Ancient corridors via Tharus (Nepal) for migrations from East Asia

Out of India through Tharus to Southeast Asia and … to Japan

Fornarino, Simon et al., Mitochondrial and Y-chromosome diversity of the Tharus (Nepal): a reservoir of genetic variation BMC Evolutionary Biology 2009, 9:154 doi:10.1186/1471-2148-9-154:

Background: Central Asia and the Indian subcontinent represent an area considered as a source and a reservoir for human genetic diversity, with many markers taking root here, most of which are the ancestral state of eastern and western haplogroups, while others are local. Between these two regions, Terai (Nepal) is a pivotal passageway allowing, in different times, multiple population interactions, although because of its highly malarial environment, it was scarcely inhabited until a few decades ago, when malaria was eradicated. One of the oldest and the largest indigenous people of Terai is represented by the malaria resistant Tharus, whose gene pool could still retain traces of ancient complex interactions. Until now, however, investigations on their genetic structure have been scarce mainly identifying East Asian signatures.

Results
High-resolution analyses of mitochondrial-DNA (including 34 complete sequences) and Y-chromosome (67 SNPs and 12 STRs) variations carried out in 173 Tharus (two groups from Central and one from Eastern Terai), and 104 Indians (Hindus from Terai and New Delhi and tribals from Andhra Pradesh) allowed the identification of three principal components: East Asian, West Eurasian and Indian, the last including both local and inter-regional sub-components, at least for the Y chromosome.

Conclusion
Although remarkable quantitative and qualitative differences appear among the various population groups and also between sexes within the same group, many mitochondrial-DNA and Y-chromosome lineages are shared or derived from ancient Indian haplogroups, thus revealing a deep shared ancestry between Tharus and Indians. Interestingly, the local Y-chromosome Indian component observed in the Andhra-Pradesh tribals is present in all Tharu groups, whereas the inter-regional component strongly prevails in the two Hindu samples and other Nepalese populations.

The complete sequencing of mtDNAs from unresolved haplogroups also provided informative markers that greatly improved the mtDNA phylogeny and allowed the identification of ancient relationships between Tharus and Malaysia, the Andaman Islands and Japan as well as between India and North and East Africa…

“Of particular interest is the link emerging between Tharus and tribals from Andhra Pradesh, as well illustrated by the Y-chromosome PCA plots (Figure 8) and by the high prevalence in these two populations of the local Y-chromosome haplogroup component (Figure 9), in comparison to the Hindus and to the other populations of Nepal [37] where the inter-regional component is clearly predominant. This further supports a deep common ancestry between Tharus and Indians, probably due to the legacy of the first settlers who arrived from the Indian coasts during the out-of-Africa dispersal.

The links between the Central Tharus and the Andaman Islanders through Northeast India (Hg M31), between the Eastern Tharus and Japan (Hg R30) and between Central Tharus and Malaysia (Hg M21), are ancient.”

“The East Asian component made up by haplogroups C(xC5), D, N, O3, Q, and K*, and mainly represented by Hg O3, is, on the whole, much more frequent among Tharus (39.8%) than among Indians (7.7%). The high Tharu frequency, mostly accounted for by the subgroup O3-M117 (83.8%), shows a wide range in the three groups with significant differences between Th-CI vs both Th-CII (P < 0.02) and Th-E (P = 0.001). Among the less represented East Asian markers of interest is Hg D that is very frequent in Tibet, absent in other Nepalese populations [37] but present in six Central Tharus: as D1-M15 in two Th-CI subjects and as D*-M174 in four Th-CII subjects. The latter, by showing the DYS392 -7 repeat allele that characterizes the D3-P47 chromosomes [37], could belong to the recently identified Hg D3* [73]. In addition, two other haplogroups were encountered: K-M9* in a single Eastern Tharus and Q1-P36 in two Tharus-CII. Hg Q, which is present in Tibetans, was seen in only one sample from Kathmandu [37]. In Indians, the very scarce East Asian component was represented by three Hg O3 (each belonging to a different sub-haplogroup and to a different Indian sample), one C3-M217 in Terai (previously observed only in a few Kathmandu and Tibetan samples [37]), two N1-LLY22g*, one in Terai and one in New Delhi and by three Q1-P36 in New Delhi. Only three East Asian haplogroups, Q1-P36, O3-M134* and O3-M117, are shared between Tharus and Indians.

The Indian subcontinent component includes lineages of haplogroups C, F, H, L, O, R and among Indians it ranges from 80% in the New Delhi sample to 85% in Terai, and to 90% in the Andhra Pradesh. Among Tharus, with the exception of an incidence of ~32% in the Th-CI group, it reaches values around 50% in the other two groups….”

“Of particular interest is the detection of haplogroups M21 and M31 (two subjects each) among the central Tharus. The Tharu M21 sequence (Figure 5) shares nine mutations with one of the three M21 lineages found in all Orang Asli groups of Malaysia [24] and in other groups from Southeast Asia [44], belonging to the sub-group M21b. The Tharu M31 sequence, together with one Megalaya mtDNA [31], clusters with one West Bengal Rajbhansi [21,27] and defines a sub-group of M31b. This subclade, together with M31a2 of the tribal Lodha, Lambadi and Chenchu populations, represents the Indian counterparts of the M31a1 Andaman lineages [27], further supporting a common ancestry of the Indian subcontinent and people of the Bengal Bay islands.

