Questioning the melting pot: Analysis of Alu inserts in three population

samples from Uruguay.

Pedro C. Hidalgo*, Patricia Mut, Elizabeth Ackermann, Gonzalo Figueiro

Monica Sans

Departamento de Antropologa Biolgica. Facultad de Humanidades y Ciencias

de la Educacin. Universidad de la Repblica, Uruguay.
*Correspondence:
Departamento de Antropologa Biolgica.
Facultad de Humanidades y Ciencias de la Educacin.
Universidad de la Repblica,
Magallanes 1577. 11200. Montevideo
Uruguay.

Tel: +(598) 2403-1004

E-mail: drpedro.hidalgo@gmail.com
KEY WORDS: ADMIXTURE, GENE DIVERSITY, POPULATION
STRUCTURE, ALU INSERTION POLYMORPHIS.
Running title: Alu polymorphisms in Uruguay

Human Biology (enviado)

ABSTRACT
The way that immigrants integrate to recipient societies has been discussed for
decades, mainly from the perspective of the social sciences. Uruguay, as other
American countries, received different waves of European immigrants, although
the details of the process of assimilation, when occurred, are unclear. In this
paper, we use genetic markers to understand the process suffered by the
Basques, one of the major migration waves that populated Uruguay, and its
relation to other immigrants as well as to Native American and African
descendants. For this purpose, we analyze the allele frequencies of ten ALU
loci (A25, ACE, APOA1, B65, F13B, PV92, TPA25, HS2.43, and HS4.65) in
three samples from Uruguay (two of Basque-descendants, one of non-Basquedescendants) from two locations: Montevideo and Trinidad. No departure from
Hardy-Weinberg expectations was observed, with the exceptions of the APO
and D1 loci in the non-Basque descendants sample. Our data show that the
maximum genetic contribution in the three samples comes from Europe (78% to
88%) with minor African (10% to 15%) and Native American (0% to 10%)
contributions to the present-day Uruguayan population. Genetic distances
reveal that Basque-descendants from Trinidad cluster with Europeans, while
both Montevideo samples cluster together and separate from other populations,
showing two different ways of integration, related to the general characteristics
of each regional population.

Introduction
According to the 2011 National Census, Uruguay has a population of
3,286,314 inhabitants, 1,319,108 of them living in the capital city, Montevideo
(INE 2011). The present population is the consequence of various processes,
several of them shared with other Latin American countries. However, there are
some differences make the Uruguayan population unique, such as the variety of
European source populations and the lack of Native American and African
descendant communities.
In general terms it is possible to recognize several origins for European
migratory waves, most of them occurring during the 19th and 20th centuries,
that superpose to the Iberians (mostly from the Canary Islands but also several
Basques, and Portuguese) that occupied the region during colonial times. The
first wave of immigrants (1825-1842) came during the early times of Uruguayan
independence and includes 40,000 to 45,000 individuals, mostly French
Basques. More Basques came in a second wave (1842-1876), in this case
mainly from Spain (Barrn and Nahum 1979; Marenales and Luzuriaga 1990).
These two foundational waves were composed mostly by males (81.8%,
compared to 18.2% females). A third, forced wave occurred during the
Spanish Civil War (1936-1939), and does also include several Basques
(Arocena and Aguiar 2007). It should be emphasized that during the 19th
century, migrants to Uruguay were mostly of Spanish, Portuguese and Italian
origin, while during the 20th century, origins diversified, adding several
thousands of Armenians, Jews from Western Europe, Central Europe and the
Middle East, Russians, Swiss, and others (Vidart and Pi Hugarte, 1969). In

