Non-coding Chloroplast Research Articles

Phylogeography of Coccoloba uvifera (Polygonaceae) Sampled across the Caribbean Basin

Coccoloba uvifera L. (seagrape) is a primarily dioecious neotropical tree species which often grows in the beach–forest transitional ecotone of coastal strand vegetation. We used five maternally inherited non-coding chloroplast regions to characterize the phytogeography of C. uvifera collected across the Caribbean Basin and Florida. Bayesian analysis revealed divergence between the Aruba–Trinidad–Tobago–Antigua–Jamaica island group and the continental Belize–Florida–US Virgin Islands (USVI) group at 1.78 million years before present (mybp), divergence between the Belize and Florida–USVI groups at 1.08 mybp, and a split of Antigua–Jamaica from Aruba–Trinidad–Tobago at 0.217 mybp. Haplotype network analysis supports the three clades, with the island group possessing the oldest haplotype. Based on geology and proximity, these clades correspond to South American (oldest), Central American, and North American (most recent). Coccoloba uvifera demographic expansion occurred during the Pleistocene epoch and peaked near the end of the last glacial maximum (ca. 0.026–0.019 mybp) when the global sea levels were 125 m lower than today. Our findings also reveal that tropical cyclones, which often impact coastal strand vegetation, did not affect genetic diversity. However, there was a positive association between latitude and the average number of substitutions, further enriching our understanding of the species’ phytogeography.

New Approach to the Systematics of the Section Psammiris (Iris, Iridaceae): What Does Chloroplast DNA Sequence Tell Us?

Iris sect. Psammiris comprises rhizomatous perennials distributed in the north temperate zone of Eurasia. The systematics of the section are currently based on morphology, and the phylogenetic relationships within it still remain unclear. In the framework of Iris systematics, we conducted molecular and morphological analyses of the currently recognized I. sect. Psammiris species to elucidate the taxonomic composition and relationships within the section. The phylogenetic reconstructions based on sequence variation of four noncoding chloroplast DNA regions support the monophyly of I. sect. Psammiris, which includes I. tigridia, while I. potaninii var. ionantha belongs to I. sect. Pseudoregelia. The proposed novel classification of I. sect. Psammiris recognizes three series: an autonymic series with I. humilis, I. bloudowii, and I. vorobievii and two unispecific series (I. ser. Potaninia with I. potaninii and I. ser. Tigridiae with I. tigridia). In addition, the taxonomic statuses of I. arenaria, I. ivanovae, I. kamelinii, I. mandshurica, I. pineticola, I. psammocola, and I. schmakovii are clarified herein. We provide a revised taxonomic treatment for I. sect. Psammiris, including notes on the types; updated information on species synonymy, distributions, habitats, and chromosome numbers; and a new identification key to the species. Three lectotypes are designated here.

Recircumscription of Utricularia leptorhyncha and U. lasiocaulis and three related new species for northern Australia

The Utricularia lasiocaulis F.Muell. complex (subg. Polypompholyx; sect. Lasiocaules) is a morphologically and ecologically variable group of closely related taxa with a mostly tropical distribution across northern Australia. A recent molecular phylogenetic study placed the recognised species U. kamienskii F.Muell. sister to a clade of accessions relegated to either U. leptorhyncha O. Schwarz or U. lasiocaulis, with the latter previously circumscribed as an assemblage of highly variable morphological forms. We have expanded the previous study to include populations representing the known distributions of all three species and have attempted to include all morphological variants; 55 ingroup accessions were used in the full phylogenetic analysis based on two non-coding chloroplast regions (rps16, trnD–T) and the nuclear ribosomal internal transcribed spacer (ITS). We found that the pink corolla form of the usually white flowered U. kamienskii is not sufficiently different, and we retain it under that species. We also found strong support for a paraphyletic U. leptorhyncha, with the smaller flowered accessions matching the type material placed sister to all other U. lasiocaulis forms, including a grouping previously assigned as a larger flowered U. leptorhyncha. Within the U. lasiocaulis clade we found that much of the variation sorts into well-supported clades that we find are sufficiently morphologically and genetically differentiated from the type clade for recognition at the specific rank, namely Utricularia brennanii R.W.Jobson & Baleeiro from Northern Territory and Queensland, Utricularia cowiei R.W.Jobson & Baleeiro from the Kimberley region of Western Australia, and U. disjuncta R.W.Jobson & Baleeiro restricted to the Darwin and Gulf region of the Northern Territory.

