Towards an optimal sampling strategy for assessing genetic variation within and among white clover (Trifolium repens L.) cultivars using AFLP

Cost reduction in plant breeding and conservation programs depends largely on correctly defining the minimal sample size required for the trustworthy assessment of intra- and inter-cultivar genetic variation. White clover, an important pasture legume, was chosen for studying this aspect. In clonal plants, such as the aforementioned, an appropriate sampling scheme eliminates the redundant analysis of identical genotypes. The aim was to define an optimal sampling strategy, i.e., the minimum sample size and appropriate sampling scheme for white clover cultivars, by using AFLP data (283 loci) from three popular types. A grid-based sampling scheme, with an interplant distance of at least 40 cm, was sufficient to avoid any excess in replicates. Simulations revealed that the number of samples substantially influenced genetic diversity parameters. When using less than 15 per cultivar, the expected heterozygosity (He) and Shannon diversity index (I) were greatly underestimated, whereas with 20, more than 95% of total intra-cultivar genetic variation was covered. Based on AMOVA, a 20-cultivar sample was apparently sufficient to accurately quantify individual genetic structuring. The recommended sampling strategy facilitates the efficient characterization of diversity in white clover, for both conservation and exploitation

Genetic diversity and fitness of forest herbs in anthropogenic landscapes.

