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.
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.
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  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  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 , could belong to the recently identified Hg D3* . 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 . 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 ), 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  and in other groups from Southeast Asia , belonging to the sub-group M21b. The Tharu M31 sequence, together with one Megalaya mtDNA , 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 , 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. 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 . The second joins a Punjab sequence  with a Japanese deep lineage  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 . 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. , 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
components, especially those of East Eurasian ancestry, and then to better understand the role of the Himalayas in peopling Nepal, we have studied the matenal genetic composition extensively, especially the East Eurasian lineages, in Nepalese and its surrounding populations. Our results revealed the closer affinity between the Nepalese and the Tibetans, specifically, the Nepalese lineages of the East Eurasian ancestry generally are phylogenetically closer with the ones from Tibet, albeit a few mitochondrial DNA haplotypes, likely resulted from recent gene flow, were shared between the Nepalese and northeast Indians.
It seems that Tibet was most likely to be the homeland for most of the East Eurasian in the Nepalese. Taking into account the previous observation on Y chromosome, now it is convincing that bearer of the East Eurasian genetic components had entered Nepal across the Himalayas around 6 kilo years ago (kya), a scenario in good agreement with the previous results from linguistics and archaeology.
The Qiangic languages in western Sichuan (WSC) are believed to be the oldest branch of the Sino-Tibetan linguistic family, and therefore, all Sino-Tibetan populations might have originated in WSC. However, very few genetic investigations have been done on Qiangic populations and no genetic evidences for the origin of Sino-Tibetan populations have been provided. By using the informative Y chromosome and mitochondrial DNA (mtDNA) markers, we analyzed the genetic structure of Qiangic populations. Our results revealed a predominantly Northern Asian-specific component in Qiangic populations, especially in maternal lineages. The Qiangic populations are an admixture of the northward migrations of East Asian initial settlers with Y chromosome haplogroup D (D1-M15 and the later originated D3a-P47) in the late Paleolithic age, and the southward Di-Qiang people with dominant haplogroup O3a2c1*-M134 and O3a2c1a-M117 in the Neolithic Age
According to the nomenclature of Y Chromosome Consortium (YCC) , , 23 SNP haplogroups were determined from the 127 male individual samples (Figure 2a, Table S1, andTable S2). Haplogroup D1-M15 and its subhaplogroups, which are widely distributed across East Asia including most of the Tibeto-Burman, Tai-Kadai and Hmong-Mien speaking populations , ,  (Figure S1 in Doc S1), are also prevalent in the four studied populations (44.44% and 12.50% in Horpa-Danba and Horpa-Daofu, respectively; 8.70% in Tibetan-Xinlong and 6.38% in Tibetan-Yajiang). Haplogroup D3a-P47 is almost exclusively distributed in Tibeto-Burman populations , ,  (Figure S1 in Doc S1) and also found highly frequent in Horpa-Daofu, Tibetan-Xinlong and Tibetan-Yajiang, but absent in Horpa-Danba. Haplogroup O1a1-P203, which occurs at high frequencies in Tai-Kadai speaking people along the southeast coast of China and Taiwan aborigines , , is also observed at a high frequency in Yajiang (21.28%) and moderate frequencies in Daofu and Xinlong (6.25% and 8.70%, respectively), but absent in Danba. The major lineages in the Indo-China Peninsula, O2a1-M95 and its subhaplogroups, are also found at moderate or relatively low levels in the four studied populations. Haplogroup O3-M122 is the most common haplogroup in China and prevalent throughout East and Southeast Asia, comprising roughly 25–37% of the studied Qiangic populations. O3a1c-002611, O3a2c1-M134, and O3a2c1a-M117 are three main subclades of O3, each accounting for 12–17% of the Han Chinese , . However, their frequencies vary a lot in Qiangic populations. O3a1c-002611 comprises 15.22% of Xinlong Tibetans, but absent in three other populations. O3a2c1*-M134 accounts for about 6% of the Horpa-Danba and Tibetans of Xinlong and Yajiang, but absent in Horpa-Daofu. Haplogroup O3a2c1a-M117, which exhibits high frequencies in other Tibeto-Burman populations, is also observed at high frequencies in Horpa-Danba and Tibetan-Yajiang (22.22% and 19.15%, respectively), and moderate frequencies in Horpa-Daofu and Tibetan-Xinlong (12.50% and 10.87%, respectively). Haplogroup C-M130 has a very wide distribution and might represent one of the earliest settlements in East Asia. Haplogroup C* (M130+, M105−, M38−, M217−, M347−, and M356−) has been found at low frequencies along the southern coast of mainland East Asia as well as throughout the islands of Southeast Asia , . In spite of the wide distribution of C*, they all have similar STR haplotypes (DYS19, 15; DYS389I, 12; DYS389b, 16; DYS390, 21; DYS391, 10; DYS392, 11; DYS393). There are two C* individuals detected in this study, one in Horpa-Danba and the other in Tibetan-Xinlong. Those two individuals also have the same STR haplotype as mentioned above. Haplogroup C3-M217 is the most widespread subclade of C-M130, and reaches the highest frequencies among the populations of Northern East Aisa, especially in Mongolians –. Haplogroup C3-M217 has also been found in Tibetan-Yajiang at a frequency of 10.64%, but totally absent in other three populations. Haplogroup N-M231 has both a unique and widespread distribution throughout northern Eurasia and reaches highest frequency among most of the Uralic populations as well as some Altaic populations. Haplogroup N1c1a-M178 is the most common subclade of N-M231 and thought to be originated in China , . N1c1a-M178 has also been detected in Horpa-Daofu and Tibetan-Xinlong at 12.50% and 2.17%, respectively. The 17-STR haplotype of N1c1a individuals in Horpa-Daofu is exactly the same with some Komi people in Russia , . However, the haplotype of N1c1a individual in Xinlong shows more similarity with samples of its surrounding populations (unpublished data). It is particularly noteworthy that Central-South Asia related haplogroups J-M304 and R2-M124  have also been detected at low frequencies in Qiangic populations.
