Y Chromosome Haplogroup Distribution in Indo-

European Speaking Tribes of Gujarat, Western India

Priyanka Khurana1., Aastha Aggarwal2, Siuli Mitra3, Yazdi M. Italia4, Kallur N. Saraswathy3,
Adimoolam Chandrasekar5, Gautam K. Kshatriya3*.
1 Department of Anthropology, School of Applied Sciences, Dr. Harisingh Gour University, Sagar, Madhya Pradesh, India, 2 South Asia Network for Chronic Disease, Public
Health Foundation of India, Delhi, India, 3 Department of Anthropology, University of Delhi, Delhi, India, 4 Valsad Raktdan Kendra, R.N.C. Free Eye Hospital Complex,
Valsad, Gujarat, India, 5 Anthropological Survey of India, Southern Regional Center, Mysore, Karnataka, India

Abstract
The present study was carried out in the Indo-European speaking tribal population groups of Southern Gujarat, India to
investigate and reconstruct their paternal population structure and population histories. The role of language, ethnicity and
geography in determining the observed pattern of Y haplogroup clustering in the study populations was also examined.
A set of 48 bi-allelic markers on the non-recombining region of Y chromosome (NRY) were analysed in 284 males;
representing nine Indo-European speaking tribal populations. The genetic structure of the populations revealed that none
of these groups was overtly admixed or completely isolated. However, elevated haplogroup diversity and FST value point
towards greater diversity and differentiation which suggests the possibility of early demographic expansion of the study
groups. The phylogenetic analysis revealed 13 paternal lineages, of which six haplogroups: C5, H1a*, H2, J2, R1a1* and R2
accounted for a major portion of the Y chromosome diversity. The higher frequency of the six haplogroups and the pattern
of clustering in the populations indicated overlapping of haplogroups with West and Central Asian populations. Other
analyses undertaken on the population affiliations revealed that the Indo-European speaking populations along with the
Dravidian speaking groups of southern India have an influence on the tribal groups of Gujarat. The vital role of geography in
determining the distribution of Y lineages was also noticed. This implies that although language plays a vital role in
determining the distribution of Y lineages, the present day linguistic affiliation of any population in India for reconstructing
the demographic history of the country should be considered with caution.

Citation: Khurana P, Aggarwal A, Mitra S, Italia YM, Saraswathy KN, et al. (2014) Y Chromosome Haplogroup Distribution in Indo-European Speaking Tribes of
Gujarat, Western India. PLoS ONE 9(3): e90414. doi:10.1371/[Link].0090414
Editor: Michael D. Petraglia, University of Oxford, United Kingdom
Received November 15, 2013; Accepted February 1, 2014; Published March 10, 2014
Copyright: ! 2014 Khurana et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: The work was funded by Department of Biotechnology (DBT) grant (vide letter BT/PR9840/MED/12/366/2007). The funders had no role in study design,
data collection and analysis, decision to publish, or preparation of the manuscript.
Competing Interests: The authors have declared that no competing interests exist.
* E-mail: g26_51@[Link]
. These authors contributed equally to this work.

Introduction with Africa in the west, Eurasia in the north and the Orient in the
east. The country’s unique location gives rise to a great variety of
India is the second most populous country in the world with a environmental conditions and associated biodiversity [12] which
population of 1.21 billion [1]. About 4,635 different population in turn has attracted people from all over the globe.
groups are spread across the country [2]. Exorbitant as it may Archaeological and paleontological evidence dating back to the
sound, existence of at least 50–60 thousand essentially endoga- middle and late Pleistocene era points towards early human
mous groups has been reported in the country [3–4]. The present occupation of the Indian subcontinent [13–16]. Similar evidence
day Indian population is divided into tribal and non-tribal groups. of antiquity of Indian populations has been established by genetic
Tribal populations constitute 8.2% of the total population [5] and marker studies. The high levels of gene diversity and coefficients of
are considered to be the indigenous populations of India [6–8]. gene differentiation obtained using autosomal markers for Indian
The tribal groups of India belong to four broad linguistic families: populations are a testimony of both, the antiquity and the complex
Austro-Asiatic, Dravidian, Indo-European and Tibeto-Burman population structure, of this massive human conglomeration
[9]. The Indo-European and Dravidian speaking populations are existing in the southern part of Asia [17–19]. Recent genetic
considered to be the major contributors to the development of studies based on mtDNA showing the high frequency of
Indian culture and society [10]. Both Austro-Asiatic and mitochondrial lineages with greater age, high diversity and wide
Dravidian speaking tribes belong to the primary pre-historic distribution amply demonstrate the prehistoric existence of
populations of India [7,11]. Tibeto-Burman speakers also include mankind on the Indian subcontinent [20–21]. Similar analyses
many tribal populations but their geographical distribution is of Y chromosome variation based on both bi-allelic and
largely restricted to the North-Eastern region of India. India thus microsatellite markers have documented the existence of substan-
exhibits an enormous genetic, cultural and linguistic diversity tially deep rooted lineages among Indian populations buttressing
which can partly be attributed to its position at the tri-junction:

