In Vitro Regeneration from Leaf Explants of Helianthus verticillatus, a Critically Endangered Sunflower.

Helianthus verticillatus (Asteraceae), a whorled sunflower, is a perennial species restricted to a few locations in the southeastern United States and is now considered endangered. Therefore, restoring and protecting H. verticillatus as a species is a priority. This study introduces a highly efficient in vitro adventitious plant regeneration system from leaf explants, utilizing five diverse specimens of H. verticillatus, each representing distinct genotypes with phenotypic variations in leaf and stem morphology. Key factors influencing in vitro morphogenesis, including genetic constitution, explant source, and plant growth regulators (PGRs), were identified. The study revealed a remarkably strong genotype-dependent impact on the regeneration efficiency of the investigated H. verticillatus genotypes, ranging from a lack of regeneration to highly effective regeneration. The selection of two genotypes with varying regeneration abilities provides valuable models for genetic analyses, offering insights into factors influencing the regeneration potential of this endangered species. Optimum adventitious shoot regeneration results were achieved using Murashige and Skoog basal media (MS) supplemented with 8.8 µM N6-benzyladenine (BA) and 1.08 µM α-naphthalene acetic acid (NAA). This combination yielded the highest adventitious shoot production. Subsequent successful rooting on ½ MS medium without PGRs further solidified the efficiency of the developed protocol. Regenerated plantlets, demonstrating robust shoots and roots, were successfully acclimatized to greenhouse conditions with a 95% survival rate. The protocol developed in this study is the first such report for this endangered species and is expected to contribute to future genetic manipulation and modification studies.

A set of SSR markers to characterize genetic diversity in all Viburnum species.

About 160 species are classified within the Viburnum genus and many of these are cultivated for horticultural purposes. The vast dispersal of Viburnum makes the genus a useful model for studying evolutionary history and inferring how species expanded into their current distributions. Simple sequence repeat (SSR) markers were previously developed for five Viburnum species that were classified within the four major clades (Laminotinus, Crenotinus, Valvatotinus, and Porphyrotinus). The ability of some of these markers to cross-amplify in Viburnum species has been scantly evaluated, but there has not been any genus-wide assessment for the markers. We evaluated a collection of 49 SSR markers for the ability to cross-amplify in 224 samples, including 46 Viburnum species, representing all 16 subclades, and five additional species in the Viburnaceae and Caprifoliaceae. A subset of 14 potentially comprehensive markers for Viburnum species was identified and evaluated for the ability to detect polymorphisms in species outside of their respective clades. The 49 markers had overall amplification success in 52% of the samples, including a 60% success rate within the Viburnum genus and 14% in other genera. The comprehensive marker set amplified alleles in 74% of all samples tested, including 85% of Viburnum samples and 19% of outgroup samples. To the best of our knowledge, this is the first comprehensive set of markers able to characterize species across an entire genus. This set of markers can be used to assess the genetic diversity and population structure of most Viburnum species and closely allied species.

Development and Characterization of 20 Genomic SSR Markers for Ornamental Cultivars of Weigela.

Weigela (Caprifoliaceae) is a genus of ornamental plants popular for its phenotypic variation and hardiness, that includes species hybridized to produce the commercially available cultivars. Despite its popularity, limited genetic resources exist for the genus. Twenty genomic simple sequence repeat (gSSR) markers distributed across the genome were developed using low coverage whole-genome sequencing data of Weigela Spilled Wine®. A cross-amplification evaluation with these 20 gSSR markers on a collection of 18 Weigela cultivars revealed a total of 111 unique alleles, including 36 private alleles. A diagrammatic key was constructed to identify cultivars using only six of the gSSR markers, demonstrating the newly developed gSSR markers are immediately useful for cultivar identification. Future uses could include breeding with marker-assisted selection, determining the history of hybridization of the current cultivated lines, aiding in the construction of genetic maps, and assessing the patterns of population genetic structure of Weigela spp.

Microsatellite Loci Reveal High Genetic Diversity, Mutation, and Migration Rates as Invasion Drivers of Callery Pear (Pyrus calleryana) in the Southeastern United States.