PCA and STR genetic distance analysis
The paternal genetic relationships among Qiangic, Tibeto-Burman, and other East Asian populations were discerned with the aid of additional published Y chromosome datasets. We used a PCA based on the distribution of Y chromosome haplogroup frequencies of 51 populations to show the overall clustering pattern (Figure 3a, Table S3). Results of PCA are presented by the plots of the first two principal components (PCs), which together account for 31.31% of the Y chromosome variation in these populations. The first PC revealed a clear north-south geographic division between Altaic and Sino-Tibetan, Tai-Kadai & Hmong-Mien. Haplogroup C3-M217, G-M201, J-P209, and R-M207 were found to contribute most to the northern pole of Altaic. Haplogroup O-M175 contributed most to the southern pole. Sino-Tibetan, Tai-Kadai and Hmong-Mien populations showed different distributions of the second PC. Horpa-Danba, Horpa-Daofu, Tibetan-Xinlong, and Tibetan-Yajiang were clustered within Sino-Tibetan group, which reflected a clear linguistic clustering pattern. Haplogroups O3a1c-002611, O3a2c1*-M134, and O3a2c1a-M117 contributed most to the Sino-Tibetan pole. Contrastingly, haplogroups O3a2b*-M7 and O2a1-M95 were concentrated at the Tai-Kadai and Hmong-Mien pole. The four western Sichuan populations clustered tightly together with other Tibeto-Burman populations, such as Qiang, Tibetan-Yunnan, Yi, and Tujia, mostly due to high frequencies of haplogroup D3a-P47, O3a2c1a-M117, D1-M15, and O3a2c1*-M134. In the STR genetic distance based neighbor-joining tree, Horpa-Daofu, Tibetan-Yajiang, and Tibetan-Xinlong also clustered tightly with Tibeto-Burman populations. However, Horpa-Danba was close related to Han and Hmong-Mien populations (Figure S2 in Doc S1). As PCA was performed from frequencies of haplogroups and genetic distance was obtained from only 6 STR markers (Table S4), the results are suggestive but not conclusive.
Network analysis and time estimation
To discern the detail relationship between the D3a-P47, O3a2c1a-M117, D1-M15, and O3a2c1*-M134 haplogroups in Tibeto-Burman and other related populations, a median-joining network was constructed based on Y-STR haplotypes of those haplogroups (Figure 4). A clear Sino-Tibetan vs. Tai-Kadai and Hmong-Mien divergence can be inferred from the network of D1-M15 though sporadic haplotype sharing exists. Furthermore, within the Sino-Tibetan populations, haplogroup D1-M15 contains distinct STR haplotypes between Qiangic populations, Northern Han, and Tibetan-Tibet, implying that D1-M15 experienced a serial of founder effects or strong bottlenecks and a secondary expansion in Sino-Tibetan populations. In the network of D3a-P47, the divergence between Qiang and Tibetan with other Tibeto-Burman populations has been observed. Other Tibeto-Burman populations only have a subset of the Qiang and Tibetan haplotypes. The star-like network of D3a-P47 also suggests population expansion in Tibetans. The network of O3a2c1*-M134 shows a clear divergence between Tibetan and northern populations (Northern Han and Altaic). Southern Han and Tai-Kadai samples constitute the center of the network and act as a bridge connected Tibetan and northern populations, which supports the southern origin and northern expansion of O3a2c1*-M134. Most of the Qiangic samples belonging to haplogroup O3a2c1*-M134 share haplotypes with northern populations, indicating a recent gene flow from northern populations to Qiangic populations. A population expansion has also been observed in the star-like network of haplogroup O3a2c1a-M117. o However, the haplotypes of O3a2c1a-M117 are extensively shared among all the East Asia populations.
To get more insights into the origin of the East Eurasian maternal components observed in the Nepalese and therefore test the two competing scenarios about how these components had been introduced into Nepal, we focused on the phylogenetic affinity between from the Tibetan, northeast and northwest Indian populations. Fig 3 illustrates the principle component analysis plot of the 43 populations under study, which was constructed based merely on the East Eurasia lineages. Among the five Nepalese populations under study, three clustered with the Tibetans (Fig 3). After we considered all the Nepalese regional populations as a while and calculated its Fst value with the populations from its neighboring regions, the smallest genetic distance was observed between the Nepalese and the Tibetans from the nodes occupied almost exclusively by the Tibetan lineages and only a few haplotypes are shared sporadically between the Nepalese and the northern Indians. Taken together, the Nepalese lineages of East Eurasian ancestry generally show closer affinity with the ones from Tibet, albeit a few mtDNA haplotypes, likely resulted from recent gene flow, were shared between the Nepalese and northern (including northeast) Indians (Figures 4 and 5).