PLOS ONE | [Link] 1 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

the claim of early habitation of humans on the Indian districts of Gujarat. A description of relevant population
subcontinent [22–25]. characteristicsof study groups along with populations from other
The unique and complex structure of the Indian population is studies is presented in Table 1. Figure 1 illustrates the sampling
attributed to the multiple waves of migration and the resultant location of study populations from Gujarat. The detailed
gene-flow which occurred in the past [12,26]. Some scholars have description of the study populations has been given previously
provided evidence to explain the origin and migration of the major [42]. Blood samples collected in EDTA coated vacutainers were
linguistic families extant in India [25,27]. It is worth mentioning subjected to DNA extraction following salting-out method [43]. A
that speakers of Austro-Asiatic language in India are exclusively total of 48 bi-allelic markers were analysed to identify the Y
tribal, which may be indicative of their being one of the oldest chromosome haplogroups. The markers were typed using primer
inhabitants of India [28,29]. The arrival of the Indo-European pairs described in Karafet et al. [44]. The PCR cycling conditions
speakers via the Northern corridor of India around 3,500–4,000 with an initial denaturation of 5 minutes at 95uC; followed by 35
years ago is believed to be responsible for one of the major influxes cycles of 1 minute at 94uC; 45 seconds at the primer-specific
of people in the Indian subcontinent [30], followed later by the annealing temperature (52–60)uC and extension at 72uC for
infiltration of colonizers from different parts of the world. Thus, 2 minutes and 30 seconds; followed by final extension of
several studies have affirmed the influence of Eurasian and Asian 7 minutes at 72uC were followed. The amplicons generated were
populations on the Indian gene pool [23–24,31–34]. subjected to sequence reactions using BigDyeTM (Applied Biosys-
Within this complex scenario, many interaction models such as tems, Foster City, USA) Terminator Cycle sequencing kit in ABI
gene-language, gene-geography and gene-ethnicity have been Prism 3730 DNA analyser following manufacturer’s protocol and
contested. Previous studies have shown inverse correlation further analysed in SeqScape software, version 2.5.
between genetic affinities and geographical distance [11,35].
Significant genetic differentiation between caste and tribal Statistical and phylogenetic analysis
populations has been reported [31,33,36], as against a model The revised Y-Chromosome phylogenetic tree [44] was referred
which suggests that there is considerable sharing of Pleistocene to for the assignment of haplogroups based on informative binary
heritage among them with a limited gene flow [34]. Similarly, markers. Haplogroup frequencies were estimated by a simple gene
congruence between language and genes has been proposed by count method. In order to determine the genetic structure of study
various scholars [9,37–38] along with a competing view support- populations, a model propounded by Harpending and Ward [45]
ing that genetic affinities may not necessarily be dependent on was applied. The model examines the relative role of genetic drift
linguistic similarities [39–41]. Although each study has contributed and gene flow in causing population differentiation. In the model,
significantly to understanding the role of language, culture and
the expected frequency of gene diversity for each population is first
geography in relation to the genetic affiliation and demographic
graphically represented with respect to the distance from the gene
history of Indian populations; a major limitation in them has been
frequency centroid, rii, which is given by the formula:
the poor representation of Indo-European speaking tribal popu-
lations. This is a critically important limitation since the Indo-
European speaking tribes provide ample opportunity for examin- (pi {P)2
rii ~
ing the influence of linguistic assimilation on the genomic diversity ½P(1{P)"
of India.
Keeping the above in view, we present an analysis based on the Where, pi and P are the frequency of the haplogroups in a
study of 48 bi-allelic markers in nine Indo-European speaking population i and in the pooled populations respectively. In the
tribal groups of Southern Gujarat which lies in the western part of second stage of the model, Harpending and Ward propose an
India. The main objectives of the study were (a) to study the island model of population structure in which there is a linear
distribution of Y haplogroups; (b) to study the relative influence of relationship between gene diversity and the distance from the
language, geography or ethnicity on the genetic structure of centroid which is calculated by the formula:
populations using the pattern of Y haplogroup clustering and
finally (c) to relate the observed pattern of Y haplogroup clustering hi ~H(1{rii )
with the Y chromosome lineages which arrived in India largely
from the Northern corridor at different points of time. To achieve Where, hi and H correspond to the gene diversity value in the
the study objectives the results were first compared with published population i and in all the populations as a whole respectively. As
studies on the Indian populations and then with available data on per the model, outlier populations that have undergone systematic
the Eurasian populations. The Indian populations were selected migrations will show greater gene diversity than predicted by the
keeping in view the availability of data, their linguistic and socio- regression line, while outlier groups that are isolated will exhibit
culture status and geographical position. Similarly, the Eurasian lower than predicted gene diversity.
populations for which published sources were available were In order to determine whether the Y chromosome haplogroup
considered for analysis since the historical migrations from these distribution among Indian populations is structured on the basis of
regions are known. ethnicity, geography or linguistic affiliation, an analysis of
molecular variance (AMOVA) based on haplogroup frequencies
Materials and Methods was computed for the various tentative categories using ARLE-
QUIN software, version 3.1 [46]. Using the same software
Collection and processing of blood samples
Slatkin’s lineraised pairwise FST values were calculated on
This study was approved by the Departmental Ethical
haplogroup frequencies and analysed through non metric multi-
Committee of the Department of Anthropology, University of
dimensional scaling (MDS) in SPSS version 16.0 using ALSCAL
Delhi. Informed written consent was obtained from all the
programme. The MDS plot was constructed in order to
participants. A 5 ml blood sample was drawn by a trained
graphically represent the nature of clustering between the study
medical practitioner from randomly chosen 284 healthy, unrelated
males from nine tribal population groups of Valsad and Surat populations and other world populations.

PLOS ONE | [Link] 2 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

Figure 1. Sampling areas; Map of India highlighting Gujarat (top); Regions of study pointed out in the map of Gujarat (bottom).
doi:10.1371/[Link].0090414.g001

