Dig-up Primers: A Pipeline for Identification of Polymorphic Microsatellites Loci within Assemblies of Related Species.

Simple sequence repeats (SSRs) have become one of the most popular molecular markers and are used in numerous fields, including conservation genetics, population genetic studies, and genetic mapping. Advances in next-generation sequencing technology and the growing amount of genomic data are driving the development of bioinformatics tools for SSR marker design. These tools work with different combinations of input data, which can be raw reads or assemblies, and with one or more input datasets. We present here a new strategy and implementation of a simple standalone pipeline that utilizes more than one assembly for the in silico design of PCR primers for microsatellite loci in more than one species. Primers are tested in silico to determine if they are polymorphic, eliminating the need to test time-consuming cross-species amplification in the laboratory. The end result is a set of markers that are in silico polymorphic in all analyzed species and have great potential for the identification of interspecies hybrids. The efficiency of the tool is demonstrated using two examples at different taxonomic levels and with different numbers of input assemblies to generate promising, high-quality SSR markers.

Revealing the Diversity and Complex Relationships of Croatian Olive Germplasm.

Olive growing in Croatia has a long tradition and is of great economic and social impact. The present study includes a set of 108 tree samples (88 samples corresponding to 60 presumed cultivars and 20 trees of unnamed ones) collected from 27 groves in the entire olive growing area, and is the most comprehensive survey to be conducted in Croatia. The genetic diversity, relationships, and structures of olive plants were studied using eight microsatellite loci. All loci were polymorphic and revealed a total of 90 alleles. A total of 74 different genotypes were detected that were subjected to further diversity and genetic relationship studies. The Fitch-Margoliash tree and Bayesian analysis of population structure revealed a complex relationship between the identified olive genotypes, which were clustered into three gene pools, indicating different origins of Croatian olive germplasms. Excluding the redundant germplasms, 44 different genotypes among the sampled trees of well-known cultivars and 16 new local germplasms were identified. In addition, we provide the etymology of 46 vernacular names, which confirms that the vast majority of traditional Croatian cultivars have common and widespread names. The results presented herein underline the importance of safeguarding local cultivars and conducting continuous surveys.

Genetic diversity and selection signatures in a gene bank panel of maize inbred lines from Southeast Europe compared with two West European panels.

Southeast Europe (SEE) is a very important maize-growing region, comparable to the Corn belt region of the United States, with similar dent germplasm (dent by dent hybrids). Historically, this region has undergone several genetic material swaps, following the trends in the US, with one of the most significant swaps related to US aid programs after WWII. The imported accessions used to make double-cross hybrids were also mixed with previously adapted germplasm originating from several more distant OPVs, supporting the transition to single cross-breeding. Many of these materials were deposited at the Maize Gene Bank of the Maize Research Institute Zemun Polje (MRIZP) between the 1960s and 1980s. A part of this Gene Bank (572 inbreds) was genotyped with Affymetrix Axiom Maize Genotyping Array with 616,201 polymorphic variants. Data were merged with two other genotyping datasets with mostly European flint (TUM dataset) and dent (DROPS dataset) germplasm. The final pan-European dataset consisted of 974 inbreds and 460,243 markers. Admixture analysis showed seven ancestral populations representing European flint, B73/B14, Lancaster, B37, Wf9/Oh07, A374, and Iodent pools. Subpanel of inbreds with SEE origin showed a lack of Iodent germplasm, marking its historical context. Several signatures of selection were identified at chromosomes 1, 3, 6, 7, 8, 9, and 10. The regions under selection were mined for protein-coding genes and were used for gene ontology (GO) analysis, showing a highly significant overrepresentation of genes involved in response to stress. Our results suggest the accumulation of favorable allelic diversity, especially in the context of changing climate in the genetic resources of SEE.

Crop breeding for a changing climate in the Pannonian region: towards integration of modern phenotyping tools.

