Why do sex chromosomes progressively lose recombination?

Progressive recombination loss is a common feature of sex chromosomes. Yet, the evolutionary drivers of this phenomenon remain a mystery. For decades, differences in trait optima between sexes (sexual antagonism) have been the favoured hypothesis, but convincing evidence is lacking. Recent years have seen a surge of alternative hypotheses to explain progressive extensions and maintenance of recombination suppression: neutral accumulation of sequence divergence, selection of nonrecombining fragments with fewer deleterious mutations than average, sheltering of recessive deleterious mutations by linkage to heterozygous alleles, early evolution of dosage compensation, and constraints on recombination restoration. Here, we explain these recent hypotheses and dissect their assumptions, mechanisms, and predictions. We also review empirical studies that have brought support to the various hypotheses.

Stepwise recombination suppression around the mating-type locus in an ascomycete fungus with self-fertile spores.

Recombination is often suppressed at sex-determining loci in plants and animals, and at self-incompatibility or mating-type loci in plants and fungi. In fungal ascomycetes, recombination suppression around the mating-type locus is associated with pseudo-homothallism, i.e. the production of self-fertile dikaryotic sexual spores carrying the two opposite mating types. This has been well studied in two species complexes from different families of Sordariales: Podospora anserina and Neurospora tetrasperma. However, it is unclear whether this intriguing association holds in other species. We show here that Schizothecium tetrasporum, a fungus from a third family in the order Sordariales, also produces mostly self-fertile dikaryotic spores carrying the two opposite mating types. This was due to a high frequency of second meiotic division segregation at the mating-type locus, indicating the occurrence of a single and systematic crossing-over event between the mating-type locus and the centromere, as in P. anserina. The mating-type locus has the typical Sordariales organization, plus a MAT1-1-1 pseudogene in the MAT1-2 haplotype. High-quality genome assemblies of opposite mating types and segregation analyses revealed a suppression of recombination in a region of 1.47 Mb around the mating-type locus. We detected three evolutionary strata, indicating a stepwise extension of recombination suppression. The three strata displayed no rearrangement or transposable element accumulation but gene losses and gene disruptions were present, and precisely at the strata margins. Our findings indicate a convergent evolution of self-fertile dikaryotic sexual spores across multiple ascomycete fungi. The particular pattern of meiotic segregation at the mating-type locus was associated with recombination suppression around this locus, that had extended stepwise. This association between pseudo-homothallism and recombination suppression across lineages and the presence of gene disruption at the strata limits are consistent with a recently proposed mechanism of sheltering deleterious alleles to explain stepwise recombination suppression.

Sheltering of deleterious mutations explains the stepwise extension of recombination suppression on sex chromosomes and other supergenes.

Many organisms have sex chromosomes with large nonrecombining regions that have expanded stepwise, generating "evolutionary strata" of differentiation. The reasons for this remain poorly understood, but the principal hypotheses proposed to date are based on antagonistic selection due to differences between sexes. However, it has proved difficult to obtain empirical evidence of a role for sexually antagonistic selection in extending recombination suppression, and antagonistic selection has been shown to be unlikely to account for the evolutionary strata observed on fungal mating-type chromosomes. We show here, by mathematical modeling and stochastic simulation, that recombination suppression on sex chromosomes and around supergenes can expand under a wide range of parameter values simply because it shelters recessive deleterious mutations, which are ubiquitous in genomes. Permanently heterozygous alleles, such as the male-determining allele in XY systems, protect linked chromosomal inversions against the expression of their recessive mutation load, leading to the successive accumulation of inversions around these alleles without antagonistic selection. Similar results were obtained with models assuming recombination-suppressing mechanisms other than chromosomal inversions and for supergenes other than sex chromosomes, including those without XY-like asymmetry, such as fungal mating-type chromosomes. However, inversions capturing a permanently heterozygous allele were found to be less likely to spread when the mutation load segregating in populations was lower (e.g., under large effective population sizes or low mutation rates). This may explain why sex chromosomes remain homomorphic in some organisms but are highly divergent in others. Here, we model a simple and testable hypothesis explaining the stepwise extensions of recombination suppression on sex chromosomes, mating-type chromosomes, and supergenes in general.

Molecular ecology of microbiomes in the wild: Common pitfalls, methodological advances and future directions.