For carrying out AMOVA on different populations secondary the haplogroups. Its frequency varied from 3.45% among
source haplogroup frequency data (Table S1) was compiled from Pavagadhi Chaudhari to 40.74% among Mota Chaudhari.
different studies [23–25,47]. For the construction of the MDS plot However, H2-Apt was found to be absent among Vasava and
data (Table S2) was compiled from various studies [48–53]. Gamit populations. While lineage H* was observed in only three
individuals, one each from Konkana, Gamit and Mota Chaudhari.
Results Haplogroup R. R1a1*a sub-clade of R haplogroup was
found to be the next most frequently occurring lineage after H1a*.
Haplogroups distribution Its frequency was found ranging from 5.56% in Gamit to 62.07%
The frequency distribution of Y chromosome haplogroups in Pavagadhi Chaudhari. Its sister sub-clade R2-M124 was
among the study populations along with the phylogenetic observed 27 times with a frequency varying from 5.56% among
relationship between them is presented in Figure 2. The side Gamit to 20.83% among the Konkana tribe. Barring Valvi
branches on the tree represent Y SNP for which the ancestral state Chaudhari, R2 was absent from all other Chaudhari groups.
was observed, while the direct branch represents Y markers for Haplogroup J. J2 with its two sub-clades J2b2* and J2a
which the mutant allelic state was observed; leading to haplogroup constituted a major portion of Haplogroup J in the current study.
designation in the particular sample. Analysis of 48 bi-allelic Except Konkana and Mota Chaudhari, either of the two J2 sub-
markers of the Y chromosome showed 13 paternal lineages that clades was present in all other groups. J2b2* sub-clade was
were distributed throughout haplogroups H, R, J, C, F, L, K and observed in 21 Y chromosomes. Its frequency was found to be
Q. Haplogroup H represented the most frequently occurring 11% in Gamit, 19% in Valvi Chaudhari and 21% in Pavagadhi
haplogroup (40.14%) followed by groups R (28.17%), J (10.21%) Chaudhari. The remaining four populations of Dhodia, Dubla,
and C (8.45%) respectively. Sparse distribution was observed for Vasava and Nana Chaudhari exhibited similar frequency values
the lineages F*, L1, Q3 and K* across all the populations. varying in a narrow range between 4% and 4.48%. Sub-clade J2a
Haplogroup H. M69 mutation, which is a characteristic of was observed only 8 times in the nine groups. It was present in four
haplogroup H was found on 114 of the total 284 Y chromosomes. of the groups with a minimum frequency of 3.17% in Dhodia to a
Haplogroup H was further segregated into three lineages, H1 by maximum frequency of 7.14% in Dubla.
the presence of M52-C allele, H2 by the presence of Apt-A allele Haplogroup C. Out of the seven sub-clades of C haplogroup,
and H* by absence of the two alleles. These haplogroups were only one sub-clade C5 with its two main derivatives C5a and C5*
further subdivided into a number of sub-clades. Lineage H1a* of was observed 21 times in all the populations except Pavagadhi
H1 group was observed 71 times and represented the most Chaudhari. C5a lineage was observed to be 62 times more
frequently occurring lineage across all the populations. Its frequent then its sister branch C5*. Its frequency varied from
frequency varied from a minimum of 3.45% among Pavagadhi 3.17% in Dhodia to 12.5% in Valvi Chaudhari, while that of C5*
Chaudhari to a maximum of 62.5% in Vasava. Lineage H2 was found to vary from 3.7% in Mota Chaudhari to 9.38% in
represented the second most frequently occurring lineage among Valvi Chaudhari.
the H haplogroup and third most common haplogroup among all

PLOS ONE | [Link] 3 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

Table 1. Geographical, Social and Linguistic description of study groups and populations included in the study for Y haplogroup
comparison.

Population Linguistic affiliationa Socio-Culture affiliationb Sample size

South Asia
India
West
Dhodia IE T 63
Dubla IE T 42
Konkana IE T 24
Vasava IE T 24
Gamit IE T 18
Valvi Chaudhari IE T 32
Nana Chaudhari IE T 25
Mota Chaudhari IE T 27
Pavagadhi Chaudhari IE T 29
Madia Gond DR T 14
Katkari IE T 19
MahadeoKoli IE T 11
Pawara IE T 16
Thakur IE T 48
Desasth Brahmin IE C 16
Maratha IE C 16
Dhangar IE C 16
Chitpavan Brahmin IE C 15
Gujrat Patel IE C 9
South
Chenchu DR T 20
Yerukula DR T 18
Kuruva DR T 10
Irular DR T 10
Naikpod Gonnd DR T 18
Andhra Brahmin DR C 15
Kamma Chaudhary DR C 15
Kappu naidu DR C 18
Komati DR C 20
Raju DR C 19
Reddy DR C 12
Bhovi DR C 13
Gowda DR C 4
Iyengar DR C 17
Lingayat DR C 10
Chakkliar DR C 9
Gounder DR C 14
Kallar DR C 9
Pallar DR C 15
Vanniyar DR C 10
East
Mahali AA T 25
Bhumij AA T 15
Birhor AA T 10
Ho AA T 7
Kharia AA T 10

PLOS ONE | [Link] 4 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

Table 1. Cont.

Population Linguistic affiliationa Socio-Culture affiliationb Sample size

Munda AA T 7
Santhal AA T 7
Juang AA T 10
Saora AA T 13
Paroja DR T 13
Bhuiyan IE T 81
Bathudi IE T 36
Kora IE T 17
Bihar brahmin IE C 18
kayasth IE C 14
bhumihar IE C 20
Baniya IE C 11
Rajput IE C 12
Yadav IE C 8
Gope IE C 16
Karan IE C 18
Oriya Brahmin IE C 24
Bauri IE C 19
Mahishiya IE C 17
Namasudra IE C 13
Central
Halba IE T 21
jaunsri IE T 6
Bhoksha IE C 10
Kanyakubj Brahmin IE C 10
Khatri IE C 7
Kurmi IE C 13
UP Thakur IE C 5
UP Kurmi IE C 6
North-East
Hamar TB T 9
Kuki TB T 7
Lai TB T 10
Lusei TB T 6
Mara TB T 5
North
Himachal Pradesh RAJPUT IE C 15

Afganistan 204
Pakistan 718
Iran 150
West Asia
Iraq 139
Jordan 146
Turkey 523
Lebanon 104
Syria 111
Central Asia
Kazakhstan 30
Alltai 98

PLOS ONE | [Link] 5 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

Table 1. Cont.

Population Linguistic affiliationa Socio-Culture affiliationb Sample size

Uzbekistan 54
Kyrgyzthan 13
Uyghursta 68
Europe
Greece 442
France 23
Netherlands 27
Germany 16
Czech and Slovakia 45
Alabina 51
Macedonia 20
Poland 55
Hungary 45
Ukraine 50
Georgia 63

a
Linguistic affiliation: Indo-European (IE), Dravidian (DR), Tibeto-Burman (TB) and Austro-Asiatic (AA);
b
Socio-cultural affiliation: Tribe (T), Caste (C).
doi:10.1371/[Link].0090414.t001