The Pannonian Plain, as the most productive region of Southeast Europe, has a long tradition of agronomic production as well as agronomic research and plant breeding. Many research institutions from the agri-food sector of this region have a significant impact on agriculture. Their well-developed and fruitful breeding programmes resulted in productive crop varieties highly adapted to the specific regional environmental conditions. Rapid climatic changes that occurred during the last decades led to even more investigations of complex interactions between plants and their environments and the creation of climate-smart and resilient crops. Plant phenotyping is an essential part of botanical, biological, agronomic, physiological, biochemical, genetic, and other omics approaches. Phenotyping tools and applied methods differ among these disciplines, but all of them are used to evaluate and measure complex traits related to growth, yield, quality, and adaptation to different environmental stresses (biotic and abiotic). During almost a century-long period of plant breeding in the Pannonian region, plant phenotyping methods have changed, from simple measurements in the field to modern plant phenotyping and high-throughput non-invasive and digital technologies. In this review, we present a short historical background and the most recent developments in the field of plant phenotyping, as well as the results accomplished so far in Croatia, Hungary, and Serbia. Current status and perspectives for further simultaneous regional development and modernization of plant phenotyping are also discussed.

Chloroplast Genome Annotation Tools: Prolegomena to the Identification of Inverted Repeats.

The development of next-generation sequencing technology and the increasing amount of sequencing data have brought the bioinformatic tools used in genome assembly into focus. The final step of the process is genome annotation, which works on assembled genome sequences to identify the location of genome features. In the case of organelle genomes, specialized annotation tools are used to identify organelle genes and structural features. Numerous annotation tools target chloroplast sequences. Most chloroplast DNA genomes have a quadripartite structure caused by two copies of a large inverted repeat. We investigated the strategies of six annotation tools (Chloë, Chloroplot, GeSeq, ORG.Annotate, PGA, Plann) for identifying inverted repeats and analyzed their success using publicly available complete chloroplast sequences of taxa belonging to the asterid and rosid clades. The annotation tools use two different approaches to identify inverted repeats, using existing general search tools or implementing stand-alone solutions. The chloroplast sequences studied show that there are different types of imperfections in the assembled data and that each tool performs better on some sequences than the others.

Essential oils and chemical diversity of southeast European populations of Salvia officinalis L.

The essential oils of 25 populations of Dalmatian sage (Salvia officinalis L.) from nine Balkan countries, including 17 indigenous populations (representing almost the entire native distribution area) and eight non-indigenous (cultivated or naturalized) populations were analyzed. Their essential-oil yield ranged from 0.25 to 3.48%. Within the total of 80 detected compounds, ten (ß-pinene, 1,8-cineole, cis-thujone, trans-thujone, camphor, borneol, trans-caryophyllene, α-humulene, viridiflorol, and manool) represented 42.60 to 85.70% of the components in the analyzed essential oils. Strong positive correlations were observed between the contents of trans-caryophyllene and α-humulene, α-humulene and viridiflorol, and viridiflorol and manool. Principal component analysis (PCA) on the basis of the contents of the ten main compounds showed that four principal components had an eigenvalue greater than 1 and explained 79.87% of the total variation. Performing cluster analysis (CA), the sage populations could be grouped into four distinct chemotypes (A-D). The essential oils of 14 out of the 25 populations of Dalmatian sage belonged to Chemotype A and were rich in cis-thujone and camphor, with low contents of trans-thujone. The correlation between the essential-oil composition and geographic variables of the indigenous populations was not significant; hence, the similarities in the essential-oil profile among populations could not be explained by the physical proximity of the populations. Additionally, the southeastern populations tended to have higher EO yields than the northwestern ones.

Chemotype diversity of indigenous Dalmatian sage (Salvia officinalis L.) populations in Montenegro.