Diversity among rare and common congeneric plant species from the Garry oak and Okanagan shrub-steppe ecosystems in British Columbia: implications for conservation

It is often assumed that the northern peripheral populations of species’ ranges are genetically depauperate due in part to founder effects from postglacial colonization. The majority of federally protected plant species are peripheral in Canada, yet we have little information about their patterns of genetic diversity and structure. In British Columbia, the majority of these protected plant species occur in two threatened habitats: the Garry oak and Okanagan shrub-steppe ecosystems. Using universal noncoding chloroplast DNA markers, we investigated genetic diversity and genetic structure in four rare and common plant species pairs inhabiting these two ecosystems. We found that rare species had lower genetic diversity than their common congeners, and detected contrasting patterns of regional diversity and structure based on ecosystem. Species from the Garry oak ecosystem showed lower genetic diversity in the northern deglaciated region and significant differentiation between regions, likely due to limited dispersal between Vancouver Island and the mainland. Species from the Okanagan shrub-steppe, however, tended to have uniform diversity across their range and lack regional structure. This study provides an important first look at the phylogeographic patterns of four rare plant species in British Columbia.

The nuanced nature of mesic refugia in arid landscapes: a tale of two peas.

Understanding how genetic diversity is distributed and maintained within species is a central tenet of evolutionary and conservation biology, yet is understudied in arid regions of the globe. In temperate, glaciated environments, high genetic diversity in plant species is frequently found in refugial areas, which are often associated with southern non-glaciated landscapes. In arid, unglaciated environments, landscape features providing mesic conditions are likely to be refugia, although our understanding needs more refinement in these biomes. We test whether refugia and nuclear diversity hotspots occur in high-elevation, topographically complex areas for co-distributed shrubs (Petalostylis labicheoides and Indigofera monophylla; Fabaceae) in the ancient, arid Pilbara bioregion of north-western Australia. We conducted extensive sampling of the Pilbara (>1400 individuals from 62 widespread populations) to detect patterns in nuclear diversity and structure based on 13-16 microsatellite loci. Evidence of historical refugia was investigated based on patterns of diversity in three non-coding chloroplast (cp) sequence regions for approx. 240 individuals per species. Haplotype relationships were defined with median-joining networks and maximum likelihood phylogenetic trees. We found cpDNA evidence for a high-elevation refugium in P. labicheoides but not for I. monophylla that instead exhibited extraordinary haplotype diversity and evidence for persistence across a widespread area. Nuclear diversity hotspots occurred in, but were not exclusive to, high-elevation locations and extended to adjacent, low-elevation riparian areas in both species. Phylogeographic refugia in arid environments may occur in high-elevation areas for some species but not all, and may be influenced by species-specific traits: a mesic montane refugium in P. labicheoides could be related to its preference for growth in water-gaining areas, while a lack of such evidence in I. monophylla could be related to maintenance of cpDNA diversity in a large soil seed bank and dynamic evolutionary history. Mesic environments created by the intersection of topographically complex landscapes with riparian zones can be contemporary reservoirs of genetic diversity in arid landscapes.

The role of seaweed extract on improvement drought tolerance of wheat revealed by osmoprotectants and DNA (cpDNA) markers

Drought stress is one of the most severe abiotic stresses affecting adversely plant growth, crop production, and various metabolic processes. Using seaweed extract in mitigating water stress adverse effects is highly important for plant production. The present study discussed the physiological role of seaweed extract (Sargassum denticulatum) in improving wheat tolerance to water stress.Water stress (40% of field capacity) caused significant decreases in wheat plant growth parameters (shoot height, fresh, and dry weights of the shoot) as well as with significant decreases in chlorophyll content and starch. Total soluble sugars, free amino acids, proline, and phenolic compounds contents increased in stressed wheat plants irrigated every three weeks compared with control plants. The foliar application of seaweed extract 2% enhanced all growth and yield parameters and more accumulation of the organic solutes in leaves of water-stressed plants. These increases correlated with significant increases in total phenolic contents as compared with control plants. The trnL intron and psbA-trnH intergenic regions of cpDNA were amplified from extracted total genomic DNA. The results indicated that the variation among psbA-trnH intergenic region was more than trnL intron region to distinct the variation of wheat treatments as responsible to water deficit.Foliar spray of seaweeds extract was effective in improving wheat performance by enhancing compatible osmolytes, antioxidant compounds and enhancing variation among non-coding chloroplast DNA (cpDNA) regions trnL intron and psbA-tnH as a response to water deficit.