Haplogroup F*, K*, L1 and Q3. Parahaplogroup F* along (p,0.001) is due to differences between the populations. In the
with the other parahaplogroup K* and two haplogroups L and Q hierarchical approach taken, at the next level populations were
accounted for 13.03% of the total haplogroups. After initial subdivided according to their affiliation into: Indo-European,
screening of M89-T allele, 11 samples failed to resolve further and Dravidian, Tibeto-Burman and Austro-Asiatic language families.
were therefore grouped under F*. Similarly, 8 individuals did not A substantially high percentage of difference between populations
exhibit any mutation except G allele for M9 and were therefore (13.21%) was observed indicating strong distinctiveness of
grouped under K*. Other two sub-clades L1-M27 and Q3-M346 populations belonging to the different linguistic families. This
were present, but in low frequencies only. was followed by the subdivision of populations by the geographical
regions they inhabit. The geographical clustering of populations
Y chromosome diversity displayed comparatively lower (6.25%) amount of differentiation
Haplogroup diversity values for each of the nine populations between the populations. Similarly, in a comparison between
along with haplogroup distribution are given in Figure 2. Indian castes and tribes a comparatively lower fraction of variance
Haplogroup diversity (h) which is equivalent to gene diversity for (5.13%) points towards relatively lesser differences between them.
haploid genomes ranged from 0.586 in Pavagadhi Chaudhari to Following this, further categories of Indo-European speaking tribal
0.899 in Valvi Chaudhari. populations of Gujarat were made for comparing with other
Indian populations, keeping in mind the effect of language,
Population Structure and Gene Flow geography and ethnicity; in that order of importance. The results
Figure 3 represents a plot of haplogroup diversity regressed showed that the lowest group variance among all categories was
against distance from gene frequency centroid (rii). The values of between Indo-European speaking groups of Gujarat and other
gene diversity (hi) and genetic distances from centroid (rii) used in Indo-European populations; it was almost 5 times lesser than the
the nine study population groups along with their standard errors variance observed between the studied populations and the
are given in Table 2. Majority of the populations exhibited higher Dravidian speaking groups of India. Interestingly, further subdi-
than predicted gene diversity combined with a low to moderate vision of Indo-European populations into tribes and castes
deviation from the theoretical line of regression and the distance revealed a lesser fraction of variability among study populations
from the gene frequency centroid. Three populations Gamit, and Indo-European caste populations (3.45%) as compared to
Vasava and Pavagadhi Chaudhari displayed lower than predicted Indo-European speaking tribes (6.57%). Moreover, as compared
gene diversity. Pavagadhi Chaudhari showed the farthest distance to Dravidian speaking caste populations, the population in Gujarat
from the gene frequency centroid. The results indicated that the showed less differentiation with Dravidian speaking tribes as
tribal groups of Gujarat are neither explicitly isolated nor reflected by low group variance values (5.3%). Subdivision of
absolutely admixed. Indo-European populations simultaneously by caste, tribe and
geographical zones (West, Central and East) indicated effective
AMOVA role of geographic distance in determining genetic distance. All the
Table 3 presents the results of the AMOVA based on different values compared were found to be highly significant.
categories of populations subdivided by language, geography and
ethnicity. Analysis of molecular variance based on haplogroup Genetic proximities
frequencies among the study groups showed that 91.6% of We compared the study populations with 24 additional world
variability is due to within population differences and 8.4% populations already published in separate studies (Figure 4). A

PLOS ONE | [Link] 6 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

Figure 2. Distribution of Y-binary halpogroups and haplogroup diversity (h) among the study populations of Gujarat. The markers
used in the study are shown on each branch.
doi:10.1371/[Link].0090414.g002

stress value of 0.18 for the MDS plot indicated a good fit between belt of Gujarat and are composed of tribal populations which are
the two dimensional graph and the original distance matrix. The linguistically classified as Indo-European. To date several studies
comparison revealed four major clusters of South Asian, Central have provided significant insights into the paternal genetic history
Asian, West Asian and European populations. All the populations of India [23–24,34,36]; however none till now had taken into
under study with the exception of Pavagadhi Chaudhari were consideration the Indo-European speaking tribal populations
found to be clustered together. The occurrence of South Asian inhabiting Gujarat in spite of their interesting geographical
populations (the current study groups, Afghanistan, Pakistan and location and cultural attributes. It is also worth mentioning that
Iran) with Central Asian populations (Kazakhstan, Altai Region, the Indo-European speaking tribes of India not only exhibit the
Uzbekistan, Kyrgyzstan, Uyghurstan) on Axis I, conversely with complexity of historical interaction between the indigenous Indian
West Asian populations (Iraq, Jordan, Turkey, Lebanon, Syria) and migratory groups but also reflect lack of one-to-one
with respect to Axis II probably indicate similarities of hap- correlation between language, mode of subsistence and social
logroups between them. system [23]. Thus, the Indo-European speaking tribes represent an
appropriate model to study the possible genetic foot prints of the
Discussion multiple waves of migrations. In addition to tracing the origin and
impact of ancient migrations, evaluation of the impact of
The North-Western corridor of India has witnessed many waves geography and social structure in shaping the genetic structure
of migrants from different parts of the world, with a majority being of the present day Indo-European speaking tribal populations of
male migrants [31]. Gujarat is located on the western most point Gujarat was considered equally important. Thus, in the subse-
of the Indian sub-continent and has acted as a significant corridor quent text we examine the paternal genetic variation of the nine
to draw outsiders -conquerors, refugees and travellers who have Indo-European speaking tribes from Gujarat using high resolution
contributed significantly to the present day gene pool of Indian Y chromosomal unique event polymorphisms (UEPs).
populations [34]. Valsad and Surat districts are part of the tribal

PLOS ONE | [Link] 7 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

Figure 3. Regression of gene diversity (hi) on distance from centroid (rii). The solid line represents the theoretical regression line.
doi:10.1371/[Link].0090414.g003