To identify how many chemotypes of Salvia officinalis exist in Montenegro, the chemical composition of the essential oils of 12 wild-growing populations was determined by GC-FID and GC/MS analyses. Among the 40 identified constituents, the most abundant were cis-thujone (16.98-40.35%), camphor (12.75-35.37%), 1,8-cineol (6.40-12.06%), trans-thujone (1.5-10.35%), camphene (2.26-9.97%), borneol (0.97-8.81%), viridiflorol (3.46-7.8%), limonene (1.8-6.47%), α-pinene (1.59-5.46%), and α-humulene (1.77-5.02%). The composition of the essential oils under study did not meet the ISO 9909 requirements, while the oils of populations P02-P04, P09, and P10 complied with the German Drug Codex. A few of the main essential-oil constituents appeared to be highly intercorrelated. Strong positive correlations were observed between α-pinene and camphene, camphene and camphor, as well as between cis-thujone and trans-thujone. Strong negative correlations were evidenced between cis-thujone and α-pinene, cis-thujone and champhene, cis-thujone and camphor, as well as between trans-thujone and camphene. Multivariate analyses allowed the grouping of the populations into three distinct chemotypes, i.e., Chemotype A, rich in total thujones, Chemotype B, with intermediate contents of thujones, α-pinene, camphene, and camphor and high borneol contents, and Chemotype C, rich in camphor, camphene, and α-pinene. The chemotypes did not significantly differ in the total essential-oil content and the cis/trans-thujone ratio.

Asymmetric Introgression and Cryptic Natural Hybridization between Two Species of Teucrium Section Polium (Lamiaceae) on the Balkan Peninsula.

In this work, we analyzed the morphology and genetic structure of Teucrium montanum, T. capitatum and their hybrid T. × rohlenae from three syntopic populations. A morphometric study showed that the parents and their hybrids exhibited continuous morphological variation, with the hybrid positioned exactly between the parents. Genetic analysis revealed that plants morphologically identified as T. × rohlenae are fertile hybrids that produce hybrid swarms dominated by later-generation hybrids. This suggests that introgression, rather than speciation, is the more likely outcome of hybridization between these plant species. The extent and direction of gene flow between the two species differed markedly between the three syntopic localities. At the Trilj locality, it was clearly unidirectional, with T. capitatum playing the dominant role. At the Sicevo locality, gene flow was slightly asymmetric, favoring the genetic background of T. capitatum, while at the Sliven site, it was completely asymmetric in the opposite direction. The extreme case of unidirectional gene flow was observed at the Trilj locality where plants morphologically identified as T. montanum could not be genetically distinguished from T. capitatum. This suggests that interspecific hybridization occurred long ago, leading to introgression and cryptic hybrids, blurring of species boundaries and generating evolutionary noise.

Local adaptation to the native environment affects pyrethrin variability in Dalmatian pyrethrum populations.

The insecticidal compound pyrethrin is synthesized in Dalmatian pyrethrum (Tanacetum cinerariifolium (Trevis.) Sch.Bip.; Asteraceae), a plant species endemic to the eastern Mediterranean. Pyrethrin is a mixture of six compounds, pyrethrin I and II, cinerin I and II, and jasmolin I and II. For this study we sampled 15 natural Dalmatian pyrethrum populations covering the entire natural distribution range of the species; Croatian coastal regions and the islands, inland Bosnia and Herzegovina and Montenegro. The plants were grown in a field experiment under uniform growing conditions to exclude a short-term response to environmental factors and instead observe variation in pyrethrin content and composition among and within populations due to genetic adaptation to the native environment. The drivers of local adaptation were explored by examining the role of bioclimatic factors as a cause of population differentiation. Pyrethrins were extracted by ultrasound-assisted extraction, and the extracts were analyzed by HPLC-UV-DAD. The populations differed significantly in the content and composition of pyrethrins. The highest levels of total pyrethrins (1.27% flower DW), were found in population P14 Budva and the significantly highest levels of pyrethrin I in population P14 Vranjske Njive, Podgorica (66.47% of total pyrethrin). Based on bioclimatic conditions of the sampling sites, populations were grouped into five bioclimatic groups (A, B, C, D, and E), which showed qualitative and quantitative variability in pyrethrin content. The most abundant bioclimatic group was bioclimatic group E, which was characterized by the highest average values for pyrethrin I (53.87% of total pyrethrin), total pyrethrin content (1.06% flower DW) and the ratio of pyrethrin I and II (1.85). The correlation analysis between the pyrethrin compounds and some of the bioclimatic variables (e. g., BIO03 Isothermality and BIO04 Temperature seasonality) showed their significant contribution in explaining the variation of pyrethrins in T. cinerariifolium. The differences in pyrethrin content and composition may be partly due to genetic adaptation to the ecological conditions of the native environment. The obtained data would enable the selection of source populations for breeding programs aimed at producing cultivars with desirable biochemical properties and adaptation to different bioclimatic conditions.

