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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.

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.

Grass pea (Lathyrus sativus L.) is an annual legume species, phylogenetically close to pea (Pisum sativum L.), that may be infected by Fusarium oxysporum f. sp. pisi (Fop), the causal agent of fusarium wilt in peas with vast worldwide yield losses. A range of responses varying from high resistance to susceptibility to this pathogen has been reported in grass pea germplasm. Nevertheless, the genetic basis of that diversity of responses is still unknown, hampering its breeding exploitation. To identify genomic regions controlling grass pea resistance to fusarium wilt, a genome-wide association study approach was applied on a grass pea worldwide collection of accessions inoculated with Fop race 2. Disease responses were scored in this collection that was also subjected to high-throughput based single nucleotide polymorphisms (SNP) screening through genotyping-by-sequencing. A total of 5,651 high-quality SNPs were considered for association mapping analysis, performed using mixed linear models accounting for population structure. Because of the absence of a fully assembled grass pea reference genome, SNP markers’ genomic positions were retrieved from the pea's reference genome v1a. In total, 17 genomic regions were associated with three fusarium wilt response traits in grass pea, anticipating an oligogenic control. Seven of these regions were located on pea chromosomes 1, 6, and 7. The candidate genes underlying these regions were putatively involved in secondary and amino acid metabolism, RNA (regulation of transcription), transport, and development. This study revealed important fusarium wilt resistance favorable grass pea SNP alleles, allowing the development of molecular tools for precision disease resistance breeding.

Lentil is an important cool season food legume. Vascular wilt incited by Fusarium oxysporum f.sp. lentis (FOL) is the most important disease of lentil worldwide. Knowledge of pathogen genetic structure is crucial to develop effective control strategies. In this study, the genetic structure of a collection of FOL isolates from Iran, Syria and Algeria was analysed using SSR markers. Eight markers were developed in this study and constitute a valuable resource for future molecular studies in FOL. Our results showed that there is a high molecular variation within regions, with isolates from North West Iran showing the highest gene diversity. Variation among regions also exists, with Iranian populations differing significantly from non-Iranian ones, having some private alleles. The clustering of isolates was well in agreement in both distance-based and model-based approaches suggesting the presence of seven ancestral FOL lineages, being three present exclusively in Iran while the others were distributed across all the regions. These results suggest that Iran could be a regional center of origin for FOL.

Lentil cultivation is threatened worldwide by Fusarium wilt, caused by Fusarium oxysporum f. sp. lentis (Fol). Knowledge on pathogenic diversity and genetic structure of Fol populations is fundamental for breeding for resistance and managing the disease. We therefore studied virulence diversity within a collection of Fol isolates.

Pea (Pisum sativum L.) is an important grain legume whose yield stability and production is constrained by drought stress in most environments. However, little is known on availability of drought adaptation sources and its genetic control in pea. The aim of this work was to study the genetic of drought adaptation in pea and identify the genomic regions controlling the trait. Towards this objective, in this work, we assessed drought symptoms and relative water content in soil (RWCS) and leaves (RWCL) along a time course of water stress on a pea Recombinant Inbreed Lines (RILs) population from two parents known to segregate for drought adaptation. Drought adaptation in this population was a quantitative trait. QTL analysis using composite interval mapping (CIM) and multiple interval mapping (MIM) allowed us to identify ten quantitative trait loci (QTLs) associated with the traits explaining individually from 9 to 33 % of the phenotypic variation depending on the variable assessed and altogether from 20 to 57 %. A set of reproducible markers linked to these QTLs (A6, AA175, AC74, AD57, AB141, AB64, Psblox2, PsAAP2_SNP4, and DipeptIV_SNP1) were identified. These markers can be used to select the individuals harbouring the desired QTLs in pea breeding programs for drought adaptation.