Genetic structure of study populations explanation for the observation of high gene diversity coupled
Our analysis based on Y chromosomal UEPs apportioned the with high-level of genetic differentiation could lie in an
study samples into 13 haplogroups representing 10 major Y overwhelming genetic admixture from different sources or in an
lineages (C5, H1a*, H2, R1a1*, R2, J2, L1, F*, K* and Q3) in early inflow of genes from a common source followed by rapid
India. Haplogroup diversity (0.772) was found to be comparable population expansion and subsequent fission into isolated,
with Dravidian speaking populations (0.723), but higher than endogamous populations. Results from Harpending and Ward
Indo-European speaking populations (0.684) in the country [54]. analysis do not appear to favour the former explanation. Thus, the
Given the number of views in support of Austro-Asiatic and most plausible explanation for the current diversity scenario in the
Dravidian speaking tribes belonging to the primary pre-historic Indian population seems to be the suggestion made by Majumder
population of India [7,11]; the high gene diversity values observed et al. [17], in which an early demographic expansion of modern
in the study populations, which are in turn comparable to humans within India during Palaeolithic period and later fission of
Dravidian speaking populations, are suggestive of greater antiq- the populations is conjectured.
uity, large effective size or role of gene flow in these groups. The
analysis of molecular variance revealed that the extent of genetic In-situ versus Ex-situ Origin of Y lineages
differentiation was high among study populations which could be The indigenous versus exogenous origin of Indian paternal
attributed to either the lower effective population size of these lineages has been widely contested. Among the study populations,
groups or the Y chromosome making them vulnerable to the effect six sub-haplogroups namely, C5, H1a*, H2, J2, R1a1* and R2
of genetic drift which further accelerates the process of differen- constituted the major paternal lineages that together accounted for
tiation between the populations. But, accentuated differentiation 85.92% of the Y chromosomes. While the indigenous origin of
due to genetic drift is expected to be accompanied with lower sub-clades C5, H1a*, H2 and R2 are accepted, the status of sub-
diversity. However, in the present study, elevated levels of gene clades J2 and R1a1* are contested as they are believed to have
diversity, rule out a major role of genetic drift in shaping the been introduced in India with the demic diffusion of Proto-
observed pattern of genetic differentiation. Nevertheless, the Dravidian Neolithic agriculturists from West Asia and the influx of
Indo-European pastorals from Central Asia. It is worth mention-
ing here that the high frequency and associated diversity of a
Table 2. Gene diversity (hi) and genetic distances from the haplogroup is correlated with the possible place of origin of a
centroid (rii) among the study populations of Gujarat. particular haplogroup [55]. In the present investigation hap-
logroup H, especially H1a, represented the most frequently
observed Y chromosomal lineage followed by H2 sub-haplogroup.
Population rii ± S.E hi± S.E Its higher frequency among the Indian tribes particularly among
Dhodia 0.02460.008 0.86960.002 the Dravidian speaking tribes of South India and its limited
Dubla 0.02460.006 0.86660.004 presence elsewhere on the Indian subcontinent had led some
scholars to denote it as a tribe-specific haplogroup [36]. However,
Konkana 0.05560.011 0.85560.009
several subsequent studies have confirmed the presence and equal
Vasava 0.08660.057 0.60560.022
prevalence of haplogroup H and its associated H1a and H2
Gamit 0.09260.038 0.69960.028 branches across linguistically and ethnically diverse populations
Valvi Chaudhari 0.06260.019 0.89960.004 and in different regions of India, except the North-Eastern region
Nana Chaudhari 0.06760.029 0.82360.010 [23–24,54]; thus ruling it out as a tribal-specific marker and
Mota Chaudhari 0.08960.043 0.79260.012 supporting the uniform distribution of haplogroup H among
Indian populations. An Indian homeland for haplogroup H can
Pavagadhi Chaudhari 0.15760.093 0.58660.017
also not be refuted keeping in view its higher microsatellite
doi:10.1371/[Link].0090414.t002 diversity among Indian populations [24]. Haplogroup H has also

PLOS ONE | [Link] 8 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

Table 3. AMOVA based on Y Chromosome haplogroup frequencies.

Among groups Among populations Within populations

Categories variance (%) within groups variance (%) variance (%)

IE speaking tribes of Gujarat 8.40 91.60

4 language groupsa 13.21 18.52 68.27
Geographyb 6.25 23.22 70.53
Castes and tribes of India 5.13 24.70 70.17
IE speaking tribes of Gujarat with IE populations of India 1.81 19.57 78.62
IE speaking tribes of Gujarat with DR populations of India 9.61 11.55 78.84
IE speaking tribes of Gujarat with AA Populations of India 28.16 7.94 63.90
IE speaking tribes of Gujarat with IE castes of India 3.45 12.97 83.58
IE speaking tribes of Gujarat with IE tribes of India 6.57 15.17 78.26
IE speaking tribes of Gujarat with DR tribes of India 5.30 11.47 83.23
IE speaking tribes of Gujarat with DR caste of India 8.54 12.51 78.95
IE speaking tribes of Gujarat with IE tribes from three 9.24 15.12 75.63
geographical regions C,W & E of India
IE speaking tribes of Gujarat with IE castes from three 7.59 9.78 82.63
geographical regions C,W & E of India

a
Language groups = Indo-European (IE), Dravidian (DR), Tibeto-Burman (TB) and Austro-Asiatic (AA),
b
Geography = Central (C), West (W), East (E), South(S) and North East (NE). All the values are significant, p,0.05.
doi:10.1371/[Link].0090414.t003

been reported from Central Asian, West Asian and Gypsy recent back migration [32,52]. On the other hand, the established
populations in Europe. However, its low frequency and prevalent Indian ancestry of gypsy populations is surely the reason for
diversity pattern in Central Asia and West Asia could be due to elevated levels of H haplogroup among them [56]. All the

Figure 4. MDS Plot showing genetic relationships particularly between the South Asian populations with the world populations.
N
The South Asian populations including the study populations of India are shown in as solid circles ( ), the European populations as open circles (o),
Central Asian populations as triangle (D) and West Asian population as cross (x). The abbreviation used are Afganistan (Afg), Pakistan (Pak), Iran (Ira),
Iraq (Irk), Jordan (Jor), Turkey (Tur), Lebanon (Leb), Syria (Syr), Kazakhstan (Kaz), Altai (Alt), Uzbekistan (Uzb), Kyrgyztan (Kyr), Uyghurstan (Uyg), Greece
(Gre), France (Fra), Netherlands (Net), Germany (Ger), Czech and Slovakia (CzandSlo), Alabina (Ala), Macedonia (Mac), Poland (Pol), Hungary (Hun),
Ukraine (Ukr), Georgia (Geo), Dhodia (Dh), Dubla (Du), Konkana (Kon), Vasava (Vas), Gamit (Gam), Valvi Chaudhari (VC), Nana Chaudhari (NC), Mota
Chaudhari (MC), Pavagadhi Chaudhari (PC).
doi:10.1371/[Link].0090414.g004