Patterns of Leaf and Fruit Morphological Variation in Marginal Populations of Acer tataricum L. subsp. tataricum.

Marginal populations are usually smaller and more isolated and grow in less favourable conditions than those at the distribution centre. The variability of these populations is of high importance, as it can support the adaptations needed for the conditions that they grow in. In this research, the morphological variability of eight Tatar maple (Acer tataricum L. subsp. tataricum) populations was analysed. Tatar maple is an insect-pollinated and wind-dispersed shrub/tree, whose northwestern distribution edge is in southeastern Europe. Morphometric methods were used to analyse the variability of the populations using leaf and fruit morphology. The research revealed significant differences between and within populations. Furthermore, differences in the distribution of the total variability were noted, which suggest that different evolutionarily factors affect different plant traits. Correlation analysis confirmed a weak dependency between the vegetative and generative traits. In addition, no evidence was found for the presence of isolation by environment (IBE). However, the Mantel test for isolation by distance (IBD) was significant for the leaf morphometric traits and non-significant for the fruit morphometric traits. Being the marginal leading-edge populations, they are younger and were less likely to have had time for adaptation to local environments, which would have resulted in the development of IBE. Overall, edge populations of Tatar maple were characterised by great morphological variability, which helps these populations in their response to the intensive selective pressures they face in their environment.

Lack of pollinators selects for increased selfing, restricted gene flow and resource allocation in the rare Mediterranean sage Salvia brachyodon.

Range contraction and habitat fragmentation can cause biodiversity loss by creating conditions that directly or indirectly affect the survival of plant populations. Fragmented habitats can alter pollinator guilds and impact their behavior, which may result in pollen/pollinator limitation and selection for increased selfing as a mechanism for reproductive assurance. We used Salvia brachyodon, a narrowly distributed and endangered sage from eastern Adriatic, to test the consequences of range contraction and habitat fragmentation. Molecular data indicate a severe and relatively recent species range reduction. While one population is reproductively almost completely isolated, moderate gene flow has been detected between the remaining two populations. The high pollen-to-ovule ratio and the results of controlled hand pollination indicate that S. brachyodon has a mixed mating system. Quantitative and qualitative differences in the community and behaviour of flower visitors resulted in limited pollination services in one population where no effective pollinator other than pollen and nectar robbers were observed. In this population, self-pollination predominated over cross-pollination. Various environmental factors, in which plant-pollinator interactions play a pivotal role, have likely created selection pressures that have led to genetic and phenotypic differentiation and different resource allocation strategies among populations.

A reference consensus genetic map for molecular markers and economically important traits in faba bean (Vicia faba L.).