Pyrus calleryana Decne. (Callery pear) is a deciduous tree native to China, Japan, Korea, and Taiwan. It is a popular ornamental tree in the United States (US) with early spring blooms and vibrant fall color. There are at least 26 cultivars of P. calleryana available in the US of which "Bradford" is the most well-known. Open-pollinated P. calleryana escapees are becoming one of the most common invasive tree species in the eastern United States. Developing better management practices for invasive P. calleryana requires detailed knowledge about reproductive biology and genetic diversity of the species, however, little is currently known about genetic variability within those open-pollinated populations. We investigated genetic diversity and population structure of non-cultivated, escaped P. calleryana populations within a ∼177 km radius in the southeastern United States. Because P. calleryana exhibits a range of morphological variation with great evolutionary potential, we hypothesized that a high genetic diversity would be manifested among escaped P. calleryana. Using 15 previously developed microsatellite loci, we genotyped 180 open-pollinated P. calleryana individuals that were collected across six naturally occurring sites in Tennessee, Georgia, and South Carolina, United States. Our results demonstrated the presence of a population structure with high genetic diversity, high gene flow, and high genetic differentiation between individuals across collection sites. Our results revealed that P. calleryana populations had differentiated shortly after the introduction to the US, most likely from specimens imported from Asia, consistent with historical records and our prior findings. The high invasive potential of the species is perhaps best underscored by transformation of P. calleryana specimens introduced from Asia into escape populations at continental scale across the United States. Our data also provided novel insight into potential issues that could be problematic for the future as P. calleryana may pose a potential threat to the economy, ecology, and native biodiversity in invaded areas.

Identification of two Eleusine indica (goosegrass) biotypes of cool-season turfgrass resistant to dithiopyr.

BACKGROUND: Turfgrass managers reported poor Eleusine indica control following applications of the mitosis-inhibiting herbicide dithiopyr in cool-season turfgrass. Field, glasshouse, and laboratory experiments were conducted to understand the response of these biotypes to dithiopyr and prodiamine. RESULTS: In field experiments at two locations with putative dithiopyr-resistant E. indica, preemergence applications of dithiopyr provided no E. indica control. Single applications of the protoporphyrinogen oxidase (PPO)-inhibitor, oxadiazon, provided > 85% control at these locations. When subjected to agar-based bioassays, root growth of putative resistant biotypes planted with 0.01 mmol L-1 dithiopyr was slightly reduced (< 25%) whereas roots were completely inhibited in the susceptible biotype. Glasshouse whole plant rate-response experiments found that the cytochrome P450 inhibitor, piperonyl butoxide (PBO), did not increase the sensitivity of these putative resistant biotypes to dithiopyr. Sequencing of α-tubulin 1 (TUA1) revealed a Leu-136-Phe substitution in both dithiopyr-resistant populations. CONCLUSION: Eleusine indica biotypes with resistance to dithiopyr are present in cool-season turfgrass systems in the United States. Resistance is possibly related to a single nucleotide polymorphism (SNP) of an α-tubulin gene. If turfgrass managers suspect resistance to dithiopyr, oxadiazon can still be an effective alternative for preemergence control. © 2021 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Microsatellite Markers from Peronospora tabacina, the Cause of Blue Mold of Tobacco, Reveal Species Origin, Population Structure, and High Gene Flow.

Peronospora tabacina is an obligate parasite that causes blue mold of tobacco. The pathogen reproduces primarily by sporangia, whereas the sexual oospores are rarely observed. A collection of 122 isolates of P. tabacina was genotyped using nine microsatellites to assess the population structure of individuals from subpopulations collected from central, southern, and western Europe; the Middle East; Central America; North America; and Australia. Genetic variations among the six subpopulations accounted for ∼8% of the total variation, including moderate levels of genetic differentiation, high gene flow among these subpopulations, and a positive correlation between geographic and genetic distance (r = 0.225; P < 0.001). Evidence of linkage disequilibrium (P < 0.001) showed that populations contained partially clonal subpopulations but that subpopulations from Australia and Mediterranean Europe did not. High genetic variation and population structure among samples could be explained by continuous gene flow across continents via infected transplant exchange and/or long-distance dispersal of sporangia via wind currents. This study analyzed the most numerous P. tabacina collection and allowed conclusions regarding the migration, mutation, and evolutionary history of this obligate biotrophic oomycete. The evidence pointed to the species origin in Australia and identified intracontinental and intercontinental migration patterns of this important pathogen.[Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Genetic diversity in North American Cercis Canadensis reveals an ancient population bottleneck that originated after the last glacial maximum.