The study of microbiomes across organisms and environments has become a prominent focus in molecular ecology. This perspective article explores common challenges, methodological advancements, and future directions in the field. Key research areas include understanding the drivers of microbiome community assembly, linking microbiome composition to host genetics, exploring microbial functions, transience and spatial partitioning, and disentangling non-bacterial components of the microbiome. Methodological advancements, such as quantifying absolute abundances, sequencing complete genomes, and utilizing novel statistical approaches, are also useful tools for understanding complex microbial diversity patterns. Our aims are to encourage robust practices in microbiome studies and inspire researchers to explore the next frontier of this rapidly changing field.

Tempo of Degeneration Across Independently Evolved Nonrecombining Regions.

Recombination is beneficial over the long term, allowing more effective selection. Despite long-term advantages of recombination, local recombination suppression can evolve and lead to genomic degeneration, in particular on sex chromosomes. Here, we investigated the tempo of degeneration in nonrecombining regions, that is, the function curve for the accumulation of deleterious mutations over time, leveraging on 22 independent events of recombination suppression identified on mating-type chromosomes of anther-smut fungi, including newly identified ones. Using previously available and newly generated high-quality genome assemblies of alternative mating types of 13 Microbotryum species, we estimated degeneration levels in terms of accumulation of nonoptimal codons and nonsynonymous substitutions in nonrecombining regions. We found a reduced frequency of optimal codons in the nonrecombining regions compared with autosomes, that was not due to less frequent GC-biased gene conversion or lower ancestral expression levels compared with recombining regions. The frequency of optimal codons rapidly decreased following recombination suppression and reached an asymptote after ca. 3 Ma. The strength of purifying selection remained virtually constant at dN/dS = 0.55, that is, at an intermediate level between purifying selection and neutral evolution. Accordingly, nonsynonymous differences between mating-type chromosomes increased linearly with stratum age, at a rate of 0.015 per My. We thus develop a method for disentangling effects of reduced selection efficacy from GC-biased gene conversion in the evolution of codon usage and we quantify the tempo of degeneration in nonrecombining regions, which is important for our knowledge on genomic evolution and on the maintenance of regions without recombination.

Size Variation of the Nonrecombining Region on the Mating-Type Chromosomes in the Fungal Podospora anserina Species Complex.

Sex chromosomes often carry large nonrecombining regions that can extend progressively over time, generating evolutionary strata of sequence divergence. However, some sex chromosomes display an incomplete suppression of recombination. Large genomic regions without recombination and evolutionary strata have also been documented around fungal mating-type loci, but have been studied in only a few fungal systems. In the model fungus Podospora anserina (Ascomycota, Sordariomycetes), the reference S strain lacks recombination across a 0.8-Mb region around the mating-type locus. The lack of recombination in this region ensures that nuclei of opposite mating types are packaged into a single ascospore (pseudohomothallic lifecycle). We found evidence for a lack of recombination around the mating-type locus in the genomes of ten P. anserina strains and six closely related pseudohomothallic Podospora species. Importantly, the size of the nonrecombining region differed between strains and species, as indicated by the heterozygosity levels around the mating-type locus and experimental selfing. The nonrecombining region is probably labile and polymorphic, differing in size and precise location within and between species, resulting in occasional, but infrequent, recombination at a given base pair. This view is also supported by the low divergence between mating types, and the lack of strong linkage disequilibrium, chromosomal rearrangements, transspecific polymorphism and genomic degeneration. We found a pattern suggestive of evolutionary strata in P. pseudocomata. The observed heterozygosity levels indicate low but nonnull outcrossing rates in nature in these pseudohomothallic fungi. This study adds to our understanding of mating-type chromosome evolution and its relationship to mating systems.

Convergent recombination cessation between mating-type genes and centromeres in selfing anther-smut fungi.