Even though we focused on the East Eurasian lineages identified in the Nepalese poplations we did observe a number of Nepalese-specific haplotypes, strongy suggesting their rather ancient origin and most plausibly de novo differentiation in Nepal. To get some hint at the arrival time of the lineages, we have focused on two clades from contain from haplogroups G2a and M9a1a2 simply because both clades contain the Nepalese haplotypes at their terminal branch or basal node and likely have differentiated in Nepal; estimating their ages would then help to date the arrival time of the migration from Tibet. In fact, time estimation results revealed that haplogroups G2a2 and M9a1a2a have very similar ages of B5.7 kya, and this age becomes a little older (B6 kya) when calibration rate proposed by Forster et al. 44 was used. To this end, the very similar ages of both haplogroups, which likely had in situ differentiated in Nepal, strongly suggest that the bearers of these East Eurasian maternal components would have arrived at Nepal no later than 5.7 kya (Table 4). In retrospect, previous work has suggested that the maternal genetic components from the northern East Eurasian was introduced into Tibet around 8.2 kya,1 and our time estimation results fit this dating frame very well. It is then conceivable that the settlement of Nepal by the bearer of the East Eurasian genetic components occurred likely before 5.7 kya, a result in good agreement with the archeological findings reporting shared the Neolithic features between Nepal and Tibet (references therein).49
Previous studies have observed substantial East Eurasian genetic components in the Nepalese populations;4,5 however, it remains controversial whether the East Eurasian lineages have been introduced into Nepal from Tibet directly (across the Himalayas)4,6 or via northeast India.5,8,50 By extensively analyzing the mtDNA variation in Nepal, Tibet, northern India populations, our observations, based on the principle component analysis, Fst and admixture estimation, revealed the closer genetic affinity between the Nepalese and the Tibetans, and this result was further substantiated by the median networks, (Figures 4 and 5) in which most of the Nepalese mtDNAs
prevalent among northern Asian populations shared the haplotypes
with the Tibetans at root level or branched off directly from the nodes
consisting almost exclusively of the Tibetan lineages. Our results
strongly suggest that most of the East Eurasian maternal components identified in the Nepalese were introduced directly from Tibet,4,6 and the time estimation results further date that this peopling scenario plausibly occurred about 6 kya. Indeed, this inference seems to be in striking accordance with the historically recorded passes (such as the Kodari and Rasuwa Passes), which bridged the Nepalese and the Tibetans since the ancient time.3 However, the observed gene flow from northeast India suggests genetic contribution, albeit limited, from this region, a scenario echoing the proposed inland dispersal route.50 In this spirit, our findings complete the understanding of the origin of the Nepalese and the way how the East Eurasian genetic components had been introduced into Nepal. Taking into account the previous observation on Y chromosome, now it is convincing that bearer of the East Eurasian genetic components had entered Nepal across the Himalayas around 6 kilo years ago (kya), a scenario in good agreement with the previous results from linguistics and archeology.
We then estimated the coalescence and expansion time of Y chromosome lineages in Qiangic populations (Table 1). The ages estimated using evolutionary rate are about two or three times higher than using genealogical rates. As the times using genealogical rates fit well with sequence-based estimates in Y chromosome lineage dating , we present results from the genealogical calculations in the following section. Haplogroup D can trace back to late Palaeolithic period, while other subhaplogroups coalescence more likely in Neolithic Time. The lineage expansion times all fall into Neolithic Time ranging from 4.2 to 7.5 kya.
397 samples were successfully assigned to mtDNA haplogroups using a combination of HVS-I sequence motifs and single nucleotide polymorphisms (SNPs) distributed around the coding region of the mtDNA genome. A total of 79 haplogroups or paragroups (unclassified lineages within a clade marked with an asterisk [*]) were identified (Figure 2b, Table S1 and Table S2), all within the two principal out-Africa macrohaplogroups: M and N (including R). Macrohaplogroup M and its subhaplogroups comprise 59.70% of the Qiangic maternal gene pool, and macrohaplogroup N and its subhaplogroups comprise the left 49.30%. The most prevalent haplogroups within macrohaplogroup M, haplogroup D and G represent 18.14% and 13.60% of all the samples. Within macrohaplogroup N, haplogroup A and F are the most common lineages, accounting for 13.60% and 10.58% of Qiangic, respectively. The majority of the mtDNA lineages belong to eastern Eurasian specific groups, including those from Northeast Asia (A, D4, D5, G, C, and Z) – and Southern China or Southeast Asia (B, F, M7, and R9) . Only two U samples in Yajiang might be traced for their origins to western or southern Eurasia, comprising 0.5% of Qiangic. The frequencies of Southern China or Southeast Asia specific haplogroups in Horpa-Danba, Horpa-Daofu, Tibetan-Xinlong, and Tibetan-Yajiang are 26.09%, 22.50%, 27.73%, and 21.35%, respectively. However, Tibetan-Yajiang, Horpa-Danba, Horpa-Daofu and, to a lesser extent, Tibetan-Xinlong, display a considerable Northeast Asian proportion of lineages (56.77%, 56.52%, 55.00%, and 43.70%, respectively). Consistent with other studied Tibetan populations on the Tibetan Plateau, Qiangic populations also showed a strong similarity with Northeast Asian populations.
We performed a PCA using the mtDNA haplogroup frequencies of Qiangic groups in this study and other 68 populations to see the detailed genetic patterns of those populations (Figure 3b,Table S3). The first PC revealed a clear geographic division between northern populations (Altaic and Northern Han) and southern populations (Southern Han, Tai-Kadai, and Hmong-Mien). Qiangic groups were clustered in the northern pole due to the high frequencies of haplogroup A and G. Han Chinese and Tibeto-Burman populations showed significantly different distributions in the second PC. Qiangic populations were clustered within Tibeto-Burman group due to the existence of haplogroup M9a’b and M13.
Phylogeography of Macrohaplogroup M.