Key message: Phenotyping of specific cellular resistance responses and improvement of previous genetic map allowed the identification of novel genomic regions controlling cellular mechanisms involved in pea resistance to ascochyta blight and provided candidate genes suitable for MAS. Didymella pinodes, causing ascochyta blight, is a major pathogen of the pea crop and is responsible for serious damage and yield losses. Resistance is inherited polygenically and several quantitative trait loci (QTLs) have been already identified. However, the position of these QTLs should be further refined to identify molecular markers more closely linked to the resistance genes. In previous works, resistance was scored visually estimating the final disease symptoms; in this study, we have conducted a more precise phenotyping of resistance evaluating specific cellular resistance responses at the histological level to perform a more accurate QTL analysis. In addition, P665 × Messire genetic map used to identify the QTLs was improved by adding 117 SNP markers located in genes. This combined approach has allowed the identification, for the first time, of genomic regions controlling cellular mechanisms directly involved in pea resistance to ascochyta blight. Furthermore, the inclusion of the gene-based SNP markers has allowed the identification of candidate genes co-located with QTLs and has provided robust markers for marker-assisted selection. © 2014 Springer-Verlag Berlin Heidelberg.

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 dif

Traditional landraces are unique genetic resources, highly valuable for yield, disease resistance and quality modern crop improvement. Common bean (Phaseolus vulgaris L.) was introduced in Portugal more than 5 centuries ago and due to its high organoleptic and nutritional value, it became a popular crop all over the country. Very diverse landraces are still widely grown at the farmersʼ fields where they are commonly found in intercropping associations with other crops, such as maize (Zea mays L.). Nevertheless, several constraints to bean yield stability exist, such as susceptibility to several major fungal diseases. Rusts and powdery mildew fungi, foliar biotrophic pathogens, are among the limiting factors of common bean production. Identification and characterization of sources and mechanisms of resistance to use in plant breeding is regarded as a cost efficient method. Understanding the detected resistance genetic basis will facilitate the resistance handling by the breeders. The study of the genetic diversity existing among the different sources of resistance will be fundamental to select the parental lines to use on the development of contrasting populations needed for the genetic resistance studies. During a field expedition to the Central region of Portugal, members of our team collected several bean landraces adapted to intercropping and low-input farming systems. From those, 45 were screened for genetic diversity with 6 microsatellite markers. Parameters of genetic variation were analyzed, and a Neighbor-joining tree generated, suggesting the existence of two genetically distinct clusters. Additionally, a collection of 92 common bean landraces (7 in common with the described molecular screening) was evaluated for resistance to rust and powdery mildew. Rust infection was analyzed 15 days after inoculation (DAI) with results revealing different types of infection and, although several accessions showed intermediate infection patterns, some landraces presented

Nowadays underused in Europe, Lathyrus species were in the past widespread in Mediterranean regions. The superior atmospheric nitrogen fixation rate and potential in crop rotation, allied to the ability to tolerate salinity, drought and flooding , make some of these Lathyrus species highly potential for sustainable agriculture. Being member of the legume family Fabaceae, Lathyrus spp. could share genes, physiological processes and defense mechanisms with economically important related crops such as pea and lupines. Likewise, the molecular tools developed on these model species and cross-amplifiable on Lathyrus spp. could be used on them for different genetic studies, such as diversity analysis. Accordingly, the genetic diversity of an Iberian collection of 45 Lathyrus sativus (grass pea) and 45 L. cicera (chickling vetch) accessions was evaluated using 17 cross -species amplifiable molecular markers (ITAP, EST -SSR and gSSR), developed for Pisum sativum, Lupinus albus and Medicago truncatula . Homology of amplified fragments was confirmed by sequencing. In L. sativus , 2 loci did not amplify, 7 were monomorphic and 8 polymorphic, with an allele average of 3.75 (range 2 -8) and a PIC average of 0.356 (range 0.024 - 0.792). For L. cicera, 7 loci were polymo rphic and 10 monomorphic, with an allele average of 2.14 (range 2 - 3) and a PIC average of 0.287 (range 0.099 - 0.435). Four loci were polymorphic in both Lathyrus species. These results confirm the feasibility of interchange molecular markers for genetic studies, revealing a higher heterozygosity and a potential higher outcrossing rate of L. sativus in relation to L. cicera.

