Ayta Negritos: Highest Levels of Denisovan DNA

Ayta Negritos: Highest Levels of Denisovan DNA Aug 27, 2021 2:21:06 GMT

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Post by Admin on Aug 27, 2021 2:21:06 GMT

The Toalean burial from Leang Panninge
The most distinctive archaeological assemblages associated with Holocene hunter-gatherers in Wallacea belong to the Toalean technocomplex (8–1.5 kya)3,4,27,28. Found only in a 10,000 km2 area of South Sulawesi3 (Fig. 1b), Toalean cultural assemblages are generally characterized by backed microliths and small stone projectiles (‘Maros points’)4 (Extended Data Fig. 1a–c). In 2015, excavations at Leang Panninge in the Mallawa district of Maros, South Sulawesi (Fig. 1b), uncovered the first relatively complete human burial from a secure Toalean context (Extended Data Figs. 1–5, Supplementary Information). The individual was interred in a flexed position29 in a rich aceramic Toalean stratum. Exposed at a depth of around 190 cm, the burial has an inferred age of 7.3–7.2 kyr cal BP obtained from 14C dating of a Canarium sp. seed (Extended Data Figs. 2, 3, Supplementary Table 1). Morphological characters indicate that this Toalean forager was a 17–18-year-old female with a broadly Australo-Melanesian affinity, although the morphology does not fall outside the range of recent Southeast Asian variation (Supplementary Information).

Genomic analysis
We extracted ancient DNA from bone powder obtained from the petrous portion of the temporal bone of the Leang Panninge individual. After library preparation, we used a DNA hybridization capture approach to enrich for approximately 3 million single-nucleotide polymorphisms (SNPs) across the human genome (1240K and archaic admixture capture panels30) as well as for the entire mitochondrial genome (mtDNA capture31). We retrieved 263,207 SNPs on the 1240K panel, 299,047 SNPs on the archaic admixture panel and the almost complete mtDNA sequence. Authenticity of the analysed ancient DNA was confirmed by short average fragment length, elevated damage patterns towards the molecule ends, and low autosomal and mtDNA contamination estimates (Supplementary Fig. 1). We confirmed that the individual was genetically of female sex. Analysis of the polymorphisms present in the reconstructed mtDNA sequence suggests a deeply divergent placement within mtDNA haplogroup M (Supplementary Table 17, Supplementary Fig. 2).

We initiated our genomic investigation by principal component analyses (PCAs), comparing the Leang Panninge genome with present-day individuals from East Asia, southeast Asia and Near Oceania (comprising Indigenous Australia, Papua New Guinea and Bougainville) genotyped on the Human Origins SNP panel18,32,33,34. The newly generated genome and relevant published genomes from ancient individuals from eastern Eurasia were then projected on the PCA1,34,35,36,37,38. Leang Panninge falls into PCA space not occupied by any present-day or ancient individuals, but is broadly located between Indigenous Australian peoples and the Onge (Fig. 2a, Extended Data Fig. 6). F3-statistics33 of the form f3 (Mbuti; Leang Panninge, X), where X is replaced with present-day Asian-Pacific groups, indicated that the new genome shares most genetic drift with Near Oceanian individuals (Fig. 2b). We confirmed these results with f4-statistics33, suggesting similar affinity of Leang Panninge and Papuan individuals to present-day Asian individuals, despite Near Oceanian groups forming a clade to the exclusion of Leang Panninge (Extended Data Fig. 7a, b). All present-day groups from the region, with the exception of the Mamanwa and the Lebbo26, carry only a minor contribution of Papuan-related ancestry (Supplementary Fig. 4).

Fig. 2: The Leang Panninge genome within the regional genetic context.

a, PCA calculated on present-day individuals from eastern Eurasia and Near Oceania, projecting key ancient individuals from the region1,34,35,36,37,38. b, Shared genetic drift of present-day groups with the Leang Panninge individual, as calculated using f3 (Mbuti; Leang Panninge, X) mapped at the geographical position of the tested group. WGA, whole genome amplification. c, The amount of introgressed Denisovan sequence in fragments longer than 0.05 cM in present-day (Simons Genome Diversity Project) individuals and longer than 0.2 cM in ancient individuals (measured with admixfrog). Each bar represents the posterior mean estimate from a single genome and the whiskers indicate 2 s.d. (estimated from 200 samples from the posterior decoding).