Substitution rate heterogeneity across hexanucleotide contexts in noncoding chloroplast DNA.

Substitutions between closely related noncoding chloroplast DNA sequences are studied with respect to the composition of the 3 bases on each side of the substitution, that is the hexanucleotide context. There is about 100-fold variation in rate, among the contexts, particularly on substitutions of A and T. Rate heterogeneity of transitions differs from that of transversions, resulting in a more than 200-fold variation in the transitions: transversion bias. The data are consistent with a CpG effect, and it is shown that both the A + T content and the arrangement of purines/pyrimidines along the same DNA strand are correlated with rate variation. Expected equilibrium A + T content ranges from 36.4% to 82.8% across contexts, while G–C skew ranges from −77.4 to 72.2 and A–T skew ranges from −63.9 to 68.2. The predicted equilibria are associated with specific features of the content of the hexanucleotide context, and also show close agreement with the observed context-dependent compositions. Finally, by controlling for the content of nucleotides closer to the substitution site, it is shown that both the third and fourth nucleotide removed on each side of the substitution directly influence substitution dynamics at that site. Overall, the results demonstrate that noncoding sites in different contexts are evolving along very different evolutionary trajectories and that substitution dynamics are far more complex than typically assumed. This has important implications for a number of types of sequence analysis, particularly analyses of natural selection, and the context-dependent substitution matrices developed here can be applied in future analyses.

Genetic Differentiation among Subspecies of Banksia nivea (Proteaceae) Associated with Expansion and Habitat Specialization

Subspecies are traditionally defined using phenotypic differences associated with different geographical areas. Yet patterns of morphological and genetic variation may not coincide and thereby fail to reflect species’ evolutionary history. The division of the shrub Banksia nivea Labill. into one widespread (B. nivea subsp. nivea) and two geographically localized subspecies (B. nivea subsp. uliginosa (A.S. George) A.R. Mast & K.R. Thiele and B. nivea subsp. Morangup (M. Pieroni 94/2)) in south-west Australia has been based mainly on variation in leaf shape and pistil length, although flowering time and habitat differences are also evident, and subsp. uliginosa occurs on a different substrate. To assess the genetic divergence of B. nivea subspecies, we genotyped representatives from each subspecies for nuclear microsatellite and non-coding chloroplast sequence variation. We used distance and parsimony-based methods to assess genetic relatedness. Patterns were consistent with the existing taxonomy of subsp. nivea and uliginosa but not subsp. Morangup. Phylogenetic analyses revealed evidence for a more recent divergence of subsp. uliginosa associated with expansion from dryer sandy soils into the winter-wet ironstone soils in the southwest of Western Australia, consistent with progressive long-term climatic drying. Nuclear microsatellites showed low to moderate diversity, high population differentiation overall, and genetic structuring of subspecies in different biogeographical areas. We propose this pattern reflects the predicted impact of a patchy distribution, small populations, and restrictions to gene flow driven by both distance and biogeographic differences in subspecies’ habitats.

Juniperus excelsa s. str. in crimea – differentiation and history inferred from genetic and morphological markers

Abstract Juniperus excelsa s. str. (Greek juniper) in Crimea is a relic species on the limits of its range, and represents the Mediterranean flora in the Sub-Mediterranean part of the peninsula. Its origin and history in this area remains unresolved. We aimed to analyze phylogeny and potential demographic expansion of the juniper in the Crimea as well as to study its morphological differentiation. We analyzed plant material from 59 trees inhabiting eight populations. Genetic variation assessments were based on the four non-coding chloroplast DNA (cpDNA) fragments and the nuclear internal transcribed spacer region ITS1-5,8S-ITS2 (ITS). To examine the morphological differentiation, eight measured/counted traits of cones, seeds, and shoots were chosen and eight ratios were calculated. Morphological parameters were compared using ANOVA, Student’s t test, discrimination analysis and Kruskal-Wallis and U Mann-Whitney tests. Two cpDNA fragments were polymorphic and, in total, 10 cpDNA haplotypes were found. Haplotype diversity (Hd) ranged from 0.0 to 0.9. Based on both cpDNA and ITS sequences variation, phylogenetic analyses revealed a close relationship of the Crimean junipers to the individuals from other parts of the species range. In general, our molecular results confirmed the low level of genetic differentiation of J. excelsa individuals inhabiting different parts of the species range, likely resulting from a common ancestry. Only slight morphological differences were found between populations with different geographic location or habitat. The analyzes showed the distinctness of the populations from the southern part of the coast. Some unique morphological and molecular features of southern coastal populations imply that they are remnants of Late Pleistocene abundant forests. We suggest that the recent fragmentation of the Juniperus populations in the Crimean Peninsula could have arisen during the Atlantic period of the Holocene.