Macrohaplogroup M and its subhaplogroups represent the majority of the Qiangic maternal lineages, with frequencies ranging from 65.22% in Horpa-Danba to 57.98% in Tibetan-Xinlong. Haplogroup D4 and G are the most frequent sub-clades of macrohaplogroup M in Qiangic populations, each comprising 13.60%. Haplogroup D4, which is prevalent throughout Central Asia , Northeast Asia , , and Southwest China , , , , represents the majority of haplogroup D samples in Horpa-Danba (17.39%), Tibetan-Yajiang (13.54%), Tibetan-Xinlong (13.45%), and Horpa-Daofu (10.00%). The haplotypes of D4* were extensively shared among Qiangic, Tibetan, Han Chinese, and Altaic (Figure 5). Specifically, sub-haplogroup D4j3 was detected in Horpa-Danba and Horpa-Daofu with considerable frequencies (4.35% and 5.00%, respectively). The age estimates generated for D4* and D4j3 in Qiangic were about 15 kya (Table 3). In addition, the population growth factor, Fu’s Fs values of haplogroups D4* and D4j3, were significantly negative (Table 4), implying post-LGM expansions of those two lineages in Qiangic.
Haplogroup G is found at high frequencies in northeastern Siberia but it is also common among populations of Japanese Archipelago and Korean Peninsula. This haplogroup also comprises an average of 20% of the maternal gene pool of the Tharus from Nepal  and accounts for more than 10% in the Tibetan populations of Nagqu, Chamdo, Lhasa, Garze, and Monba . In this study, haplogroup G and subhaplogroups G2a, G2b1b, G3, and G3a1 account for 20% of Horpa-Daofu and reach frequencies greater than 10% in three other Qiangic populations. Subhaplogroup G2a is represented as four distinct HVS-I motif types: 16129–16223–16278–16362 (I), frequent in Tibetan and Southern Han but nearly absent in Altaics; 16223–16227–16278–16362 (II), frequent in all the above three populations and probably experienced population expansion in Altaics (Figure 5); 16193–16223–16278–16362 (III), exclusive in South Asia. All of the G2a samples in Horpa-Daofu harbor haplotype II but add one more mutation at site 16304. However, most of Tibetan-Xinlong samples belong to haplotype I (50%). Subhaplogroup G2b1b was first reported as a novel haplogroup in northeast India and has low frequency distribution in Tibet and surrounding regions , . This haplogroup accounts for 4.69%, 2.50%, and 0.84 of Tibetan-Yajiang, Horpa-Daofu, and Tibetan-Xinlong. Compared with other Tibetan samples, 72.73% of Qiangic G2b1b samples were detected with a mutation at site 16356, thus forming some exclusive clades in the network (Figure 5). Subhaplogroup G3 comprises 6.77%, 5.00%, 3.36%, and 2.17% of Tibetan-Yajiang, Horpa-Daofu, Tibetan-Xinlong, and Horpa-Danba, respectively. Two Yajiang samples are further defined as G3a1 by a mutation at site 16215. In addition, we have found two Horpa-Danba G2a samples bearing both G2a (16278) and G3 (16274) characteristic mutations and thus we could not tell the exact haplogroup classification of those two samples. The coalescence time estimates of G*, G2b1b, and G3 were all around 20 kya and the age of G2a even reached about 34 kya (Table 3). However, it is noteworthy that the arrival time of these haplogroups at the Tibetan Plateau might be somewhat more recent than their coalescent ages would indicate, because nearly all these haplogroups (except G2b1b) had already differentiated before their arrival on the plateau (Figure 5). The exclusive clades in the network (Figure 5) and the significant negative Fu’s Fs values (Table 4) of G2a and G3 suggest the probable isolation and secondary population expansion of the two lineages.
Haplogroup M8 has two sublineages, haplogroup C and Z. Haplogroup C is a common lineage, which is widespread in East Asia and Siberia and is one of the founder lineages among Native Americans . Haplogroup C comprises 8–10% of Horpa-Danba and Tibetan-Yajiang, but was detected at a very low frequency or even absent in Tibetan-Xinlong and Horpa-Daofu. Almost 60% of the C samples in present study harbored a specific HVS-I motif 16093–16298–16327 and were assigned as C4d. One Horpa-Danba individual with HVS-I motif 16298–16327 is also classified as C4d through complete sequencing (Doc S2). Haplogroup C4d has been supposed to be Tibetan specific, frequencies ranging from 1.6% to 5.0% in populations of Tibet . However, the frequency of C4d in Tibetan-Yajiang even reaches 6.25%. In addition, all the reported C4d samples in Tibet and Qinghai have the same motif as above mentioned. However, 25% of the C4d samples in Yajiang share another mutation at site 16111. About 23% of C samples in Qiangic with a mutation at site 16357 might be assigned as C4a2′3′4, which is also restricted to Tibeto-Burman populations. Haplogroup Z is observed at relatively low frequencies in Qiangic populations.
M9a’b is widely distributed in mainland East Asia  and Japan, and reaches its greatest frequency and diversity in Tibet ,  and its surrounding regions, including Nepal  and northeast India , . It has been proposed recently that haplogroup M9’b had most likely originated in southern China and/or mainland Southeast Asia. After the LGM, M9a’b might be involved in some northward migrations in mainland East Asia . In the present study, the frequencies of M9a’b in Horpa-Danba, Horpa-Daofu, Tibetan-Xinlong, and Tibetan-Yajiang are 4.35%, 10%, 13.45%, and 6.77%, respectively. Most M9a* samples (62.5%) of Qiangic shared the main haplotype that clustered in the central largest clade with other Tibeto-Burman populations in the network. However, the estimated age of M9a* is relatively young at about 7 kya. M9b is largely restricted to the non-Tibetans in southern China and southwest China . We have detected low frequencies of M9b in Horpa-Danba and Tibetan-Xinlong (2.17% and 0.84%, respectively). In the networks of M9a1a and M9a1b, most of the Qiangic samples shared the descent types, giving a clear signal of out of Tibet migrations of those haplogroups. The age estimates generated for M9a1a and M9a1b1 in Qiangic were around 12–13 kya (Table 3), consistent with proposed post-glacial dispersal of the M9a’b lineages.
