High SNP diversity in the non-toxic indigenous Jatropha curcas germplasm widens the potential of this upcoming major biofuel crop species.

BACKGROUND AND AIMS: Jatropha curcas (jatropha) is an oil crop cultivated in (sub)tropical regions around the world, and holds great promise as a renewable energy source. However, efforts to fully commercialize jatropha are currently hampered by the lack of genetic diversity in the extant breeding germplasm, and by the toxicity of its seeds meaning that its seed cake cannot be used as a protein source in animal feed, among other constraints. In Mexico, the species' native range, there are jatropha plants whose seeds are used to prepare traditional meals. This non-toxic jatropha 'type' is considered to harbour low genetic variation due to a presumed domestication bottleneck and therefore to be of limited breeding value; yet, very little is known regarding its origin and genetic diversity. METHODS: Using genotyping-by-sequencing (GBS), we extensively genotyped both indigenous toxic and non-toxic jatropha collected along roads and home gardens throughout southern Mexico. KEY RESULTS: Single nucleotide polymorphism diversity in non-toxic jatropha is relatively high, particularly in northern Veracruz state, the probable origin of this germplasm. Genetic differences between toxic and non-toxic indigenous genotypes are overall quite small. A a genome-wide association study supported a genomic region (on LG 8, scaffold NW_012130064), probably involved in the suppression of seed toxicity. CONCLUSIONS: Conservation actions are urgently needed to preserve this non-toxic indigenous, relatively wild germplasm, having potential as a fuel feedstock, animal feed and food source among other uses. More generally, this work demonstrates the value of conservation genomic research on the indigenous gene pool of economically important plant species.

Evolution, plasticity and evolving plasticity of phenology in the tree species Alnus glutinosa.

Both traits and the plasticity of these traits are subject to evolutionary change and therefore affect the long-term persistence of populations and their role in local communities. We subjected clones from 12 different populations of Alnus glutinosa, located along a latitudinal gradient, to two different temperature treatments, to disentangle the distribution of genetic variation in timing of bud burst and bud burst plasticity within and among genotypes, populations, and regions. We calculated heritability and evolvability estimates for bud burst and bud burst plasticity and assessed the influence of divergent selection relative to neutral drift. We observed higher levels of heritability and evolvability for bud burst than for its plasticity, whereas the total phenological heritability and evolvability (i.e. combining timing of bud burst and bud burst plasticity) suggest substantial evolutionary potential with respect to phenology. Earlier bud burst was observed for the low-latitudinal populations than for the populations from higher latitudes, whereas the high-latitudinal populations did not show the expected delayed bud burst. This countergradient variation can be due to evolution towards increased phenological plasticity at higher latitudes. However, because we found little evidence for adaptive differences in phenological plasticity across the latitudinal gradient, we suggest differential frost tolerance as the most likely explanation for the observed phenological patterns in A. glutinosa.

The population genomic signature of environmental selection in the widespread insect-pollinated tree species Frangula alnus at different geographical scales.

The evaluation of the molecular signatures of selection in species lacking an available closely related reference genome remains challenging, yet it may provide valuable fundamental insights into the capacity of populations to respond to environmental cues. We screened 25 native populations of the tree species Frangula alnus subsp. alnus (Rhamnaceae), covering three different geographical scales, for 183 annotated single-nucleotide polymorphisms (SNPs). Standard population genomic outlier screens were combined with individual-based and multivariate landscape genomic approaches to examine the strength of selection relative to neutral processes in shaping genomic variation, and to identify the main environmental agents driving selection. Our results demonstrate a more distinct signature of selection with increasing geographical distance, as indicated by the proportion of SNPs (i) showing exceptional patterns of genetic diversity and differentiation (outliers) and (ii) associated with climate. Both temperature and precipitation have an important role as selective agents in shaping adaptive genomic differentiation in F. alnus subsp. alnus, although their relative importance differed among spatial scales. At the 'intermediate' and 'regional' scales, where limited genetic clustering and high population diversity were observed, some indications of natural selection may suggest a major role for gene flow in safeguarding adaptability. High genetic diversity at loci under selection in particular, indicated considerable adaptive potential, which may nevertheless be compromised by the combined effects of climate change and habitat fragmentation.

Recolonization after habitat restoration leads to decreased genetic variation in populations of a terrestrial orchid.