PLOS ONE | [Link] 9 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

observations, therefore clearly point towards in-situ origin of three Y lineages based on their possible origin. These lineages
haplogroup H among Indian populations. include Central Asian, West Asian and indigenous Indian Y
Haplogroup C is widely distributed in Eastern and Central Asia, lineages.
Oceania and Australia [44]. As expected, all the Y chromosomes
under C haplogroup belonged to the C5 sub-clade. It was Determinants of population Clustering
observed in a frequency of 8.45% which is the highest ever Lack of any major exogenous contribution of Y lineages and
reported frequency for C5 in India and whose spread is existence of high haplogroup diversity indicate the possible male
circumscribed along the coastal belt of India [24,54]. High STR assimilation from other neighbouring populations of India among
diversity in India and South-East Asia in the backdrop of the study groups. Analysis of molecular variance (Table 3)
haplogroup C has also been observed [34,57]. Interestingly none performed on the basis of haplogroup frequencies revealed some
of the C haplogroup derivatives frequent in South-East Asia have interesting patterns. The quantification of variance into different
been reported from India. Therefore, the possibility of introduc- categories of population showed the highest variance between the
tion of C5 haplogroup from South-East Asia as a result of back four linguistic groups of India followed by the variance between
migration to India appears doubtful and its indigenous origin geographical regions and finally the caste versus tribe categories.
appears to be more probable [13]. The least percentage of variance was observed between the study
Haplogroup J is predominantly found among the populations of populations and other Indo-European speaking caste populations.
West Asia, North Africa, Europe, Central Asia, Pakistan, and However, the geographical partitioning of the Indo-European
India [44] and widely linked with the spread of agriculture from speaking tribal and caste populations into three major zones: West,
the Fertile Crescent that extends from Israel to Western Iraq. In Central and East showed the important effect of geography in
the Indian subcontinent two sub-clades of J - J2a and J2b have shaping the Y diversity pattern since on this basis the zonal
been reported. Consistent with the previous studies, a higher percentage of variance was observed to elevate between the study
proportion of J2 in West India as compared to North and South populations and other Indo-European groups of India. Thus,
regions of India has been recorded in the present study [24,54]. pattern of Y chromosome clustering of Indian populations reflects
Two models pertaining to the homeland for Indian J2 sub-clades the major role of geography over language and ethnicity as far as
have been contested, West Asia and Central Asia. Cordaux et al. Y chromosomal lineages are concerned. After Indo-European
[35] had proposed the Central Asian homeland for Indian J2 sub-
speaking caste populations, the lowest variance of Gujarat
clade mainly because of higher frequency of J2 in Central Asia. It
populations was observed with Dravidian speaking tribal popula-
is worth mentioning here that J1 sub-clade which appears in
tions. These similarities suggest either shared paternal ancestries of
appreciable frequency in Central Asia is largely absent from the
linguistically dissimilar groups or the influence over the indigenous
Indian as well as most of the West Asian populations [44].
tribal groups of India of the Indo-European speakers, who arrived
Haplogroup R is represented by two sub-clades R1a1* and R2
later.
among the study populations. After haplogroup H1a, haplogroup
The present investigation indicates that tribes of Gujarat show
R1a1* represented the most frequently occurring haplogroup.
both high genetic diversity and genetic differentiation. It is
Haplogroup R is widely distributed in Central Asia, Eastern
therefore conceivable that these groups, with a fairly large
Europe, West Asia and the Indian subcontinent [58–59]. The
higher frequency of haplogroup R1a, up to 63% in Central Asia population size, have passed through a long evolutionary history
and its relatively lower occurrence in other regions has been linked experiencing an early demographic expansion and later fission of
with Central Asian origin of R1a clade [36]. However, later the populations. Further affinities of study groups with Indo-
studies [24,54] showing higher prevalence of R1a sub-clade along European speaking non tribes followed by Dravidian speaking
with high microsatellite diversity among the tribal populations of tribal groups suggest the possibility that these native tribal
India lend support to probable South Asian origin of R1a sub- population groups of Western India might have adopted the
clade as suggested by Kivisild et al. [34]. This is further Indo-European language during the process of cultural assimila-
substantiated by almost the complete absence of other derivatives tion and absorption while still retaining their genetic links with the
of haplogroup R1 among the Indian populations, which is Dravidian speaking tribal populations. Thus, it is recommended
expected in case of the inflow from Central Asia [23–24,41]. that the present day linguistic affiliation of any Indian population
In the present investigation sub-clade R2 occurred with a should be considered with caution while reconstructing the
frequency of 9.51%, which is similar to its frequency reported from demographic history of the country. In conclusion, a study based
other Indian populations [24,34,36,54]. The frequency of R2 on the recently discovered bi-allelic loci and microsatellite loci in
decreases as one goes further west from India and its frequency is the populations can shed light on the possible explanation of the
almost negligible in Europe. Moreover, its frequent occurrence overlapping of haplogroup distribution between tribes of Gujarat
among Dravidian speaking groups as compared to Indo-European with other Asian populations and further deepen the understand-
or Austro-Asiatic speaking groups of India can be attributed to its ing of the population history of India.
indigenous Indian origin.
The MDS plot (Figure 4) also reflects the closeness of South Supporting Information
Asian populations with West Asian and Central Asian populations
Table S1 Y chromosome haplogroup frequencies data
possibly due to overlapping of haplogroups. In comparison to the
among study populations and other populations of India
world populations the overall mean haplogroup diversity among
considered for AMOVA.
the study populations was relatively higher than in the European
(XLS)
or East Asian populations [32,48] whereas it was found to be lower
than that of Central Asian and West Asian populations [49,51]. As Table S2 Y chromosome haplogroup frequencies data
mentioned earlier the high frequency and diversity of a among study populations and other populations of
haplogroup is indicator of the possible place of origin of a World considered for MDS analyses.
particular haplogroup [55]. Consequently, the observed hap- (XLS)
logroups in the present investigation could be apportioned into

PLOS ONE | [Link] 10 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

Acknowledgments the Department of Anthropology, University of Delhi, Delhi for granting us

the ethical clearance to carry out our work.
We would like to thank all the individuals who volunteered for this study on
genetic variation and provided their blood samples. We would like to
extend our gratitude towards the Valsad Raktdan Kendra (Centre for Blood
Author Contributions
Donation, Valsad), Valsad, Gujarat for helping us collect the blood Conceived and designed the experiments: GKK KNS YMI PK. Performed
samples. We would also like to thank the Anthropological Survey of India the experiments: PK SM AA. Analyzed the data: PK AA AC GKK.
for their collaboration and for giving the permission to undertake our Contributed reagents/materials/analysis tools: AC GKK YMI KNS.
laboratory work in their Southern Regional Centre. We also wish to thank Wrote the paper: PK AC AA GKK.

