Double-digest Restriction Site-associated DNA Sequencing Research Articles

Genomic signatures of hybridization between Ixodes ricinus and Ixodes persulcatus in natural populations.

Identifying hybridization between common pathogen vectors is essential due to the major public health implications through risks associated with hybrid's enhanced pathogen transmission potential. The hard-ticks Ixodes ricinus and Ixodes persulcatus are the two most common vectors of tick-borne pathogens that affect human and animal health in Europe. Ixodes ricinus is a known native species in Finland with a well-known distribution, whereas I. persulcatus has expanded in range and abundance over the past 60 years, and currently it appears the most common tick species in certain areas in Finland. Here we used double-digest restriction site-associated DNA (ddRAD) sequencing on 186 ticks (morphologically identified as 92 I. ricinus, and 94 I. persulcatus) collected across Finland to investigate whether RAD generated single nucleotide polymorphisms (SNPs) can discriminate tick species and identify potential hybridization events. Two different clustering methods were used to assign specific species based on how they clustered and identified hybrids among them. We were able to discriminate between the two tick species and identified 11 putative hybrids with admixed genomic proportions ranging from approximately 24 to 76 percent. Four of these hybrids were morphologically identified as I. ricinus while the remaining seven were identified as I. persulcatus. Our results thus indicate that RAD SNPs are robust in identifying both species of the ticks as well as putative hybrids. These results further suggest ongoing hybridization between I. ricinus and I. persulcatus in their natural populations in Finland. The unique ability of RAD markers to discriminate between tick species and hybrids adds a useful aspect to tick evolutionary studies. Our findings align with previous studies and suggest a shared evolutionary history between the species, with instances of individuals possessing a considerable proportion of the other species' genome. This study is a significant step in understanding the formation of hybridization zones due to range expansion potentially associated with climate change.

Shared phylogeographic patterns and environmental responses of co-distributed soybean pests: Insights from comparative phylogeographic studies of Riptortus pedestris and Riptortus linearis in the subtropics of East Asia

Comparative phylogeographic studies of closely related species sharing co-distribution areas can elucidate the role of shared historical factors and environmental changes in shaping their phylogeographic pattern. The bean bugs, Riptortus pedestris and Riptortus linearis, which both inhabit subtropical regions in East Asia, are recognized as highly destructive soybean pests. Many previous studies have investigated the biological characteristics, pheromones, chemicals and control mechanisms of these two pests, but few studies have explored their phylogeographic patterns and underlying factors. In this study, we generated a double-digest restriction site-associated DNA sequencing (ddRAD-seq) dataset to investigate phylogeographic patterns and construct ecological niche models (ENM) for both Riptortus species. Our findings revealed similar niche occupancies and population genetic structures between the two species, with each comprising two phylogeographic lineages (i.e., the mainland China and the Indochina Peninsula clades) that diverged approximately 0.1 and 0.3 million years ago, respectively. This divergence likely resulted from the combined effects of temperatures variation and geographical barriers in the mountainous regions of Southwest China. Further demographic history and ENM analyses suggested that both pests underwent rapid expansion prior to the Last Glacial Maximum (LGM). Furthermore, ENM predicts a northward shift of both pests into new soybean-producing regions due to global warming. Our study indicated that co-distribution soybean pests with overlapping ecological niches and similar life histories in subtropical regions of East Asia exhibit congruent phylogeographic and demographic patterns in response to shared historical biogeographic drivers.

Phylogenetic insights into Central European Chorthippus and Pseudochorthippus (Orthoptera: Acrididae) species using ddRADseq data