As for the R haplogroups, R7 and R30 are of particular interest. Very informative for the structure and for the age evaluation of haplogroup R7 is the Andhra Pradesh sequence #56 (Figure 5) that defines an extremely deep branch of the R7 in India. This branch shares with the root of the phylogeny of Chaubey et al.[54] only the mutations 13105, 16319 and, in addition, it does not display the 16260 and 16261 mutations characterizing the R7a and R7b branches observed in different R samples from Indian groups [11,52,54-57] and, interestingly, in one R7 Tutsi from Rwanda (unpublished data). Two Tharu mtDNAs, one from Chitwan and one from Eastern Terai, belong to the R30 haplogroup. The first is closely related to two Indian sequences, one from Andhra Pradesh and the other from Uttar Pradesh, and contributes to define a sub-clade of the R30a [54]. The second joins a Punjab sequence [54] with a Japanese deep lineage [22] indicating an ancient link between India and Japan. A more recent connection with Japan is, in turn, revealed by the F1d haplogroup showing a tight linkage between an Eastern Tharu sequence and two Japanese mtDNAs.

Ancient Hg O2-M95

“The T deletion further characterizes the HgO2-M95 clade that is considered a genetic footprint of the earliest Palaeolithic Austro-Asiatic settlers in the Indian subcontinent [14,71,74], and also as an autochthonous Indian Austro-Asiatic population marker…[Tdel, was first noticed in haplogroup O2-P31 while typing the P31 marker and was confirmed by sequencing. This is due to a T deletion in the 6T stretch starting at np 127, adjacent to the P31 T to C transition [63]. The T deletion, not found in the other examined Hg O derivatives, is always present in our O2 samples (all tribals; four of the Eastern Tharus and one from Andhra Pradesh). Taking into account that this haplogroup is often recognized through markers different from P31 and that in other studies, where the P31 was examined [64,65], a technique not detecting Tdel was employed, additional DHPLC/sequencing analyses of P31 chromosomes are necessary to evaluate the extent of the contemporary presence of the two mutations. It is worth noting that these samples were also all positive for the PK4 marker recently observed in four Pakistani Pathans...] The remaining endogenous haplogroups include haplogroup C5-M356, shared between Indians and Tharus (two in the Terai Hindus and one in the Tharus-CII), haplogroup F-M89* and its new derivative F5-M481, both considered as tribal markers and observed in Andhra Pradesh (10.3%).”

The Nepalese populations in Tharus “examined by Gayden et al. [37], apart from the homogeneous Tamang sample that displays almost exclusively the East Asian haplogroup O3-M134, the Newar and Kathmandu groups, like Tharus, show an important Indian component. However, whereas in the first two, the inter-regional haplogroups are most represented, in the Tharus the local ones are prevalent (Figure 9). Both quantitative and qualitative differences emerge from the East Asian component: on the whole it is most frequent and heterogeneous among Tharus, especially in the Chitwan groups which, in addition to the frequent Hg-O3-M117, show the Hgs D and Q, reflecting a Tibetan influence.

The analyses carried out on the mtDNA and Y chromosome of the Tharus, one of the oldest and the largest indigenous people of Terai, have shown a complex genetic structure within which are identifiable: i) a deep common ancestry between Tharus and Indians, not previously reported, more evident for mtDNA but also revealed by the prevalence of the local Indian Y-chromosome subcomponent, as in the tribals of Andhra Pradesh; ii) a significant East Asian genetic contribution both in the male and female gene pool; iii) a western heritage, clearly evident for the Y-chromosome; iv) a remarkable heterogeneity of the Tharu population (with the Eastern Tharus more dissimilar to the others) ascribable both to various exogenous influences and to subgroup specific lineages stemming from a shared genetic background with Indians.

Particularly informative has been the complete mtDNA sequencing that further supports a deep differentiation of mtDNA haplogroups in the Indian subcontinent, indicating that some branches are geographically or socially specific, while others are widespread. The improvement in the mtDNA phylogeny has also allowed the identification of ancient relationships between Tharus, not only with the Indian subcontinent area, including Pakistan, but also with the Andaman Islands, Malaysia, and Japan, as well as between India and North and East Africa

Gayden, Tenzin et al., The Himalayas: barrier and conduit for gene flow. Am J Phys Anthropol. 2013 Jun;151(2):169-82. doi: 10.1002/ajpa.22240. Epub 2013 Apr 12.

“Although previous Y-chromosome studies indicate that the Himalayas served as a natural barrier for gene flow from the south to the Tibetan plateau, this region is believed to have played an important role as a corridor for human migrations between East and West Eurasia along the ancient Silk Road.” The analysis of  mitochondrial DNA variation in 344 samples from three Nepalese collections (Newar, Kathmandu and Tamang) and a general population of Tibet “revealed a predominantly East Asian-specific component in Tibet and Tamang, whereas Newar and Kathmandu are both characterized by a combination of East and South Central Asian lineages. Newar and Kathmandu harbor several deep-rooted Indian lineages, including M2, R5, and U2, whose coalescent times from this study (U2, >40 kya) and previous reports (M2 and R5, >50 kya) suggest that Nepal was inhabited during the initial peopling of South Central Asia.”

The study confirmed “that while the Himalayas acted as a geographic barrier for human movement from the Indian subcontinent to the Tibetan highland, it also served as a conduit for gene flow between Central and East Asia.”

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