Lene Rostgaard Nielsen

Towards domestication of Jatropha curcas

Jatropha curcas L. attracts a lot of interest as a biofuel crop, triggering large investments and rapid expansion of cultivation areas, and yet, it should still be considered as a (semi-)wild, undomesticated plant. To use the full potential of Jatropha and to support further expansion and systematic selection, breeding and domestication are a prerequisite. This review reveals and identifies gaps in knowledge that still impede domestication of Jatropha. Prebreeding knowledge is limited. In particular, the regeneration ecology and the degree of genetic diversity among and within natural populations in and outside the center of origin are poorly studied. There is only a limited understanding of the Jatropha breeding system and the effect of inbreeding and outbreeding. This review presents all currently available and relevant information on the species distribution, site requirements, regeneration ecology, genetic diversity, advances in selection, development of varieties and hybridization. It also describes possible routes to a better Jatropha germplasm, gives recommendations for tackling current problems and provides guidance for future research. We also discuss the participatory domestication strategy of Jatropha integration in agroforestry.

Presence of natural genetic resistance in Fraxinus excelsior (Oleraceae) to Chalara fraxinea (Ascomycota): an emerging infectious disease

Fraxinus excelsior, common ash native to Europe, is threatened by a recently identified pathogenic fungus Chalara fraxinea, which causes extensive damage on ash trees across Europe. In Denmark, most stands are severely affected leaving many trees with dead crowns. However, single trees show notably fewer symptoms. In this study, the impact of the emerging infectious disease on native Danish ash trees is assessed by estimating presence of inherent resistance in natural populations. Disease symptoms were assessed from 2007 to 2009 at two different sites with grafted ramets of 39 selected clones representing native F. excelsior trees. A strong genetic variation in susceptibility to C. fraxinea infections was observed. No genetic or geographic structure can explain the differences, but strong genetic correlations to leaf senescence were observed. The results suggest that a small fraction of trees in the Danish population of ash possess substantial resistance against the damage. Though this fraction is probably too low to avoid population collapse in most natural or managed ash forests, the observed presence of putative resistance against the emerging infectious disease in natural stands is likely to be of evolutionary importance. This provides prospects of future maintenance of the species through natural or artificial selection in favour of remaining healthy individuals.

Adaptive potential of ash (Fraxinus excelsior) populations against the novel emerging pathogen Hymenoscyphus pseudoalbidus.

An emerging infectious pathogen Hymenoscyphus pseudoalbidus has spread across much of Europe within recent years causing devastating damage on European common ash trees (Fraxinus excelsior) and associated plant communities. The present study demonstrates the presence of additive genetic variation in susceptibility of natural F. excelsior populations to the new invasive disease. We observe high levels of additive variation in the degree of susceptibility with relatively low influence of environmental factors (narrow‐sense heritability = 0.37–0.52). Most native trees are found to be highly susceptible, and we estimate that only around 1% has the potential of producing offspring with expected crown damage of <10% under the present disease pressure. The results suggest that the presence of additive genetic diversity in natural F. excelsior populations can confer the species with important ability to recover, but that low resistance within natural European populations is to be expected because of a low frequency of the hypo‐sensitive trees. Large effective population sizes will be required to avoid genetic bottlenecks. The role of artificial selection and breeding for protection of the species is discussed based on the findings.

Genome sequence and genetic diversity of European ash trees

Ash trees (genus Fraxinus, family Oleaceae) are widespread throughout the Northern Hemisphere, but are being devastated in Europe by the fungus Hymenoscyphus fraxineus, causing ash dieback, and in North America by the herbivorous beetle Agrilus planipennis. Here we sequence the genome of a low-heterozygosity Fraxinus excelsior tree from Gloucestershire, UK, annotating 38,852 protein-coding genes of which 25% appear ash specific when compared with the genomes of ten other plant species. Analyses of paralogous genes suggest a whole-genome duplication shared with olive (Olea europaea, Oleaceae). We also re-sequence 37 F. excelsior trees from Europe, finding evidence for apparent long-term decline in effective population size. Using our reference sequence, we re-analyse association transcriptomic data, yielding improved markers for reduced susceptibility to ash dieback. Surveys of these markers in British populations suggest that reduced susceptibility to ash dieback may be more widespread in Great Britain than in Denmark. We also present evidence that susceptibility of trees to H. fraxineus is associated with their iridoid glycoside levels. This rapid, integrated, multidisciplinary research response to an emerging health threat in a non-model organism opens the way for mitigation of the epidemic.

Molecular markers for tolerance of European ash (Fraxinus excelsior) to dieback disease identified using Associative Transcriptomics.