The evolution of several orthopteran groups, especially within the grasshopper family Acrididae, remains poorly understood. This is particularly true for the subfamily Gomphocerinae, which comprises cryptic sympatric and syntopic species. Previous mitochondrial studies have highlighted major discrepancies between taxonomic and phylogenetic hypotheses, thereby emphasizing the necessity of genome-wide approaches.In this study, we employ double-digest restriction site-associated DNA sequencing (ddRADseq) to reconstruct the evolution of Central European Chorthippus and Pseudochorthippus species, especially C.smardai, P.tatrae and the C.biguttulus group.Our phylogenomic analyses recovered deep discordance with mitochondrial DNA barcoding, emphasizing its unreliability in Gomphocerinae grasshoppers. Specifically, our data robustly distinguished the C.biguttulus group and confirmed the distinctiveness of C.eisentrauti, also shedding light on its presence in the Berchtesgaden Alps. Moreover, our results support the reclassification of C.smardai to the genus Pseudochorthippus and of P.tatrae to the genus Chorthippus.Our study demonstrates the efficiency of high-throughput genomic methods such as RADseq without prior optimization to elucidate the complex evolution of grasshopper radiations with direct taxonomic implications. While RADseq has predominantly been utilized for population genomics and within-genus phylogenomics, its application extends to resolve relationships between deeply-diverged clades representative of distinct genera.

DdRAD Sequencing and DNA Barcoding.

Double-digest restriction site-associated DNA sequencing is a library preparation protocol that enables capturing variable sites across the genome including single-nucleotide polymorphisms (SNPs). These SNPs can be utilized to gain evolutionary insights into patterns observed in DNA barcodes, to infer population structure and phylogenies, to detect gene flow and introgression, and to perform species delimitation analyses. The protocol includes chemically shearing genomic DNA with restriction enzymes, unique tagging, size selection, and amplification of the resulting DNA fragments. Here we provide a detailed description of each step of the protocol, as well as information on essential equipment and common issues encountered during laboratory work.

Rapid seasonal changes in phenotypes in a wild Drosophila population

Seasonal environmental change is one of the most rapid and striking environmental variables. Although relatively rapid adaptation to environmental changes over several years or several decades has been described in many taxa, rapid responses to seasonal environments are delicate, and therefore, the detection of the evolutionary responses requires sensitive methods. In this study, we examined seasonal changes in phenotypes related to thermal tolerance and morphological traits of Drosophila lutescens collected at the spring and autumn periods from a single location. We first demonstrated that flies in the two seasonal periods were almost genetically identical using double-digest restriction site-associated DNA sequencing and analysis. Using an experimental design to eliminate the effect of possible confounding factors that influence phenotypes (i.e., maternal effects and the environmental conditions in which each phenotype was analyzed), we showed that the heat tolerance of D. lutescens was significantly higher in the autumn population than in the spring population. Furthermore, cold tolerance was slightly higher in the spring population than in the autumn one. Although wing length and thorax length did not change significantly between seasons, the ratio of wing length to thorax length changed significantly between them. These results suggest that seasonal environmental heterogeneity induces rapid phenotypic changes within a year. Finally, we discuss the possibility of rapid evolutionary responses to seasonal changes.

Optimizing ddRAD sequencing for population genomic studies with ddgRADer.

Double-digest Restriction-site Associated DNA sequencing (ddRADseq) is widely used to generate genomic data for non-model organisms in evolutionary and ecological studies. Along with affordable paired-end sequencing, this method makes population genomic analyses more accessible. However, multiple factors should be considered when designing a ddRADseq experiment, which can be challenging for new users. The generated data often suffer from substantial read overlaps and adaptor contamination, severely reducing sequencing efficiency and affecting data quality. Here, we analyse diverse datasets from the literature and carry out controlled experiments to understand the effects of enzyme choice and size selection on sequencing efficiency. The empirical data reveal that size selection is imprecise and has limited efficacy. In certain scenarios, a substantial proportion of short fragments pass below the lower size-selection cut-off resulting in low sequencing efficiency. However, enzyme choice can considerably mitigate inadvertent inclusion of these shorter fragments. A simple model based on these experiments is implemented to predict the number of genomic fragments generated after digestion and size selection, number of SNPs genotyped, number of samples that can be multiplexed and the expected sequencing efficiency. We developed ddgRADer - http://ddgrader.haifa.ac.il/ - a user-friendly webtool and incorporated these calculations to aid in ddRADseq experimental design while optimizing sequencing efficiency. This tool can also be used for single enzyme protocols such as Genotyping-by-Sequencing. Given user-defined study goals, ddgRADer recommends enzyme pairs and allows users to compare and choose enzymes and size-selection criteria. ddgRADer improves the accessibility and ease of designing ddRADseq experiments and increases the probability of success of the first population genomic study conducted in labs with no prior experience in genomics.