In gematigde streken werden uitgestrekte bossen omgezet in de huidige do or de mens gedomineerde landschappen, waardoor de biodiversiteit onder d ruk is komen te staan. Zowel de sterk versnipperde landschapsstructuur a ls de verminderde habitatkwaliteit kunnen in principe de genetische vera rming van de overblijvende bosplantenpopulaties bewerkstelligen, wat de leefbaarheid ( fitness ) van deze populaties op korte termijn en hun ada ptatievermogen op lange termijn kan beperken. Ruimtelijk isolatie kan de uitwisseling van genen via pollen en zaad verhinderen, waardoor de gene tische differentiatie tussen populaties toeneemt en de genetische divers iteit van populaties afneemt, door genetische drift, inteelt en stichtin gseffecten in geïsoleerde nieuwe bosfragmenten. Ook een verminderd voort plantingssucces onder suboptimale omstandigheden kan leiden tot genetisc he verarming. Om conservatiemiddelen efficiënt aan te wenden is het cruciaal de invloe d van heterogeniteit in ruimte, bosleeftijd en omgeving op de genetische diversiteit en de fitness van bosplantenpopulaties beter te begrijpen. Drie vrij algemene bosplanten, verschillend in mate van klonaliteit en b estuivingssysteem, werden uitvoerig bestudeerd in Vlaamse anthropogene l andschappen. Genetische diversiteit en stuctuur werd bepaald met genetis che merkertechnieken (AFLP en SSR). Relaties tussen genetische diversite it en fitness en de fitness gevolgen van kruisingen binnen en tussen pop ulaties werden onderzocht. Populatiegenetische patronen van Geel nagelkruid (Geum urbanum), een nie t-klonaal, relatief goed verbreidend kruid, werden het best verklaard do or bosfragmentatie. De invloed van bosleeftijd en bosomgeving was niet b eduidend. De genenstroom naar minder goed geconnecteerde fragmenten leek te beperkt om de genetische erosie en divergentie van kleine populaties , vermoedelijk door genetische drift, tegen te gaan. Een afgenomen genet ische diversiteit leek de fitness van het nageslacht evenwel niet te bep erken. De fitness van kleine en grote populaties verschilde niet beduide nd en uitkruisen leidde niet tot een fitnesstoename ten opzichte van zel fbestuiving. Uitkruisen over grote afstand leidde zelfs tot een matige f itness-afname, zoals verwacht indien de negatieve gevolgen van inteelt a fwezig zijn of niet opwegen tegen de gevolgen van een uitkruisingsdepres sie. De genetische diversiteit van Bosbingelkruid (Mercurialis perennis) en M eiklokje (Convallaria majalis) leek daarentegen voornamelijk bepaald doo r variatie in de lokale omgeving ten gevolge van natuurlijke variatie en menselijk ingrijpen in de oude bossen waar deze soorten voorkomen. Beid e soorten planten zich klonaal voort via rhizomen. Vrij grote populaties van de tweehuizige M. perennis in relatief ongestoorde, donkere omstand igheden hadden meer kans een evenwichtige sekse-ratio en hoge genotypisc he diversiteit te behouden door herhaaldelijke recrutering van zaad. De zaadzetting van deze soort was in beperkte mate gerelateerd aan de plaat selijke genetische diversiteit en de afstand tot mannelijke planten in o vereenstemming met een korte pollendispersieafstand. In enkele populatie s die uit één slechts mannelijke kloon bestonden lijkt geslachtelij ke voortplanting niet meer mogelijk. Uitkruisen tussen populaties verhoo gde de zaadzetting en -kieming niet ten opzichte van kruisingen binnen p opulaties. Het merendeel van de bestudeerde C. majlis populaties bestond uit ampe r één kloon. Populaties bestaande uit meerdere genotypes werden vak er aangetroffen op plaatsen met een dunnere strooisellaag, wat een omgev ingsinvloed via geslachtelijke voortplanting doet vermoeden. In overeens temming met een zelf-incompatibel karakter was vruchtzetting in monoklon ale populaties vrijwel onbestaand. Een gebrekkige geslachtelijke voortpl anting heeft vermoedelijk geleid tot monoklonale of welhaast monoklonale populaties, wat het voortbestaan van deze overblijvende popu laties in het gedrang kan brengen. Hoewel waarchijnlijk initieel gerelat eerd aan de lokale omgeving, heeft ruimtelijke isolatie dit proces mogel ijks versterkt door de instroom van pollen uit naburige populaties te be lemmeren. Deze resulaten suggereren de invloed van soortkenmerken op de impact van habitatfragmentatie. De genetische diversiteit van de twee bestudeerde klonale bosplanten met een vermoedelijk van nature gefragmenteerde struc tuur is waarschijnlijk voornamelijk beïnvloed door de lokale omgeving. Z elf-incompatibiliteit lijkt negatieve fitnessgevolgen gekoppeld aan een te lage genetische diversiteit te versterken terwijl geen relatie tussen genetische diversiteit en fitness in de onderzochte populaties van een hoofdzakelijk zelfbestuivend kruid gevonden werd.TABLE OF CONTENTS Chapter 1. General introduction 1 1.1 GENERAL CONSEQUENCES OF HABITAT DESTRUCTION AND DEGRADATION 3 1.2 FOREST HERBS IN CULTURAL LANDSCAPES 4 1.3 GENETIC DIVERSITY AND FITNESS OF PLANT POPULATIONS IN HUMAN-ALTERED AND DEGRADED LANDSCAPES 7 1.4 SCOPE, AIMS AND THESIS OUTLINE 14 Chapter 2. Landscape genetics of Geum urbanum: effects of habitat age, fragmentation and local environment 21 2.1 SUMMARY 23 2.2 INTRODUCTION 24 2.3 MATERIALS AND METHODS 27 2.4 RESULTS 35 2.5 DISCUSSION 40 2.6 CONCLUSION 44 2.7 ACKNOWLEDGMENTS 44 Chapter 3. Fitness effects of outcrossing in fragmented populations of Geum urbanum 45 3.1 SUMMARY 47 3.2 INTRODUCTION 47 3.3 MATERIALS AND METHODS 50 3.4 DISCUSSION 61 3.5 ACKNOWLEDGEMENTS 67 Chapter 4. Fine-scale genetic and clonal structure of Mercurialis perennis under various degrees of canopy closure 68 4.1 SUMMARY 70 4.2 INTRODUCTION 71 4.3 MATERIALS AND METHODS 73 4.4 RESULTS 82 4.5 DISCUSSION 89 4.6 CONCLUSION 94 4.7 ACKNOWLEDGEMENTS 95 Chapter 5. Pattern of sex ratio variation and genetic diversity in Mercurialis perennis 96 5.1 SUMMARY 98 5.2 INTRODUCTION 99 5.3 MATERIALS AND METHODS 101 5.4 RESULTS 108 5.5 DISCUSSION 111 5.6 CONCLUSIONS 114 5.7 ACKNOWLEDGEMENTS 114 Chapter 6. Reproductive consequences of mate quantity versus mate diversity in Mercurialis perennis 117 6.1 SUMMARY 119 6.2 INTRODUCTION 119 6.3 MATERIALS AND METHODS 122 6.4 RESULTS 129 6.5 DISCUSSION 133 6.6 CONCLUSION 136 6.7 ACKNOWLEDGEMENTS 136 Chapter 7. Genotypic diversity and reproductive sucess of Lilly of the valley and the role of the local forest environment 139 7.1 SUMMARY 141 7.2 INTRODUCTION 142 7.3 MATERIALS AND METHODS 144 7.4 RESULTS 151 7.5 DISCUSSION 155 7.6 CONCLUSIONS 158 7.7 ACKNOWLEDGEMENTS 159 Chapter 8. General discussion and conclusions 160 8.1 OUTLINE OF MAIN RESULTS 161 8.2 INFLUENCE OF CLONALITY 165 8.3 INFLUENCE OF DISPERSAL ABILITY 166 8.4 INFLUENCE OF MATING SYSTEM 168 8.5 CONSERVING THE GENETIC DIVERSITY OF FOREST HERBS? 169 8.6 SHORTCOMINGS AND RESEARCH PERSPECTIVES 171 References 174 Publication list 201 PEER REVIEWED PUBLICATIONS 201 UNDER REVIEW 202nrpages: 204status: publishe