The degree of selfing has major impacts on adaptability and is often controlled by molecular mechanisms determining mating compatibility. Changes in compatibility systems are therefore important evolutionary events, but their underlying genomic mechanisms are often poorly understood. Fungi display frequent shifts in compatibility systems, and their small genomes facilitate elucidation of the mechanisms involved. In particular, linkage between the pre- and postmating compatibility loci has evolved repeatedly, increasing the odds of gamete compatibility under selfing. Here, we studied the mating-type chromosomes of two anther-smut fungi with unlinked mating-type loci despite a self-fertilization mating system. Segregation analyses and comparisons of high-quality genome assemblies revealed that these two species displayed linkage between mating-type loci and their respective centromeres. This arrangement renders the same improved odds of gamete compatibility as direct linkage of the two mating-type loci under the automictic mating (intratetrad selfing) of anther-smut fungi. Recombination cessation was found associated with a large inversion in only one of the four linkage events. The lack of trans-specific polymorphism at genes located in nonrecombining regions and linkage date estimates indicated that the events of recombination cessation occurred independently in the two sister species. Our study shows that natural selection can repeatedly lead to similar genomic patterns and phenotypes, and that different evolutionary paths can lead to distinct yet equally beneficial responses to selection. Our study further highlights that automixis and gene linkage to centromeres have important genetic and evolutionary consequences, while being poorly recognized despite being present in a broad range of taxa.

Evidence of an additional centre of apple domestication in Iran, with contributions from the Caucasian crab apple Malus orientalis Uglitzk. to the cultivated apple gene pool.

Divergence processes in crop-wild fruit tree complexes in pivotal regions for plant domestication such as the Caucasus and Iran remain little studied. We investigated anthropogenic and natural divergence processes in apples in these regions using 26 microsatellite markers amplified in 550 wild and cultivated samples. We found two genetically distinct cultivated populations in Iran that are differentiated from Malus domestica, the standard cultivated apple worldwide. Coalescent-based inferences showed that these two cultivated populations originated from specific domestication events of Malus orientalis in Iran. We found evidence of substantial wild-crop and crop-crop gene flow in the Caucasus and Iran, as has been described in apple in Europe. In addition, we identified seven genetically differentiated populations of wild apple (M. orientalis), not introgressed by the cultivated apple. Niche modelling combined with genetic diversity estimates indicated that these wild populations likely resulted from range changes during past glaciations. This study identifies Iran as a key region in the domestication of apple and M. orientalis as an additional contributor to the cultivated apple gene pool. Domestication of the apple tree therefore involved multiple origins of domestication in different geographic locations and substantial crop-wild hybridization, as found in other fruit trees. This study also highlights the impact of climate change on the natural divergence of a wild fruit tree and provides a starting point for apple conservation and breeding programmes in the Caucasus and Iran.

Onset and stepwise extensions of recombination suppression are common in mating-type chromosomes of Microbotryum anther-smut fungi.

Sex chromosomes and mating-type chromosomes can display large genomic regions without recombination. Recombination suppression often extended stepwise with time away from the sex- or mating-type-determining genes, generating evolutionary strata of differentiation between alternative sex or mating-type chromosomes. In anther-smut fungi of the Microbotryum genus, recombination suppression evolved repeatedly, linking the two mating-type loci and extended multiple times in regions distal to the mating-type genes. Here, we obtained high-quality genome assemblies of alternative mating types for four Microbotryum fungi. We found an additional event of independent chromosomal rearrangements bringing the two mating-type loci on the same chromosome followed by recombination suppression linking them. We also found, in a new clade analysed here, that recombination suppression between the two mating-type loci occurred in several steps, with first an ancestral recombination suppression between one of the mating-type locus and its centromere; later, completion of recombination suppression up to the second mating-type locus occurred independently in three species. The estimated dates of recombination suppression between the mating-type loci ranged from 0.15 to 3.58 million years ago. In total, this makes at least nine independent events of linkage between the mating-type loci across the Microbotryum genus. Several mating-type locus linkage events occurred through the same types of chromosomal rearrangements, where similar chromosome fissions at centromeres represent convergence in the genomic changes leading to the phenotypic convergence. These findings further highlight Microbotryum fungi as excellent models to study the evolution of recombination suppression.

Higher Gene Flow in Sex-Related Chromosomes than in Autosomes during Fungal Divergence.