Genomic signatures of ecological divergence between savanna and forest populations of a Neotropical tree.

In eastern Neotropical South America, the Cerrado, a large savanna vegetation, and the Atlantic Forest harbour high biodiversity levels, and their habitats are rather different from each other. The biomes have intrinsic evolutionary relationships, with high lineage exchange that can be attributed, in part, to a large contact zone between them. The genomic study of ecotypes, i.e. populations adapted to divergent habitats, can be a model to study the genomic signatures of ecological divergence. Here, we investigated two ecotypes of the tree Plathymenia reticulata, one from the Cerrado and the other from the Atlantic Forest, which have a hybrid zone in the ecotonal zone of Atlantic Forest-Cerrado. The ecotypes were sampled in the two biomes and their ecotone. The evolutionary history of the divergence of the species was analysed with double-digest restriction site-associated DNA sequencing. The genetic structure and the genotypic composition of the hybrid zone were determined. Genotype-association analyses were performed, and the loci under putative selection and their functions were investigated. High divergence between the two ecotypes was found, and only early-generation hybrids were found in the hybrid zone, suggesting a partial reproductive barrier. Ancient introgression between the Cerrado and Atlantic Forest was not detected. The soil and climate were associated with genetic divergence in Plathymenia ecotypes and outlier loci were found to be associated with the stress response, with stomatal and root development and with reproduction. The high genomic, ecological and morphophysiological divergence between ecotypes, coupled with partial reproductive isolation, indicate that the ecotypes represent two species and should be managed as different evolutionary lineages. We advise that the forest species should be re-evaluated and restated as vulnerable. Our results provide insights into the genomic mechanisms underlying the diversification of species across savanna and forest habitats and the evolutionary forces acting in the species diversification in the Neotropics.

Retained chromosomal integrity following CRISPR-Cas9-based mutational correction in human embryos

Human germline gene correction by targeted nucleases holds great promise for reducing mutation transmission. However, recent studies have reported concerning observations in CRISPR-Cas9-targeted human embryos, including mosaicism and loss of heterozygosity (LOH). The latter has been associated with either gene conversion or (partial) chromosome loss events. In this study, we aimed to correct a heterozygous basepair substitution in PLCZ1, related to infertility. In 36% of the targeted embryos that originated from mutant sperm, only wild-type alleles were observed. By performing genome-wide double-digest restriction site-associated DNA sequencing, integrity of the targeted chromosome (i.e., no deletions larger than 3 Mb or chromosome loss) was confirmed in all seven targetedGENType-analyzed embryos (mutant editing and absence of mutation), while short-range LOH events (shorter than 10 Mb) were clearly observed by single-nucleotide polymorphism assessment in two of these embryos. These results fuel the currently ongoing discussion on double-strand break repair in early human embryos, making a case for the occurrence of gene conversion events or partial template-based homology-directed repair.

Construction of a high-density genetic linkage map and QTL mapping for bioenergy-related traits in sweet sorghum [Sorghum bicolor (L.) Moench