Understanding of the present-day genetic diversity, population structure, and evolutionary history of tree species can inform resource management and conservation activities, including response to pressures presented by a changing climate. Cercis canadensis (Eastern Redbud) is an economically valuable understory tree species native to the United States (U.S.) that is also important for forest ecosystem and wildlife health. Here, we document and explain the population genetics and evolutionary history of this deciduous tree species across its distributed range. In this study, we used twelve microsatellite markers to investigate 691 wild-type trees sampled at 74 collection sites from 23 Eastern U.S. states. High genetic diversity and limited gene flow were revealed in wild, natural stands of C. canadensis with populations that are explained by two major genetic clusters. These findings indicate that an ancient population bottleneck occurred coinciding with the last glacial maximum (LGM) in North America. The structure in current populations likely originated from an ancient population in the eastern U.S. that survived LGM and then later diverged into two contemporary clusters. Data suggests that populations have expanded since the last glaciation event from one into several post-glacial refugia that now occupy this species' current geographic range. Our enhanced understanding benchmarks the genetic variation preserved within this species and can direct future efforts in conservation, and resource utilization of adaptively resilient populations that present the greatest genetic and structural diversity.

"Jumping Jack": Genomic Microsatellites Underscore the Distinctiveness of Closely Related Pseudoperonospora cubensis and Pseudoperonospora humuli and Provide New Insights Into Their Evolutionary Past.

Downy mildews caused by obligate biotrophic oomycetes result in severe crop losses worldwide. Among these pathogens, Pseudoperonospora cubensis and P. humuli, two closely related oomycetes, adversely affect cucurbits and hop, respectively. Discordant hypotheses concerning their taxonomic relationships have been proposed based on host-pathogen interactions and specificity evidence and gene sequences of a few individuals, but population genetics evidence supporting these scenarios is missing. Furthermore, nuclear and mitochondrial regions of both pathogens have been analyzed using microsatellites and phylogenetically informative molecular markers, but extensive comparative population genetics research has not been done. Here, we genotyped 138 current and historical herbarium specimens of those two taxa using microsatellites (SSRs). Our goals were to assess genetic diversity and spatial distribution, to infer the evolutionary history of P. cubensis and P. humuli, and to visualize genome-scale organizational relationship between both pathogens. High genetic diversity, modest gene flow, and presence of population structure, particularly in P. cubensis, were observed. When tested for cross-amplification, 20 out of 27 P. cubensis-derived gSSRs cross-amplified DNA of P. humuli individuals, but few amplified DNA of downy mildew pathogens from related genera. Collectively, our analyses provided a definite argument for the hypothesis that both pathogens are distinct species, and suggested further speciation in the P. cubensis complex.

Propagation Methods for the Conservation and Preservation of the Endangered Whorled Sunflower (Helianthus verticillatus).