Nonrecombining sex chromosomes are widely found to be more differentiated than autosomes among closely related species, due to smaller effective population size and/or to a disproportionally large-X effect in reproductive isolation. Although fungal mating-type chromosomes can also display large nonrecombining regions, their levels of differentiation compared with autosomes have been little studied. Anther-smut fungi from the Microbotryum genus are castrating pathogens of Caryophyllaceae plants with largely nonrecombining mating-type chromosomes. Using whole genome sequences of 40 fungal strains, we quantified genetic differentiation among strains isolated from the geographically overlapping North American species and subspecies of Silene virginica and S. caroliniana. We inferred that gene flow likely occurred at the early stages of divergence and then completely stopped. We identified large autosomal genomic regions with chromosomal inversions, with higher genetic divergence than the rest of the genomes and highly enriched in selective sweeps, supporting a role of rearrangements in preventing gene flow in genomic regions involved in ecological divergence. Unexpectedly, the nonrecombining mating-type chromosomes showed lower divergence than autosomes due to higher gene flow, which may be promoted by adaptive introgressions of less degenerated mating-type chromosomes. The fact that both mating-type chromosomes are always heterozygous and nonrecombining may explain such patterns that oppose to those found for XY or ZW sex chromosomes. The specific features of mating-type chromosomes may also apply to the UV sex chromosomes determining sexes at the haploid stage in algae and bryophytes and may help test general hypotheses on the evolutionary specificities of sex-related chromosomes.

Recombination suppression and evolutionary strata around mating-type loci in fungi: documenting patterns and understanding evolutionary and mechanistic causes.

Genomic regions determining sexual compatibility often display recombination suppression, as occurs in sex chromosomes, plant self-incompatibility loci and fungal mating-type loci. Regions lacking recombination can extend beyond the genes determining sexes or mating types, by several successive steps of recombination suppression. Here we review the evidence for recombination suppression around mating-type loci in fungi, sometimes encompassing vast regions of the mating-type chromosomes. The suppression of recombination at mating-type loci in fungi has long been recognized and maintains the multiallelic combinations required for correct compatibility determination. We review more recent evidence for expansions of recombination suppression beyond mating-type genes in fungi ('evolutionary strata'), which have been little studied and may be more pervasive than commonly thought. We discuss testable hypotheses for the ultimate (evolutionary) and proximate (mechanistic) causes for such expansions of recombination suppression, including (1) antagonistic selection, (2) association of additional functions to mating-type, such as uniparental mitochondria inheritance, (3) accumulation in the margin of nonrecombining regions of various factors, including deleterious mutations or transposable elements resulting from relaxed selection, or neutral rearrangements resulting from genetic drift. The study of recombination suppression in fungi could thus contribute to our understanding of recombination suppression expansion across a broader range of organisms.

Pattern and causes of the establishment of the invasive bacterial potato pathogen Dickeya solani and of the maintenance of the resident pathogen D. dianthicola.

Invasive pathogens can be a threat when they affect human health, food production or ecosystem services, by displacing resident species, and we need to understand the cause of their establishment. We studied the patterns and causes of the establishment of the pathogen Dickeya solani that recently invaded potato agrosystems in Europe by assessing its invasion dynamics and its competitive ability against the closely related resident D. dianthicola species. Epidemiological records over one decade in France revealed the establishment of D. solani and the maintenance of the resident D. dianthicola in potato fields exhibiting blackleg symptoms. Using experimentations, we showed that D. dianthicola caused a higher symptom incidence on aerial parts of potato plants than D. solani, while D. solani was more aggressive on tubers (i.e. with more severe symptoms). In co-infection assays, D. dianthicola outcompeted D. solani in aerial parts, while the two species co-existed in tubers. A comparison of 76 D. solani genomes (56 of which have been sequenced here) revealed balanced frequencies of two previously uncharacterized alleles, VfmBPro and VfmBSer , at the vfmB virulence gene. Experimental inoculations showed that the VfmBSer population was more aggressive on tubers, while the VfmBPro population outcompeted the VfmBSer population in stem lesions, suggesting an important role of the vfmB virulence gene in the ecology of the pathogens. This study thus brings novel insights allowing a better understanding of the pattern and causes of the D.solani invasion into potato production agrosystems, and the reasons why the endemic D. dianthicola nevertheless persisted.

Population Genomics Reveals Molecular Determinants of Specialization to Tomato in the Polyphagous Fungal Pathogen Botrytis cinerea in France.