Sorghum is an important but arguably undervalued cereal crop, grown in large areas in Asia and Africa due to its natural resilience to drought and heat. There is growing demand for sweet sorghum as a source of bioethanol as well as food and feed. The improvement of bioenergy-related traits directly affects bioethanol production from sweet sorghum; therefore, understanding the genetic basis of these traits would enable new cultivars to be developed for bioenergy production. In order to reveal the genetic architecture behind bioenergy-related traits, we generated an F2 population from a cross between sweet sorghum cv. 'Erdurmus' and grain sorghum cv. 'Ogretmenoglu'. This was used to construct a genetic map from SNPs discovered by double-digest restriction-site associated DNA sequencing (ddRAD-seq). F3 lines derived from each F2 individual were phenotyped for bioenergy-related traits in two different locations and their genotypes were analyzed with the SNPs to identify QTL regions. On chromosomes 1, 7, and 9, three major plant height (PH) QTLs (qPH1.1, qPH7.1, and qPH9.1) were identified, with phenotypic variation explained (PVE) ranging from 10.8 to 34.8%. One major QTL (qPJ6.1) on chromosome 6 was associated with the plant juice trait (PJ) and explained 35.2% of its phenotypic variation. For fresh biomass weight (FBW), four major QTLs (qFBW1.1, qFBW6.1, qFBW7.1, and qFBW9.1) were determined on chromosomes 1, 6, 7, and 9, which explained 12.3, 14.5, 10.6, and 11.9% of the phenotypic variation, respectively. Moreover, two minor QTLs (qBX3.1 and qBX7.1) of Brix (BX) were mapped on chromosomes 3 and 7, explaining 8.6 and 9.7% of the phenotypic variation, respectively. The QTLs in two clusters (qPH7.1/qBX7.1 and qPH7.1/qFBW7.1) overlapped for PH, FBW and BX. The QTL, qFBW6.1, has not been previously reported. In addition, eight SNPs were converted into cleaved amplified polymorphic sequences (CAPS) markers, which can be easily detected by agarose gel electrophoresis. These QTLs and molecular markers can be used for pyramiding and marker-assisted selection studies in sorghum, to develop advanced lines that include desirable bioenergy-related traits.

A community genomics approach to natural hybridization.

Hybridization is a complicated, oft-misunderstood process. Once deemed unnatural and uncommon, hybridization is now recognized as ubiquitous among species. But hybridization rates within and among communities are poorly understood despite the relevance to ecology, evolution and conservation. To clarify, we examined hybridization across 75 freshwater fish communities within the Ozarks of the North American Interior Highlands (USA) by single nucleotide polymorphism (SNP) genotyping 33 species (N = 2865 individuals; double-digest restriction site-associated DNA sequencing (ddRAD)). We found evidence of hybridization (70 putative hybrids; 2.4% of individuals) among 18 species-pairs involving 73% (24/33) of study species, with the majority being concentrated within one family (Leuciscidae/minnows; 15 species; 66 hybrids). Interspecific genetic exchange-or introgression-was evident from 24 backcrossed individuals (10/18 species-pairs). Hybrids occurred within 42 of 75 communities (56%). Four selected environmental variables (species richness, protected area extent, precipitation (May and annually)) exhibited 73-78% accuracy in predicting hybrid occurrence via random forest classification. Our community-level assessment identified hybridization as spatially widespread and environmentally dependent (albeit predominantly within one diverse, omnipresent family). Our approach provides a more holistic survey of natural hybridization by testing a wide range of species-pairs, thus contrasting with more conventional evaluations.

The Genetic Diversity of Mink (Neovison vison) Populations in China.

The American mink (Neovison vison) is a semiaquatic species of Mustelid native to North America that is now widespread in China. However, the knowledge of genetic diversity of mink in China is still limited. In this study, we investigated the genetic diversity and identified significant single nucleotide polymorphisms (SNPs) in mink populations of five different color types in three different mink farms in China. Using double-digest restriction site-associated DNA sequencing, we identified a total of 1.3 million SNPs. After filtering the SNPs, phylogenetic tree, Fst, principal component, and population structure analyses were performed. The results demonstrated that red mink and black mink grouped, with separate clustering of all other color types. The population divergence index (Fst) study confirmed that different mink populations were distinct (K = 4). Two populations with different coat colors were subjected to the selection signature analysis, and 2300 genes were found to have a clear selection signature. The genes with a selection signature were subjected to Gene Ontology (GO) categorization and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis, the results revealed that the genes with a selection signature were enriched in the melanogenesis pathway. These study's findings have set the stage for improved breeding and conservation of genetic resources in real-world practical mink farming.

Genetic Assessment of Fertile F1 Hybrids between Two Hercules Beetles, Dynastes maya Hardy and D. grantii Horn (Scarabaeidae).

