Biogeography and assembly processes of abundant and rare soil microbial taxa in the southern part of the Qilian Mountain National Park, China.

Soil microorganisms play vital roles in regulating multiple ecosystem functions. Recent studies have revealed that the rare microbial taxa (with extremely low relative abundances, which are still largely ignored) are also crucial in maintaining the health and biodiversity of the soil and may respond differently to environmental pressure. However, little is known about the soil community structures of abundant and rare taxa and their assembly processes in different soil layers on the Qinghai-Tibet Plateau (QTP). The present study investigated the community structure and assembly processes of soil abundant and rare microbial taxa on the northeastern edge of the QTP. Soil microbial abundance was defined by abundant taxa, whereas rare taxa contributed to soil microbial diversity. The results of null model show that the stochastic process ruled the assembly processes of all sub-communities. Dispersal limitation contributed more to the assembly of abundant microbial taxa in the different soil layers. In contrast, drift played a more critical role in the assembly processes of the rare microbial taxa. In addition, in contrast to previous studies, the abundant taxa played more important roles in co-occurrence networks, most likely because of the heterogeneity of the soil, the sparsity of amplicon sequencing, the sampling strategy, and the limited samples in the present study. The results of this study improve our understanding of soil microbiome assemblies on the QTP and highlight the role of abundant taxa in sustaining the stability of microbial co-occurrence networks in different soil layers.

Hybridization and divergent climatic preferences drive divergence of two allopatric Gentiana species on the Qinghai-Tibet Plateau.

BACKGROUND AND AIMS: Exploring how species diverge is vital for understanding the drivers of speciation. Factors such as geographical separation and ecological selection, hybridization, polyploidization and shifts in mating system are all major mechanisms of plant speciation, but their contributions to divergence are rarely well understood. Here we test these mechanisms in two plant species, Gentiana lhassica and G. hoae, with the goal of understanding recent allopatric species divergence on the Qinghai-Tibet Plateau (QTP). METHODS: We performed Bayesian clustering, phylogenetic analysis and estimates of hybridization using 561 302 nuclear genomic single nucleotide polymorphisms (SNPs). We performed redundancy analysis, and identified and annotated species-specific SNPs (ssSNPs) to explore the association between climatic preference and genetic divergence. We also estimated genome sizes using flow cytometry to test for overlooked polyploidy. KEY RESULTS: Genomic evidence confirms that G. lhassica and G. hoae are closely related but distinct species, while genome size estimates show divergence occurred without polyploidy. Gentiana hoae has significantly higher average FIS values than G. lhassica. Population clustering based on genomic SNPs shows no signature of recent hybridization, but each species is characterized by a distinct history of hybridization with congeners that has shaped genome-wide variation. Gentiana lhassica has captured the chloroplast and experienced introgression with a divergent gentian species, while G. hoae has experienced recurrent hybridization with related taxa. Species distribution modelling suggested range overlap in the Last Interglacial Period, while redundancy analysis showed that precipitation and temperature are the major climatic differences explaining the separation of the species. The species differ by 2993 ssSNPs, with genome annotation showing missense variants in genes involved in stress resistance. CONCLUSIONS: This study suggests that the distinctiveness of these species on the QTP is driven by a combination of hybridization, geographical isolation, mating system differences and evolution of divergent climatic preferences.

Polypeptide from Moschus Suppresses Lipopolysaccharide-Induced Inflammation by Inhibiting NF-κ B-ROS/NLRP3 Pathway.