References
1. Census of India (2011) Registrar General and Census commissioner, 29. Pattanayak DP (1998) In: The Indian Human Heritage. Balasubramanian D,
Government of India. Rao NA, editors. Universities Press, Hyderabad. pp. 95–99.
2. Singh KS (1993) People of India: The profile of national project (1985–92). Curr 30. Cavalli-Sforza LL, Menozzi P, Piazza A (1994) The history and geography of
Sci 64: 5–10. human genes. Princeton, NJ: Princeton University Press.
3. Gadgil M, Malhotra KC (1983) Adaptive significance of the Indian caste system: 31. Bamshad M, Kivisild T, Watkins WS, Dixon ME, Ricker CE, et al. (2001)
an ecological perspective. Ann Hum Biol 10: 465–478. Genetic evidence on the origins of Indian caste populations. Genome Res 11:
4. Joshi NV, Gadgil M, Patil S (1993) Exploring cultural diversity of the people of 994–1004.
India. Curr Sci 64: 10–17. 32. Wells RS, Yuldasheva N, Ruzibakiev R, Underhill PA, Evseeva I, et al. (2001)
5. Census of India (2001) Registrar General and Census commissioner, The Eurasian heartland: a continental perspective on Y-chromosome diversity.
Government of India. Proc Natl Acad Sci USA 98: 10244–10249.
6. Fuchs S (1964) Anthropometry of the Gujarat tribes. In: Shah PG, editor. Tribal 33. Basu A, Mukherjee N, Roy S, Sengupta S, Banerjee S, et al. (2003) Ethnic India:
life in Gujarat. Bombay: Gujarat Research Society. pp. 263–275. a genomic view, with special reference to peopling and structure. Genome Res
7. Thapar R (1966) A history of India. Volume 1. Middlesex, UK: Penguin. 13: 2277–2290.
8. Ray N (1973) Nationalism in India. Aligarh, India: Aligarh Muslim University. 34. Kivisild T, Rootsi S, Metspalu M, Mastana S, Kaldma K, et al. (2003) The
9. Majumder PP (1998) People of India: biological diversity and affinities. Evol genetic heritage of earliest settlers persist in both the Indian tribal and caste
Anthrop 6: 100–110. populations. Am J Hum Genet 72: 313–332.
10. Meenakshi K (1995) Linguistics and the study of early Indian history. In: Thapar 35. Cordaux R, Saha N, Bentley GR, Aunger R, Sirajuddin SM, et al. (2003)
R, editor. Recent perspectives of early Indian history. Mumbai, India: Popular Mitochondrial DNA analysis reveals diverse histories of tribal populations from
Prakashan. pp 53–79. India. Eur J Hum Genet 11: 253–264.
11. Malhotra KC, Vasulu TS (1993) Structure of human populationsin India. In: 36. Cordaux R, Aunger R, Bentley GR, Nasidze I, Sirajuddin SM, et al. (2004)
Majumdar PP, editor. Human population genetics: a centennial tribute to J.B.S. Independent origins of Indian caste and tribal paternal lineages. Curr Biol 14:
Haldane. New York: Plenum Press. pp. 207–233. 231–235.
12. Gadgil M, Joshi NV, Prasad UVS, Manoharan S, Patil S (1997) Peopling of 37. Cavalli-Sforza LL, Minch E, Mountain JL (1992) Coevolution of genes and
India. In: Balasubramanian D, Rao NN, editors. The Indian human heritage. languages revisited. Proc Natl Acad Sci USA 89: 5620–5624.
Hyderabad: University Press. pp. 100–129. 38. Roychoudhury S, Roy S, Basu A, Banerjee R, Vishwanathan H, et al. (2001)
Genomic structure and population histories of linguistically distinct tribal groups
13. Sonakia A (1985) Skull cap of an early man from the Narmada valley alluvium
of India. Hum Genet 109: 339–350.
(Pleistocene) of central India. Am Anthropol 87: 612–615.
39. Chaubey G, Metspalu M, Karmin M, Thangaraj K, Rootsi S, et al. (2008)
14. Kennedy KAR, Deraniyagala SU (1989) Fossil remains of 28,000-year-old
Language shift by indigenous population: A model genetic study in South Asia.
hominids from Sri Lanka. Curr Anthropol 30: 394–399.
Int J Hum Genet 8: 41–50.
15. Misra VN (1992) Stone age in India: an ecological perspective. Man Environ 14:
40. Kshatriya GK, Aggarwal A, Khurana P, Italia YM (2011) Genomic congruence
17–64
of Indo-European speaking tribes of Western India with Dravidian-speaking
16. Cameron D, Patnaik R, Sahni A (2004) The phylogenetic significance of the populations of southern India: A study of 20 autosomal DNA markers. Ann
Middle Pleistocene Narmada cranium from Central India. Int J of Osteo 14: Hum Biol 38: 583–591.
419–447.
41. Sharma G, Tamang R, Chaudhary R, Singh VK, Shah AM, et al. (2012)
17. Majumder PP, Roy B, Banerjee S, Chakraborty M, Dey B, et al. (1999) Human- Genetic affinities of the Central Indian tribal populations. PLoS One 7: e32546.
specific insertion/deletion polymorphisms in Indian populations and their 42. Kshatriya GK, Aggarwal A, Khurana P, Singh HS, Italia YM, et al. (2010)
possible evolutionary implications. Eur J Hum Genet 7: 435–446. Genomic and linguistic affinities: a study of allelic and haplotype diversity at
18. Indian Genome Variation Consortium (2008) Genetic landscape of the people of DRD2 locus among the tribes of Gujarat, Western India. Genet Test Mol
India: a canvas for disease gene exploration. J Genet 87: 3–20. Biomarkers 14: 215–223.
19. Reich D, Thangaraj K, Patterson N, Price AL, Singh L (2009) Reconstructing 43. Miller SA, Dykes DD, Polesky HF (1988) A simple salting out procedure for
Indian population history. Nature 461: 489–494. extracting DNA from human nucleated cells. Nucleic Acids Res 16: 1215.
20. Chandrasekar A, Kumar S, Sreenath J, Sarkar BN, Urade SP, et al. (2009) 44. Karafet TM, Mendez FL, Meilerman MB, Underhill PA, Zegura SL, et al.
Updating phylogeny of Mitochondrial DNA macrohaplogroup M in India: (2008) New binary polymorphisms reshape and increase resolution of the human
dispersal of modern human in South Asian corridor. PLoS One 4: e7447. Y chromosomal haplogroup tree. Genome Res 18: 830–838.
21. Thangaraj K, Singh L, Reddy AG, Rao VR, Sehgal SC, et al. (2003) Genetic 45. Harpending HC, Ward R (1982) Chemical systematics and human evolution.
Affinities of the Andaman Islanders, a Vanishing Human Population. Curr Biol, In: Nitecki M, editor. Biochemical aspects of evolutionary biology. Chicago:
13: 86–93. University of Chicago Press. pp. 213–256.
22. Underhill PA, Shen P, Lin AA, Jin L, Passarino G (2000) Y chromosome 46. Excoffier L, Laval G, Schneider S (2005) Arlequin: An integrated software for
sequence variation and the history of human populations. Nat Genet 26: 358– population genetics data analysis. Evol Bioinform Online 1: 47–50.
361. 47. Thangaraj K, Naidu BP, Crivellaro F, Tamang R, Upadhyay S, et al. (2010).
23. Sahoo S, Singh A, Himabindu G, Banerjee J, Sitalaximi T, et al. (2006) A The influence of natural barriers in shaping the genetic structure of Maharashtra
prehistory of Indian Y chromosomes: evaluating demic diffusion scenarios. Proc populations. PloS one 5: e15283.
Natl Acad Sci USA 103: 843–848. 48. Semino O, Passarino G, Oefner PJ, Lin AA, Arbuzova S, et al. (2000) The
24. Sengupta S, Zhivotovsky LA, King R, Mehdi SQ, Edmonds CA, et al. (2006) genetic legacy of Paleolithic Homo sapiens sapiens in extant Europeans: a Y
Polarity and temporality of high-resolution Y-chromosome distributions in India chromosome perspective. Science 290: 1155–9.
identify both indigenous and exogenous expansions and reveal minor genetic 49. Karafet TM, Osipova LP, Gubina MA, Posukh OL, et al. (2002) High levels of
influence of central Asian pastoralists. Am J Hum Genet 78: 202–221. Y-chromosome differentiation among native Siberian populations and the
25. Kumar V, Reddy AN, Babu JP, Rao TN, Langstieh BT, et al. (2007) Y- genetic signature of a boreal hunter-gatherer way of life. Hum Biol 74: 761–89.
chromosome evidence suggests a common paternal heritage of Austro-Asiatic 50. Qamar R, Ayub Q, Mohyuddin A, Helgason A, Mazhar K, et al. (2002) Y-
populations. BMC Evol Biol 7: 47. chromosomal DNA variation in Pakistan. Am J Hum Genet 70: 1107–1124.
26. Ratnagar S (1995) Archaeological perspectives of early Indian societies. In: 51. Flores C, Maca-Meyer N, Larruga JM, Cabrera VM, Karadsheh N, et al. (2005)
Thapar R editor. Recent perspectives of early Indian history Bombay, India: Isolates in a corridor of migrations: a high-resolution analysis of Y-chromosome
Popular Prakashan. pp. 1–52. variation in Jordan. J Hum Genet 50: 435–41.
27. Chaubey G, Metspalu M, Choi Y, Magi R, Romero IG, et al. (2011) Population 52. Regueiro M, Cadenas AM, Gayden T, Underhill PA, Herrera RJ (2006) Iran:
genetic structure in Indian Austroasiatic speakers the role of landscape barriers tricontinental nexus for Y-chromosome driven migration. Hum Hered 61:132–
and sex-specific admixture. Mol Biol Evol 28: 1013–1024 43.
28. Gadgil M., Joshi NV, Manoharan S, Patil S, Prasad UVS (1998) In: 53. Haber M, Platt DE, Ashrafian Bonab M, Youhanna SC, Soria-Hernaz DF, et al.
Balasubramanian, D, Rao NA, editors. The Indian Human Heritage. (2012) Afghanistan’s ethnic groups share a Y-chromosomal heritage structured
Hyderabad: Hyderabad University press. pp. 100–129. by historical events. PloS One 7: e34288.