1908 the proportion of foreigners in the population was approximately 18%,

decreasing to 8% in 1963 and to a present-day minimum of 3% (INE 2012).
The process of assimilation, when occurred, is unclear, and the
coexistence in a common social space, shared by the immigrants and other
ethnic groups (Africans, Natives, and their descendants), does not necessary
imply a melting pot, a model inspired in I. Zangwills play about immigrants in
the United States of America, in which immigrants quickly amalgamated with
the receiver population. This model is opposed to cultural pluralism, in which
ethnic groups retained some individuality. According to Yinger (1985), the
process of assimilation includes four threads, which can be termed as
structural, cultural, psychological, and biological (amalgamation). The
amalgamation, which results in the gene pools being indistinguishable, depends
on the characteristics of both the migrants and the recipients, and various
barriers have been observed (linguistic, religious, or "racial") that can restrain
the process. However, in general terms, the coexistence of different groups
does not occur in conditions of equality. Nowadays in Uruguay, Native
American-descendants and Afro-descendants claim for cultural diversity. As
Olivera Chirimini (2004) said, Natives American as well as African descendants
have been forgotten, occulted, and erased from the history of the Rio de la
Plata region.
Since the mid-1980s the Uruguayan population has been studied in
terms of their genetic structure and how it reflects the prehistoric and historic
processes of the population. The tri-hybrid origin composed of Native
Americans, Africans, and Europeans has been proven through blood
polymorphisms and DNA, with estimates of around 10% (1% to 20%) Native
American and 6% (7% to 15%) African contribution (Sans et al. 1997, 2006;

Hidalgo et al. 2005). Different studies have shown the matrilineal contributions
to be between 20% and 62% Native American and between 7% and 15%
African in different parts of the country (Bonilla et al. 2004, Gascue et al. 2005,
Pagano et al. 2005a, Sans et al. 2006). Few studies have analyzed the Ychromosome, showing low proportions (<7%) of Native American and African
contribution (Bertoni et al. 2005, Pagano et al 2005b). These data are poorly
correlated with the information given in the last Census (INE, 2011), as only
4.9% of the population believes to have at least one Native American ancestor,
and 7.8% believe to have at least an African one.
Recently, we analyzed the maternal contribution in a self-defined
Basque-descent sample from the city of Trinidad, Flores, Uruguay (Sans et al.
2011). Unexpectedly, it showed that the Native American contribution was
similar to the one found in a general sample of mixed ancestry from Montevideo
(20%), with a smaller African contribution (2%). The analyses of surnames
showed a different picture, with 91% of the individuals having a Basque paternal
surname, while 41% had a Basque maternal surname.
To improve the comprehension of the ways of integration of European
ethnic groups, we analyzed nuclear DNA markers in the above mentioned self
identified Basque-descendant subpopulation from Trinidad, and we extended
the study to Basque-descendants and non-Basque descendants from
Montevideo, to answer two research questions: 1) is it possible to observe
significant differences among Basque descendants living in different areas of
Uruguay? 2) are there differences between Basques and non-Basques living in
the same area or does amalgamation occur?
For this purpose we selected Alu inserts, which comprise approximately
10% of human DNA (Rowold and Herrera 2000; Batzer and Deininger 2002). In

the course of human evolution, the number of Alu type transposons has grown
through a mechanism of self-copy (Batzer and Deininger 2002), being a
testimony of both ancient and recent evolutionary processes. As the insertion of
Alu elements is produced by a single mutational event, it is possible to establish
the ancestral state (Rowold and Herrera 2000). Alu elements have the
advantage of being easily identified and some of them have been characterized
in many populations, including the Basque (Batzer et al. 1994; Comas et al.
2000; Gonzlez-Prez et al. 2003; Garca-Obregn et al. 2007; Gmez-Prez
et al. 2007; Stoneking et al. 1997; Terreros et al. 2005). In addition, Alu
insertions have been validated for the estimation of the degree of mixing in
hybrid populations, based on yielding a correct classification of 91.5% of
individuals using discriminant functions (Gmez-Prez et al. 2007).

Materials and Methods

The first sample is composed by subjects originally ascertained for
anthropological studies, all of them descendant from Basques living in the city
of Trinidad (BTD), Department of Flores, located 200 kilometers away from
Montevideo in the southwest of Uruguay (described in Sans et al 2011). A
second sample is composed by 55 individuals of self-defined Basque descent
from the city of Montevideo (BMVD), having at least one Basque grandparent or
with a recognized Basque surname. The third sample is a subset of 70
individuals of European ancestry but without known Basque ancestry, that is,
without a known grandparent of Basque origin and lacking surnames identified
as such, also taken from Montevideo (MVD). All individuals were asked for
information regarding parental and grandparental place of birth. None of the