Pseudomonas syringae is the main pathogen responsible for bacterial blight disease in pea and can cause yield losses of 70%. P. syringae pv. pisi is prevalent in most countries but the importance of P. syringae pv. syringae (Psy) is increasing. Several sources of resistance to Psy have been identified but genetics of the resistance is unknown. In this study the inheritance of resistance to Psy was studied in the pea recombinant inbred line population P665 × 'Messire'. Results suggest a polygenic control of the resistance and two quantitative trait loci (QTL) associated with resistance, Psy1 and Psy2, were identified. The QTL explained individually 22.2 and 8.6% of the phenotypic variation, respectively. In addition 21 SSR markers were included in the P665 × 'Messire' map, of which six had not been mapped on the pea genome in previous studies. © 2011 Springer Science+Business Media B.V.

This study presents the development of an enhanced map in faba bean. The map contains 258 loci, mostly gene-based markers, organized in 16 linkage groups that expand 1,875 cM, with an average inter-marker distance of 7.26 cM. The combination of EST-derived markers with a number of markers physically located or previously ascribed to chromosomes by trisomic segregation, allowed the allocation of eight linkage groups (229 markers), to specific chromosomes. Moreover, this approach provided anchor points to establish a global homology among the faba bean chromosomes and those of closely-related legumes species. The map was used to identify and validate, for the first time, QTLs controlling five flowering and reproductive traits: days to flowering, flowering length, pod length, number of seeds per pod and number of ovules per pod. Twelve QTLs stable in the 2 years of evaluation were identified in chromosomes II, V and VI. Comparative mapping suggested the conservation of one of the faba bean genomic regions controlling the character days to flowering in other five legume species (Medicago, Lotus, pea, lupine, chickpea). Additional syntenic co-localizations of QTLs controlling pod length and number of seeds per pod between faba bean and Lotus japonicus are likely. The new genetic map opens the way for further translational studies between faba bean and related legume species, and provides an efficient tool for breeding applications such as QTL analysis and marker-assisted selection. © 2012 Springer-Verlag.

The present study reports the setting up of Diversity Arrays Technology (DArT) markers in olive (Olea europaea L.). Two genomic representations were generated using the PstI/. TaqI combination. A first one was aimed to cover the variability available at the World Olive Germplasm Bank (WOGB) and it was based on DNA from 87 olive cultivars carefully selected from the WOGB to represent the olive variability. The second one was obtained from DNA of the parents of a mapping population, 'Picual' and 'Arbequina', and was used to increase the number of markers segregating within them. A total of 2031 and 1630 markers were used for diversity and mapping analyses, respectively to test the utility of DArT markers in olive. A set of 62 cultivars was genotyped with olive-specific DArT markers. A dendrogram was constructed confirming the relationships among olive cultivars reported in previous works. Besides, DArT markers permitted the unambiguous discrimination of all 62 genotypes confirming thus their utility for identification studies, an important issue for management of germplasm collections. DArT markers also allowed the construction of a genetic map of olive using the population derived from the cross 'Picual'. ×. 'Arbequina' and the pseudo-testcross mapping strategy. This map will be used as a framework map to add new markers derived from olive sequencing originating from the project OLEAGEN. In conclusion, olive-specific DArT markers will enhance identification and genetic studies in olive given their high throughput and low cost per data. © 2011 Elsevier B.V.

DNA sequence analysis of the nuclear ribosomal internal transcribed spacer region (ITS) was performed to determine phylogenetic relationship between 49 isolates of rusts infecting grain and forage legumes. Isolates were collected from different hosts and distinct geographic origins and represent eight species of Uromyces: U. anthyllidis, U. appendiculatus, U. ciceris-arietini, U. minor, U. pisi, U. striatus, U. viciae-fabae and U. vignae. ITS sequences revealed length polymorphisms and variation in DNA sequence that were used to characterize phylogenetic relationships by maximum parsimony, maximum likelihood and Bayesian analyses which in general agreed revealing the presence of four clearly distinct clades. Clade one included the isolates causing rust on chickpea, fenugreek and alfalfa. Clade two was composed by rust isolates of field clover and pea plants, while the third clade was formed by bean and cowpea isolates. Clade four was the largest and included all the rust isolates infecting faba bean. Within this clade, the highly supported subclusters of U. viciae-fabae collected on Lens culinaris, U. viciae-fabae collected on Vicia sativa and U. viciae-fabae collected on Lathyrus palustris suggest an ongoing process of host specialization. © 2010 Blackwell Verlag GmbH.