PLOS ONE | [Link] 11 March 2014 | Volume 9 | Issue 3 | e90414

 Y Haplogroups in Tribes of Western India

54. Trivedi R, Sahoo S, Singh A, Bindu GH, Banerjee J (2008) Genetic imprints of 57. Zhong H, Shi H, Qi XB, Xiao CJ, Jin L, et al. (2010) Global distribution of Y-
pleistocene prigin of Indian Ppopulations: A comprehensive phylogeographic chromosome haplogroup C reveals the prehistoric migration routes of African
sketch of Indian populations. Int J Hum Genet 8: 97–118. exodus and early settlement in East Asia. Hum Genet 55: 428–35.
55. Barbujani G (2000) Geographic patterns: how to identify them and why. Hum 58. Karafet TM, Zegura SL, Posukh O, Osipova L, Bergen A, et al. (1999) Ancestral
Biol 72:133–153. Asian source(s) of new world Y-chromosome founder haplotypes. Am J Hum
56. Rai N, Chaubey G, Tamang R, Pathak AK, Singh VK, et al. (2012) The Genet 64:817–831.
phylogeography of Y-chromosome haplogroup h1a1a-82 reveals the likely 59. Underhill P, Passarino G, Lin AA, Shen P, Mirazón Lahr M, et al. (2001) The
Indian origin of the European Romani populations. PLoS One 7: e48477. phylogeography of the Y chromosome binary haplotypes and the origins of
modern human populations. Ann Hum Genet 65: 43–62.

PLOS ONE | [Link] 12 March 2014 | Volume 9 | Issue 3 | e90414