individuals were close biological relatives at least at third grade, and all have at
least two generations born in the country.
After a complete description of the study was provided, written informed
consent was obtained from all subjects, and confidentiality was assured. The
project was approved by the Facultad de Humanidades (Universidad de la
Repblica) Ethics Review Board.
DNA typing
The genetic characterization of the Uruguay population was carried out
by analyzing the polymorphism of 10 autosomal Alu insertions (A25, ACE,
APOA1, B65, D1, F13B, PV92, TPA25, HS2.43, and HS4.65), typed using DNA
isolated from peripheral venous blood, buccal swab and hair. The extraction of
genomic DNA from the leukocyte fraction of the blood and buccal swab samples
was carried out using the Miller et al. (1988) salting out technique, while DNA
from hair was extracted following the protocol of Hidalgo et al. (2009). The
extracted DNA was stored at -20C.
PCR amplification
The 10 Alu insertions were typed according to the conditions previously
established for A25, ACE, APOA1, B65, F13B, PV92, and TPA25 loci by
Garca-Obregn et al. (2006), while for HS2.43 and HS4.65, the conditions
published by Ray et al. (2005) were used. The genotyping was carried out
through electrophoresis in 2% agarose gel and staining using ethidium bromide,
and visualizing the gels under UV light. In some cases the results were
confirmed in polyacrylamide gel electrophoresis (PAGE) with silver staining.
Population Affinities
For the analysis of the genetic affinities of our samples, 16 populations
from a broad continental context, typed for the same ten Alu loci used in our

study, were selected from the literature (Table 1). We also included two
admixed populations from the United States of America for comparative
purposes.
Statistical Methods
Allele and genotype frequencies of the Alu-insertion loci were obtained
by direct gene counting (Li, 1976). Divergence from the Hardy-Weinberg
equilibrium (HWE) was determined at each locus by Chi-square, Levenes
correction and exact test with the Biosys 2 program (Swofford and Selander
1997). For each sample the observed and expected heterozygosity were
calculated using GenAlEx 6.5 software (Peakall and Smouse 2012).
Polymorphic information content (PIC) value for Alu loci was calculated on the
basis of observed allele frequencies (Botstein et al., 1980), using Cervus
software (Marshall, 2000).
The heterogeneity in allele frequencies among Uruguayan samples were
performed with an exact test using the Genepop package (Rousset, 2008).
Genetic differentiation was measured by GST (Nei, 1973; 1987) by using the
gene diversity for the entire population (H T) and the average gene diversity for
samples (HS) with the Dispan computer program (Ota, 1983). The significance
of the allele heterogeneity in each locus among Uruguayan samples was
calculated according to Srikumari et al (1986).
Analysis of molecular variance (AMOVA; Excoffier et al., 1992) was
performed to test the presence of genetic structure among samples with the
GenAlEx 6.5 software (Peakall and Smouse 2012).
Genetic admixture was estimated by Elstons method (1971). We used
the average allele frequencies for the European, African and Native American
populations listed in Table 1. Allele frequencies for Native Americans were

complemented using data for seven loci in Amazonian Indians (Battilana et al.
2006). The allele frequencies for each individual locus were pooled and
weighted by arithmetical means.
Neis DA distance (Nei et al. 1983; Nei 1987 ) and Reynolds et al. (1983)
FST distance among Uruguayan samples were computed. We also estimated
Neis DA genetic distances between the Uruguayan samples and the
populations listed in Table 1. DA genetic distances were represented in
Unweighted Pair Group Method with Arithmetic Mean (UPGMA) trees (Sneath
and Sokal 1973) by means of the Neighbor program, generating 1,000
bootstrap replicates with Seqboot, and a consensus tree was obtained with
Consense as implemented in the PHYLIP package (Felsenstein 2002).
The Neis genetic distance matrix obtained was then represented in a
two-dimensional space by applying nonmetric multidimensional scaling (MDS)
analysis, using the SPSS v.21 (SPSS Inc., Chicago, IL) statistical package.

Results
The distribution of Alu insertion frequencies in the ten loci typed for the
three Uruguayan samples is shown in Table 2. The highest allele frequency
corresponds to the APOA1 locus in all samples. The lowest frequencies were
observed for the HS2.43 and the HS4.65 loci. Table 3 shows the departure from
HWE. Significant deviations from HWE (p<0.05) were observed for the loci
APOA1, D1 and TPA25 in MVD, for locus D1 in BMVD, and for loci B65, D1 and
F13B in BTD. Significant deviation from HWE using the Dunn-Sidak correction
(Sokal and Rohlf 1995) were only found for APO and D1 loci in the MVD
sample, at the 0.16% level.