BACKGROUND: Faba bean (Vicia faba L.) is among the earliest domesticated crops from the Near East. Today this legume is a key protein feed and food worldwide and continues to serve an important role in culinary traditions throughout Middle East, Mediterranean region, China and Ethiopia. Adapted to a wide range of soil types, the main faba bean breeding objectives are to improve yield, resistance to biotic and abiotic stresses, seed quality and other agronomic traits. Genomic approaches aimed at enhancing faba bean breeding programs require high-quality genetic linkage maps to facilitate quantitative trait locus analysis and gene tagging for use in a marker-assisted selection. The objective of this study was to construct a reference consensus map in faba bean by joining the information from the most relevant maps reported so far in this crop. RESULTS: A combination of two approaches, increasing the number of anchor loci in diverse mapping populations and joining the corresponding genetic maps, was used to develop a reference consensus map in faba bean. The map was constructed from three main recombinant inbreed populations derived from four parental lines, incorporates 729 markers and is based on 69 common loci. It spans 4,602 cM with a range from 323 to 1041 loci in six main linkage groups or chromosomes, and an average marker density of one locus every 6 cM. Locus order is generally well maintained between the consensus map and the individual maps. CONCLUSION: We have constructed a reliable and fairly dense consensus genetic linkage map that will serve as a basis for genomic approaches in faba bean research and breeding. The core map contains a larger number of markers than any previous individual map, covers existing gaps and achieves a wider coverage of the large faba bean genome as a whole. This tool can be used as a reference resource for studies in different genetic backgrounds, and provides a framework for transferring genetic information when using different marker technologies. Combined with syntenic approaches, the consensus map will increase marker density in selected genomic regions and will be useful for future faba bean molecular breeding applications.

Phylogenetic relationships in Brassicaceae tribe Alysseae inferred from nuclear ribosomal and chloroplast DNA sequence data.

Numerous molecular systematic studies within Brassicaceae have resulted in a strongly improved classification of the family, as morphologically defined units at and above the generic level were often found to poorly reflect phylogenetic relationships. Here, we focus on tribe Alysseae, which despite its size (accounting for about 7% of all species) has only received limited coverage in previous phylogenetic studies. Specifically, we want to test phylogenetic hypotheses implied by current tribal and generic circumscriptions and to put diversification within tribe Alysseae into a temporal context. To this end, sequence data from the nrDNA ITS and two plastid regions (ndhF gene, trnL-F intergenic spacer) were obtained for 176 accessions, representing 16 out of 17 currently recognized genera of the tribe, and were phylogenetically analysed, among others, using a relaxed molecular clock. Due to large discrepancies with respect to published ages of Brassicaceae, age estimates concerning Alysseae are, however, burdened with considerable uncertainty. The tribe is monophyletic and contains four strongly supported major clades and Alyssum homalocarpum, whose relationships among each other remain uncertain due to incongruences between nuclear and plastid DNA markers. The largest genus of the tribe, Alyssum, is not monophyletic and contains, apart from A. homalocarpum, two distinct lineages, corresponding to sections Alyssum, Psilonema, Gamosepalum and to sections Odontarrhena and Meniocus, respectively. Clypeola, whose monophyly is supported only by the plastid data, is very closely related to and possibly nested within the second Alyssum lineage. Species of the genus Fibigia intermingle with those of Alyssoides, Clastopus, Degenia, and Physoptychis, rendering Fibigia polyphyletic. The monotypic genera Leptoplax and Physocardamum are embedded in Bornmuellera.

Identification of quantitative trait loci for resistance to Verticillium wilt and yield parameters in hop (Humulus lupulus L.).

Verticillium wilt (VW) can cause substantial yield loss in hop particularly with the outbreaks of the lethal strain of Verticillium albo-atrum. To elucidate genetic control of VW resistance in hop, an F1 mapping population derived from a cross of cultivar Wye Target, with the predicted genetic basis of resistance, and susceptible male breeding line BL2/1 was developed to assess wilting symptoms and to perform QTL mapping. The genetic linkage map, constructed with 203 markers of various types using a pseudo-testcross strategy, formed ten major linkage groups (LG) of the maternal and paternal maps, covering 552.98 and 441.1 cM, respectively. A significant QTL for VW resistance was detected at LOD 7 on a single chromosomal region on LG03 of both parental maps, accounting for 24.2-26.0 % of the phenotypic variance. QTL analysis for alpha-acid content and yield parameters was also performed on this map. QTLs for these traits were also detected and confirmed our previously detected QTLs in a different pedigree and environment. The work provides the basis for exploration of QTL flanking markers for possible use in marker-assisted selection.