Helianthus verticillatus Small, the whorled sunflower, is a perennial species only found at a few sites in the southeastern United States and was declared federally (USA) endangered in 2014. The species spreads locally via rhizomes and can produce copious seeds when sexually compatible genotypes are present. Vegetative propagation of the species via cuttings and the optimum conditions for seed germination have not been determined. To investigate asexual propagation via cuttings, stem sections were harvested in late May, June, and July in Knoxville, Tennessee (USA) and trimmed to a minimum of two nodes. The base of the cuttings was treated with either auxin or water, and grown in a Promix BX potting medium with intermittent mist and 50% shade for one month. Seeds were harvested from a population of multiple genotypes in Maryville, Tennessee and used to determine viability and the range of temperatures suitable for germination. A clonal population was developed and used for three years to assess sexual compatibility at three locations in Knoxville, Tennessee. Ninety-five percent of the cuttings from May rooted in two-to-three weeks and formed more than 20 adventitious roots per cutting with auxin and 18 with water treatments. The ability of cuttings to root decreased in June to about 20%, and none rooted in July with either water or auxin pretreatments. Pre-germination tetrazolium tests indicated that about 91% of seeds (achenes) were viable. Subsequent germination tests revealed high germination at varying temperatures (96 to 99% of seeds (achenes) germinated at 22/11, 27/15, and 29/19 °C), whereas germination was significantly inhibited by 33/24 °C. Fifty percent of the seeds germinated at 22/11 °C in 7.5 days, whereas only 2.0 to 2.5 days were required for 50% germination at 27/15, 29/19, and 33/24 °C. Seeds were not produced at any of the clonal planting locations during the three years. Vegetative propagation via rooted cuttings was successful in the mid-spring, seed germination was possible over a wide range of temperatures, and self-incompatibility was evident in this species. The results of this study will permit fast and efficient propagation of multiple and selected genotypes for conservation, commerce, and breeding of elite cultivars with disease resistance or other desirable attributes.

Microsatellite Loci Reveal Genetic Diversity of Asian Callery Pear (Pyrus calleryana) in the Species Native Range and in the North American Cultivars.

Pyrus calleryana Decne. (Callery pear) includes cultivars that in the United States are popular ornamentals in commercial and residential landscapes. Last few decades, this species has increasingly naturalized across portions of the eastern and southern US. However, the mechanisms behind this plant's spread are not well understood. The genetic relationship of present-day P.calleryana trees with their Asian P. calleryana forebears (native trees from China, Japan, and Korea) and the original specimens of US cultivars are unknown. We developed and used 18 microsatellite markers to analyze 147 Pyrus source samples and to articulate the status of genetic diversity within Asian P. calleryana and US cultivars. We hypothesized that Asian P. calleryana specimens and US cultivars would be genetically diverse and would show genetic relatedness. Our data revealed high genetic diversity, high gene flow, and presence of population structure in P. calleryana, potentially relating to the highly invasive capability of this species. Strong evidence for genetic relatedness between Asian P. calleryana specimens and US cultivars was also demonstrated. Our data suggest the source for P. calleryana that have become naturalized in US was China. These results will help understand the genetic complexity of invasive P. calleryana when developing management for escaped populations: In follow-up studies, we use the gSSRs developed here to analyze P. calleryana escape populations from across US.

Differential expression of genes associated with non-target site resistance in Poa annua with target site resistance to acetolactate synthase inhibitors.

BACKGROUND: Poa annua is a pervasive grassy, self-pollinating, weed that has evolved resistance to 10 different herbicide modes-of-action, third most of all weed species. We investigated constitutive overexpression of genes associated with non-target site resistance (NTSR) in POAAN-R3 and the response of those genes when treated with trifloxysulfuron despite the biotype having a known target site mutation in acetolactate synthase (ALS). RESULTS: Despite having an ALS target site mutation, POAAN-R3 still had a transcriptomic response to herbicide application that differed from a susceptible biotype. We observed differential expression of genes associated with transmembrane transport and oxidation-reduction activities, with differences being most pronounced prior to herbicide treatment. CONCLUSIONS: In the P. annua biotype we studied with confirmed target site resistance to ALS inhibitors, we also observed constitutive expression of genes regulating transmembrane transport, as well as differential expression of genes associated with oxidative stress after treatment with trifloxysulfuron. This accumulation of mechanisms, in addition to the manifestation of target site resistance, could potentially increase the chance of survival when plants are challenged by different modes of action.

Development and Characterization of 15 Novel Genomic SSRs for Viburnum farreri.