The transition to agricultural cultivation of neo-crops may fail to account for wild genetic diversity patterns: insights from the Cape Floristic Region.

AimThe global increase in the cultivation of native wild plants has raised concerns regarding potential risks associated with translocating genetic lineages beyond their natural range. This study aimed to investigate whether agricultural cultivation of neo-crops (a) accounts for the levels of genetic diversity present in wild populations, and whether (b) cultivated populations are genetically divergent from wild populations and thus pose a potential threat to wild genetic diversity.LocationThe Cape Floristic Region (CFR), located along the southern Cape of South Africa.MethodsHigh Resolution Melt analysis (HRM) coupled with Sanger sequencing was used to screen three non-coding chloroplast DNA loci in Cyclopia Vent. (Fabaceae), a CFR endemic neo-crop cultivated for the production of a herbal infusion referred to as Honeybush tea. Wild and cultivated populations for three of three widely cultivated Honeybush species (C. intermedia, C. longifolia, and C. subternata) were screened. Genetic diversity and differentiation were measured and compared between wild and cultivated groups.ResultsAcross all asseccions, a total of 17 haplotypes were detected, four of which were shared between wild and cultivated populations, while the remaining 13 were only detected in wild populations. Genetic diversity and differentiation was significantly higher in wild populations than in cultivated populations.ConclusionsIf no guidelines exist to facilitate the introduction of native wild plant taxa to a cultivated setting, wild genetic diversity patterns are likely to be compromised by cultivated populations. In the case presented here, cultivation represents a genetic bottleneck, failing to account for rare haplotypes, and may have disrupted species boundaries by initiating interspecific hybridization. More empirical work is required to evaluate the extent to which neo-crop cultivation poses a risk to wild genetic resources in the CFR and globally.

How co-distribution of two related azaleas (Rhododendron) developed in the Japanese archipelago: insights from evolutionary and demographic analyses

Plants growing in similar climates with extensively overlapping distributions may exhibit similar phylogeographic patterns, due to similarities in range shifts during past climatic oscillations. We tested and exploited this expectation in a comparison of the evolutionary and demographic histories of two related and co-distributed species endemic to Japan—Rhododendron pentaphyllum and R. quinquefolium. Genetic variation in sequences of noncoding chloroplast DNA regions and hundreds of nuclear single nucleotide polymorphisms were used to investigate 191 individuals from 13 populations of R. pentaphyllum and 142 individuals from 10 populations of R. quinquefolium. We found that the two species were not each other’s closest relative, and we found different times to the most recent common ancestor of extant populations based on both chloroplast DNA and nuclear SNPs. Rhododendron pentaphyllum exhibited higher genetic variation between populations than R. quinquefolium, while the two species had similar patterns of genetic divergence between regions with slightly different divergence times. These lines of evidence suggest that the similar ranges of the two species are the result of repeated range shifts since speciation during climatic oscillations.

Aegilops tauschii Coss. molecular phylogeny: nuclear gene Got2 versus chloroplast DNA data

Aegilops tauschii Coss. phylogenetic tree based on DNA sequences of nuclear gene Got2, about 3000 b.p., studied in 114 accessions of the species was constructed and compared with the tree previously built using non-coding chloroplast DNA (cpDNA), about 3000 b.p. in total, in 111 Ae. tauschii accessions. (110 accessions are the same on both trees.) The two trees differ greatly, implying that despite of Ae. tauschii being mostly self-pollinating species, cross pollination played an important role in the species evolutionary history. Got2 tree rather well corresponds to Ae. tauschii intraspecies systematics based on spike morphology, while cpDNA tree is in contradiction with this systematics. Ae. tauschii phylogeographic patterns essentially differ in Ae. tauschii subsp. tauschii and Ae. tauschii subsp. strangulata. As revealed by Got2 DNA data, subsp. strangulata presents five major common phylogenetic lineages sharing the subspecies range between each other, and minor relict lineages in addition. Subspecies tauschii presents single major lineage occupying all the subspecies range, and additional relict lineages.