OBJECTIVE: To examine the anti-inflammatory effects and potential mechanisms of polypeptide from Moschus (PPM) in lipopolysaccharide (LPS)-induced THP-1 macrophages and BALB/c mice. METHODS: The polypeptide was extracted from Moschus and analyzed by high-performance liquid chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Subsequently, LPS was used to induce inflammation in THP-1 macrophages and BALB/c mice. In LPS-treated or untreated THP-1 macrophages, cell viability was observed by cell counting kit 8 and lactate dehydrogenase release assays; the proinflammatory cytokines and reactive oxygen species (ROS) were measured by enzyme-linked immunosorbent assay and flow cytometry, respectively; and protein and mRNA levels were measured by Western blot and real-time quantitative polymerase chain reaction (qRT-PCR), respectively. In LPS-induced BALB/c mice, the proinflammatory cytokines were measured, and lung histology and cytokines were observed by hematoxylin and eosin (HE) and immunohistochemical (IHC) staining, respectively. RESULTS: The SDS-PAGE results suggested that the molecular weight of purified PPM was in the range of 10-26 kD. In vitro, PPM reduced the production of interleukin 1ß (IL-1ß), IL-18, tumor necrosis factor α (TNF-α), IL-6 and ROS in LPS-induced THP-1 macrophages (P<0.01). Western blot analysis demonstrated that PPM inhibited LPS-induced nuclear factor κB (NF-κB) pathway and thioredoxin interacting protein (TXNIP)/nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain containing 3 (NLRP3) inflammasome pathway by reducing protein expression of phospho-NF-κB p65, phospho-inhibitors of NF-κB (Iκ Bs) kinase α/ß (IKKα/ß), TXNIP, NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), and pro-caspase-1 (P<0.05 or P<0.01). In addition, qRT-PCR revealed the inhibitory effects of PPM on the mRNA levels of TXNIP, NLRP3, ASC, and caspase-1 (P<0.05 or P<0.01). Furthermore, in LPS-induced BALB/c mice, PPM reduced TNF-α and IL-6 levels in serum (P<0.05 or P<0.01), decreased IL-1ß and IL-18 levels in the lungs (P<0.01) and alleviated pathological injury to the lungs. CONCLUSION: PPM could attenuate LPS-induced inflammation by inhibiting the NF-κB-ROS/NLRP3 pathway, and may be a novel potential candidate drug for treating inflammation and inflammation-related diseases.

Bacterial communities associated with mushrooms in the Qinghai-Tibet Plateau are shaped by soil parameters / Las comunidades bacterianas asociadas con los hongos en la meseta de Qinghai-Tíbet están formadas por parámetros del suelo

Fungi capable of producing fruit bodies are essential food and medicine resources. Despite recent advances in the study of microbial communities in mycorrhizospheres, little is known about the bacterial communities contained in fruit bodies. Using high-throughput sequencing, we investigated the bacterial communities in four species of mushrooms located on the alpine meadow and saline-alkali soil of the Qinghai-Tibet Plateau (QTP). Proteobacteria (51.7% on average) and Actinobacteria (28.2% on average) were the dominant phyla in all of the sampled fairy ring fruit bodies, and Acidobacteria (27.5% on average) and Proteobacteria (25.7% on average) dominated their adjacent soils. For the Agria. Bitorquis, Actinobacteria was the dominant phylum in its fruit body (67.5% on average) and adjacent soils (65.9% on average). The alpha diversity (i.e., Chao1, Shannon, Richness, and Simpson indexes) of the bacterial communities in the fruit bodies were significantly lower than those in the soil samples. All of the fungi shared more than half of their bacterial phyla and 16.2% of their total operational taxonomic units (OTUs) with their adjacent soil. Moreover, NH4+ and pH were the key factors associated with bacterial communities in the fruit bodies and soils, respectively. These results indicate that the fungi tend to create a unique niche that selects for specific members of the bacterial community. Using culture-dependent methods, we also isolated 27 bacterial species belonging to three phyla and five classes from fruit bodies and soils. The strains isolated will be useful for future research on interactions between mushroom-forming fungi and their bacterial endosymbionts.(AU)

Bacterial communities associated with mushrooms in the Qinghai-Tibet Plateau are shaped by soil parameters.

Fungi capable of producing fruit bodies are essential food and medicine resources. Despite recent advances in the study of microbial communities in mycorrhizospheres, little is known about the bacterial communities contained in fruit bodies. Using high-throughput sequencing, we investigated the bacterial communities in four species of mushrooms located on the alpine meadow and saline-alkali soil of the Qinghai-Tibet Plateau (QTP). Proteobacteria (51.7% on average) and Actinobacteria (28.2% on average) were the dominant phyla in all of the sampled fairy ring fruit bodies, and Acidobacteria (27.5% on average) and Proteobacteria (25.7% on average) dominated their adjacent soils. For the Agria. Bitorquis, Actinobacteria was the dominant phylum in its fruit body (67.5% on average) and adjacent soils (65.9% on average). The alpha diversity (i.e., Chao1, Shannon, Richness, and Simpson indexes) of the bacterial communities in the fruit bodies were significantly lower than those in the soil samples. All of the fungi shared more than half of their bacterial phyla and 16.2% of their total operational taxonomic units (OTUs) with their adjacent soil. Moreover, NH4+ and pH were the key factors associated with bacterial communities in the fruit bodies and soils, respectively. These results indicate that the fungi tend to create a unique niche that selects for specific members of the bacterial community. Using culture-dependent methods, we also isolated 27 bacterial species belonging to three phyla and five classes from fruit bodies and soils. The strains isolated will be useful for future research on interactions between mushroom-forming fungi and their bacterial endosymbionts.