Table 4 gives the observed and expected heterozygosity for each sample. The
highest average expected heterozygosity was found in the BTD sample (33%)
and the lowest was found in BMVD (27%). There are also broad patterns of
genetic variation within the locus by sample (Table 4). We found differences
among the three samples in the mean heterozygosity (F statistic) with an
excess of homozygote excess of almost 11% in MVD sample relative to HardyWeinberg expectation (Table 4). The average polymorphic information content
(PIC) ranges from 0.223 to 0.259, being the highest and the lowest PIC
recorded in BTD and BMVD respectively (Table 4).
For the ten Alu loci genetic heterogeneity for each paired sample was
calculated, showing both samples from Montevideo (MVD and BMVD)
significant differences with respect to Basques from Trinidad, but not between
one another (Table 5).
The gene diversity and the coefficient of differentiation analysis are given
in Table 6. The intrapopulational gene diversity (Hs ) for the Uruguayan
samples ranged from 0.041 (HS4.65) to 0.458 (F13B) with a mean value of
0.302. The average value of Dst a measure of the gene diversity among
samples, is 0.01 with large variation among loci (0.001 to 0.028). The gene
differentiation coefficient relative to the total population (G ST) shows large
variation from locus to locus (0.004 to 0.063), being the highest differentiation
found for the B65 locus, and the least for PV92. The mean G ST over all 10 loci is
3.1% (slightly lower than Wrights FST = 0.034), with significant differences in
five loci. The mean estimate of GST indicates modest but definite differentiation
in Alu polymorphisms.
The result of the GST indicates that the fraction of the total gene diversity
is fundamentally attributable to differences within the samples. Our analysis

indicates that even with ten loci, reliable estimation of genetic variation could be
obtained. Hierarchical analysis of molecular variance (AMOVA) across
individuals from the three samples revealed most of the genetic diversity to be
due to the variation within samples (88%), while the variation among samples
(12%) was small but significant (Table 7).
Genetic distances, based on Neis DA and using the 10 Alu loci, range
from 0.87% to 3.6%, being the highest value between BTD and the Montevideo
samples (Table 8). Similar results were obtained using Reynoldss distance
(Table 8). When constructing a UPGMA dendrogram based on Neis DA
distances the results revealed two distinct clusters, one consisting of MVD and
BMVD and the other including BTD alone (Fig 1).
Using Elstons (1971) admixture method we estimated an average of 4%
Native contribution and 12% African contribution (Table 9). The level of
admixture estimated indicates a substantial European ancestry in the samples.
In addition, these data show that the percentage of African alleles ranges from
9.6% to 15% in the Uruguayan samples with a relatively low level of Native
American contribution, albeit higher in BTD.
The groups formed using MDS analysis (Fig. 2) confirms the
relationships among populations found in the admixture analysis. The stress
value of the MDS is 13.2 %, which is acceptable according to Kruskal and Wish
(1991), and the R 2 value is 0.93, indicating that the model has a high goodness
of fit (Schiffman et al. 1981). The Uruguayan samples are placed in the
European cluster of populations, and separated from the African and the Native
American clusters. The dominant feature along the first dimension (D1)
separates Native Americans and Hispanics from the United States from
Europeans and Africans. The second dimension (D2) separates Africans/Afro-

Americans and Europeans from other populations. In both, it is evident that

Uruguayan samples exhibit a typically European profile. This pattern is repeated
in the UPGMA tree based on the DA distances (Fig. 3), where the Uruguayan
samples fit within the main European cluster, separate from both Native
American and Africans. However, the Montevideo samples (MVD and BMVD)
appear separate within the European cluster, while the BTD sample is more
related to the Spanish populations.
Discussion
The pattern of genetic variation defined by the Alu insertion/deletion
polymorphism in the Uruguayan population shows higher differentiation than the
one observed when considering other systems as HLA, ABO and Rh (CDE) loci
in the Uruguayan population as a whole (Hidalgo et al. 2007). Three statistics
showed significant departure from HWE in several comparisons (7 in 30),
leading to the conclusion that loci APO and D1 are not in HWE in the MVD
sample. One possible explanation could be the occurrence of microevolutionary
process that affected Uruguayan populations, such as founder effects, external
and internal migrations, and consanguinity, as discussed below.
The Native American contribution found in Montevideo is similar to the
one found by Sans et al (1997) based on classical markers, while the estimate
for Trinidad is close to the estimates for Uruguay as a whole according to the
data by Hidalgo et al (2005) based on Polymarkers. However, the African
contribution determined using Alu insertions is slightly higher than the one found
in previous work (Sans et al 1997; Hidalgo et al. 2005). Differences in African
contribution can be due to different factors, as sampling, lack of adequate data
on parental populations for the estimates and/or changes in the frequencies in

