogroup C (33.3 %), with A (20.9 %) coming next and D (8.3 %) coming last. Similar proportion of frequencies are observed in the southern and southeastern regions of Brazil [23]. Therefore, the major founder haplotypes of the Native American haplogroups A–D [42] are present in Brazil and shared with many other American populations [25, 43]. In relation to European maternal ancestry, the haplogroup H is the most frequent found (65.6 %) in the population. A strong correlation was also observed between those haplogroups found in the population of Rio de Janeiro and the Iberian Peninsula ones [23, 44, 45]. Pairwise genetic distance analysis was performed between the study sample and previously published data concerning five Brazilian populations [24–27, 38] and those representatives of continental population groups: Europe [46], Africa [47], Asia [48] and Native American (Silva DA, personal communication). In comparison with populations from other Brazilian regions, significant differences were observed when comparing Rio de Janeiro (RJ) to Santa Catarina (SC) and Rio Grande do Sul (RS). For the other Brazilian populations: Alagoas (AL), Amazonas (AM) and Sa˜o Paulo (SP), a significant genetic difference was not identified. Genetic distance among Rio de Janeiro state and other population groups is shown in Table 3. In relation to the ancestral groups that potentially contribute to the formation of the Brazilian population, the MDS plot of Fig. 1, indicates the sample from Rio de Janeiro plotted closer to the African (AF) and it has shown no significant genetic differentiation between them. This is
Table 3 FST genetic distances between the studied population (RJ) and other populations RJ
AF
TE
PT
CH
AL
AM
SC
RS
RJ
–
AF
0.03159
–
TE PT
0.14945* 0.22524*
0.26905* 0.3172*
– 0.34375
–
CH
0.14945*
0.22406*
0.14021*
0.20979*
–
AL
0.01048
0.06162*
0.10817*
0.17225*
0.10594*
–
AM
0.02378
0.10851*
0.07073*
0.25653*
0.10403*
0.02276*
–
SC
0.08377*
0.16654*
0.17058*
0.05694*
0.10514*
0.04528*
0.09485*
RS
0.06399*
0.14865*
0.124*
0.13116*
0.08633*
0.02948*
0.05193*
0.01141
–
SP
0.00773
0.06146*
0.13073*
0.14578*
0.10745*
0.00311
0.02428*
0.03183*
0.02628*
SP
– –
RJ Rio de Janeiro, AF Africa, TE Terena, PT Portugal, CH China, AL Alagoas, AM Amazonas, SC Santa Catarina, RS Rio Grande do Sul, SP Sa˜o Paulo * Significant genetic distance values after applying Bonferroni correction in FST P value
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References
Fig. 1 MDS plot of the pairwise genetic distances among six Brazilian samples [Rio de Janeiro population of present study (RJ); Alagoas (AL); Sa˜o Paulo (SP); Amazonas (AM), Santa Catarina (SC); Rio Grande do Sul (RS)] and four reference samples from Africa (Angola-AF), Europe (Portugal-PT), Asia (China-CH) and Native American (Terena-TE)
explained by the prevalence of African haplogroups in the study sample. Significant genetic differentiation was observed among Rio de Janeiro state, Native American (Terena-TE), European (Portugal-PT) and Asian (ChinaCH) populations.
Conclusions In conclusion, we have shown that 57.9 % of the mtDNA lineages from the admixed population of Rio de Janeiro state are associated to African ancestry. The results confirm that the population derived from crosses among Native Americans interethnic Africans and Europeans for over 500 years but there was an asymmetry favouring crosses among male Europeans and Africans or Amerindian women as evidenced by historical, anthropological and genetic studies already carried out with mitochondrial DNA and the Y-chromosome lineage markers. The study demonstrates, therefore, that admixed population cannot be adequately classified taking into consideration just arbitrary parameters, as skin color, because of the heterogeneity of our population. Currently, there are sociological discussions happening in the Brazilian political life concerning the introduction of public policies favoring the non-white population in a population demonstrably mixed. Moreover, not only the application of phylogenetic methods to our data, but also the comparison with closely related sequences will provide valuable information for forensic and population genetic analysis in the admixed population of Rio de Janeiro state.
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