Browsing by Subject "Genetic diversity"

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Anthracnose in white lupin : genetic diversity, virulence and host resistance
(2022) Alkemade, Joris A.; Vögele, Ralf
White lupin (Lupinus albus L.) is a grain legume that is known for its high protein content, nutritional quality, efficient nitrogen fixation and unique ability to form specialized cluster roots that support phosphorus uptake. Despite a severe production decline at the end of the past century, white lupin has seen a recent revival to sustain the demand for plant-based protein and reduce Europe’s dependency on imported soybean. A major problem in (white) lupin cultivation is anthracnose disease, caused by the globally dispersed, seed- and air-borne fungal pathogen Colletotrichum lupini. This PhD thesis aims to provide insights on how to sustainably control anthracnose disease in white lupin in order to support its re-introduction into European cultivation systems. It describes (i) the genetic diversity, phylogeography and virulence of C. lupini, (ii) the development of a high-throughput phenotyping protocol to reliably screen white lupin germplasm, (iii) a genome-wide association study identifying resistance candidate genes and (iv) the exploration of effective seed treatments to reduce the primary pathogen inoculum. Multi-locus phylogeny and morphological characterization of 39 C. lupini isolates showed that diversity is greater than previously reported, distinguishing a total of six genetic groups and ten distinct morphotypes. Highest diversity was found across the South American Andes, indicating it to be the center of origin of C. lupini. Results reveal that the current pandemic is caused by strains belonging to genetic group II, which are spread globally, and are genetically and morphologically uniform. Group II isolates were shown to be highly aggressive on tested white and Andean lupin accessions. Isolates belonging to the other five genetic groups were mostly found locally and showed distinct virulence patterns on both white and Andean lupin accessions. Despite its uniformity, it was shown that two highly virulent group II isolates from Chile could overcome resistance of elite white lupin breeding material, stressing the need to implement phytosanitary protocols for international seed transports. A reliable high-throughput phenotyping tool was developed to identify anthracnose resistance in white lupin germplasm and study pathogen-host interactions. Phenotyping under controlled conditions, performing stem wound inoculation on seedlings, showed to be applicable for high-throughput and its disease scores strongly correlated with two-year Swiss field disease assessments (r = 0.95) and yield (r = -0.64). Phenotyping a diverse set of 40 white lupin accessions revealed eight accessions with improved resistance to anthracnose, which can be incorporated into white lupin breeding programs. A genome-wide association study (GWAS) was done to dissect the genetic architecture for anthracnose resistance in white lupin. White lupin genotypes, collected from the center of domestication and traditional cultivation regions, were screened for anthracnose resistance and characterized through genotyping-by-sequencing (GBS). GWAS revealed two significant SNPs associated with anthracnose resistance on gene Lalb_Chr05_g0216161, encoding a RING zinc-finger E3 ubiquitin ligase potentially involved in plant immunity. Further validation experiments are now required to confirm involvement in plant resistance. Population analysis showed a remarkably fast linkage disequilibrium (LD) decay, weak population structure and grouping of commercial varieties with landraces, corresponding to the slow domestication history and scarce breeding efforts in white lupin. A total of eleven different seed treatments was tested in field trials in Switzerland between 2018 and 2021 to identify treatments that reduce C. lupini infection levels in white lupin. Treatments consisted of hot water, steam, electron, long term storage, vinegar, plant extracts and biological control agents (BCAs). The BCAs were tested under controlled conditions for potential antagonistic activity against C. lupini during white lupin infection prior to field trials. Results showed that long term storage and vinegar treatments can successfully reduce disease incidence and increase yield to levels similar to those observed for certified seeds. In order to sustainably and effectively control anthracnose disease in white lupin, an integrative approach, including modern breeding efforts, disease prevention strategies and mixed cropping systems, is recommended. Further research is required to increase our understanding on white lupin-C. lupini interaction and to identify genetic regions involved in resistance or virulence, respectively, which could greatly support white lupin breeding. This thesis provides the basis to further explore C. lupini population dynamics, virulence and host-speciation, white lupin resistance mechanisms and sustainable ways to control anthracnose disease in order to further facilitate successful white lupin cultivation.
Assessing the Genetic Diversity in Crops with Molecular Markers: Theory and Experimental Results with CIMMYT Wheat and Maize Elite Germplasm and Genetic Resources
(2004) Reif, Jochen Christoph; Melchinger, Albrecht E.