In a previous study a genetic map had been developed using a RIL population derived from a cross between the Pisum sativum ssp. syriacum accession P665 and the P. sativum ssp. sativum cv. Messire. This population segregated for several agricultural important traits and was successfully used to identify QTLs (Quantitative Trait Loci) controlling resistance to Mycosphaerella pinodes and Orobanche crenata, earliness, root length and aerial biomass. However, this map contained only a few markers in common with the international pea consensus map, hampering comparison with other pea maps. The objective of this study was to incorporate a set of common transferable and reproducible markers into the P665 × Messire map to favour comparative mapping and QTL validation. Seventy-eight out of the 248 SSRs assayed resulted polymorphic in the parental lines. Thirty-eight of them, uniformly distributed all over the genome, were genotyped in the whole population and included in the map. This SSR enriched map allowed identification of six new QTLs (three for resistance to M. pinodes, two for resistance to broomrape and one for root length). Inclusion of the SSRs confirmed the homology of some of the QTLs identified in the population P665 × Messire with other QTLs associated with related traits in different pea genetic backgrounds. © 2011 Springer Science+Business Media B.V.

The genetic diversity of wild olive trees sampled from eight different sites from two provinces of Andalusia (Cadiz and Jaen), Southern Spain, was evaluated by means of agro-morphological traits and SSR markers. The agro-morphological traits showed a high variation between genotypes and significant correlation coefficients were obtained among the values recorded in two consecutive years, 2007/08 and 2008/09 (r= 0.59-0.78). Traits such as fruit symmetry (position A), stone shape, fruit flesh/stone ratio and olive oil content on both wet and dry matter, showed the highest values of diversity and were very useful in genotype discrimination. As expected, average values obtained in wild olives for fruit size and oil content were lower than the previously reported in cultivated material. However, it is noticeable that wild trees with fruit weights (1.3. g) and olive oil percentage in dry matter (33.8%) comparable to the values found in some olive cultivars, were also found. With both DNA-based and agro-morphological descriptors, higher levels of variability were found within each site than between sites. Genetic variation observed among the wild olive germplasm at the DNA level was higher than the agro-morphological traits, indicating the efficiency of SSR markers for detecting genetic diversity among wild olive genotypes and their relationships. The lack of consistency between the relationship studies performed with molecular and morphological markers could indicate that each marker system measures different aspects of the wild olive genetic variability. Molecular data obtained by SSR markers together with morphological and agronomical characterization of olive trees confirmed the high diversity within the wild populations and their potential use for olive breeding. © 2011 Elsevier B.V.

This article documents the addition of 411 microsatellite marker loci and 15 pairs of Single Nucleotide Polymorphism (SNP) sequencing primers to the Molecular Ecology Resources Database. Loci were developed for the following species: Acanthopagrus schlegeli, Anopheles lesteri, Aspergillus clavatus, Aspergillus flavus, Aspergillus fumigatus, Aspergillus oryzae, Aspergillus terreus, Branchiostoma japonicum, Branchiostoma belcheri, Colias behrii, Coryphopterus personatus, Cynogolssus semilaevis, Cynoglossus semilaevis, Dendrobium officinale, Dendrobium officinale, Dysoxylum malabaricum, Metrioptera roeselii, Myrmeciza exsul, Ochotona thibetana, Neosartorya fischeri, Nothofagus pumilio, Onychodactylus fischeri, Phoenicopterus roseus, Salvia officinalis L., Scylla paramamosain, Silene latifo, Sula sula, and Vulpes vulpes. These loci were cross-tested on the following species: Aspergillus giganteus, Colias pelidne, Colias interior, Colias meadii, Colias eurytheme, Coryphopterus lipernes, Coryphopterus glaucofrenum, Coryphopterus eidolon, Gnatholepis thompsoni, Elacatinus evelynae, Dendrobium loddigesii Dendrobium devonianum, Dysoxylum binectariferum, Nothofagus antarctica, Nothofagus dombeyii, Nothofagus nervosa, Nothofagus obliqua, Sula nebouxii, and Sula variegata. This article also documents the addition of 39 sequencing primer pairs and 15 allele specific primers or probes for Paralithodes camtschaticus.

