An in-depth analysis of the mitochondrial phylogenetic landscape of Cambodia

Anita Kloss-Brandstätter,Florian Kronenberg,Julia Raschenberger,Torpong Sanguansermsri,David Horst,Hansi Weissensteiner,Jürgen Horst,Basil Horst,Gertraud Streiter,Monika Summerer

doi:10.1038/s41598-021-90145-2

Abstract

Cambodia harbours a variety of human aboriginal populations that have scarcely been studied in terms of genetic diversity of entire mitochondrial genomes. Here we present the matrilineal gene pool of 299 Cambodian refugees from three different ethnic groups (Cham, Khmer, and Khmer Loeu) deriving from 16 Cambodian districts. After establishing a DNA-saving high-throughput strategy for mitochondrial whole-genome Sanger sequencing, a HaploGrep based workflow was used for quality control, haplogroup classification and phylogenetic reconstruction. The application of diverse phylogenetic algorithms revealed an exciting picture of the genetic diversity of Cambodia, especially in relation to populations from Southeast Asia and from the whole world. A total of 224 unique haplotypes were identified, which were mostly classified under haplogroups B5a1, F1a1, or categorized as newly defined basal haplogroups or basal sub-branches of R, N and M clades. The presence of autochthonous maternal lineages could be confirmed as reported in previous studies. The exceptional homogeneity observed between and within the three investigated Cambodian ethnic groups indicates genetic isolation of the whole population. Between ethnicities, genetic barriers were not detected. The mtDNA data presented here increases the phylogenetic resolution in Cambodia significantly, thereby highlighting the need for an update of the current human mtDNA phylogeny.

Highlights

Cambodia harbours a variety of human aboriginal populations that have scarcely been studied in terms of genetic diversity of entire mitochondrial genomes
Being one of the most ancient language families in eastern Asia, Austro-Asiatic is spoken in India, Bangladesh and South-western China, implying that the Austro-Asiatic speaking populations may represent the descendants of the earliest settlers of modern humans who migrated from Africa and entered into eastern Asia about 60,000 years a go[2,3,4]
Phylogenetic trees were directly generated within HaploGrep 2, data exported in multiple alignment format for maximum likelihood phylogenetic tree inference with RamXL-NG23, Bayesian evolutionary analysis with BEAST 224, Median-Joining Networks with Network.exe (v.4)[25] as well as Maximum Parsimony trees

Summary

Introduction

Cambodia harbours a variety of human aboriginal populations that have scarcely been studied in terms of genetic diversity of entire mitochondrial genomes. Besides various control region sequencing projects in populations from Southeast Asia (for example, populations from T hailand15, Laos16, Vietnam17, Burma[18], and other Southeast Asian c ountries19–22), only one publication sequenced the mtDNA HVS-I (range 16,038–16,462) and HVS-II (range 65–417) from 1,054 unrelated Cambodians[2] From these partial sequences, Zhang and colleagues analysed the complete mitochondrial genome from 98 Cambodian individuals and identified eight new and basal mitochondrial DNA haplogroups, which led to the conclusion that present-day Cambodians still carry ancient genetic polymorphisms in their. The aim of our study was to provide a high quality mtDNA data set from Cambodia in order to understand the complex history of Southeast Asian mtDNA evolution and diversity This was accomplished by genotyping the so far largest population set of complete mitochondrial genomes of 299 Cambodian individuals, with a cost-efficient, DNA-saving, and reliable high-throughput strategy for mitochondrial whole-genome Sanger sequencing, and by distilling ~ 7,000 mtDNA samples from studies on neighbouring- and global populations. Phylogenetic trees were directly generated within HaploGrep 2, data exported in multiple alignment format for maximum likelihood phylogenetic tree inference with RamXL-NG23, Bayesian evolutionary analysis with BEAST 224, Median-Joining Networks with Network.exe (v.4)[25] as well as Maximum Parsimony trees

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Conclusion