Haplogroup M13a has been found at its greatest frequency and diversity in Tibet, but it has also been detected at very low frequencies in Siberian Buryat, Yakut, Altaian Kazakh, and Ewenki, and central Asian Kirghizs  as well as Barghuts , , . The frequency of haplogroup M13a in Qiangic populations is remarkable, accounting for 3.27% of all samples. In the network of haplogroup M13a1 and M13a2, Qiangic and Tibetan-Burman samples formed some almost exclusive clades. This strongly suggests that these specific lineages have de novo origins within Tibetans. Specially, 70% of subhaplogroup M13a1b samples in Qiangic share the same haplotype. A coalescence time estimate for M13a1b corresponded to 5.7 kya (Table 3), suggesting a relatively recent Neolithic expansion out of Tibet and even more recent arrival into northern Asia of this lineage.
Qiangic populations also exhibit some basal Eurasian mtDNA lineages. Haplogroup M62, for example, was first reported in Northeast India  and since then has been reported in several populations at low frequency throughout Tibet , . Zhao et al. suggested that M62 might represent the genetic relics of the initial Late Paleolithic settlers (>21 kya) on the Tibetan Plateau. In this study, we observed haplogroup M62b in three Yajiang Tibetans. The haplotype of those three individuals is different from all other reported M62 samples with a mutation at site 16305. Likewise, haplogroup M74a was detected in one Xinlong Tibetan, and the haplotype of which bearing a distinctive mutation at site 16274 only shared with one Maonan individual, one Zhuang individual, and one Hainan Han Chinese . Haplogroup M33c was found in a Tibetan sample from Yajiang with a similar haplotype as some Hmong-Mien samples .
Phylogeography of Macrohaplogroup N.
Haplogroup R and its subhaplogroups (B and F) represent the majority of the lineages branching from the basal N trunk, accounting for 26.09%, 22.50%, 28.57%, and 23.44% of the maternal diversity in Horpa-Danba, Horpa-Daofu, Tibetan-Xinlong, and Tibetan-Yajiang, respectively. Subhaplogroup B4* is the most frequent lineage of haplogroup B in Qiangic, comprising 4.53% of all the samples. In the network of B4*, the root clade composed almost exclusively of non-Tibetan-Burman samples, however, the Tibetan-Burman samples only formed some small clusters or shared the terminal types, suggesting that B4* had already differentiated before its arrival in Tibet. Subhaplogroup F1* is the most frequent lineage of haplogroup F in Qiangic, accounting for 5.54% of all the samples, and even comprising as high as 12.5% of Horpa-Daofu. Age estimate generated for F1* in Qiangic was around 5 kya (Table 3). The exclusive Qiangic cluster of F1* in the network suggests a strong bottleneck or founder effect in its Neolithic migration towards the plateau. The significant negative values of the growth factor estimates (Table 4) suggest a secondary expansion and probable selection of F1* lineage during its adaptation in the plateau.
Haplogroup N* is almost exclusively represented by haplogroup A in our samples. Haplogroup A is widely distributed in northern and eastern Asia, occurring at frequencies of 5%–10% in different populations . Haplogroup A also has an average frequency of nearly 9% on the plateau . Subhaplogroup A4*, which is mainly found in Central, Northeast and Southwest Asia, is the most frequent sublineage of haplogroup A in Qiangic, accounting for 2.17%, 5.00%, 4.20%, and 12.50% of Horpa-Danba, Horpa-Daofu, Tibetan-Xinlong, and Tibetan-Yajiang, respectively. Network analysis of haplogroup A4* revealed a star-like pattern and thus showed a signal of population expansion on the plateau (Figure 5). The probable population expansion was also confirmed by growth summary statistics in this lineage (Table 4). Subhaplogroup A11 split from the root of haplogroup A very early and formed a distinct lineage. A11a and A11b, the two sublineages of A11, have the different distribution pattern. Most of the A11 samples in Tibet belong to A11* or A11a and only a few have a control-region substitution at site 16234, assigned as A11b. However, almost all the A11 samples in the Tibetan-Burman and Han Chinese of Yunnan belong to A11b. In the present study, three of five A11 samples belonged to A11* and the other two were assigned as A11b.
The Sino-Tibetan linguistic family comprises some 460 languages distributed in East Asia, Southeast Asia, and parts of South Asia, including the Chinese and Tibeto-Burman subfamilies. Despite intense linguistic, archaeological, and genetic researches, where the Sino-Tibetan speakers came from, how they dispersed remain major open questions. One widely accepted hypothesis states that the ancestors of the Sino-Tibetan population were originally from the Neolithic Age Di-Qiang people in the upper and middle Yellow River basin. Di people have gradually developed into Han Chinese and Qiangic populations since the collapse of Later Liang dynasty (one of the Sixteen Kingdoms dynasty, AD 386–403). Here, we integrated the Y chromosome and mtDNA evidence of Qiangic populations to provide a broader framework for reconstructing the history of Sino-Tibetan.
From the paternal Y chromosome perspective, haplogroup D1-M15 originated from D*-M174 during its migration into mainland East Asia . Around 50–60 kya, a subgroup of haplogroup D*-M174 and D1-M15 started their northward migration through WSC corridor into nowadays Qinghai province, and then probably moved along the well-known route, called the Tibeto-Burman corridor, to enter the Himalayas . Haplogroup D*-M174 probably gave birth to D3a-P47 in Tibet . Haplogroup D3a-P47 experienced recent population expansion on the Tibetan Plateau, and then probably migrated southward via the WSC corridor and gradually became the main genetic component of Tibeto-Burman populations in nowadays Sichuan, Yunnan, and Guangxi province. Y chromosome haplogroup D might give the evidences of the late Palaeolithic human activity on the plateau. The genetic relics of late Palaeolithic age have also been detected in the maternal side, for example, haplogroup M62b. In addition, a number of Paleolithic sites have been excavated crossing the Tibetan Plateau –, documenting the earliest human presence on the plateau dated to 20–30 kya.