Ayta Negritos: Highest Levels of Denisovan DNA Aug 27, 2021 18:32:23 GMT

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Post by Admin on Aug 27, 2021 18:32:23 GMT

To investigate the presence and distribution of genetic contributions attributable to Denisovan-related groups39, we calculated the statistic f4 (Mbuti, Denisova; Leang Panninge, X), where X are groups from present-day Island Southeast Asia, Near Oceania and the Andamans, as well as ancient Asian-Pacific individuals1,37,38. Positive values calculated for Near Oceanian groups suggest higher proportions of Denisovan-related ancestry than the Leang Panninge individual (z-scores of >3.19), while the Onge and the remaining ancient individuals returned negative values, indicating a lower proportion of Denisovan-related ancestry (Extended Data Fig. 7c, Supplementary Fig. 3). We also calculated f4-ratio statistics to estimate the Denisovan proportion using SNPs from the 1240K capture panel and Han individuals from East Asia as a baseline18. We confirmed that Indigenous Australian and Papuan individuals carry a similar amount of Denisovan ancestry (approximately 2.9%)18,21,40, whereas the Leang Panninge individual has a lower value of approximately 2.2 ± 0.5% (Supplementary Tables 18–20). The Denisovan admixture proportion in the Leang Panninge individual is higher than in the Hòabìnhian individuals from Pha Faen and Gua Cha1, suggesting that groups ancestral to hunter-gatherers from Wallacea and Sunda were involved in different introgression events with archaic hominins. In addition, we performed D-statistics on a set of SNPs designed to measure the contribution of archaic ancestry in modern humans (archaic admixture capture panel). The Leang Panninge individual shares fewer Denisovan-related alleles with Papuan individuals, but has more such alleles than most tested groups, including the Tianyuan individual from Late Pleistocene China38. Neanderthal allele sharing is similar across all tested present-day non-African groups (Supplementary Tables 21–23). Finally, we ran admixfrog41 on the set of archaic admixture SNPs and measured 22.4 Mb (±1.9 Mb) of Denisovan-related ancestry in 33 fragments distributed across the Leang Panninge genome. This contribution accounts for around half of what is found in Papuan groups, but there is a significant correlation between the Denisovan fragments in the Leang Panninge genome and those in present-day Near Oceanian groups, suggesting shared introgression events (Fig. 2c, Extended Data Fig. 8, Supplementary Fig. 5).

To investigate whether the apparent PCA shift of Leang Panninge away from Near Oceanian groups is due to genetic drift alone, we performed a multidimensional scaling plot based on genetic similarities measured as 1 − f3 (Mbuti; Leang Panninge, X). The multidimensional scaling positioning of the Leang Panninge individual recapitulates the PCA with an intermediate placement between Papuan and Asian individuals (Extended Data Fig. 9). We then used f4-statistics and qpWave33 to formally test for the presence of additional genetic sources in Leang Panninge other than the Papuan-related ancestry. This identified a marginal affinity towards ancient Asian genomes (Extended Data Fig. 7d), and a minimum of two streams of ancestry when Denisova37 and/or ancient Asian groups1,37,38 were included in the qpWave reference groups (Supplementary Table 24). On the basis of these results, we used qpAdm33 to identify potential sources for an Asian-related ancestry in the genome alongside the Papuan-related component (Supplementary Table 25). Using a rotating approach among different Asian groups1,37,38, we were able to model the Leang Panninge genome as a mixture between Papuan and Tianyuan (51 ± 11%) or Onge (43 ± 9%) (Fig. 3a, Supplementary Table 26). Further exploration with admixture graphs built in qpGraph33 and TreeMix42, including present-day groups and relevant ancient individuals37,38,40,43, provided evidence for the presence of deep Asian ancestry (Fig. 3b, c, Supplementary Figs. 6–11). In TreeMix, the first admixture edge represents archaic introgression from Denisovan-related groups into the common ancestor of Leang Panninge and present-day Near Oceanian peoples. This is followed by an East Asian-related gene flow into Leang Panninge departing basally from the Qihe lineage, an early Neolithic genome from southeastern China37 (Fig. 3b, Extended Data Fig. 10, Supplementary Fig. 6). The qpGraph analysis confirmed this branching pattern, with the Leang Panninge individual branching off from the Near Oceanian clade after the Denisovan gene flow, although with the most supported topology indicating around 50% of a basal East Asian component contributing to the Leang Panninge genome (Fig. 3c, Supplementary Figs. 7–11).