Chemical diversity of the natural populations of Dalmatian pyrethrum (Tanacetum cinerariifolium (TREVIR.) SCH.BIP.) in Croatia.

Dalmatian pyrethrum (Tanacetum cinerariifolium (TREVIR.) SCH.BIP.) is a plant species endemic to the east Adriatic coast. The bioactive substance of Dalmatian pyrethrum is a natural insecticide, pyrethrin, a mixture of six active components (pyrethrins I and II, cinerins I and II, and jasmolins I and II). The insecticidal potential of pyrethrin was recognized decades ago, and dried and ground flowers have traditionally been used in Croatian agriculture and households. A total of 25 Dalmatian pyrethrum populations from Croatia were studied to determine the pyrethrin content and composition, and to identify distinct chemotypes. The total pyrethrin content ranged from 0.36 to 1.30% (dry flower weight; DW) and the pyrethrin I/pyrethrin II ratio ranged from 0.64 to 3.33%. The statistical analyses revealed that the correlations between the percentage of pyrethrin I and of all the other components were significant and negative. The total pyrethrin content was positively correlated with the percentage of pyrethrin I and negatively correlated with cinerin II. The multivariate analysis of the chemical variability enabled the identification of five chemotypes among 25 Dalmatian pyrethrum populations. The chemical characterization of indigenous Dalmatian pyrethrum populations may serve as a good background for future breeding and agricultural exploitation.

Variation in Chloroplast Genome Size: Biological Phenomena and Technological Artifacts.

The development of bioinformatic solutions is guided by biological knowledge of the subject. In some cases, we use unambiguous biological models, while in others we rely on assumptions. A commonly used assumption for genomes is that related species have similar genome sequences. This is even more obvious in the case of chloroplast genomes due to their slow evolution. We investigated whether the lengths of complete chloroplast sequences are closely related to the taxonomic proximity of the species. The study was performed using all available RefSeq sequences from the asterid and rosid clades. In general, chloroplast length distributions are narrow at both the family and genus levels. In addition, clear biological explanations have already been reported for families and genera that exhibit particularly wide distributions. The main factors responsible for the length variations are parasitic life forms, IR loss, IR expansions and contractions, and polyphyly. However, the presence of outliers in the distribution at the genus level is a strong indication of possible inaccuracies in sequence assembly.

New insights in the Spanish gene pool of olive (Olea europaea L.) preserved ex situ and in situ based on high-throughput molecular markers.

In Spain, several local studies have highlighted the likely presence of unknown olive cultivars distinct from the approximately 260 ones previously described in the literature. Furthermore, recent advancements in identification techniques have significantly enhanced in terms of efficacy and precision. This scenario motivated a new nationwide prospecting effort aimed at recovering and characterizing new cultivated germplasm using high-throughput molecular markers. In the present study, the use of 96 EST-SNP markers allowed the identification of a considerable amount of new material (173 new genotypes) coming from areas with low intensification of production in different regions of Spain. As a result, the number of distinct national genotypes documented in the World Olive Germplasm Bank of IFAPA, Córdoba (WOGBC-ESP046) increased to 427. Likewise, 65 and 24 new synonymy and homonymy cases were identified, respectively. This rise in the number of different national cultivars allowed to deepen the knowledge about the underlying genetic structure. The great genetic variability of Spanish germplasm was confirmed, and a new hot spot of diversity was identified in the northern regions of La Rioja and Aragon. Analysis of the genetic structure showed a clear separation between the germplasm of southern and northern-northeastern Spain and indicated a significantly higher level of admixture in the latter. Given the expansion of modern olive cultivation with only a few cultivars, this cryptic germplasm is in great danger of disappearing. This underlines the fact that maintaining as many cultivars as possible will increase the genetic variability of the olive gene pool to meet the future challenges of olive cultivation.