Isolation by 454-sequencing and characterization of polymorphic microsatellite markers in the tetraploid perennial herb Campanula rotundifolia

© 2015, Springer Science+Business Media Dordrecht. Campanula rotundifolia is a perennial polyploid herb with a wide circumpolar distribution growing in a range of grassland, heathland and dune communities. The species is showing signs of strong decline in most parts of its range due to habitat degradation. Eight polymorphic microsatellites were developed using a 454-sequencing approach and subsequently characterized in 323 tetraploid individuals from 15 populations in a fragmented landscape in central Sweden. Between 7 and 26 alleles were observed per locus with observed and expected heterozygosity ranging between 0.71–0.98 and 0.71–0.93, respectively. C. rotundifolia may serve as a model species for studying the relative importance of life-history traits in genetic diversity responses to environmental changes.status: publishe

Population genetic diversity of the clonal self-incompatible herbaceous plant species Linaria vulgaris along an urbanization gradient

How increasing urbanization affects biodiversity is one of the most understudied aspects of global change biology. It is, however, known that it may negatively affect plant population genetic diversity in numerous ways, for example through its negative effects on plant population size, between‐population connectivity, and reproductive success. Therefore, it is important to investigate to what extent different levels of urbanization result in these negative phenomena. Here we used microsatellite markers to investigate urbanization effects on the population genetic structure of 23 populations of the self‐incompatible, partially clonal herb Linaria vulgaris which were sampled across a gradient of urbanization. Clonal diversity as measured by the Pareto‐parameter varied between 1.11 and 2.97 and was negatively correlated to both the degree of urbanization and to population size. Urbanization and population size were not interrelated. The least clonally rich populations also experienced significantly reduced seed set. Irrespective of the degree of urbanization, L. vulgaris populations exhibited strong genetic differentiation (FST = 0.33) and there was no significant correlation between genetic and geographic distances, suggesting low gene flow among populations. In conclusion, we showed that urbanization negatively affected fitness of L. vulgaris populations through decreasing their clonal diversity and reproductive success, an effect that may be exacerbated by the low gene flow between populations. Although the effect was modest, the results could probably be extrapolated to bigger cities where it would be considerably more pronounced.status: publishe

