MS and HO conceived and designed the experiments. HO and BP performed the experiments. MS, HO, BP, GW, and AvH analyzed the data. HO, SP, WSI, DT, and TI contributed reagents/materials/analysis tools. MS, HO, and BP wrote the paper.
¤Current address: Department of Genetics, Yale University School of Medicine, New Haven, Connecticut, United States of America
Deceased 13 July 2004.
The authors have declared that no competing interests exist.
Contemporary hunter–gatherer groups are often thought to serve as models of an ancient lifestyle that was typical of human populations prior to the development of agriculture. Patterns of genetic variation in hunter–gatherer groups such as the !Kung and African Pygmies are consistent with this view, as they exhibit low genetic diversity coupled with high frequencies of divergent mtDNA types not found in surrounding agricultural groups, suggesting long-term isolation and small population sizes. We report here genetic evidence concerning the origins of the Mlabri, an enigmatic hunter–gatherer group from northern Thailand. The Mlabri have no mtDNA diversity, and the genetic diversity at Y-chromosome and autosomal loci are also extraordinarily reduced in the Mlabri. Genetic, linguistic, and cultural data all suggest that the Mlabri were recently founded, 500–800 y ago, from a very small number of individuals. Moreover, the Mlabri appear to have originated from an agricultural group and then adopted a hunting–gathering subsistence mode. This example of cultural reversion from agriculture to a hunting–gathering lifestyle indicates that contemporary hunter–gatherer groups do not necessarily reflect a pre-agricultural lifestyle.
Genes, language and culture reveal that the Mlabri reverted from an agricultural to a hunter-gatherer lifestyle, suggesting that hunter-gatherer groups might not always represent the pre- agricultural lifestyle of humans.
The Mlabri are an enigmatic group of about 300 people who nowadays range across the Nan, Phrae, and Phayao provinces of north and northeastern Thailand and the Sayaburi province of western Laos [
The origins of the Mlabri are controversial. Some investigators have assumed that there is a direct connection between the Mlabri and the ancient Hoabinhian hunting–gathering culture of Southeast Asia [
We report here the results of an investigation of genetic diversity in the Mlabri, to see whether patterns of genetic variation might provide further insights into the question of an agricultural versus hunting–gathering origin for the Mlabri. The rationale for using genetic analyses to investigate this question is that previous work has shown that hunter–gatherer groups typically differ from their agricultural neighbors in having reduced genetic diversity and high frequencies of unique mtDNA types [
We analyzed 360 bp of the first hypervariable segment (HV1) of the mtDNA control region in 58 Mlabri; surprisingly, all of the sequences were identical, with the following differences from the reference sequence [
Genetic diversity based on mtDNA HV1 sequences, Y-STR haplotypes, and autosomal STR (A-STR) genotypes in the Mlabri, compared to the average genetic diversity for six other hill tribes. The haplotype diversity is indicated for the mtDNA and Y-STR data, while the average heterozygosity is indicated for the autosomal STR loci.
Diversity in the Mlabri is significantly lower than the average for the other groups for all three genetic systems, based on
a Probability of the observed heterozygosity excess under the stepwise mutation model, Wilcoxon one-tailed test
We analyzed nine short tandem repeat (STR) loci on the Y chromosome in 54 Mlabri, and again found significantly reduced variation in the Mlabri compared to the other six hill tribes (
The number of repeats for the allele at each locus in the four haplotypes is given
We analyzed nine autosomal STR loci in the Mlabri and the other six hill tribes, and again found significantly reduced variation in the Mlabri (
Probability values of the likelihood ratio test of association versus no association for nine unlinked autosomal STR loci in the Mlabri and six other hill tribes (average probability over the 36 pairs of loci in parentheses).