Population genomics reveal deep divergence and strong geographical structure in gentians in the Hengduan Mountains.

Understanding the evolutionary and ecological processes driving population differentiation and speciation can provide critical insights into the formation of biodiversity. Here, we examine the link between population genetic processes and biogeographic history underlying the generation of diversity in the Hengduan Mountains (HM), a region harboring a rich and dynamic flora. We used restriction site-associated DNA sequencing to generate 1,907 single-nucleotide polymorphisms (SNPs) and four-kb of plastid sequence in species of the Gentiana hexaphylla complex (Gentianaceae). We performed genetic clustering with spatial and non-spatial models, phylogenetic reconstructions, and ancestral range estimation, with the aim of addressing the processes influencing diversification of G. hexaphylla in the HM. We find the G. hexaphylla complex is characterized by geographic genetic structure with clusters corresponding to the South, North and the central HM. Phylogenetic reconstruction and pairwise F ST analyses showed deep differentiation between Southern and Northern populations in the HM. The population in Mount Taibai exhibited the highest genetic similarity to the North HM. Ancestral range estimation indicated that the G. hexaphylla complex originated in the central HM and then diverged in the Pliocene and the Early Pleistocene, before dispersing widely, resulting in the current distinct lineages. Overall, we found deep genomic differentiation in the G. hexaphylla complex corresponds to geographic barriers to dispersal in the HM and highlights a critical role of the uplift of the Daxue Mountains and subsequent climatic fluctuations underlying diversification. The colonization of G. hexaphylla in the Mount Taibai region suggests directional dispersal between the alpine flora of the Qinling Mountains and the HM.

Strong plastid degradation is consistent within section Chondrophyllae, the most speciose lineage of Gentiana.

Recovering phylogenetic relationships in lineages experiencing intense diversification has always been a persistent challenge in evolutionary studies, including in Gentiana section Chondrophyllae sensu lato (s.l.). Indeed, this subcosmopolitan taxon encompasses more than 180 mostly annual species distributed around the world. We sequenced and assembled 22 new plastomes representing 21 species in section Chondrophyllae s.l. In addition to previously released plastome data, our study includes all main lineages within the section. We reconstructed their phylogenetic relationships based on protein-coding genes and recombinant DNA (rDNA) cistron sequences, and then investigated plastome structural evolution as well as divergence time. Despite an admittedly humble species cover overall, we recovered a well-supported phylogenetic tree based on plastome data, and found significant discordance between phylogenetic relationships and taxonomic treatments. Our results show that G. capitata and G. leucomelaena diverged early within the section, which is then further divided into two clades. The divergence time estimation showed that section Chondrophyllae s.l. evolved in the second half of the Oligocene. We found that section Chondrophyllae s.l. had the smallest average plastome size (128 KB) in tribe Gentianeae (Gentianaceae), with frequent gene and sequence losses such as the ndh complex and its flanking regions. In addition, we detected both expansion and contraction of the inverted repeat (IR) regions. Our study suggests that plastome degradation parallels the diversification of this group, and illustrates the strong discordance between phylogenetic relationships and taxonomic treatments, which now need to be carefully revised.

Reassessment of the Phylogeny and Systematics of Chinese Parnassia (Celastraceae): A Thorough Investigation Using Whole Plastomes and Nuclear Ribosomal DNA.