Extremely low genotypic diversity and sexual reproduction in isolated populations of the self-incompatible lily-of-the-valley (Convallaria majalis) and the role of the local forest environment

Background and Aims: Clonal growth is a common phenomenon in plants and allows them to persist when sexual life-cycle completion is impeded. Very low levels of recruitment from seed will ultimately result in low levels of genotypic diversity. The situation can be expected to be exacerbated in spatially isolated populations of obligated allogamous species, as low genotypic diversities will result in low availability of compatible genotypes and low reproductive success. Populations of the self-incompatible forest herb lily-of-the-valley (Convallaria majalis) were studied with the aim of inferring the relative importance of sexual and asexual recruitment. Then the aim was to establish a relationship between genotypic diversity, sexual reproduction and the local forest environment. Methods: Highly polymorphic microsatellite markers were used to investigate clonal diversities and population genetic structure of 20 populations of C. majalis in central Belgium. Key: Results Most of the populations studied consisted of a single genotype and linkage disequilibrium within populations was high, manifesting clonal growth as the main mode of reproduction. A population consisting of multiple genotypes mainly occurred in locations with a thin litter layer and high soil phosphorus levels, suggesting environment-mediated sporadic recruitment from seed. Highly significant genetic differentiation indicated that populations are reproductively isolated. In agreement with the self-incompatibility of C. majalis, monoclonal populations showed very low or even absent fruit set. Conclusions: Lack of sexual recruitment in spatially isolated C. majalis populations has resulted in almost monoclonal populations with reduced or absent sexual reproduction, potentially constraining their long-term persistence. The local forest environment may play an important role in mediating sexual recruitment in clonal forest plant species

Boosting perceived customer orientation as a driver of patient satisfaction

Objective: Several hospitals have embraced customer orientation as a strategy to better meet patients' needs, desires, and wishes. This study therefore investigates how hospitals can boost the extent to which patients feel treated in a customer-oriented way by staff (hereafter, “perceived customer orientation”) and its implications for their satisfaction with the hospital. Design: A cross-sectional study of hospital staff's interpersonal skills (interpersonal quality) and the atmosphere and physical features of the hospital (environmental quality) as drivers of perceived customer orientation and its satisfaction implications. Participants and Setting: Two hundred eighty-nine patients in seven surgery wards and two day-surgery departments of a Belgian hospital. Main Outcome Measures: Perceived customer orientation and patient satisfaction. Results: Our results show that interpersonal and environmental qualities have a positive impact on perceived customer orientation, with the interpersonal skills of nursing staff having a greater effect than those of physicians. Perceived customer orientation, however, matters most for patient satisfaction when the nature of the service involves high-contact frequency. Conclusions: Interpersonal and environmental qualities may help to ensure that patients feel treated in a customer-oriented way, which improves patient satisfaction with the hospital

Rapid loss of genetic variation in a founding population of Primula elatior (Primulaceae) after colonization

Land-use changes and associated extinction/colonization dynamics can have a large impact on population genetic diversity of plant species. The aim of this study was to investigate genetic diversity in a founding population of the self-incompatible forest herb Primula elatior and to elucidate the processes that affect genetic diversity shortly after colonization. AFLP markers were used to analyse genetic diversity across three age classes and spatial genetic structure within a founding population of P. elatior in a recently established stand in central Belgium. Parentage analyses were used to assess the amount of gene flow from outside the population and to investigate the contribution of mother plants to future generations. The genetic diversity of second and third generation plants was significantly reduced compared with that of first generation plants. Significant spatial genetic structure was observed. Parentage analyses showed that 50 % of first and second generation plants did not contribute to seedling recruitment. These results suggest that a small effective population size and genetic drift can lead to rapid decline of genetic diversity of offspring in founding populations shortly after colonization. This multigenerational study also highlights that considerable amounts of gene flow seem to be required to counterbalance genetic drift and to sustain high levels of genetic diversity after colonization in recently established stands

Data from: Rapid buildup of genetic diversity in founder populations of the gynodioecious plant species Origanum vulgare after semi-natural grassland restoration