Although hybridization may complicate taxonomic practices, it can be common between animal species. Animal hybridization not only can help with generating phenotypic and species diversity in nature, but also with understanding the genetic and genomic basis of phenotypic evolution in the laboratory. We assessed the genetic composition of captive bred F1 hybrids between two Hercules beetle species using mitochondrial CO1 and nuclear loci from a double-digest restriction-site associated DNA sequencing (ddRADseq) library. We showed that the F1 hybrids were genetically clustered with samples from the maternal species, D. grantii, based on CO1 data. Nuclear genome data, on the other hand, clearly showed that the F1 individuals were genetically intermediate between D. maya, the paternal species, and D. grantii, based on a principal component analysis. Our results also revealed that sampling design may have a major impact on the inferred genetic structure and hybrid individuals using ddRADseq data sets. We discuss the importance and potential from studying the genomics of this hybrid progeny in terms of understanding the origin and maintenance of both intraspecific and interspecific phenotypic divergence and convergence.

Spotted-wing drosophila, Drosophila suzukii (Diptera: Drosophilidae), exhibits large-scale spatial genetic structure across Canada but not fruit host–associated genetic structure

Abstract Spotted-wing drosophila, Drosophila suzukii, is a global pest of soft fruits that is capable of reproducing on a wide range of cultivated and wild plant species. In Canada, D. suzukii was first reported in British Columbia in 2009 and is now widespread across the country. Understanding the genetic structure of D. suzukii populations could be important for pest management if there are phenotypic differences between genetically distinct populations. For example, insect pest populations could respond differently to directional selection imposed by insecticides, differ in their host plant preferences, and vary in their susceptibility to biological control agents. Here, we used double-digest restriction site–associated DNA sequencing to examine large- and fine-scale patterns of the genetic structure of D. suzukii reared from fruit hosts in Canada. We found that this species has a large-scale spatial genetic structure; the flies collected formed two distinct genetic clusters, one of which was distinct to western Canada and the other to eastern Canada. At the local scale, D. suzukii populations showed no evidence of host-associated structuring in British Columbia, suggesting that pest management tactics may be best applied at the landscape level. Our results highlight the need to investigate phenotypic differences between western and eastern D. suzukii populations in Canada.

Detecting a novel locus associated with pellicle peelability in Japanese chestnut populations preselected for a major peelability gene

• The distribution of phenotypic values of peelability in Japanese chestnut populations preselected for a major peelability gene was biased toward maximum values. • Use of rank-based inverse normal transformation and categorization of peelability phenotypes was applied for GWAS to further identify QTLs associated with peelability. • A single QTL for peelability was identified at CCR1.0F_56177061 on chromosome F. Peelability is one of the most important traits in Japanese chestnut ( Castanea crenata Sieb. et Zucc.) breeding. Previous studies revealed that the peelability of Japanese chestnut is controlled by a major recessive peeling gene (MRPG). This locus has been used effectively for marker-assisted selection in chestnut breeding programs, but there are also likely to be minor loci that would further improve the efficiency of breeding for peelability. We conducted a genome-wide association study (GWAS) based on a variational approximation version of BayesB using 27 cultivars and MRPG-preselected populations consisting of 520 F 1 seedlings from 22 families. Using double-digest restriction site-associated DNA sequencing (ddRAD-Seq), we obtained 9164 tag single-nucleotide polymorphisms. Because the phenotypic values of peelability were distributed very asymmetrically, with a peak at the high end due to the preselection for MRPG, we converted the original data set to both inverse normal transformed values and categorical scores, and we analyzed both sets of converted data by GWAS. In both analyses, a quantitative trait locus (QTL) for peelability at CCR1.0F_56177061 was identified with posterior probability values of 0.68 and 1.00, respectively. The average values of genotypes at CCR1.0F_56177061 for cultivars and seedlings in this study were 60.4% (peelability rate) for CC, 63.7% for CT, and 86.5% for TT. This analysis reveals a significant association between CCR1.0F_56177061 genotype and peelability, indicating that this newly identified QTL will be useful for improving peelability in chestnut breeding programs.