Parnassia L., a perennial herbaceous genus in the family Celastraceae, consists of about 60 species and is mainly distributed in the Pan-Himalayan and surrounding mountainous regions. The taxonomic position and phylogenetic relationships of the genus are still controversial. Herein, we reassessed the taxonomic status of Parnassia and its intra- and inter-generic phylogeny within Celastraceae. To that end, we sequenced and assembled the whole plastid genomes and nuclear ribosomal DNA (nrDNA) of 48 species (74 individuals), including 25 species of Parnassia and 23 species from other genera of Celastraceae. We integrated high throughput sequence data with advanced statistical toolkits and performed the analyses. Our results supported the Angiosperm Phylogeny Group IV (APG IV) taxonomy which kept the genus to the family Celastraceae. Although there were topological conflicts between plastid and nrDNA phylogenetic trees, Parnassia was fully supported as a monophyletic group in all cases. We presented a first attempt to estimate the divergence of Parnassia, and molecular clock analysis indicated that the diversification occurred during the Eocene. The molecular phylogenetic results confirmed numerous taxonomic revisions, revealing that the morphological characters used in Parnassia taxonomy and systematics might have evolved multiple times. In addition, we speculated that hybridization/introgression might exist during genus evolution, which needs to be further studied. Similarly, more in-depth studies will clarify the diversification of characters and species evolution models of this genus.

Reduced microbial stability in the active layer is associated with carbon loss under alpine permafrost degradation.

Permafrost degradation may induce soil carbon (C) loss, critical for global C cycling, and be mediated by microbes. Despite larger C stored within the active layer of permafrost regions, which are more affected by warming, and the critical roles of Qinghai-Tibet Plateau in C cycling, most previous studies focused on the permafrost layer and in high-latitude areas. We demonstrate in situ that permafrost degradation alters the diversity and potentially decreases the stability of active layer microbial communities. These changes are associated with soil C loss and potentially a positive C feedback. This study provides insights into microbial-mediated mechanisms responsible for C loss within the active layer in degraded permafrost, aiding in the modeling of C emission under future scenarios.

Lineage-specific plastid degradation in subtribe Gentianinae (Gentianaceae).

The structure and sequence of plastid genomes is highly conserved across most land plants, except for a minority of lineages that show gene loss and genome degradation. Understanding the early stages of plastome degradation may provide crucial insights into the repeatability and predictability of genomic evolutionary trends. We investigated these trends in subtribe Gentianinae of the Gentianaceae, which encompasses ca. 450 species distributed around the world, particularly in alpine and subalpine environments. We sequenced, assembled, and annotated the plastomes of 41 species, representing all six genera in subtribe Gentianinae and all main sections of the species-rich genus Gentiana L. We reconstructed the phylogeny, estimated divergence times, investigated the phylogenetic distribution of putative gene losses, and related these to substitution rate shifts and species' habitats. We obtained a strongly supported topology consistent with earlier studies, with all six genera in Gentianinae recovered as monophyletic and all main sections of Gentiana having full support. While closely related species have very similar plastomes in terms of size and structure, independent gene losses, particularly of the ndh complex, have occurred in multiple clades across the phylogeny. Gene loss was usually associated with a shift in the boundaries of the small single-copy and inverted repeat regions. Substitution rates were variable between clades, with evidence for both elevated and decelerated rate shifts. Independent lineage-specific loss of ndh genes occurred at a wide range of times, from Eocene to Pliocene. Our study illustrates that diverse degradation patterns shape the evolution of the plastid in this species-rich plant group.

Environmental filtering affects fungal communities more than dispersal limitation in a high-elevation hyperarid basin on Qinghai-Tibet Plateau.

The Qaidam Basin is the most extensive (120 000 km2) basin on the Qinghai-Tibet Plataea (QTP). Recent studies have shown that environmental selection and dispersal limitation influence the soil fungal community significantly in a large-scale distance. However, less is known about large-scale soil fungal community assemblages and its response to the elevation gradient in the high-elevation basin ecosystems. We studied fungal assemblages using Illumina sequencing of the ITS1 region from 35 sites of the Qaidam Basin. As the increase of elevation, fungal species richness and Chao1 index also increased. The Ascomycota was the most abundant phylum (more than 70% of total sequences), and six of the 10 most abundance fungal family was detected in all 35 soil samples. The key factors influencing the soil fungal community composition in the Qaidam Basin were environmental filtering (soil properties and climate factors). The Mantel test showed no significant relationship between geographic distance and community similarity (r = 0.05; p = 0.81). The absence of the distance effect might be caused by lacking dispersal limitation for the soil fungal community.

Recurrent hybridization underlies the evolution of novelty in Gentiana (Gentianaceae) in the Qinghai-Tibetan Plateau.