Tree disease epidemics are a global problem, impacting food security, biodiversity and national economies. The potential for conservation and breeding in trees is hampered by complex genomes and long lifecycles, with most species lacking genomic resources. The European Ash tree Fraxinus excelsior is being devastated by the fungal pathogen Hymenoscyphus fraxineus, which causes ash dieback disease. Taking this system as an example and utilizing Associative Transcriptomics for the first time in a plant pathology study, we discovered gene sequence and gene expression variants across a genetic diversity panel scored for disease symptoms and identified markers strongly associated with canopy damage in infected trees. Using these markers we predicted phenotypes in a test panel of unrelated trees, successfully identifying individuals with a low level of susceptibility to the disease. Co-expression analysis suggested that pre-priming of defence responses may underlie reduced susceptibility to ash dieback.

Interspecific differentiation and hybridization in Vanilla species (Orchidaceae)

Vanilla claviculata, V. barbellata and V. dilloniana are distributed throughout the Caribbean islands and are all found in Puerto Rico. The vegetative parts of the species are similar; however, their conspicuous flowers easily distinguish them. Electrophoresis of seven polymorphic enzymes revealed that the genetic composition of the three species is also very similar: they deviate mainly from each other in allele frequencies rather than by specific alleles. A hierarchical analysis of genetic differentiation showed that the between-species component is slightly higher (FSG=0.237) than the component between populations within species (FPS=0.141). Nevertheless, they are efficiently recognized by their genotypic compositions. In V. barbellata and V. claviculata 97–99% of all individuals were assigned to the correct species. Assignment to a wrong species occurred only with individuals at localities where species coexist. This suggests that the species may hybridize.

Hybridization between Quercus robur and Q. petraea in a mixed oak stand in Denmark

Abstract• Hybridization and mating pattern between Quercus robur and Q. petraea was studied in a 5.8 ha mixed forest stand in Jutland, Denmark which comprises in total 135 Quercus robur and 230 Q. petraea trees. Classification of the oak trees into species was performed using canonical discriminant analysis of a range of leaf morphological traits. Adult trees (365) and offspring (582) were genotyped with eight microsatellite markers. Seedlings were sampled in 2003 and acorns were collected in 2004.• Mating patterns of Q. robur and Q. petraea are expected to be different in the northern range of the distribution area and a larger hybridization rate is expected. It is further expected, that pollination from outside sources will be relatively less in small fragmented forest management systems compared to large scale oak forest. The conclusions should be verified through repeated year to year analysis of the mating pattern.• Phenological studies revealed that there was no difference in flowering time between species. Data for the adult trees revealed no significant departures from Hardy-Weinberg proportions and there was weak, but significant spatial genetic structure, which supports the idea that the stand is of natural origin. Spatial genetic structure in the first distance class is stronger for Q. petraea. The genetic composition of the offspring was remarkably consistent from year to year. Paternity analysis revealed that, on average, 85% pollination came from fathers within the stand. The direction of the pollen flow varied from year to year. Inter-specific hybridization was high and ranged from 15–17% and from 48–55% for Q. petraea and Q. robur mothers respectively. Paternity analysis revealed that the population was basically outcrossing and only 3.7% of the analysed progeny were the product of selfing. Over the two years of study, approximately 200 trees contributed to the paternity of the next generations.• The study confirms earlier studies with a greater tendency for Q. robur mothers to produce hybrid seeds than Q. petraea mothers. The rate of hybridization is higher in this Danish stand than in comparable studies elsewhere in Europe. Gene flow from outside sources are relatively low.Résumé• L’hybridation et le schéma de croisement entre Quercus robur et Q. petraea ont été étudiés dans une forêt mélangée de 5,8 ha dans le Jutland au Danemark. Cette forêt comprenait 135 arbres de Quercus robur et 230 de Q. petraea. Le classement des arbres dans les différentes espèces a été réalisé à partir de caractères morphologiques des feuilles grâce à une analyse canonique discriminante. Les arbres adultes (365) et leurs descendances ont été génotypés à l’aide de 8 marqueurs microsatellites. Les semis ont été échantillonnés en 2003 et les glands ont été récoltés en 2004.• Le schéma de croisement entre Q. robur et Q. petraea est suspecté différent dans la partie nord de l’aire de distribution où un plus fort taux d’hybridation est également attendu. De plus, il est probable que la pollinisation à partir de pollen étranger soit plus faible dans des forêts fragmentées que dans des forêts continues de chênes. Des observations répétées d’année en année sont nécessaires pour valider ces hypothèses.• Des études phénologiques montrent qu’il n’y a pas de différence entre les espèces pour la période de floraison. Les données sur arbres adultes ne révèlent aucun écart significatif par rapport aux proportions attendues de la loi de Hardy-Weinberg. Une structuration spatiale faible mais significative a été mise en évidence, qui suggère l’idée que le peuplement est d’origine naturelle. Cette structuration est plus forte pour Q. petraea. La composition génétique des descendances est remarquablement stable d’une année sur l’autre. Les analyses de paternité révèlent qu’en moyenne 85 % de la pollinisation provient de pères du peuplement. Le sens de pollinisation varie d’une année sur l’autre. Le taux d’hybrides interspécifiques est élevé et varie de 15–17 % et de 48–55 % respectivement pour Q. petraea et Q. robur. Les études de paternité montrent que les inter-croisements sont la règle et que seulement 3,7 % des descendants analysés sont issus d’autofécondation. Durant les deux années de l’étude, approximativement 200 arbres sont intervenus comme père pour les générations suivantes.• Cette étude confirme des études précédentes montrant que Q. robur produit plus de semences hybrides que Q. petraea. Le taux d’hybridation est plus élevé dans ce peuplement danois que dans d’autres études similaires en Europe. Les flux de gènes à partir de sources extérieures sont faibles.