the parental populations during time. It is interesting to note that Montevideo as

a whole as well as Basque descendants from Trinidad have been studied for
Native maternal contributions showing similar results (21% and 20% Native,
respectively) (Gascue et al 2005, Sans et al 2012).
The two samples from Montevideo form a tight cluster that could be
explained by different patterns of mating and amalgamation inside this city,
concordant with the concept of melting pot. On the other side, the sample of
Basque-descendants from Trinidad forms a separate cluster. Moreover,
comparisons of Neis and Reynolds distances showed that the distances
between the MVD and BMVD samples are not significant but the distances
between BTD and MVD, as well as between BTD and BMVD, are statistically
significant. When including different world populations, the three samples
formed a large cluster with the European populations, Trinidad being closer to
West European populations, and all three samples apart from United States
Hispanics, as well as from African and Native American populations. As
mentioned before, the magnitude of Native American or African contributions
cannot be the explanation, as Trinidad has more non-European contribution
than the other two samples. Other reasons need therefore to be explored.
Local genetic structures are determined not only by the amount of gene
flow but also by the characteristics, including size, of the populations that get in
contact. During the last part of the 19th Century and first half of the 20th Century,
Uruguay received a large amount of migrants from Europe and surrounding
regions, process that ended soon after the end of World War II (Vidart and Pi
1969). However, after 1910, internal migrations had an important role, being
mainly from the countryside to the capital city, Montevideo (Campiglia, 1968).
When analyzing the population growth during the 20 th Century, while from 1908

to 1996 Montevideo multiplied its population by 4.3 (probably as an effect of

both types of migration), the population of Trinidad multiplied by 2.4, and a
pronounced decline in growth rate is observed when considering years 1963 to
1996, when external migrations had finished (data from INE, 2013): during this
period, the population of Trinidad multiplied by 2.1 and 1.3 respectively. It is
then possible to hypothesize that Montevideo became more exogamous due to
incoming migrations, while Trinidad continued being relatively endogamous. A
study about patterns of endogamy in Montevideo showed that Italians and
French (mostly Basques) have the highest rates of endogamy, followed by the
Spaniards, including in this last group a broad variety of origins. In all three
cases, the endogamy of these groups is higher than in the population of
Montevideo taken as a whole (Barreto and Sans, 2000).
As a conclusion, it is possible to answer the two questions asked in the
introduction of this study. The first question, on the possibility of observing
significant differences among Basque descendants living in different areas of
Uruguay, it has been proven that there are differences between the ones living
in Trinidad related to the ones living in Montevideo. The second question,
relative to differences between Basques and non-Basques living in the same
area and the occurrence of amalgamation, it is possible to say that, in
Montevideo, amalgamation has indeed occurred. However, we are not able to
either confirm or dismiss the occurrence of amalgamation in Trinidad, as there
is no data available for non-Basque descendants in the city or the surrounding
region. Moreover, our data show that each population, with a different history
and inhabiting a different region, need to be studied, as two populations with an
apparent same origin (Basques) were found to differ in their present-day genetic
structure. Consequently, as a final consideration, we emphasize the importance

of prospective analyses of the ancestral characteristics of the populations to be

studied, especially when admixture occurs as it is the case in Latin America.
Multiculturalism is the real definition for Uruguay. Finally, we strongly suggest
the necessity of developing holistic views to deal with tri-hybrid population
complexities. Multiculturalism is not only related with sociocultural aspects, but it
also affects the genetic structure of the populations (Harrison and Boyce 1972;
Stobart 2005; Stone and Lurquin 2007)

Acknowledgements
This work was made possible thanks to the collaboration of the Basque
descendants living in Trinidad and Montevideo, as well as other inhabitants of
Montevideo that generously participated in this project. We especially thank the
association Haize Hegoa from Montevideo, and their members M. Bengoa, E.
Poittevin-Gilmet, D. Maytia and R. Fernndez. We wish to thank G. Dorado for
her technical assistance.
The project was supported by CSIC (Universidad de la Repblica, Uruguay).
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Table 1. Populations selected with the ten Alu loci typed in our study.
Region / Population