Many fungal plant pathogens encompass multiple populations specialized on different plant species. Understanding the factors underlying pathogen adaptation to their hosts is a major challenge of evolutionary microbiology, and it should help to prevent the emergence of new specialized pathogens on novel hosts. Previous studies have shown that French populations of the gray mold pathogen Botrytis cinerea parasitizing tomato and grapevine are differentiated from each other, and have higher aggressiveness on their host of origin than on other hosts, indicating some degree of host specialization in this polyphagous pathogen. Here, we aimed at identifying the genomic features underlying the specialization of B. cinerea populations to tomato and grapevine. Based on whole genome sequences of 32 isolates, we confirmed the subdivision of B. cinerea pathogens into two genetic clusters on grapevine and another, single cluster on tomato. Levels of genetic variation in the different clusters were similar, suggesting that the tomato-specific cluster has not recently emerged following a bottleneck. Using genome scans for selective sweeps and divergent selection, tests of positive selection based on polymorphism and divergence at synonymous and nonsynonymous sites, and analyses of presence and absence variation, we identified several candidate genes that represent possible determinants of host specialization in the tomato-associated population. This work deepens our understanding of the genomic changes underlying the specialization of fungal pathogen populations.

Threat to Asian wild apple trees posed by gene flow from domesticated apple trees and their "pestified" pathogens.

Secondary contact between crops and their wild relatives poses a threat to wild species, not only through gene flow between plants, but also through the dispersal of crop pathogens and genetic exchanges involving these pathogens, particularly those that have become more virulent by indirect selection on resistant crops, a phenomenon known as "pestification." Joint analyses of wild and domesticated hosts and their pathogens are essential to address this issue, but such analyses remain rare. We used population genetics approaches, demographic inference and pathogenicity tests on host-pathogen pairs of wild or domesticated apple trees from Central Asia and their main fungal pathogen, Venturia inaequalis, which itself has differentiated agricultural and wild-type populations. We confirmed the occurrence of gene flow from cultivated (Malus domestica) to wild (Malus sieversii) apple trees in Asian forests, potentially threatening the persistence of Asian wild apple trees. Pathogenicity tests demonstrated the pestification of V. inaequalis, the agricultural-type population being more virulent on both wild and domesticated trees. Single nucleotide polymorphism (SNP) markers and the demographic modelling of pathogen populations revealed hybridization following secondary contact between agricultural and wild-type fungal populations, and dispersal of the agricultural-type pathogen population in wild forests, increasing the threat of disease in the wild apple species. We detected an SNP potentially involved in pathogen pestification, generating an early stop codon in a gene encoding a small secreted protein in the agricultural-type fungal population. Our findings, based on joint analyses of paired host and pathogen data sets, highlight the threat posed by cultivating a crop near its centre of origin, in terms of pestified pathogen invasions in wild plant populations and introgression in the wild-type pathogen population.

Congruent population genetic structures and divergence histories in anther-smut fungi and their host plants Silene italica and the Silene nutans species complex.

Study of the congruence of population genetic structure between hosts and pathogens gives important insights into their shared phylogeographical and coevolutionary histories. We studied the population genetic structure of castrating anther-smut fungi (genus Microbotryum) and of their host plants, the Silene nutans species complex, and the morphologically and genetically closely related Silene italica, which can be found in sympatry. Phylogeographical population genetic structure related to persistence in separate glacial refugia has been recently revealed in the S. nutans plant species complex across Western Europe, identifying several distinct lineages. We genotyped 171 associated plant-pathogen pairs of anther-smut fungi and their host plant individuals using microsatellite markers and plant chloroplastic single nucleotide polymorphisms. We found clear differentiation between fungal populations parasitizing S. nutans and S. italica plants. The population genetic structure of fungal strains parasitizing the S. nutans plant species complex mirrored the host plant genetic structure, suggesting that the pathogen was isolated in glacial refugia together with its host and/or that it has specialized on the plant genetic lineages. Using random forest approximate Bayesian computation (ABC-RF), we found that the divergence history of the fungal lineages on S. nutans was congruent with that previously inferred for the host plant and probably occurred with ancient but no recent gene flow. Genome sequences confirmed the genetic structure and the absence of recent gene flow between fungal genetic lineages. Our analyses of individual host-pathogen pairs contribute to a better understanding of co-evolutionary histories between hosts and pathogens in natural ecosystems, in which such studies remain scarce.

Independent domestication events in the blue-cheese fungus Penicillium roqueforti.