Colonization is crucial to habitat restoration projects that rely on the spontaneous regeneration of the original vegetation. However, as a previously declining plant species spreads again, the likelihood of founder effects increases through recurrent population founding and associated serial bottlenecks. We related Amplified Fragment Length Polymorphism markers genetic variation and fitness to colonization history for all extant populations of the outcrossing terrestrial orchid Dactylorhiza incarnata in an isolated coastal dune complex. Around 1970, D. incarnata suffered a severe bottleneck yet ultimately persisted and gradually spread throughout the spatially segregated dune slacks, aided by the restoration of an open vegetation. Genetic assignment demonstrated dispersal to vacant sites from few nearby extant populations and very limited inflow from outside the spatially isolated reserve. Results further indicated that recurrent founding from few local sources resulted in the loss of genetic diversity and promoted genetic divergence (F(ST) = 0.35) among populations, but did not influence population fitness. The few source populations initially available and the limited inflow of genes from outside the study reserve, as a consequence of habitat degradation and spatial isolation, may have magnified the genetic effects of recurrent population founding.

The consequences of mating over a range of parental genetic similarity in a selfing allopolyploid plant species.

In diploids, F(1) offspring performance is expected to increase with increasing genetic dissimilarity between the parents until an optimum is reached because outbreeding mitigates inbreeding depression and maximizes heterosis. However, many flowering plant species are derived through allopolyploidization, i.e. interspecific hybridization with genome doubling. This mode of plant speciation can be expected to considerably alter the consequences of inbreeding and outbreeding. We investigated the F1 fitness consequences of mating over a range of (genetic) distances in the allohexaploid plant species Geum urbanum. Offspring was raised under controlled conditions (632 plants). The performance of outcrossed progeny was not significantly better than that of their selfed half-siblings and did not increase with parental genetic dissimilarity (0-0.83). Our findings support low, if any, inbreeding depression and heterosis. We attribute this to the peculiar state of quasi-permanent heterozygosity in allopolyploids and frequent selfing.

A direct assessment of realized seed and pollen flow within and between two isolated populations of the food-deceptive orchid Orchis mascula.

Gene flow can counteract the loss of genetic diversity caused by genetic drift in small populations. For this reason, clearly understanding gene flow patterns is of the highest importance across fragmented landscapes. However, gene flow patterns are not only dependent upon the degree of spatial isolation of fragmented populations, but are also dependent upon the life-history traits of the species. Indeed, habitat fragmentation effects appear especially unpredictable for food-deceptive orchid species, because of their highly specialised seed and pollen dispersal mechanisms. In this study we used amplified fragment length polymorphism markers and subsequent parentage and spatial autocorrelation analysis to quantify the extent and the patterns of realized gene flow within and between two adjacent fragmented populations of the food-deceptive Orchis mascula. We observed considerable gene flow between both populations, occurring mainly through pollen dispersal. Seed dispersal, on the other hand, was mainly limited to the first few meters from the mother plant in both populations, although at least one among-population seed dispersal event was observed. This, in turn, resulted in a significant spatial genetic structure for both populations. Although genetic diversity was high in both populations and mainly outcrossing occurred, reproductive output was strongly skewed toward a limited number of successful adult plants. These observed patterns are likely due to the different pollinator behaviour associated with food-deceptive plants. We conclude that these populations can be considered viable under their current fragmented state.

Continuous ambulatory peritoneal dialysis (CAPD) in a patient with glucose-6-phosphatase deficiency.

The metabolic disturbances in glucose-6-phosphatase deficiency (von Gierke's disease) are the consequence of hypoglycemia, occurring mostly during the night. Continuous provision of glucose is the aim of every recently introduced treatment procedure. We studied the influence of continuous ambulatory peritoneal dialysis (CAPD) on the metabolic disturbances in a 42-year-old female patient with von Gierke's disease and end-stage renal disease. During six months of CAPD, there were no dialysis-related complications. The metabolic acidosis didn't worsen: arterial bicarbonate and lactate were not changed. Mean glycemia was 118.6 +/- 14.4 mg%. Total lipemia, cholesterol and triglycerides were not different from those before CAPD, despite the fact that all hypolipidaemic drugs were stopped. Three different exchange procedures were compared during the night: no dialysis, one exchange with a 2 L solution without buffer containing glucose 15 g/L and containing glucose 42.5 g/L. The results show that the 4.25% glucose solution prevents hypoglycaemia, and diminishes the increase in lactate and pyruvate concentration. Intraperitoneal glucose normalizes the plasma free fatty acid concentration. A very important result is the disappearance of hypo-insulinism. We conclude that, from a clinical point of view, CAPD is a well-tolerated treatment in von Gierke's disease. The limited results provide some evidence that the use of a 4.25% glucose solution as an overnight exchange, instead of the usual 1.5% solution, can prevent at least partly the glycogenolysis and consequently the metabolic disturbances of von Gierke's disease.