Code

Reference________

Europeans
Catalans
Andalusians
Galicians
Basques
Frenchs
Armenians
Italians

CAT
ANL
GAL
BAS
FRE
ARM
ITAL

Comas et al. 2000

Comas et al. 2000
Terreros et al. 2009
Comas et al. 2000
Romualdi et al. 2002
Romualdi et al. 2002
Selvaggi et al. 2010

BEN
KEN
RWA
CAM
BAN
NGU

Terreros et al. 2009

Terreros et al. 2009
Terreros et al. 2009
Terreros et al. 2009
Romualdi et al. 2002
Romualdi et al. 2002

YAN
MAY
WAO

Romualdi et al. 2002

Romualdi et al. 2002
Gomez-Perez et al. 2011

HAM
AAM

Romualdi et al. 2002

Romualdi et al. 2002

Africans
Benin
Kenya
Rwanda
Cameroon
Bantu
Nguni
Native Americans
Yanomami
Maya
Waorinai
USA

Hispanic
African Americans

Table 2. Allele Frequencies and Sample Sizes.

Locus Allele

MVD

BMVD

A25

64

49

58

insertion

0.117

0.082

0.138

deletion

0.883

0.918

0.862

67

46

58

insertion

0.343

0.283

0.379

deletion

0.657

0.717

0.621

58

44

58

insertion

0.845

0.773

0.94

deletion

0.155

0.227

0.06

67

49

58

insertion

0.336

0.184

0.474

deletion

0.664

0.816

0.526

49

27

57

insertion

0.296

0.278

0.482

deletion

0.704

0.722

0.518

35

19

58

insertion

0.429

0.263

0.457

deletion

0.571

0.737

0.543

67

48

58

insertion

0.149

0.146

0.198

deletion

0.851

0.854

0.802

63

48

58

insertion

0.278

0.292

0.509

deletion

0.722

0.708

0.491

66

47

58

insertion

0.068

0.011

0.043

deletion

0.932

0.989

0.957

51

47

58

insertion

0.010

0.011

0.043

deletion

0.990

0.989

0.957

ACE

APO

B65

D1

F13B

PV92

TPA25 N

HS2.43 N

HS4.65

BTD

Table 3. Test of Hardy-Weinberg equilibrium of allele frequencies.

Locus

MVD

BMVD

BTD

Prob.

Prob.

Prob.

ChiSquare

0.1756 ns

0.5338 ns

0.223

ns

Levene

0.147

ns

0.562

ns

0.240

ns

Exact Test

0.189

ns

1.000

ns

0.579

ns

ChiSquare

0.5495 ns

0.3349 ns

0.8475 ns

Levene

0.508

ns

0.296

ns

0.900

ns

Exact Test

0.590

ns

0.463

ns

1.000

ns

ChiSquare

0.0003 ***

0.138

ns

0.625

ns

Levene

0.0002 ***

0.115

ns

0.652

ns

Exact Test

0.002

0.187

ns

1.000

ns

ChiSquare

0.0514 ns

0.741

ns

0.008

**

Levene

0.590

ns

0.683

ns

0.010

Exact Test

0.097

ns

0.648

ns

0.017

ChiSquare

0.0012 **

0.0457 *

0.0477 *

Levene

0.001

**

0.055

ns

0.041

Exact Test

0.002

**

0.138

ns

0.062

ns

ChiSquare

0.693

ns

0.709

ns

0.039

Levene

0.631

ns

0.795

ns

0.033

Exact Test

0.734

ns

1.000 ns

0.039

ChiSquare

0.151

ns

0.2379 ns

0.155

ns

Levene

0.163

ns

0.256

ns

0.133

ns

Exact Test

0.335

ns

0.571

ns

0.204

ns

TPA25 ChiSquare

0.009

**

0.180

ns

0.114

ns

Levene

0.008

**

0.156

ns

0.130

ns

Exact Test

0.012

0.176

ns

0.188

ns

HS2.43 ChiSquare

0.552

ns

0.941

ns

0.732

ns

Levene

0.577

ns

1.000

ns

0.760

ns

Exact Test

1.000

ns

1.000

ns

1.000

ns

A25

ACE

APO

B65

D1

F13B

PV92

HS4.65 ChiSquare

0.944

ns

0.941

ns

0.732

ns

Levene

1.000

ns

1.000

ns

0.760

ns

Exact Test

1.000

ns

1.000

ns

1.000

ns

ns: no significant; p<0,05 *; p<0,01 **; p<0,001 ***

Table 4. Genetic parameters in Uruguayan samples .