The Qinghai-Tibetan Plateau (QTP) and adjacent areas are centres of diversity for several alpine groups. Although it is known that the QTP acted as a source area for diversification of the alpine genus Gentiana, the evolutionary processes underlying diversity in this genus, especially the formation of narrow endemics, are still poorly understood. Hybridization has been proposed as a driver of plant endemism in the QTP but few cases have been documented with genetic data. Here, we describe a new endemic species in Gentiana section Cruciata as G. hoae sp. nov., and explore its evolutionary history with complete plastid genomes and nuclear ribosomal internal transcribed spacer sequence data. Genetic divergence within G. hoae ~3 million years ago was followed by postglacial expansion on the QTP, suggesting Pleistocene glaciations as a key factor shaping the population history of G. hoae. Furthermore, a mismatch between plastid and nuclear data suggest that G. hoae participated in historical hybridization, while population sequencing show this species continues to hybridize with the co-occurring congener G. straminea in three locations. Our results indicate that hybridization may be a common process in the evolution of Gentiana and may be widespread among recently diverged taxa of the QTP.

Morphology, genetic characterization and molecular phylogeny of pinworm Skrjabinema longicaudatum n. sp. (Oxyurida: Oxyuridae) from the endangered Tibetan antelope Pantholops hodgsonii (Abel) (Artiodactyla: Bovidae).

BACKGROUND: The Tibetan antelope Pantholops hodgsonii (Abel) (Artiodactyla: Bovidae) is an endangered species of mammal endemic to the Qinghai-Tibetan Plateau. Parasites and parasitic diseases are considered to be important threats in the conservation of the Tibetan antelope. However, our present knowledge of the composition of the parasites of the Tibetan antelope remains limited. METHODS: Large numbers of nematode parasites were collected from a dead Tibetan antelope. The morphology of these nematode specimens was observed using light and scanning electron microscopy. The nuclear and mitochondrial DNA sequences, i.e. small subunit ribosomal DNA (18S), large subunit ribosomal DNA (28S), internal transcribed spacer (ITS) and cytochrome c oxidase subunit 1 (cox1), were amplified and sequenced for molecular identification. Moreover, phylogenetic analyses were performed using maximum likelihood (ML) inference based on 28S and 18S + 28S + cox1 sequence data, respectively, in order to clarify the systematic status of these nematodes. RESULTS: Integrated morphological and genetic evidence reveals these nematode specimens to be a new species of pinworm Skrjabinema longicaudatum (Oxyurida: Oxyuridae). There was no intraspecific nucleotide variation between different individuals of S. longicaudatum n. sp. in the partial 18S, 28S, ITS and cox1 sequences. However, a high level of nucleotide divergence was revealed between the new species and its congeners in 28S (8.36%) and ITS (20.3-23.7%) regions, respectively. Molecular phylogenetic results suggest that the genus Skrjabinema should belong to the subfamily Oxyurinae (Oxyuroidea: Oxyuridae), instead of the subfamily Syphaciidae or Skrjabinemiinae in the traditional classification, as it formed a sister relationship to the genus Oxyuris. CONCLUSIONS: A new species of pinworm Skrjabinema longicaudatum n. sp. (Oxyurida: Oxyuridae) is described. Skrjabinema longicaudatum n. sp. represents the first species of Oxyurida (pinworm) and the fourth nematode species reported from the Tibetan antelope. Our results contribute to the knowledge of the species diversity of parasites from the Tibetan antelope, and clarify the systematic position of the genus Skrjabinema.

Population subdivision and hybridization in a species complex of Gentiana in the Qinghai-Tibetan Plateau.