Forest cover and geographical distance influence fine-scale genetic structure of leaf-toed geckos in the tropical dry forests of western Mexico

Abstract The biodiversity within tropical dry forests (TDFs) is astounding and yet poorly catalogued due to inadequate sampling and the presence of cryptic species. In the Mexican TDF, endemic species are common, and the landscape has been continually altered by geological and anthropogenic changes. To understand how landscape and environmental variables have shaped the population structure of endemic species, we studied the recently described species of leaf-toed gecko, Phyllodactylus benedettii, in coastal western Mexico. Using double-digest restriction site-associated DNA sequencing data, we first explore population structure and estimate the number of ancestral populations. The results indicate a high degree of genetic structure with little admixture, and patterns corresponding to both latitudinal and altitudinal gradients. We find that genetic structure cannot be explained purely by geographical distance, and that ecological corridors may facilitate dispersal and gene flow. We then model the spatial distribution of P. benedettii in the TDF through time and find that the coastline has been climatically suitable for the species since the Last Glacial Maximum. Landscape genetic analyses suggest that the combination of isolation by distance (IBD) and isolation by resistance (IBR; forest cover) has influenced the spatial genetic structure of the species. Overall, our genomic data demonstrate fine-scale population structure in TDF habitat, a complex colonization history, and spatial patterns consistent with both IBD and other ecological factors. These results further highlight the Mexican TDF as a diversity hotspot and suggest that continued anthropogenic changes are likely to affect native fauna.

A genetic linkage map of the Pacific white shrimp (Litopenaeus vannamei): QTL mapping for low-temperature tolerance and growth-related traits and identification of the candidate genes

Low water temperature is a prevailing abiotic stress which adversely impacts global shrimp growth and production. In this study, a high-density genetic linkage map of L. vannamei was constructed using double-digest restriction site-associated DNA sequencing (ddRAD-seq) with an F1 full-sib family mapping panel comprising 148 progenies and 2 parents. A total of 2528 single-nucleotide polymorphisms (SNPs) were assigned to 48 linkage groups (LGs), with a total genetic map length of 3227.383 centimorgans (cM) and an inter-locus density of 0.783 cM/locus. The number of markers per LG ranged from 12 to 190, with a mean of 56. The length of LGs varied from 15.07 to 158.314 cM, with a mean length of 67.237 cM. Five quantitative trait loci (QTLs) for low-temperature tolerance with a logarithm of odds (LOD) threshold≥2.5 were detected, of which only one was identified based on a linkage group-wide significant LOD threshold. Notably, two growth-trait (body weight) related QTLs were located on LG36 with a genome-wide significant LOD threshold. According to the genome and relevant annotations of L. vannamei, a major growth-related candidate gene, tRNA methyltransferase 10 homolog A (TRMT10A), was identified from genomic scaffolds. Further analysis demonstrated that four linked missense mutation SNPs within TRMT10A were closely related to the body weight. Taken together, the constructed genetic linkage map under low-temperature stress provides a valuable reference for QTL mapping and future genomic studies. Its application can facilitate comparative mapping with other populations, as well as marker-assisted selection (MAS) breeding of L. vannamei.

Population genomics reveals a single semi-continuous population of a commercially exploited marine gastropod

Buccinum undatum is a commercially important marine gastropod with limited dispersal capabilities. Previous genetic studies utilising microsatellites and Double-digest Restriction-site Associated DNA sequencing (ddRADseq) provided evidence that B. undatum exhibits fine-scale genetic structure. Using ddRADseq, 128 individuals from the southern North Sea, English Channel, and the Irish Sea were genotyped using 7015 filtered single nucleotide polymorphisms (SNPs), 19 of which were identified as being under putative selection. Multiple genetic clustering methods – Discriminant analysis of principal components, Principal component analysis, and Sparse non-negative matrix factorisation,– were used to investigate population structure. Spatially explicit genetic structure was investigated using an Estimated Effective Migration Surface analysis and a Mantel correlogram. A single genetic population was found using neutral SNPs, with weak within-population structuring. Global FST was low (0.0046, p < 0.001), and pairwise FST estimates between sampling locations were between 0.0004 and 0.0224. There was a significant trend of isolation-by-distance across all sampling locations (r = 0.743, p < 0.001). Positive spatial autocorrelation indicated whelks located ≤ 50 km of one another were significantly more related than by chance (r = 0.12, p = 0.003), further emphasising the low dispersal capabilities of B. undatum. Finally, two barriers of lower-than-average dispersal were discovered; the Thames estuary and across the English Channel. Management implications are discussed for the sustainability of whelks from inshore fisheries.