An unsuspected cause of diarrhoea and gastrointestinal bleeding during corticosteroid therapy.

We report on a fatal case of disseminated strongyloidiasis during corticosteroid treatment presenting with abdominal pain, diarrhoea and lower gastrointestinal bleeding. The patient emigrated from Thailand 16 years before the current hospitalisation. Complicated strongyloidiasis is a relatively unrecognized complication of corticosteroid therapy in non-endemic areas. In individuals who have resided in endemic areas, even decades before treatment, strongyloidiasis should be excluded before initiation of immunosuppressants.

SNP discovery using Paired-End RAD-tag sequencing on pooled genomic DNA of Sisymbrium austriacum (Brassicaceae).

Single nucleotide polymorphisms SNPs are rapidly replacing anonymous markers in population genomic studies, but their use in non model organisms is hampered by the scarcity of cost-effective approaches to uncover genome-wide variation in a comprehensive subset of individuals. The screening of one or only a few individuals induces ascertainment bias. To discover SNPs for a population genomic study of the Pyrenean rocket (Sisymbrium austriacum subsp. chrysanthum), we undertook a pooled RAD-PE (Restriction site Associated DNA Paired-End sequencing) approach. RAD tags were generated from the PstI-digested pooled genomic DNA of 12 individuals sampled across the species distribution range and paired-end sequenced using Illumina technology to produce ~24.5 Mb of sequences, covering ~7% of the specie's genome. Sequences were assembled into ~76 000 contigs with a mean length of 323 bp (N(50)  = 357 bp, sequencing depth = 24x). In all, >15 000 SNPs were called, of which 47% were annotated in putative genic regions based on homology with the Arabidopsis thaliana genome. Gene ontology (GO) slim categorization demonstrated that the identified SNPs covered extant genic variation well. The validation of 300 SNPs on a larger set of individuals using a KASPar assay underpinned the utility of pooled RAD-PE as an inexpensive genome-wide SNP discovery technique (success rate: 87%). In addition to SNPs, we discovered >600 putative SSR markers.

Landscape genetics of the self-compatible forest herb Geum urbanum: effects of habitat age, fragmentation and local environment.

To investigate the role of habitat fragmentation, fragment age and local environment in shaping the genetics of plant populations, we examined the genetic structure of the self-compatible forest herb Geum urbanum using microsatellite markers. A historical land-use reconstruction assigned the studied populations to two age classes: populations in primary forest fragments, and populations in secondary fragments. Local environmental conditions were quantified on the basis of the herb-layer community composition. A stepwise general linear model revealed that levels of within-population genetic diversity were best explained by population size, landscape connectivity and the interaction between both. Connectivity was positively correlated with the genetic diversity of small populations, but did not significantly affect the diversity of large populations. Contrary to what we expected, secondary-forest populations showed lower divergence relative to populations located in primary patches. Small populations were genetically more diverged compared to large populations. Mantel tests showed no significant isolation by distance and no significant correlation between habitat similarity and genetic differentiation. We conclude that gene flow has probably prevented founder events from being reflected in the present genetic structure of G. urbanum. Gene flow towards low-connectivity populations, however, seemed to be insufficient to counteract the effects of drift in small populations.

Azathioprine hypersensitivity in a renal transplant recipient.

Hypersensitive reactions to azathioprine have been reported infrequently, and always in nontransplanted patients. Here, a renal transplant recipient with a severe hypersensitive reaction to azathioprine is described. We suggest that, until recently, hypersensitivity to azathioprine was suppressed in transplant recipients by the association of high doses of corticosteroids. Since the introduction of cyclosporin, azathioprine therapy is usually associated with corticosteroids in a much lower dose, so an increasing occurrence of azathioprine hypersensitivity in transplanted patients might be expected.