Around 20–40 kya, a population with dominant haplogroup O3-M122 Y chromosomes (haplogroup O3a1c-002611, O3a2c1*-M134, O3a2c1a-M117, and probably other O3 lineages) finally reached the upper and middle Yellow River basin and formed the Di-Qiang populations. During the Neolithic period, the Di-Qiang people experienced relatively huge population expansion. A subgroup of the Di-Qiang people with dominant haplogroup O3a2c1*-M134 and O3a2c1a-M117, now called the Proto-Tibeto-Burman people left their Yellow River homeland, probably also moved along the Tibeto-Burman corridor, embarking on large-scale westward migrations to nowadays Qinghai province and then southward to the Himalayas, or southward migration directly via the WSC corridor to Yunnan and Guangxi, where they mixed with D-M174 linages and developed into Tibeto-Burman populations. However, haplogroup O3a2c1*-M134 might have already reached Tibet predated the above southward migration together with O3a2c1a-M117, judging from the high diversity in the network of O3a2c1*-M134 (Figure 4). In addition, another branch of the Di-Qiang people, the proto-Chinese, with dominant haplogroup O3a1c-002611 migrated eastward to the central China plain area, the middle and lower Yellow River Valley, and integrated gradually with the natives (probably populations with haplogroup C-M130 or D-M174) around 5–6 kya. Subsequently, the Di-Qiang people that resided in upper and middle Yellow River basin with haplogroup O3a2c1*-M134 and O3a2c1a-M117 formed the well-known Yan-Huang tribe (Hot Emperor and Yellow Emperor), and the eastward branch with O3a1c-002611 developed into the Dong Yi tribe. The Yan-Huang tribe together with the Dong Yi tribe gradually developed into a large population known as Han Chinese. With the expansion of Han Chinese, especially southward, this group became the largest one of the 56 officially recognized ethnic populations in China.
The role of haplogroup O3-M122 lineages played in the origin of Tibeto-Burman populations has suggested extensive genetic input from northern Asians. This suggestion has been supported by previous studies employing autosomal STR , , Y chromosome , , and mtDNA –. It is not surprising that the maternal variation of Qiangic populations was also largely contributed by northern Asian-prevalent haplogroups, including haplogroups A, C, D, and G. In addition, cultural features of the upper Yellow River basin, such as painted pottery, millet agriculture, and urn burial, are prevalent in the Neolithic sites of WSC, probably due to the demic diffusion via the genetic corridor . However, we still could not rule out the possibility that the complex genetic structure of Qiangic populations might be due to repeated admixture from surrounding populations, which provides directions for future work.
The Himalayan mountain range has played a dual role in shaping the genetic landscape of the region by (1) delineating east–west migrations including the Silk Road and (2) restricting human dispersals, especially from the Indian subcontinent into the Tibetan plateau. In this study, 15 hypervariable autosomal STR loci were employed to evaluate the genetic relationships of three populations from Nepal (Kathmandu, Newar and Tamang) and a general collection from Tibet. These Himalayan groups were compared to geographically targeted worldwide populations as well as Tibeto-Burman (TB) speaking groups from Northeast India. Our results suggest a Northeast Asian origin for the Himalayan populations with subsequent gene flow from South Asia into the Kathmandu valley and the Newar population, corroborating a previous Y-chromosome study. In contrast, Tamang and Tibet exhibit limited genetic contributions from South Asia, possibly due to the orographic obstacle presented by the Himalayan massif. The TB groups from Northeast India are genetically distinct compared to their counterparts from the Himalayas probably resulting from prolonged isolation and/or founder effects.
Close genetic ties have been reported between the Tamang and Tibet.1 It is likely that Tamangs are descendants of Tibetans who migrated south and settled in the southern region of the Himalayan range.1 This affinity is reflected in both CA plots (Figures 2 and 3) and NJ dendrograms (Figures 4 and 5). The Tibetan connection to the Tamang is also evident in their shared cultural and religious practices. The partitioning of these two populations with Bhutan and their proximity to the general collection from Nepal (Figures 2, 3, 4 and 5) may be associated with Neolithic migrants carrying Y-haplogroup O3a5-M134, an East Asian-specific marker, shared among TB populations.1, 3, 4, 9, 60 The Himalayan populations, with the exception of Newar and Kathmandu, segregate close to the Northeast Asian cluster in agreement with the admixture analyses results (Table 3). Northeast Asia is the major contributor to both Tibet (63.4%) and Tamang (59.7%) whereas Newar (44.7%) and Bhutan (41.1%) received equivalent percentages, followed by Kathmandu (22.3%). These results corroborate studies indicating a shared common ancestry between Tibet and the Northeast Asian collections of Japan and Korea by a variety of marker systems, including classical,61, 62 autosomal,63 Y-chromosome1, 12, 64, 65 and mtDNA.12, 64, 66, 67
More than half of the Tibetan men possess the YAP polymorphic Alu insertion in their Y-chromosome which is believed to have originated in Central Asia,1, 4, 11, 14although its source remains highly debated.64, 68, 69 In this study, however, given the lack of representative Central Asian populations due to the paucity of the data available from the region, no clear connections were made between Tibet and its possible Central Asian genetic contributors. Afghanistan is the sole Central Asian collection included in the analyses and appears to make no contributions to any of the Himalayan groups except for a minor influence in Kathmandu (12.9%).