The Viburnum genus is of particular interest to horticulturalists, phylogeneticists, and biogeographers. Despite its popularity, there are few existing molecular markers to investigate genetic diversity in this large genus, which includes over 160 species. There are also few polymorphic molecular tools that can delineate closely related species within the genus. Viburnum farreri, a member of the Solenotinus subclade and one of the centers of diversity for Viburnum, was selected for DNA sequencing and development of genomic simple sequence repeats (gSSRs). In this study, 15 polymorphic gSSRs were developed and characterized for a collection of 19 V. farreri samples. Number of alleles per locus ranged from two- to- eight and nine loci had four or more alleles. Observed heterozygosity ranged from 0 to 0.84 and expected heterozygosity ranged from 0.10 to 0.80 for the 15 loci. Shannon diversity index values across these loci ranged from 0.21 to 1.62. The markers developed in this study add to the existing molecular toolkit for the genus and will be used in future studies investigating cross-transferability, genetic variation, and species and cultivar delimitation in the Viburnum genus and closely allied genera in the Adoxaceae and Caprifoliaceae.

Conventional Gel Electrophoresis and TaqMan Probes Enable Rapid Confirmation of Thousand Cankers Disease from Diagnostic Samples.

Thousand cankers disease (TCD) is caused by the fungal pathogen Geosmithia morbida and vectored by the walnut twig beetle Pityophthorus juglandis. In infected walnut and butternut (Juglans spp.) hosts and wingnut species (Pterocarya spp.) hosts, tree decline and death results in ecological disruption and economic losses. A rapid molecular detection protocol for TCD using microsatellite markers can confirm the presence of insect vector or fungal pathogen DNA, but it requires specialized expensive equipment and technical expertise. Using four different experimental approaches, capillary and conventional gel electrophoresis, and traditional polymerase chain reaction (PCR) and quantitative PCR (qPCR), we describe simplified and inexpensive processes for diagnostic confirmation of TCD. The improved and rapid detection protocols reported in this study reduce time and equipment costs associated with detection of molecular pest and pathogen DNA by (1) using conventional gel electrophoresis or TaqMan molecular probes to elucidate the detection limits for G. morbida and P. juglandis DNA and (2) identifying resources that allow visualization of positive test results for infected host plant tissue samples. Conventional gel electrophoresis and TaqMan molecular probe protocols detected presence of DNA from TCD-associated fungal and insect samples. These procedural improvements can be readily adopted by diagnostic end-users and adapted for use with other complex disease systems to enable rapid pest and pathogen detection.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Species diversity and phylogeography of Cornus kousa (Asian dogwood) captured by genomic and genic microsatellites.

Cornus kousa (Asian dogwood), an East Asia native tree, is the most economically important species of the dogwood genus, owing to its desirable horticultural traits and ability to hybridize with North America-native dogwoods. To assess the species genetic diversity and to better inform the ongoing and future breeding efforts, we assembled an herbarium and arboretum collection of 131 noncultivated C. kousa specimens. Genotyping and capillary electrophoresis analyses of our C. kousa collection with the newly developed genic and published nuclear genomic microsatellites permitted assessment of genetic diversity and evolutionary history of the species. Regardless of the microsatellite type used, the study yielded generally similar insights into the C. kousa diversity with subtle differences deriving from and underlining the marker used. The accrued evidence pointed to the species distinct genetic pools related to the plant country of origin. This can be helpful in the development of the commercial cultivars for this important ornamental crop with increased pyramided utility traits. Analyses of the C. kousa evolutionary history using the accrued genotyping datasets pointed to an unsampled ancestor population, possibly now extinct, as per the phylogeography of the region. To our knowledge, there are few studies utilizing the same gDNA collection to compare performance of genomic and genic microsatellites. This is the first detailed report on C. kousa species diversity and evolutionary history inference.

In Vitro Propagation of an Endangered Helianthus Verticillatus by Axillary Bud Proliferation.