Fig. 3: Admixture signals detected in the Leang Panninge genome.

a, Admixture proportions modelling Leang Panninge as a combination of Papuan49 and Tianyuan38 or Onge49 groups as estimated by qpAdm33 using Mbuti, Denisovan39, Kostenki 14 (ref. 50) and ancient Asian individuals1,37 as rotating reference groups (Supplementary Table 26). The error bars denote standard errors as calculated with block jacknife in the qpAdm software. b, c, Admixture graphs placing Leang Panninge on the branch with the present-day Near Oceanian clade41 and showing the admixture with a deep Asian-related ancestry in TreeMix42 (b) (Extended Data Fig. 10, Supplementary Fig. 6) and qpGraph (c) (worst z-score of −2.194; Supplementary Figs. 7–11)33,37,38,39,43. In b, ‘1’ and ‘2’ refer to the order in which the TreeMix software added ‘migration events’ (indicated by the arrows) to the graph. When plotting qpGraph results (c), the dotted arrows indicate admixture edges.

Discussion
Genome-wide analyses of the Leang Panninge individual show that most genetic drift is shared with present-day groups from New Guinea and Aboriginal Australia (Fig. 2b, Extended Data Fig. 7a). However, this Toalean-associated genome represents a previously undescribed ancestry profile, one that branched off after Onge-related and Hòabìnhian-related lineages but around the time that Papuan and Indigenous Australian groups split (Fig. 3b, c, Extended Data Fig. 8, Supplementary Figs. 6–9). It is possible that this Toalean individual carries a local ancestry that was present in Sulawesi before the initial peopling of Sahul at least 50 kya9, although whether this population produced the Late Pleistocene rock art in the south of the island11,44,45 is unknown.

The Toalean individual carries substantial Denisovan-related ancestry, probably sharing the archaic admixture event with present-day Near Oceanian groups (Fig. 2c, Extended Data Figs. 7c, 10, Supplementary Fig. 5, Supplementary Tables 21–23). This provides strong support for the main Denisovan-related gene flow happening before modern humans reached Sahul, making both Wallacea and Sunda equally likely locations for this archaic introgression event. However, previously published hunter-gatherer genomes from Sunda carry almost no Denisovan-related ancestry (Supplementary Tables 18–20), suggesting either a Hòabìnhian-related spread into southeast Asia after the aforementioned gene flow or that Wallacea was indeed the crucial meeting point between archaic and modern humans. The apparent presence of a long-established population of archaic hominins in southwestern Sulawesi46 provides a possible source for the introgression event. Two previous studies have suggested that two deeply divergent Denisovan lineages admixed into the ancestors of Papuan individuals17,22, but our genomic data currently do not have enough resolution to distinguish among one or multiple introgression pulses.

The lower amount of Denisovan ancestry in the Leang Panninge individual than in Papuan and Indigenous Australian individuals could result from: (1) an additional admixture with Denisovan ancestry into the common ancestors of Near Oceanian groups, or (2) a dilution of the Denisovan-related ancestry in the Leang Panninge genome through admixture with lineages carrying less or no such ancestry. Our allele frequency-based analyses do not support the first scenario (Supplementary Fig. 11), but they do favour the latter. The scarcity of pre-Neolithic genomes from across Asia prevents us from defining the exact source and admixture proportions of this gene flow event. It is noteworthy, however, that despite the reconstructed population trees (TreeMix and qpGraph) suggesting a genetic influence on middle Holocene Sulawesi from mainland East Asia, our qpAdm modelling cannot rule out a southeast Asian contribution from a group related to present-day Andamanese peoples (Fig. 3, Supplementary Figs. 6–11, Supplementary Table 26). This is consistent with a recent study that describes widespread admixtures across Asia between Onge-related and Tianyuan-related ancient populations47. However, the presence of this type of ancestry in a middle Holocene forager from Wallacea suggests that the Asian-related admixture could have taken place long before the expansion of Austronesian societies into the region.

We could not detect evidence for the Leang Panninge ancestry in any tested present-day groups (Supplementary Fig. 4). This could be owing to the overall limited proportion of Near Oceanian-related ancestry in Wallacea or large-scale genetic discontinuity between earlier hunter-gatherers and modern groups. The latter scenario would suggest that any genetic signal related to the Leang Panninge individual was obscured by later demographic processes, including the Austronesian expansion1,20,26,48. Higher coverage genetic data from present-day populations in Sulawesi, and additional Toalean ancient genomes, are needed to further investigate this unique ancestry profile and the genetic diversity of hunter-gatherers from Wallacea more generally.