Re-evaluating the Systematics of Dendrolycopodium Using Restriction-Site Associated DNA-Sequencing.

The genus Dendrolycopodium (Lycopodiaceae) includes four to five species across North America and East Asia. Species identification in Dendrolycopodium is difficult due to limited or inconsistent characters. In addition, plants with intermediate morphologies regularly occur, potentially indicative of interspecific hybridization. To determine the species relationships in Dendrolycopodium and investigate the existence of hybrids, we generated a draft genome assembly for D. obscurum and carried out double-digest restriction-site associated DNA sequencing (RADSeq) on 86 Dendrolycopodium specimens. Our sampling includes all the described species and 11 individuals with intermediate morphology. We find that the genus can be divided into four clades that largely correspond to the described taxa, as well as evidence of interspecific hybridization. Within these clades, our STRUCTURE analysis suggests that there are multiple finer subgroups, with evidence of hybridization and introgression between these subgroups. Given the limited availability of specimens collected from Asia, the status of the various Asian species remains uncertain and will require further study. In summary, our study confirms several hybrid relationships in Dendrolycopodium and provides a clear phylogenetic framework for future taxonomic revision.

Landscape Genomics Provides Evidence of Ecotypic Adaptation and a Barrier to Gene Flow at Treeline for the Arctic Foundation Species Eriophorum vaginatum.

Global climate change has resulted in geographic range shifts of flora and fauna at a global scale. Extreme environments, like the Arctic, are seeing some of the most pronounced changes. This region covers 14% of the Earth’s land area, and while many arctic species are widespread, understanding ecotypic variation at the genomic level will be important for elucidating how range shifts will affect ecological processes. Tussock cottongrass (Eriophorum vaginatum L.) is a foundation species of the moist acidic tundra, whose potential decline due to competition from shrubs may affect ecosystem stability in the Arctic. We used double-digest Restriction Site-Associated DNA sequencing to identify genomic variation in 273 individuals of E. vaginatum from 17 sites along a latitudinal gradient in north central Alaska. These sites have been part of 30 + years of ecological research and are inclusive of a region that was part of the Beringian refugium. The data analyses included genomic population structure, demographic models, and genotype by environment association. Genome-wide SNP investigation revealed environmentally associated variation and population structure across the sampled range of E. vaginatum, including a genetic break between populations north and south of treeline. This structure is likely the result of subrefugial isolation, contemporary isolation by resistance, and adaptation. Forty-five candidate loci were identified with genotype-environment association (GEA) analyses, with most identified genes related to abiotic stress. Our results support a hypothesis of limited gene flow based on spatial and environmental factors for E. vaginatum, which in combination with life history traits could limit range expansion of southern ecotypes northward as the tundra warms. This has implications for lower competitive attributes of northern plants of this foundation species likely resulting in changes in ecosystem productivity.

Genome-wide SNPs reveal the fine-scale population structure of Laodelphax striatellus in China using double-digest restriction site-associated DNA sequencing

The small brown planthopper (SBPH), Laodelphax striatellus (Fallén) is one of the most destructive rice pests and has caused serious economic losses in China. To clarify the genetic differentiation and population genetic structure of this insect pest, we investigated the genomic polymorphisms, genetic differentiation, and phylogeography of 31 SBPH populations from 28 sampling sites from three climatic zones of China using double-digest restriction site-associated DNA sequencing (ddRADseq). In total, 2,813,221,369 high-quality paired-end reads from 306 individuals and 1925 single nucleotide polymorphisms (SNPs) were obtained. Low levels of genetic diversity and significant genetic differentiation were observed among the SBPH populations, and three genetic clusters were detected in China. Neutrality tests and bottleneck analysis provided strong evidence for recent rapid expansion with a severe bottleneck in most populations. Our work provides new insights into the genetics of the SBPH and will contribute to the development of effective management strategies for this pest.