Phylogeographic and demographic modeling analyses of the multiple origins of the rheophytic goldenrod Solidago yokusaiana Makino.

Understanding adaptation mechanisms is important in evolutionary biology. Parallel adaptation provides good opportunities to investigate adaptive evolution. To confirm parallel adaptation, it is effective to examine whether the phenotypic similarity has one or multiple origins and to use demographic modeling to consider the gene flow between ecotypes. Solidago yokusaiana is a rheophyte endemic to the Japanese Archipelago that diverged from Solidago virgaurea. This study examined the parallel origins of S. yokusaiana by distinguishing between multiple and single origins and subsequent gene flow. The haplotypes of noncoding chloroplast DNA and genotypes at 14 nuclear simple sequence repeat (nSSR) loci and single-nucleotide polymorphisms (SNPs) revealed by double-digest restriction-associated DNA sequencing (ddRADseq) were used for phylogeographic analysis; the SNPs were also used to model population demographics. Some chloroplast haplotypes were common to S. yokusaiana and its ancestor S. virgaurea. Also, the population genetic structures revealed by nSSR and SNPs did not correspond to the taxonomic species. The demographic modeling supported the multiple origins of S. yokusaiana in at least four districts and rejected a single origin with ongoing gene flow between the two species, implying that S. yokusaiana independently and repeatedly adapted to frequently flooding riversides.

Dispersal as a result of asymmetrical hybridization between two closely related oak species in China

Hybridization has played an important role in plant evolution. Less attention has been paid, however, to its role in dispersal. In this study, historical divergence and hybridization were investigated in two closely related Chinese oaks, Quercus mongolica and Q. liaotungensis, to estimate the role that hybridization played in their dispersal. We genotyped 27 Q. mongolica and Q. liaotungensis populations throughout the distributional range of the two oak species, using 14 single-copy nuclear genes and four noncoding chloroplast DNA regions. Bayesian cluster and population tree analyses indicated that there were three groups over all oak populations, namely, Q. mongolica, northwest–northern China (NW-NC) Q. liaotungensis, and northeastern China (NEC) Q. liaotungensis. Approximate Bayesian computation simulation supported an asymmetrical hybridization origin of NEC Q. liaotungensis, after a previous divergence between NW-NC Q. liaotungensis and Q. mongolica. IMa3 analyses suggested that Q. liaotungensis and Q. mongolica diverged in the NW-NC and NEC regions, respectively, and that NEC Q. liaotungensis arose from Q. mongolica, not from NW-NC Q. liaotungensis, and was greatly introgressed by NW-NC Q. liaotungensis. Oak populations in NW-NC and NEC regions held different chloroplast DNA haplotypes, and Q. liaotungensis in NEC shared most haplotypes with Q. mongolica populations, but none with NW-NC Q. liaotungensis populations, suggesting the maternal origin of NEC Q. liaotungensis from Q. mongolica. This study found clear signals of isolation divergence of Q. liaotungensis in NW-NC and Q. mongolica in NEC, and the results suggest that asymmetrical hybridization and introgression from Q. liaotungensis to Q. mongolica, mostly likely via pollen flow, facilitated Q. liaotungensis dispersal to NEC.