One explanation for the reduced diversity at mtDNA, Y-STR loci, and autosomal STR loci, and the significant number of pairs of unlinked autosomal STR loci in LD, is a severe reduction in population size in the Mlabri. Following such an event, the number of alleles is reduced more than the heterozygosity, leading to an excess of observed heterozygosity compared to that expected for the observed number of alleles under mutation–drift equilibrium [
Assuming that there was a reduction in population size in the Mlabri that set the mtDNA and Y-chromosome diversity near or equal to zero, the coalescence times for the Mlabri mtDNA and Y-STR haplotypes provide an upper estimate as to when the population reduction occurred. We therefore applied Bayesian-based coalescence analysis [
Posterior probability distribution of the time back to the most recent common ancestor for the mtDNA (A) and Y-STR haplotype (B) data for the Mlabri and six other hill tribes.
Both the mtDNA and the Y-STR data therefore indicate that the Mlabri underwent a substantial reduction in population size about 500–800 y ago (and not more than about 1,300 y ago, if the mtDNA and Y-chromosome data reflect the same event). There are two possible scenarios: (1) a bottleneck, in which the Mlabri were reduced from a formerly large population to a much smaller population size, which then increased to the current level of about 300 individuals; or (2) a founder event, in which the Mlabri began as a very small number of individuals, became isolated, and then increased over time to their present size. Similar reductions in genetic diversity are predicted under either scenario, so the genetic data cannot distinguish between these. But some information can be obtained by considering the magnitude of the reduction in population size needed to completely eliminate mtDNA diversity in the Mlabri.
The amount of population size reduction needed to completely eliminate mtDNA diversity in the Mlabri depends on how much mtDNA diversity was present prior to the size reduction. We assumed that the ancestral Mlabri population would have the same mtDNA diversity as one of the other hill tribes and then estimated the amount of population size reduction needed to completely eliminate mtDNA diversity by resampling with replacement various numbers of mtDNA types from the ancestral (pre-bottleneck) population. For example, we started with an ancestral population with the same distribution of mtDNA types as the Akha. We then sampled two mtDNA types (with replacement) from this ancestral population, repeated this procedure 1,000 times, and found that 243 out of the 1,000 resamples of size two had no mtDNA diversity; thus, the probability is 0.243 that a reduction to just two individuals would eliminate mtDNA diversity in an ancestral population that started with the same mtDNA diversity as the Akha. We then repeated this procedure, sampling three mtDNA types (with replacement), and obtained a probability of 0.007 that there would be no mtDNA diversity following a reduction to three individuals. Therefore, if the Mlabri were derived from a population with the same mtDNA diversity as the Akha, the population had to be reduced to not more than two unrelated females, in order to completely eliminate mtDNA diversity.
This resampling analysis was carried out six times, with the putative ancestral mtDNA diversity corresponding to each of the six hill tribes. The results of this analysis were that for five of the ancestral populations, resampling three (or more) individuals gave a probability of no mtDNA diversity of less than 0.05; for the remaining ancestral population (which had the same starting mtDNA diversity as the Red Karen), resampling four (or more) individuals gave a probability of no mtDNA diversity of less than 0.05.
We also carried out a similar analysis for the Y-STR types in the Mlabri. Here we again assumed an ancestral population with the same Y-STR haplotype diversity as one of the other hill tribes, then determined the maximum number of individuals that could be sampled at random that would have not more than two Y-STR types (since the four Y-STR types in the Mlabri consist of two pairs that differ by a single-step mutation at a single locus). The results of this analysis were that at most 3–6 individuals (depending on which hill tribe the ancestral population resembled most in terms of Y-STR diversity) could have been present after the size reduction, otherwise, with greater than 95% probability, more than two Y-STR types would have been retained.