Helianthus verticillatus (Asteraceae), whorled sunflower, is a perennial species restricted to a few locations in the Southeastern United States. Habitat loss has caused H. verticillatus to become rare, and since 2014, it has been federally listed as an endangered species. As a part of the recovery plan for the restoration and protection of H. verticillatus, an efficient micropropagation protocol based on axillary shoot proliferation was developed. Various concentrations of 6-benzylaminopurine (BAP; 0 to 4.44 µM) were examined for their morphogenetic potential in the regeneration of six genotypes of H. verticillatus from the nodal explants derived from greenhouse-grown plants. Both the BAP concentration and genotype had significant effects on the regeneration capacity of H. verticillatus. Although the induced buds were observed on ½-strength Murashige and Skoog medium without plant growth regulators, a higher rate of induction and bud development were achieved on media with either 0.88 or 2.22 µM BAP, regardless of the genotype. Successful rooting of the induced shoots was achieved within four weeks after the transfer from the induction medium to the fresh ½-strength MS medium, but the rooting efficiency was dependent on the plant's genetic background. Regenerated plantlets, with well-developed shoots and roots, were acclimatized successfully to greenhouse conditions with a 97% survival rate. Simple sequence repeats (SSRs) markers were employed to assess the genetic uniformity of the micropropagated plants of H. verticillatus. No extraneous bands were detected between regenerants and their respective donor plants, confirming the genetic fidelity and stability of regenerated plants. To our knowledge, the protocol developed in this study is the first such report for this endangered species.

Habitat fragmentation influences genetic diversity and differentiation: Fine-scale population structure of Cercis canadensis (eastern redbud).

Forest fragmentation may negatively affect plants through reduced genetic diversity and increased population structure due to habitat isolation, decreased population size, and disturbance of pollen-seed dispersal mechanisms. However, in the case of tree species, effective pollen-seed dispersal, mating system, and ecological dynamics may help the species overcome the negative effect of forest fragmentation. A fine-scale population genetics study can shed light on the postfragmentation genetic diversity and structure of a species. Here, we present the genetic diversity and population structure of Cercis canadensis L. (eastern redbud) wild populations on a fine scale within fragmented areas centered around the borders of Georgia-Tennessee, USA. We hypothesized high genetic diversity among the collections of C. canadensis distributed across smaller geographical ranges. Fifteen microsatellite loci were used to genotype 172 individuals from 18 unmanaged and naturally occurring collection sites. Our results indicated presence of population structure, overall high genetic diversity (H E = 0.63, H O = 0.34), and moderate genetic differentiation (F ST = 0.14) among the collection sites. Two major genetic clusters within the smaller geographical distribution were revealed by STRUCTURE. Our data suggest that native C. canadensis populations in the fragmented area around the Georgia-Tennessee border were able to maintain high levels of genetic diversity, despite the presence of considerable spatial genetic structure. As habitat isolation may negatively affect gene flow of outcrossing species across time, consequences of habitat fragmentation should be regularly monitored for this and other forest species. This study also has important implications for habitat management efforts and future breeding programs.

Low Genetic Diversity Suggests the Recent Introduction of Dogwood Powdery Mildew to North America.

Cornus florida (flowering dogwood) is a popular understory tree endemic to the eastern hardwood forests of the United States. In 1996, dogwood powdery mildew caused by Erysiphe pulchra, an obligate biotrophic fungus of large bracted dogwoods, reached epidemic levels throughout the C. florida growing region. In the late 1990s, both sexual and asexual stages of E. pulchra were regularly observed; thereafter, the sexual stage was found less frequently. We examined the genetic diversity and population structure of 167 E. pulchra samples on C. florida leaves using 15 microsatellite loci. Samples were organized into two separate collection zone data sets, separated as eight zones and two zones, for the subsequent analysis of microsatellite allele length data. Clone correction analysis reduced the sample size to 90 multilocus haplotypes. Our study indicated low genetic diversity, a lack of definitive population structure, low genetic distance among multilocus haplotypes, and significant linkage disequilibrium among zones. Evidence of a population bottleneck was also detected. The results of our study indicated a high probability that E. pulchra reproduces predominately via asexual conidia and lend support to the hypothesis that E. pulchra is an exotic pathogen to North America.[Formula: see text] Copyright © 2019 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Taraxacum kok-saghyz (rubber dandelion) genomic microsatellite loci reveal modest genetic diversity and cross-amplify broadly to related species.