Insights from Chloroplast DNA into the Progenitor-Derivative Relationship Between Campanula punctata and C. takesimana (Campanulaceae) in Korea

Of the nearly 40 endemics present on Ulleung Island, Campanula takesimana Nakai represents an anagenetically derived lineage of a continental progenitor, C. punctata Lam. Other than its low morphological divergence, little is known about the genetic diversity and population structure of the species pair or the geographical origin of C. takesimana. We sampled a total of 240 accessions in 22 populations, including one Dokdo Island population, of the two species, sequenced four noncoding chloroplast regions (rps16-trnK, trnQ-rps16, psbD-trnT, and psbM-trnD; 4482 bp), and assessed the genetic consequences of anagenetic speciation. Based on chloroplast DNA, we found substantially lower genetic diversity statistics for C. takesimana compared to its progenitor, significant population genetic structuring in insular derivative species, and significant molecular divergence between C. punctata and C. takesimana. Mutually exclusive haplotypes were found in the two species, and the haplotype network suggested that Ulleung Island haplotypes were derived from the Dokdo Island haplotype, which was originally derived from a C. punctata population in Bonghwa-gun, Gyeongsangbuk-do Province. This study pinpoints a very narrow geographical source area and suggests the potentially important role of Dokdo Island as an initial stepping stone for Ulleung Island endemics.

Applied phylogeography of Cyclopia intermedia (Fabaceae) highlights the need for 'duty of care' when cultivating honeybush.

BackgroundThe current cultivation and plant breeding of Honeybush tea (produced from members of CyclopiaVent.) do not consider the genetic diversity nor structuring of wild populations. Thus, wild populations may be at risk of genetic contamination if cultivated plants are grown in the same landscape. Here, we investigate the spatial distribution of genetic diversity within Cyclopia intermedia E. Mey.—this species is widespread and endemic in the Cape Floristic Region (CFR) and used in the production of Honeybush tea.MethodsWe applied High Resolution Melt analysis (HRM), with confirmation Sanger sequencing, to screen two non-coding chloroplast DNA regions (two fragments from the atpI-aptH intergenic spacer and one from the ndhA intron) in wild C. intermedia populations. A total of 156 individuals from 17 populations were analyzed for phylogeographic structuring. Statistical tests included analyses of molecular variance and isolation-by-distance, while relationships among haplotypes were ascertained using a statistical parsimony network.ResultsPopulations were found to exhibit high levels of genetic structuring, with 62.8% of genetic variation partitioned within mountain ranges. An additional 9% of genetic variation was located amongst populations within mountains, suggesting limited seed exchange among neighboring populations. Despite this phylogeographic structuring, no isolation-by-distance was detected (p > 0.05) as nucleotide variation among haplotypes did not increase linearly with geographic distance; this is not surprising given that the configuration of mountain ranges dictates available habitats and, we assume, seed dispersal kernels.ConclusionsOur findings support concerns that the unmonitored redistribution of Cyclopia genetic material may pose a threat to the genetic diversity of wild populations, and ultimately the genetic resources within the species. We argue that ‘duty of care’ principles be used when cultivating Honeybush and that seed should not be translocated outside of the mountain range of origin. Secondarily, given the genetic uniqueness of wild populations, cultivated populations should occur at distance from wild populations that is sufficient to prevent unintended gene flow; however, further research is needed to assess gene flow within mountain ranges.

Interspecific Hybridization and Introgression Influence Biodiversity—Based on Genetic Diversity of Central European Viola epipsila-V. palustris Complex

The Viola epipsila-V. palustris complex is a highly taxonomically complicated group of species in its entire circumboreal range of distribution. Habitat loss, forest flooding, and hybridization could lead to the extinction of V. epipsila. A hybrid index and principal component analysis (PCA) were used to select qualitative and quantitative morphological features to distinguish parent species and hybrids, inter simple sequence repeat (ISSR) markers to determine the genetic diversity of the populations, flow cytometry to estimate the genome size (GS), and non-coding chloroplast DNA (cpDNA) regions to indicate the directions of crosses. All taxa are very morphologically variable, and their features can change within a season. The most stable feature is the distance of the bracts on the pedicel from the rhizome. The genetic diversity of all taxa populations is low and highly influenced by selfing and vegetative propagation. The population structure is differentiated: populations of V. epipsila or V. palustris, mixed populations with both parent species, F1 hybrids and populations with introgressive forms occur in different regions. The interspecific GS variation corresponds to the ploidy level (4x = 2.52 pg, 8x = 4.26 pg, 6x = 3.42 pg). Viola epipsila is the mother plant of the hybrids. Research has shown the risk of V. epipsila extinction in Central Europe and the importance of local populations in studying the role of hybridization in reducing/maintaining/increasing biodiversity.