To evaluate the genetic relationships between the Himalayan collections and the neighboring TB-speaking populations at the regional level, six Northeast Indian TB groups were included in the phylogenetic and statistical analyses performed using the 13 core CODIS STR loci. These Northeast Indian TB groups map distantly from both the Himalayan and East Asian populations in the CA graph (Figure 3), inconsistent with previous Y-chromosome and mtDNA studies which report a high degree of genetic homogeneity between Himalayan and Northeast Indian TB groups.3, 4, 9, 70 The discrepancy observed between Y-chromosome and microsatellite polymorphisms in the Northeast Indian TB groups may be explained by a male founder effect from Northeast Asia and their subsequent genetic isolation for an extended period of time following their arrival.9
Altogether, our results suggest a Northeast Asian ancestry for the Himalayan populations with subsequent genetic admixture in Kathmandu and Newar populations from South Asia. South Asian influences in Tibet and Tamang are negligible most likely due to the natural barrier presented by the Himalayas.1Tamang, Tibet and Bhutan display close genetic affiliations in all analyses possibly indicating a shared common ancestry. The biparental markers examined in this study reveal unique genetic profiles for the Northeast Indian TB groups, which are distinct from their Himalayan counterparts implying limited gene flow, geographic isolation and/or founder effects.
A subsequent expansion of Tai-Kadai speakers during the early second millennium AD from their homeland in South China into Thailand and Laos replaced Austro-Asiatic speakers in large parts of Southeast Asia that previously belonged to the Khmer empire , , . Subsequently, Tai-Kadai is found from South China over Thailand to the Malay Peninsula and Myanmar.
In historic times, parts of Southeast Asia have repeatedly been ruled by colonial forces, but there has never been overall occupation , . The Han Chinese invaded North Vietnam (Tonkin) in the 1st century BC and stayed for nearly a millennium, after which Vietnamese dynasties from North Vietnam conquered central Vietnam (Annam) and South Vietnam (Cochin China). The French occupied the same area (Tonkin, Annam, Cochin China) during a far shorter period (1863–1953), and added present day Cambodia and Laos to their colonial French Indochina. Both of these colonial episodes excluded Siam (Thailand), the only country in Southeast Asia never colonized by a European power.
Archaeology suggests an ancient close connection between India and the Thailand/Cambodia region through settlement , , , , accompanied by an increasing exposure to Indian culture from about 300 BC. Early states-like societies from Southeast Asia called by the Sanskrit term “mandala” had in common the adoption of Indian forms of religion (Hinduism), the Sanskrit language and aspects of government (Funan mandala from 100 to 550 AD, Chenla mandala from 550 to 802 AD and Angkorian mandala from 802 to 1431 AD) . However, the Indian influence in Southeast Asia was not supported by human mitochondrial DNA (mtDNA) data , , .
In previous studies, we have used housekeeping gene sequences of a bacterial parasite which infects the stomach of most humans, Helicobacter pylori, to elucidate the patterns of human prehistory. H. pylori accompanied modern humans during their migrations out of Africa ca. 60,000 years ago , and subsequent geographic separation plus founder effects have resulted in genetic populations of bacterial strains that are specific for large continental areas. In all, 7 bacterial genetic populations have been described… The specific geographic distribution and ethnic association of the H. pylori populations reflects numerous ancient and historic human migrations which established H. pylori sequences as a useful genetic marker to unravel debated topics in human population history. For example, the genetic variation in H. pylori has showed more discriminatory power in determining the ancient sources of human migrations in the Ladakh region of Northern India  and in the Pacific (Austronesian expansion)  than traditional human genetic markers such as the hypervariable region (HSV1) of mtDNA. Therefore, we analysed H. pylori sequences from Cambodia which borders Thailand to its west and northwest, Vietnam to its east and southeast and Laos to its north, to gain additional insights into the human population history in continental Southeast Asia.
“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.”
If this scenario is correct, it is likely that the passage to Taiwan did not exhaust the pre-An population of the Fujian coast. More likely, this population continued expanding along the coast in a south-westerly direction towards the Pearl River delta, even after a group of them had crossed to Taiwan. Their archaeological traces SW of Fujian are perhaps seen in the Pearl river delta, although direct evidence of agriculture there has so far not appeared; Hedang in the Pearl River delta, with tooth evulsion (Higham 1996:84), c. 3000-2000 BCE, may be one such site. In Taiwan, Tsang (in press) describes the newly excavated site of Nan-kuan-li near Tainan in south-west Taiwan, where a team led by him recently discovered a neolithic culture having rice, millet, and practicing ritual tooth ablation around 5000-4500 BP. In the same paper he argues that the Ta-Pen-K’eng culture, as seen in Nan-kuan-li near Tainan, “has close affinities with the Neolithic cultures of Hong Kong and the Pearl River Delta”. I disagree with Tsang when he concludes that “The Pearl River Delta of Kuangtung is most probably the source area of the Tapenkeng Culture in Taiwan”. I think it more likely that both cultures are descended from a common precursor on the Fujian coast. Pearl River delta sites having affinities to Taiwan TPK like Hedang are also probably too early and too far east to be ancestral to the Tai-Kadai-speaking cultures.
I have presented an explicit account of the early phylogeny of the
Austronesian family. The new phylogeny is tree-like. A salient characteristic is that out of a majority of nodes, only one branch leads to further branching (Table 4). This makes Formosan phylogeny similar to Malayo-Polynesian phylogeny. Non-branching nodes can be associated with stay-at-homes, and branching ones with out-migrating groups. PMP has been shown to be part of a taxon that also includes languages of the NE Formosan Coast, as well as Tai-Kadai (as proposed in Sagart 2001; in press, a). That taxon itself is part of a larger taxon including languages of the East coast and south Taiwan.