BACKGROUND AND AIMS: Hosting several global biodiversity hotspots, the region of the Qinghai-Tibetan Plateau (QTP) is exceptionally species-rich and harbours a remarkable level of endemism. Yet, despite a growing number of studies, factors fostering divergence, speciation and ultimately diversity remain poorly understood for QTP alpine plants. This is particularly the case for the role of hybridization. Here, we explored the evolutionary history of three closely related Gentiana endemic species, and tested whether our results supported the mountain geo-biodiversity hypothesis (MGH). METHODS: We genotyped 69 populations across the QTP with one chloroplast marker and 12 nuclear microsatellite loci. We performed phylogeographical analysis, Bayesian clustering, approximate Bayesian computation and principal components analysis to explore their genetic relationship and evolutionary history. In addition, we modelled their distribution under different climates. KEY RESULTS: Each species was composed of two geographically distinct clades, corresponding to the south-eastern and north-western parts of their distribution. Thus Gentiana veitchiorum and G. lawrencei var. farreri, which diverged recently, appear to have shared at least refugia in the past, from which their range expanded later on. Indeed, climatic niche modelling showed that both species went through continuous expansion from the Last Interglacial Maximum to the present day. Moreover, we have evidence of hybridization in the northwest clade of G. lawrencei var. farreri, which probably occurred in the refugium located on the plateau platform. Furthermore, phylogenetic and population genetic analyses suggested that G. dolichocalyx should be a geographically limited distinct species with low genetic differentiation from G. lawrencei var. farreri. CONCLUSIONS: Climatic fluctuations in the region of the QTP have played an important role in shaping the current genetic structure of G. lawrencei var. farreri and G. veitchiorum. We argue that a species pump effect did occur prior to the Last Interglacial Maximum, thus lending support to the MGH. However, our results do depart from expectations as suggested in the MGH for more recent distribution range and hybridization dynamics.

Large-scale distribution of bacterial communities in the Qaidam Basin of the Qinghai-Tibet Plateau.

Many studies have investigated patterns of soil microbial communities over large spatial scales. However, these studies mainly focused on a few sites. Here, we studied the near-surface (0-30 cm) soil microbial communities of 35 soil samples collected from most of the areas of the Qaidam Basin, which is the largest basin on the Qinghai-Tibet Plateau. A total of 32 phyla and 838 genera were detected from all the samples, in which Actinobacteria, Proteobacteria, Bacteroidetes, and Acidobacteria were the most dominant and cosmopolitan phyla. The most abundant phyla (relative abundance > 5%) detected in all 35 soil samples were also the most dominant, which could be explained by their great dispersal ability. The microbial community structures correlated strongly with variations in pH and Mg2+ and were distinct between the high Mg2+ content (>20 g/kg) samples and other samples (Acidobacteria, Actinobacteria, and Chloroflexi were significantly less abundant in the high Mg2+ content group, but the abundance of Firmicutes was significantly greater). Finally, the microbial spatial pattern was influenced by both the local environment and spatial distance, but environmental factors were the primary drivers of microbial spatial patterns in the Qaidam Basin.

Development of EST-SSR markers in Saxifraga sinomontana (Saxifragaceae) and cross-amplification in three related species.

PREMISE: Saxifraga sinomontana (Saxifragaceae) is a widespread alpine species in the Qinghai-Tibetan Plateau and its flanking mountains. We developed a set of expressed sequence tag-simple sequence repeat (EST-SSR) markers to investigate the genetic diversity and evolutionary history of the species. METHODS AND RESULTS: We initially designed 50 EST-SSR markers based on transcriptome data of S. sinomontana. Nineteen of 50 loci (38%) were successfully amplified, 13 of which were polymorphic. These were tested on 71 individuals from four populations. Three to 18 alleles per locus were detected, and the levels of observed and expected heterozygosity ranged from 0.2817 to 0.9155 and 0.2585 to 0.8495, respectively. In addition, cross-amplification was successful for all 13 loci in three congeneric species, S. tangutica, S. heleonastes, and S. congestiflora. CONCLUSIONS: These EST-SSR markers will be useful for studying the genetic diversity of S. sinomontana and disentangling the phylogenetic relationships of related species.

Out of Refugia: Population Genetic Structure and Evolutionary History of the Alpine Medicinal Plant Gentiana lawrencei var. farreri (Gentianaceae).

Understanding the genetic structure and evolutionary history of plants contributes to their conservation and utilization and helps to predict their response to environmental changes. The wildflower and traditional Chinese and Tibetan medicinal plant Gentiana lawrencei var. farreri is endemic to the Qinghai-Tibetan Plateau (QTP). To explore its genetic structure and evolutionary history, the genetic diversity, divergence, and demographics were analyzed in individuals from 31 locations across the QTP using 1 chloroplast marker and 10 nuclear microsatellite loci. High genetic diversity was detected in G. lawrencei var. farreri, and most of the genetic variance was found within populations. Values of F ST in G. lawrencei var. farreri from nuclear microsatellite and chloroplast data were 0.1757 and 0.739, respectively. The data indicated the presence of isolation by distance. The southeast edge of the QTP was the main refugium for G. lawrencei var. farreri, and one microrefugium was also detected in the plateau platform of the QTP. Both nuclear microsatellite and chloroplast data indicated that the populations were divided into two geographically structured groups, a southeast group and a northwest group. The current genetic pattern was mainly formed through recolonization from the two independent refugia. Significant melt was detected at the adjacent area of the two geographically structured groups. Approximate Bayesian computation showed that the northwest group had diverged from the southeast group, which then underwent population expansion. Our results suggest that the two-refugia pattern had a significant impact on the genetic structure and evolutionary history of G. lawrencei var. farreri.