Taraxacum kok-saghyz (TKS) carries great potential as alternative natural rubber source. To better inform future breeding efforts with TKS and gain a deeper understanding of its genetic diversity, we utilized de novo sequencing to generate novel genomic simple sequence repeats markers (gSSRs). We utilized 25 gSSRs on a collection of genomic DNA (gDNA) samples from germplasm bank, and two gDNA samples from historical herbarium specimens. PCR coupled with capillary electrophoresis and an array of population genetics tools were employed to analyze the dataset of our study as well as a dataset of the recently published genic SSRs (eSSRs) generated on the same germplasm. Our results using both gSSRs and eSSRs revealed that TKS has low- to- moderate genetic diversity with most of it partitioned to the individuals and individuals within populations, whereas the species lacked population structure. Nineteen of the 25 gSSR markers cross-amplified to other Taraxacum spp. collected from Southeastern United States and identified as T. officinale by ITS sequencing. We used a subset of 14 gSSRs to estimate the genetic diversity of the T. officinale gDNA collection. In contrast to the obligatory outcrossing TKS, T. officinale presented evidence for population structure and clonal reproduction, which agreed with the species biology. We mapped the molecular markers sequences from this study and several others to the well-annotated sunflower genome. Our gSSRs present a functional tool for the biodiversity analyses in Taraxacum, but also in the related genera, as well as in the closely related tribes of the Asteraceae.

Haplotyping of Cornus florida and C. kousa chloroplasts: Insights into species-level differences and patterns of plastic DNA variation in cultivars.

Chloroplast DNA is a part of plant non-nuclear genome, and is of particular interest for lineage studies. Moreover, the non-coding regions of cpDNA display higher mutation rates than the conserved coding cpDNA, which has been employed for phylogenetic and population research. We analyzed the cpDNA of 332 gDNA samples from collections of Cornus florida and C. kousa (commercial cultivars, breeding selections, and wild kousa accessions from Asia), using the chlorotyping system developed on North America-native, wild accessions of C. florida. Our results indicated significant differences in chlorotype frequencies between the two species. Cornus florida samples were represented by all major chlorotypes previously described, whereas all C. kousa samples analyzed had only one of the chlorotype patterns shown by C. florida. The chlorotyping analytic panel was then expanded by sequencing the targeted three non-coding cpDNA regions. Results indicated a major difference in the maternally-inherited cpDNA between the two closely related Big-Bracted Cornus species. Chlorotype diversity and differences in the proportion of informative sites in the cpDNA regions of focus emphasized the importance of proper loci choice for cpDNA-based comparative studies between the closely related dogwood species. Phylogenetic analyses of the retrieved sequences for the other species of Cornus provided information on the relative utility of the cpDNA regions studied and helped delineate the groups (Big-Bracted, Cornelian Cherries, Blue/White-Fruited) within the genus. Genealogical relationships based on the cpDNA sequences and the inferred chlorotype networks indicated the need for continued analyses across further non-coding cpDNA regions to improve the phylogenetic resolution of dogwoods.

Mycobiota associated with insect galleries in walnut with thousand cankers disease reveals a potential natural enemy against Geosmithia morbida.

Thousand Cankers Disease (TCD) affects Juglans and Pterocarya species. This disease poses not only a major threat to the nut and timber industries but also to native stands of walnut trees. Galleries created by Pityophthorus juglandis (vector) are colonized by the fungus Geosmithia morbida (causal agent of necrosis). It is unknown if other fungi colonizing these galleries might act antagonistically towards G. morbida. The objectives of this study were to: (1) characterize the fungal community associated with TCD-infected trees and (2) develop a pilot study addressing their potential antagonism towards G. morbida. We collected non-Geosmithia fungi from ten TCD-infected walnut trees from California and Tennessee. Four hundred and fifty-seven isolates, representing sixty-five Operational Taxonomic Units (99 % ITS similarity) were obtained. Fungal communities were found to be highly diverse. Ophiostoma dominated the communities associated with TCD-compromised trees from California, whereas Trichoderma dominated TCD-compromised trees in Tennessee. Six Trichoderma isolates showed varying levels of antagonism against three isolates of G. morbida, suggesting potential applications for the biological control of TCD. Furthermore, results from this study contribute to the growing knowledge about the observed differential disease development between the western and eastern USA and could overall impact our understanding of TCD etiology.