A quantitative trait loci (QTL) associated with resistance to pea rust, caused by the fungus Uromyces pisi (Pers.) Wint., has been identified in a F2 population derived from an intraspecific cross between two wild pea (Pisum fulvum L.) accessions, IFPI3260 (resistant) and IFPI3251 (susceptible). Both parental lines and all the segregating population displayed a fully compatible interaction (high infection type), which indicates absence of hypersensitive response. Nevertheless, differences on the percentage of symptomatic area of the whole plant (disease severity) were observed. A genetic map was developed covering 1283.3 cM and including 146 markers (144 random amplified polymorphic DNA (RAPDs) and two sequence tagged sites (STSs) markers) distributed in 9 linkage groups. A QTL explaining 63% of the total phenotypic variation was located in linkage group 3. RAPDs markers (OPY111316 and OPV171078) flanking this QTL should allow, after their conversion in SCARs, a reliable marker-assisted selection for rust resistance. © 2010 Springer Science+Business Media B.V.

Crenate broomrape (Orobanche crenata) is the major constraint for pea cultivation in the Mediterranean Basin and Middle East. Cultivation of resistant varieties would be the most efficient, economical and environmentally friendly way to control this parasite. However, little resistance is available within cultivated pea. Promising sources of resistance have been identified in wild peas but their use in breeding programs is hampered by the polygenic nature of the resistance. The identification of molecular markers linked to the resistance would allow tracking of the underlying genes, facilitating their introgression into pea cultivars. The main objective of this study was the identification of genomic regions associated with resistance to O. crenata. A RIL (Recombinant Inbred Lines) population derived from a cross between a resistant accession of the wild pea Pisum sativum ssp. syriacum, and a susceptible pea variety was screened for resistance to O. crenata under field conditions during two seasons. In addition, resistance reactions at different stages of the O. crenata infection cycle were assessed using a Petri dish method. The approach allowed the identification of four Quantitative Trait Loci (QTL) associated with field resistance, assessed as the number of emerged broomrape shoots per pea plant under field conditions. These identified QTLs explained individually from 10 to 17% of the phenotypic variation. In addition QTLs governing specific mechanisms of resistance, such as low induction of O. crenata seed germination, lower number of established tubercles per host root length unit, and slower development of tubercles were also identified. Identified QTLs explained individually from 8 to 37% of the variation observed depending on the trait. Host plant aerial biomass and root length were also assessed and mapped. Both traits were correlated with the level of O. crenata infection and three out of the four QTLs controlling resistance under field conditions co-loca

Genetic variation of 30 different Uromyces isolates collected on faba bean, lentil, common vetch, pea, chickpea, alfalfa, cowpea and lupin was studied. Random Amplified Polymorphic DNA markers were used showing clear differences among Uromyces species. Uromyces viciae-fabae isolates clustered according to the host, with a clear cluster including all U. viciae-fabae ex Vicia faba isolates. The U. viciae-fabae ex Lens culinaris isolate was the nearest to the cluster of U. viciae faba ex V. faba isolates, followed by U. pisi from Canada and U. viciae-fabae ex V. sativa. No association was found among molecular diversity and virulence or geographic origin within U. vicia-fabae ex V. faba isolates. Among the three U. pisi isolates considered, a great variability was observed and no grouping could be established. The most different isolate from the rest of species considered was U. striatus, followed by U. vignae. The two U. ciceris-arietini isolates clustered together and so did the two U. lupinicolus isolates.