A critical assumption is the amount of genetic diversity present in the ancestral Mlabri population prior to the size reduction. The estimates used in the above analysis are based on agricultural populations, which in general have more mtDNA diversity than hunter–gatherer populations. We therefore also constructed putative ancestral populations with frequency distributions of mtDNA types identical to those found in the !Kung and in African Pygmies [
Another assumption of this analysis is that the event that led to the population size reduction completely eliminated the mtDNA diversity. Alternatively, some mtDNA diversity may have been present after the population size reduction, but was subsequently lost because of drift. Loss of mtDNA diversity due to subsequent drift is not likely if there was a single event reducing the Mlabri population size that was followed by population growth, since mtDNA diversity is retained in growing populations [
To investigate this further, we employed a Bayesian approach, following the procedure previously used to estimate the number of founders for the Maoris [
Posterior probability distribution for the number of founding individuals
The group that gave rise to the founder event that established the Mlabri could have been either a hunter–gatherer group, in which case the Mlabri maintained their hunting–gathering lifestyle from before, or an agricultural group, in which case the Mlabri subsequently adopted their current hunting–gathering lifestyle. While the genetic data cannot unequivocally distinguish between these two possibilities, they do suggest the latter. Other hunter–gatherer groups typically share few, if any, mtDNA types with neighboring agricultural groups, consistent with long-term isolation of the hunter–gatherer groups. For example, !Kung, African Pygmies, Andamanese Islanders, and south Indian hunter–gatherer groups can readily be distinguished from nearby agricultural groups on the basis of their mtDNA sequences [
The widespread sharing of mtDNA, Y-STR, and autosomal STR alleles between the Mlabri and agricultural groups in Southeast Asia are not expected if the Mlabri have always been hunter–gatherers. Instead, the genetic data suggest that the Mlabri are derived from an agricultural group. Moreover, the Mlabri vocabulary and folklore also give some evidence of ancient familiarity with agriculture coexisting with hunting and gathering (J. Rischel, personal communication). While preliminary in nature, the available linguistic evidence suggests that the present-day Mlabri language arose after some speakers of a Khmuic language, most likely Tin, became isolated and subsequently experienced intensive contact with speakers of some other, presently unknown language [
Other data that may shed light on the origins of the Mlabri, such as historical information, are scarce, since the Mlabri do not have a written language and the first recorded contact was only in 1936. However, the Tin Prai have an oral tradition concerning the origin of the Mlabri (J. Rischel, personal communication), in which several hundred years ago, Tin Prai villagers expelled two children and sent them downriver on a raft. They survived and escaped into the forest, turning to a foraging lifestyle and thus becoming the Mlabri. Although it is difficult to know how to evaluate such oral traditions, this story nevertheless intriguingly parallels the genetic and linguistic evidence concerning the origins of the Mlabri.
In sum, genetic, linguistic, and cultural data all suggest a founding event in the Mlabri, involving a single maternal lineage and 1–4 paternal lineages some 500–1,000 y ago, from an ancestral agricultural population. The Mlabri then subsequently adopted their present hunting and gathering lifestyle, possibly because the group size at the time of founding was too small to support an agricultural lifestyle. Other examples of such cultural reversion are rare; probably the best known involves Polynesian hunter–gatherers on the Chatham Islands and the South Island of New Zealand [
In any event, our conclusion that the Mlabri, a present-day group of hunters and gatherers, was founded recently and in all probability from an agricultural group further supports the contention that contemporary hunter–gatherer groups cannot be automatically assumed to represent the pre-agricultural lifestyle of human populations, descended unchanged from the Stone Age [
There are three linguistically distinct subgroups of Mlabri [
The first hypervariable segment (HV1) of the mtDNA control region (nucleotide positions 16,024–16,385) was amplified and sequenced directly, as described previously [
Genetic diversity, heterozygosity, and tests for goodness of fit to Hardy–Weinberg expectations were calculated with Arlequin 2.000 [
We thank S. Brauer for technical assistance, M. Wood for assistance with the resampling analyses, and R. Cordaux, D. Gil, and M. Kayser for useful discussion. We are especially grateful to J. Rischel for permission to cite his unpublished observations, and for discussions of the linguistic and cultural evidence concerning the Mlabri. HO was supported by a fellowship from the Japan Society for the Promotion of Science (JSPS). Sample collection was supported by funds from the JSPS and the Ministry of Education, Science, and Culture, Japan; genetic research was supported by the Max Planck Society, Germany. We dedicate this paper to the memory of Surin Pookajorn.
linkage disequilibrium
short tandem repeat
Y-chromosome short tandem repeat