Rapid Intraspecific Diversification of the Alpine Species Saxifraga sinomontana (Saxifragaceae) in the Qinghai-Tibetan Plateau and Himalayas.

An increasing number of phylogeographic studies have been conducted for plant species in the Qinghai-Tibetan Plateau (QTP) and its flanking mountains. However, these studies have mainly focused on the determination of glacial refugia and routes of inter-/post-glacial expansions. Rapid intraspecific diversification of plants in this region have not been thoroughly discussed. Herein, we investigate the effects of the Quaternary climate changes on population genetic structure and diversifications of a herbaceous alpine species, Saxifraga sinomontana, which may have an evolutionary time scale <5 million years in the QTP and Himalayan regions. Using a total of 350 individuals from 29 populations, we studied the evolutionary history of S. sinomontana by analyzing cpDNA trnL-trnF, rpl16 and nrDNA ITS sequences. A total of 89 haplotypes and 158 genotypes were detected for cpDNA and ITS sequences, respectively. Only a few haplotypes/genotypes were widespread, while an extremely large number of haplotypes/genotypes were restricted to single populations, which were scattered throughout the current geographical range of S. sinomontana. This suggests the existence of microrefugia of this species during the Quaternary glaciations. In addition, the relationships of the haplotypes/genotypes were almost completely not resolved by phylogenetic reconstruction. Combining characteristics in terms of high haplotype richness, large proportion of private haplotypes, and shallow haplotype divergence, we speculate that recent intraspecific diversification has occurred in S. sinomontana. Molecular clock analysis estimated that the onset diversification within S. sinomontana to be 1.09 Ma (95% HPD = 0.80-1.45), coinciding with the extensive Quaternary glaciations on the QTP which started ca. 1.17 Ma. The Quaternary climatic oscillations may have triggered rapid intraspecific diversification in this QTP-Himalayan species. However, large niche breadth, as well as introgression/hybridization between the studied species and its closely related sympatric saxifrages, may also played a role to some extent on the current genetic structure of S. sinomontana, which need to be further studied.

Westwards and northwards dispersal of Triosteum himalayanum (Caprifoliaceae) from the Hengduan Mountains region based on chloroplast DNA phylogeography.

The varying topography and environment that resulted from paleoorogeny and climate fluctuations of the Himalaya-Hengduan Mountains (HHM) areas had a considerable impact on the evolution of biota during the Quaternary. To understand the phylogeographic pattern and historical dynamics of Triosteum himalayanum (Caprifoliaceae), we sequenced three chloroplast DNA fragments (rbcL-accD, rps15-ycf1, and trnH-psbA) from 238 individuals representing 20 populations. Nineteen haplotypes (H1-H19) were identified based on 23 single-site mutations and eight indels. Most haplotypes were restricted to a single population or neighboring populations. Analysis of molecular variance revealed that variations among populations were much higher than that within populations for the overall gene pool, as well as for the East Himalayan group (EH group) and the North Hengduan group (NHM group), but not for the Hengduan Mountains group (HM group). Ecoregions representing relatively high genetic diversity or high frequencies of private haplotypes were discovered, suggesting that this alpine herbaceous plant underwent enhanced allopatric divergence in isolated and fragmented locations during the Quaternary glaciations. The current phylogeographic structure of T. himalayanum might be due to heterogeneous habitats and Quaternary climatic oscillations. Based on the phylogeographic structure of T. himalayanum populations, the phylogenetic relationship of identified haplotypes and palaeodistributional reconstruction, we postulated both westwards and northwards expansion from the HM group for this species. The westwards dispersal corridor could be long, narrow mountain areas and/or the Yarlung Zangbo Valley, while the northwards movement path could be south-north oriented mountains and low-elevation valleys.