High diversity of indigenous populations of dalmatian sage (Salvia officinalis L.) in essential-oil composition.

Essential oils of 25 indigenous populations of Dalmatian sage (Salvia officinalis L.) that represent nearly half of native distribution area of the species were analyzed. Plantlets collected from wild populations were grown in the same field under the same environmental conditions and then sampled for essential-oil analysis. The yield of essential oil ranged from 1.93 to 3.70% with average of 2.83%. Among the 62 compounds detected, eight (cis-thujone, camphor, trans-thujone, 1,8-cineole, ß-pinene, camphene, borneol, and bornyl acetate) formed 78.13-87.33% of essential oils of individual populations. Strong positive correlations were observed between camphor and ß-pinene, ß-pinene and borneol, as well as between borneol and bornyl acetate. The strongest negative correlation was detected between camphor and trans-thujone. Principal component analysis (PCA) on the basis of eight main compounds showed that first main component separated populations with high thujone content, from those rich in camphor, while the second component separated populations rich in cis-thujone from those rich in trans-thujone. Cluster analysis (CA) led to the identification of three chemotypes of S. officinalis populations: cis-thujone; trans-tujone, and camphor/ß-pinene/borneol/bornyl acetate. We propose that differences in essential oils of 25 populations are mostly genetically controlled, since potential environmental factors were controlled in this study.

Development of new microsatellite markers for Salvia officinalis L. and its potential use in conservation-genetic studies of narrow endemic Salvia brachyodon Vandas.

Nine new microsatellite markers (SSR) were isolated from Salvia officinalis L. A total of 125 alleles, with 8 to 21 alleles per locus, were detected in a natural population from the east Adriatic coast. The observed heterozygosity, expected heterozygosity, and polymorphic information content ranged from 0.46 to 0.83, 0.73 to 0.93 and 0.70 to 0.92, respectively. New microsatellite markers, as well as previously published markers, were tested for cross-amplification in Salvia brachyodon Vandas, a narrow endemic species known to be present in only two localities on the Balkan Peninsula. Out of 30 microsatellite markers tested on the natural S. brachyodon population, 15 were successfully amplified. To obtain evidence of recent bottleneck events in the populations of both species, observed genetic diversity (H(E)) was compared to the expected genetic diversity at mutation-drift equilibrium (H(EQ)) and calculated from the observed number of alleles using a two-phased mutation model (TPM). Recent bottleneck events were detected only in the S. brachyodon population. This result suggests the need to reconsider the current threat category of this endemic species.

The complete chloroplast genome of dalmatian pyrethrum (Tanacetum cinerariifolium (Trevir.) Sch. Bip. (Asteraceae)), the source of the natural insecticide pyrethrin.

Tanacetum cinerariifolium is an endemic species of the eastern Adriatic coast that synthesizes the natural insecticide pyrethrin. We have characterized the complete chloroplast genome of the species and analyzed its phylogeny within the Asteraceae family. The complete chloroplast genome of T. cinerariifolium has a size of 150,136 bp, including a large single-copy (LSC) region of 82,717 bp, a small single-copy (SSC) region of 18,411bp, and a pair of inverted repeats (IRs) of 24,504 bp. The chloroplast genome of T. cinerariifolium encodes 108 genes, including 77 protein-coding genes (PCGs), 27 tRNA genes, and 4 rRNA genes. Phylogenetic analyses based on the complete chloroplast genomes placed T. cinerariifolium in a sister position to species of the genera Artemisia and Chrysanthemum.