Selective advantage of ray florets in Scalesia affinis and S. pedunculata (Asteraceae), two endemic species from the Galápagos

The presence of neuter ray florets in species within Asteraceae is generally believed to increase pollinator attraction. In the endemic Galápagos genus Scalesia (Asteraceae) a natural variation in the presence/absence of neuter ray florets is found. To evaluate whether the presence of ray florets plays a selective role on female reproductive success we chose two species of Scalesia, Scalesia affinis that carries ray florets and S. pedunculata that is rayless. On Santa Cruz Island capitula of S. pedunculata were equipped with fake ray florets while others were untouched. On Isabela Island ray florets were removed on half of the capitula of S. affinis. In S. affinis rayed capitula received more pollinators and more pollen, which resulted in a significantly higher embryo production. In S. pedunculata no effect on embryo production was found. The disagreement between the two species may correspond to a difference in visitation frequency, S. pedunculata receiving many more visit than S. affinis. Thus, ray floret development proved beneficial in pollinator-restricted localities.

The Use of DNA Barcoding in Identification and Conservation of Rosewood (Dalbergia spp.)

The genus Dalbergia contains many valuable timber species threatened by illegal logging and deforestation, but knowledge on distributions and threats is often limited and accurate species identification difficult. The aim of this study was to apply DNA barcoding methods to support conservation efforts of Dalbergia species in Indochina. We used the recommended rbcL, matK and ITS barcoding markers on 95 samples covering 31 species of Dalbergia, and tested their discrimination ability with both traditional distance-based as well as different model-based machine learning methods. We specifically tested whether the markers could be used to solve taxonomic confusion concerning the timber species Dalbergia oliveri, and to identify the CITES-listed Dalbergia cochinchinensis. We also applied the barcoding markers to 14 samples of unknown identity. In general, we found that the barcoding markers discriminated among Dalbergia species with high accuracy. We found that ITS yielded the single highest discrimination rate (100%), but due to difficulties in obtaining high-quality sequences from degraded material, the better overall choice for Dalbergia seems to be the standard rbcL+matK barcode, as this yielded discrimination rates close to 90% and amplified well. The distance-based method TaxonDNA showed the highest identification rates overall, although a more complete specimen sampling is needed to conclude on the best analytic method. We found strong support for a monophyletic Dalbergia oliveri and encourage that this name is used consistently in Indochina. The CITES-listed Dalbergia cochinchinensis was successfully identified, and a species-specific assay can be developed from the data generated in this study for the identification of illegally traded timber. We suggest that the use of DNA barcoding is integrated into the work flow during floristic studies and at national herbaria in the region, as this could significantly increase the number of identified specimens and improve knowledge about species distributions.

Inbreeding depression in the partially self-incompatible endemic plant species Scalesia affinis (Asteraceae) from Galápagos islands

A previous study showed that some individuals of the tetraploid Galápagos endemic Scalesia affinis were able to produce offspring after selfing. The present study compares the fitness of self-pollinated offspring with the fitness of cross-pollinated offspring. Germination success, seedling survival, and four different growth parameters was measured. In most of the studied characters selfed offspring were significantly inferior to outcrossed progeny. The effect was very clear in germination and survival. Outcrossed embryos were 3.4 times more likely to germinate than those that were selfed-fertilized, and the mortality was 84% higher among selfed individuals. Also, there was no genetic variation in inbreeding depression. The present study is based on material from a large population on Isabela Island, Galápagos. At other localities in the archipelago, populations have been through recent dramatic bottlenecks due to the grazing of introduced mammals. Considering the significant inbreeding depression found in the large population and the presence of a partial self-incompatibility system, these small populations are likely to be highly vulnerable and their future survival critically threatened.