In most landscapes the success of habitat restoration is largely dependent on spontaneous colonization of plant species. This colonization process, and the outcome of restoration practices, can only be considered successful if the genetic makeup of founding populations is not eroded through founder effects and subsequent genetic drift. Here we used 10 microsatellite markers to investigate the genetic effects of recent colonization of the long-lived gynodioecious species Origanum vulgare in restored semi-natural grassland patches. We compared the genetic diversity and differentiation of fourteen recent populations with that of thirteen old, putative source populations, and we evaluated the effects of spatial configuration of the populations on colonization patterns. We did not observe decreased genetic diversity in recent populations, or inflated genetic differentiation among them. Nevertheless, a significantly higher inbreeding coefficient was observed in recent populations, although this was not associated with negative fitness effects. Overall population genetic differentiation was low (FST = 0.040). Individuals of restored populations were assigned to on average 6.1 different source populations (likely following the ‘migrant pool’ model). Gene flow was, however, affected by the spatial configuration of the grasslands, with gene flow into the recent populations mainly originating from nearby source populations. This study demonstrates how spontaneous colonization after habitat restoration can lead to viable populations in a relatively short time, overcoming pronounced founder effects, when several source populations are nearby. Restored populations can therefore rapidly act as stepping stones and sources of genetic diversity, likely increasing overall metapopulation viability of the study species

Data from: Rapid buildup of genetic diversity in founder populations of the gynodioecious plant species Origanum vulgare after semi-natural grassland restoration

In most landscapes the success of habitat restoration is largely dependent on spontaneous colonization of plant species. This colonization process, and the outcome of restoration practices, can only be considered successful if the genetic makeup of founding populations is not eroded through founder effects and subsequent genetic drift. Here we used 10 microsatellite markers to investigate the genetic effects of recent colonization of the long-lived gynodioecious species Origanum vulgare in restored semi-natural grassland patches. We compared the genetic diversity and differentiation of fourteen recent populations with that of thirteen old, putative source populations, and we evaluated the effects of spatial configuration of the populations on colonization patterns. We did not observe decreased genetic diversity in recent populations, or inflated genetic differentiation among them. Nevertheless, a significantly higher inbreeding coefficient was observed in recent populations, although this was not associated with negative fitness effects. Overall population genetic differentiation was low (FST = 0.040). Individuals of restored populations were assigned to on average 6.1 different source populations (likely following the ‘migrant pool’ model). Gene flow was, however, affected by the spatial configuration of the grasslands, with gene flow into the recent populations mainly originating from nearby source populations. This study demonstrates how spontaneous colonization after habitat restoration can lead to viable populations in a relatively short time, overcoming pronounced founder effects, when several source populations are nearby. Restored populations can therefore rapidly act as stepping stones and sources of genetic diversity, likely increasing overall metapopulation viability of the study species

Effect of Weave and Weft Type on Mechanical and Comfort Properties of Hemp–Linen Fabrics

In this study the influence of fabric weave (plain, twill, and panama) and weft type (flax and hemp yarns) on selected mechanical and comfort properties of six fabrics was analyzed. The results showed that tear and abrasion properties were most affected by the weave. The tensile properties of the linen fabrics were not significantly different when weft hemp yarns were used instead of flax. Fabrics with the same weave seemed to be equally resilient to abrasion regardless of the type of weft. By contrast, the hemp weft yarns favorized the physical and comfort properties of the investigated fabrics. For the same weave, the hemp–linen fabrics were slightly lighter and exhibited lower bulk density, significantly larger air permeability, and improved moisture management properties. Although the results of maximum thermal flux (Qmax) suggested a cooler sensation of the linen fabrics with panama and twill, the hemp–linen fabric with a plain weave seemed to be the optimal choice when a cool sensation was desired. Higher thermal conductivity values also suggested slightly better heat transfer properties of the hemp–linen fabrics, and these were significantly influenced by the weave type. The results clearly indicated the advantages of using hemp for improving physical and specific comfort properties of linen fabrics