Apparently the main direction of movement was along the coastal plains.
This implies that, given a choice, the early Austronesians preferred to expand into the coastal plains. This pattern is consistent with what archaeology and linguistics tell us about their mode of subsistence, which combined exploitation of marine resources, including fishing, with hunting and gathering and cultivation of rice and millet. We may suppose that population movements into the mountains, as with the Saisiats, Atayalics, Thaos, Tsouics and Bununs, were generally late, and made under pressure. Such indeed is the pattern observed in the rest of the Austronesian world (Blust 1999:53). The pattern of progression from the west to the east coast is moreover consistent with archaeological dates for Ta-Pen-K’eng sites, which are older on the west coast than on the east coast.
Bing Su et al., Y-Chromosome Evidence for a Northward Migration of Modern Humans into Eastern Asia during the Last Ice Age, Am J Hum Genet. Dec 1999; 65(6): 1718–1724. Published online Nov 2, 1999. doi: 10.1086/302680
previously performed using limited number of autosomal genotypes. It also complements studies of mitochondrial and Y chromosome haplogroups as well as classical markers that provide important information with respect to part of the history of particular EAS ethnic groups [14–20]. Our study expands on previous analyses using HGDP population groups  by
examining additional parameters of population structure/diversity…
The graphic representation of the first two PCs showed close correspondence to the historical geographical location and/or sample collection site for most of the EAS population groups. Thus, despite admixture and perhaps uncertain migration patterns, overall the largest component of genotypic variation that is discernable by reducing high order data (all genotypes) to lower order variations (PCs) is consistent with the population geography. This finding supports hypotheses that the relationships among the EAS populations are largely explained by clines formed by demic expansion(s). We speculate that the inclusion of many different related ethnic groups has recapitulated the most common events that separated these ethnic groups. The first PC axis accounting for the largest variation has a north/south orientation. One major part of this pattern forms a line from Siberia (Yakut) to Mongolia to Eastern China (Figure 1). The PCA analyses also suggest that at least two separate clines originating or terminating in eastern China at one end and Cambodia and the Philippines at the other end. In addition there is another cline extending from Eastern China to the Korean peninsular and Japan.
Multiple previous studies have examined the relationship between and possible origins of different EAS population groups. Analysis of mitochondrial and Y chromosome haplogroups as well as a limited numbers of classical markers and microsatellite polymorphisms have also provided results that are generally consistent with a north/south orientation of relationships between different EAS population groups –. However, there are exceptions with some studies failing to show this relationship , . Summarized by a recent review  there are three different postulates regarding the origins of EAS population groups: 1) South East Asian origin –, 2) North Asian origin  and 3) a combination of northern and southern origin, . However, the majority of studies have supported a South-East Asian origin for most EAS populations and include detailed analyses of the age of specific mitochondrial haplogroups, Y chromosome sequences as well as limited marker studies . In contrast, hierarchical trees in the recent HGDP study  show branching points consistent with a Yakut derivation. Recent studies using a novel copying model statistical approach appear to suggest an initial northern and southern origin (Cambodians, Mongolians, Xibo, Yi , Tu, Daur, and Naxi receiving large contributions from central-Asian populations) that contribute to Han ancestry . These studies also provide data supporting the derivation of many other EAS groups from a Han expansion (including She, Japanese, Dai, Lahu and Miao). While the current study does not strongly support any of these hypotheses, it does suggest that eastern China is central to the events shaping the population groups in this region. …
The Himalayas have been suggested as a natural barrier for human migrations, especially the northward dispersals from the Indian Subcontinent to Tibetan Plateau. However, although the majority of Sherpa have a Tibeto-Burman origin, considerable genetic components from Indian Subcontinent have been observed in Sherpa people living in Tibet. The western Y chromosomal haplogroups R1a1a-M17, J-M304, and F*-M89 comprise almost 17% of Sherpa paternal gene pool. In the maternal side, M5c2, M21d, and U from the west also count up to 8% of Sherpa people. Those lineages with South Asian origin indicate that the Himalayas have been permeable to bidirectional gene flow.
In haplogroup O3a2c1a-M117, most of the Tibetan populations cluster tightly together in the NJ tree, along with Sherpa and Tamang of Nepal. However, more haplotypes of Sherpa samples share ancestry with Tibetan and other Tibeto-Burman populations from East Asia other than from Nepal (Figure 3b). Similarly, haplotypes of D1-M15 of Sherpa share ancestry with Tibetan, northwestern Han, and Zhuang (included in Tai-Kadai) populations from East Asia, although Sherpa has tended to be segregated away from the Tibetan cluster in the NJ tree. Similarly with R1a1a-M17, the haplotype diversity of D1-M15 samples in Sherpa is very low, and the haplotypes of Sherpa are only a small subset of thos e of Tibetan populations, probably also due to the founder effect when Sherpa was formed (Figure 3c). As we have mentioned above, haplogroup D3-P99 and D3a-P47 are almost exclusively distributed in Tibeto-Burman populations; but not only that, haplotypes of D3 also show strong similarities among different populations with distinctive and specific seven repeats at locus DYS392 (Figure 3d and Table S1). The haplotypes of J-M304 samples have already given us sufficient information to infer their origin and diffusion, although without typing downstream markers for those samples. The J-M304 samples should be probably assigned into J2 haplogroup and the haplotypes of those samples show strong similarities with those of Indian (in Southwest India) (Chennakrishnaiah et al.,2013), Malaysian Indian (Pamjav et al., 2011), and Lebanesesamples (Zalloua et al., 2008). One Sherpa sample has beenassigned as paragroup F*, which is observed only infrequently and primarily on the Indian subcontinent (Karafet et al., 2008).