Domestication provides an excellent framework for studying adaptive divergence. Using population genomics and phenotypic assays, we reconstructed the domestication history of the blue cheese mould Penicillium roqueforti. We showed that this fungus was domesticated twice independently. The population used in Roquefort originated from an old domestication event associated with weak bottlenecks and exhibited traits beneficial for pre-industrial cheese production (slower growth in cheese and greater spore production on bread, the traditional multiplication medium). The other cheese population originated more recently from the selection of a single clonal lineage, was associated with all types of blue cheese worldwide except Roquefort, and displayed phenotypes more suited for industrial cheese production (high lipolytic activity, efficient cheese cavity colonization ability and salt tolerance). We detected genomic regions affected by recent positive selection and putative horizontal gene transfers. This study sheds light on the processes of rapid adaptation and raises questions about genetic resource conservation.

Evolutionary strata on young mating-type chromosomes despite the lack of sexual antagonism.

Sex chromosomes can display successive steps of recombination suppression known as "evolutionary strata," which are thought to result from the successive linkage of sexually antagonistic genes to sex-determining genes. However, there is little evidence to support this explanation. Here we investigate whether evolutionary strata can evolve without sexual antagonism using fungi that display suppressed recombination extending beyond loci determining mating compatibility despite lack of male/female roles associated with their mating types. By comparing full-length chromosome assemblies from five anther-smut fungi with or without recombination suppression in their mating-type chromosomes, we inferred the ancestral gene order and derived chromosomal arrangements in this group. This approach shed light on the chromosomal fusion underlying the linkage of mating-type loci in fungi and provided evidence for multiple clearly resolved evolutionary strata over a range of ages (0.9-2.1 million years) in mating-type chromosomes. Several evolutionary strata did not include genes involved in mating-type determination. The existence of strata devoid of mating-type genes, despite the lack of sexual antagonism, calls for a unified theory of sex-related chromosome evolution, incorporating, for example, the influence of partially linked deleterious mutations and the maintenance of neutral rearrangement polymorphism due to balancing selection on sexes and mating types.

The complex evolutionary history of apricots: Species divergence, gene flow and multiple domestication events.

Domestication is an excellent model to study diversification and this evolutionary process can be different in perennial plants, such as fruit trees, compared to annual crops. Here, we inferred the history of wild apricot species divergence and of apricot domestication history across Eurasia, with a special focus on Central and Eastern Asia, based on microsatellite markers and approximate Bayesian computation. We significantly extended our previous sampling of apricots in Europe and Central Asia towards Eastern Asia, resulting in a total sample of 271 cultivated samples and 306 wild apricots across Eurasia, mainly Prunus armeniaca and Prunus sibirica, with some Prunus mume and Prunus mandshurica. We recovered wild Chinese species as genetically differentiated clusters, with P. sibirica being divided into two clusters, one possibly resulting from hybridization with P. armeniaca. Central Asia also appeared as a diversification centre of wild apricots. We further revealed at least three domestication events, without bottlenecks, that gave rise to European, Southern Central Asian and Chinese cultivated apricots, with ancient gene flow among them. The domestication event in China possibly resulted from ancient hybridization between wild populations from Central and Eastern Asia. We also detected extensive footprints of recent admixture in all groups of cultivated apricots. Our results thus show that apricot is an excellent model for studying speciation and domestication in long-lived perennial fruit trees.

Population genomics revealed cryptic species within host-specific zombie-ant fungi (Ophiocordyceps unilateralis).

The identification and delimitation of species boundaries are essential for understanding speciation and adaptation processes and for the management of biodiversity as well as development for applications. Ophiocordyceps unilateralis sensu lato is a complex of fungal pathogens parasitizing Formicine ants, inducing zombie behaviors in their hosts. Previous taxonomic works with limited numbers of samples and markers led to the "one ant-one fungus" paradigm, resulting in the use of ant species as a proxy for fungal identification. Here, a population genomics study with sampling on three ant species across Thailand supported the existence of host-specific species in O. unilateralis s.l. with no footprints of long term introgression despite occasional host shifts and first-generation hybrids. We further detected genetic clusters within the previously delimited fungal species, with each little footprints of recombination, suggesting high levels of inbreeding. The clusters within each of O. camponoti-leonardi and O. camponoti-saundersi were supported by differentiation throughout the genome, suggesting they may constitute further cryptic species parasitizing the same host, challenging the one ant-one fungus paradigm. These genetic clusters had different geographical ranges, supporting different biogeographic influences between the north/center and the south of Thailand, reinforcing the scenario in which Thailand endured compartmentation during the latest Pleistocene glacial cycles.