MVD

BMVD
He

PIC

Ho

BTD

Locus

Ho

He

PIC

Ho

He

PIC

A25

0.172 0.207 0.185 0.169

0.163 0.150 0.139 -0.089

0.276 0.238 0.210 -0.160

ACE

0.418 0.451 0.349 0.073

0.348 0.405 0.323 0.142

0.483 0.471 0.360 -0.025

APO

0.138 0.262 0.228 0.474

0.273 0.351 0.289 0.224

0.121 0.113 0.106 -0.06

B65

0.552 0.446 0.347 -0.238

0.286 0.300 0.255 0.047

0.672 0.499 0.370 -0.348

D1

0.224 0.417 0.330 0.461

0.556 0.401 0.321 -0.385

0.368 0.499 0.374 0.262

F13B

0.457 0.490 0.370 0.067

0.421 0.388 0.312 -0.086 0.362 0.496 0.373 0.270

PV92

0.299 0.254 0.221 -0.175

0.292 0.249 0.218 -0.171

0.259 0.318 0.267 0.187

TPA25

0.270 0.401 0.321 0.327

0.333 0.413 0.328 0.193

0.603 0.500 0.375 -0.207

HS2.43

0.136 0.127 0.119 -0.073

0.021 0.021 0.021 -0.011

0.086 0.082 0.079 -0.045

HS4.65

0.020 0.019 0.020 -0.010

0.021 0.021 0.021 -0.011

0.086 0.082 0.079 -0.045

Media
SD

0.269 0.307 0.249 0.108

0.157 0.150 0.110 0.079

0.271 0.270 0.223 -0.015 0.332 0.330 0.259 -0.018

0.158 0.147 0.115 0.058 0.197 0.177 0.124 0.064

Ho: observed heterozygosity; He: expected heterozygosity; UHe: unbiased

expected heterozygosity; PIC: polymorphic information content; SD: Standard
deviation.

Table 5. Genic differentiation for each population pair. P-value for each
population pair across all loci (Fisher's method)
Population pair

df

p-value

MVD
x BMVD
26.81
20
0.1406
MVD
x BTD
49.16
20
0.0004**
BMVD
x BMVD
81.05
20
0.0000**
__________________________________________
*** p<0.001

Table 6. Heterozygosity and Gene Diversity Analysis

Locus

Ht

A25

Dst

Gst

0.199 0.198

0.001

0.005

1.71

ACE

0.446 0.442

0.004

0.007

2.39

APO

0.251 0.242

0.009

0.037

11.84**

B65

0.443 0.415

0.028

0.063

21.94**

D1

0.456 0.439

0.017

0.037

9.84**

F13B

0.473 0.046

0.015

0.031

6.94*

PV92

0.275 0.273

0.002

0.004

1.38

TPA25

0.461

0.438

0.023

0.048

16.22**

HS2.43

0.078

0.077

0.001

0.014

4.79

HS4.65

0.042

0.041

0.001

0.011

3.43

Mean

0.312

0.302

0.010

0.032

* p < 0.05, **p<0.01

Hs

2 (df=2)

Table 7. Genetic distances among Uruguayan samples.

MVD
BMVD
BTD

MVD
**
0.009
0.018

BMVD
0.219
**
0.036

BTD
0.037
0.079
**

Upper: Reynolds distances. Lower: Neis distances

Table 8. Analysis of molecular variance (AMOVA) performed to

hierarchical partitioning of genetic variation among the three Uruguay samples.
Source
Among Samples
Within Samples
Total

df
2
174
176

SS
93.086
893.818
986.904

MS
46.543
5.137

Statistical testing by random permutation:

Stat
Value
P (rand >= data)
PT
0.121
0.010

Est. Var.
%
0.709
12
5.137
88
5.846
100

Table 9. Admixture calculations in the three Uruguay samples.

European
BTD
MVD
BMVD

0.782
0.847
0.883

African

Native American

0.120
0.151
0.096

0.098
0.002
0.021

Figure 1.UPGMA tree depicting genetic variation among BMVD and MVD with
BTD.

Figure 2. Nonmetric multidimensional scaling (MDS) plot representing a Neis

Da genetic distance matrix for different populations . Genetic distances were
computed based on the ten Alu insertion frequencies. Sample codes in Table 1.

Figure 3. UPGMA tree of the genetic distance matrix for the ten Alu insertions
in the ten loci analyzed. Sample codes in Table 1.