Population Genetic Structure and Natural Establishment of Hybrids Between Sarracenia flava and Sarracenia minor in Francis Marion National Forest

We assessed population genetic structure and rate of hybridization in Sarracenia flava and S. minor in Francis Marion National Forest. The forest has an abundance of potential habitat for our study species and has suffered less human mediated disturbance than much of the county's longleaf pine savanna and wet pine savanna ecosystems. We examined 63 S. flava and 62 S. minor individuals, as well as one hybrid across a 486.02 km2 study site. We used eight nuclear microsatellite loci and one non-coding chloroplast marker to assess population genetic structure and describe the parentage of the hybrid. We found relatively little population genetic structure across many distinct field sites, even when those sites were separated by unsuitable habitat. In very small populations surviving in shrub-dominated longleaf pine savanna, genetic divergence was greater than in larger populations with more suitable habitat. The single hybrid discovered was an F1 hybrid for which S. flava served as the maternal parent. We found no signs of introgression. We hope that these data can help inform conservation decisions regarding Sarracenia species, as the low genetic structure seems to suggest a high degree of connectivity between geographically distinct populations of plants.

Application of High Resolution Melt analysis (HRM) for screening haplotype variation in a non-model plant genus: Cyclopia (Honeybush)

AimThis study has three broad aims: to (a) develop genus-specific primers for High Resolution Melt analysis (HRM) of members of Cyclopia Vent., (b) test the haplotype discrimination of HRM compared to Sanger sequencing, and (c) provide an example of using HRM to detect novel haplotype variation in wild C. subternata Vogel. populations.LocationThe Cape Floristic Region (CFR), located along the southern Cape of South Africa.MethodsPolymorphic loci were detected through a screening process of sequencing 12 non-coding chloroplast DNA segments across 14 Cyclopia species. Twelve genus-specific primer combinations were designed around variable cpDNA loci, four of which failed to amplify under PCR; the eight remaining were applied to test the specificity, sensitivity and accuracy of HRM. The three top performing HRM Primer combinations were then applied to detect novel haplotypes in wild C. subternata populations, and phylogeographic patterns of C. subternata were explored.ResultsWe present a framework for applying HRM to non-model systems. HRM accuracy varied across the PCR products screened using the genus-specific primers developed, ranging between 56 and 100%. The nucleotide variation failing to produce distinct melt curves is discussed. The top three performing regions, having 100% specificity (i.e. different haplotypes were never grouped into the same cluster, no false negatives), were able to detect novel haplotypes in wild C. subternata populations with high accuracy (96%). Sensitivity below 100% (i.e. a single haplotype being clustered into multiple unique groups during HRM curve analysis, false positives) was resolved through sequence confirmation of each cluster resulting in a final accuracy of 100%. Phylogeographic analyses revealed that wild C. subternata populations tend to exhibit phylogeographic structuring across mountain ranges (accounting for 73.8% of genetic variation base on an AMOVA), and genetic differentiation between populations increases with distance (p < 0.05 for IBD analyses).ConclusionsAfter screening for regions with high HRM clustering specificity—akin to the screening process associated with most PCR based markers—the technology was found to be a high throughput tool for detecting genetic variation in non-model plants.

&lt;p&gt;&lt;strong&gt;Genetic diversity and species limits in &lt;em&gt;Veltheimia&lt;/em&gt; (Asparagaceae: Scilloideae): insights from noncoding cpDNA sequence data&lt;/strong&gt;&lt;/p&gt;

Veltheimia is a genus of two species of bulbous flowering angiosperms restricted to southern Africa. Both parsimony and Bayesian analysis of 1829 nucleotides of DNA sequence data, from three non-coding chloroplast regions from multiple samples of both species, indicates that specimens of V. bracteata form a monophyletic clade together with three specimens named as V. capensis. This clade receives low support and is embedded within a paraphyletic grade of specimens of V. capensis. A Median Joining Network analysis of a subset of samples revealed nine haplotypes, the relationships of which mirrored the relationships resolved by the Bayesian analysis. With the exception of the V. capensis members of the V. bracteata clade, the remaining specimens are from the summer rainfall thicket and coastal forests. The samples of the basal V. capensis grade are found in the winter rainfall regions of southern Africa. These results indicate that there is limited genetic diversity within the genus, and there is no clear distinction between these two species based on plastid non-coding DNA data. This may be due to their recent divergence and incomplete lineage sorting, or recent or ongoing hybridisation and / or introgression.