Genetic diversity is a valuable natural resource and plays a key role in future breeding progress. Germplasm collections as a source of genetic diversity must be well-characterized for an efficient management and effective exploitation. The advent of PCR-based molecular markers such as sim-ple sequence repeats (SSRs) has created an opportunity for fine-scale genetic characterization of germplasm collections. The objective of this research was to optimize the utilization of genetic re-sources conserved at the International Wheat and Maize Improvement Center (CIMMYT), with the aid of DNA markers. Choice of suitable dissimilarity measures is important to facilitate the interpretation of findings from DNA marker studies on a theoretically sound basis. The objective of a theoretical study was to examine 10 dissimilarity coefficients widely used in germplasm surveys, with special focus on applications in plant breeding and seed banks. The distance and Euclidean properties of the dissimi-larity coefficients were investigated as well as the underlying genetic models. Application areas for different coefficients were suggested on the basis of the theoretical findings. It has been claimed that plant breeding reduces genetic diversity in elite germplasm, which could seriously jeopardize the continued ability to improve crops. The objectives of the presented ex-perimental study with wheat were to examine the loss of genetic diversity during (i) domestication of the species, (ii) change from traditional landrace cultivars (LC) to modern breeding varieties, and (iii) intensive selection over 50 years of international breeding. A sample of 253 CIMMYT or CIMMYT-related modern wheat cultivars, LC, and Triticum tauschii accessions were characterized with up to 90 SSR markers covering the entire wheat genome. A loss of genetic diversity was ob-served from T. tauschii to LC and from LC to the elite breeding germplasm. Wheat genetic diver-sity was narrowed from 1950 to 1989, but was enhanced from 1990 to 1997. The results indicate that breeders averted the narrowing of the wheat germplasm base and subsequently increased the genetic diversity through the introgression of novel materials. The LC and T. tauschii contain nu-merous unique alleles that were absent in modern wheat cultivars. Consequently, both LC and T. tauschii represent useful sources for broadening the genetic base of elite wheat breeding germ-plasm. In the 1980's, CIMMYT generated more than 100 maize populations and pools but little is known about the genetic diversity of this germplasm. The objective of the study with 23 CIMMYT maize populations was to characterize their population genetic structure with SSRs. The populations adapted to tropical, subtropical intermediate-maturity, subtropical early-maturity, and temperate mega-environments (ME) were fingerprinted with 83 SSR markers. Estimates of genetic differen-tiation between populations revealed that most of the molecular variation was found within the populations. Principal coordinate analysis based on allele frequencies of the populations revealed that populations adapted to the same ME clustered together and, thus, supported clearly the ME structure. Novel strategies were suggested to optimize the conservation of the genetic diversity within and among the populations. Heterotic groups and patterns are of fundamental importance in hybrid breeding. The objective of the presented study with a subset of 20 out of the 23 maize populations was to investigate the rela-tionship between heterosis and genetic distance determined with SSR markers. The published data of three diallels and one factorial trial evaluated for grain yield were re-analyzed to calculate het-erosis in population hybrids. Correlations of squared modified Rogers distance and heterosis were mostly positive and significant, but adaption problems caused deviations in some cases. For popu-lations adapted to the target regions, genetic distance can be used as a further criterion in the search for promising heterotic patterns and groups. For intermediate- and early-maturity subtropical germ-plasm, two heterotic groups were suggested, consisting of a flint and dent composite. For the tropi-cal germplasm, it was possible to assign population (Pop29) to the established heterotic group A and propose new heterotic groups (Pop25, Pop43). Our experimental results corroborate that SSRs are a powerful tool to (i) detect relationships among different germplasm, (ii) assess the level of genetic diversity present in germplasm pools and its flux over time, and (iii) search for promising heterotic groups for hybrid breeding in complementa-tion to field trials.
Design and assessment of breeding strategies for hybrid wheat in Europe
(2018) Boeven, Philipp Hans Günter; Würschum, Tobias
Wheat is one of the top three global staple crops, possesses the largest global cultivation area, and plays a key role for the world’s future food security. However, its projected yield increase is insufficient to meet the future food and feed demand of an ever-growing world population. Consequently, the rate of breeding progress and productivity of wheat must be increased. Unfortunately, current wheat line breeding has a low return on investment mainly due to high levels of farm saved seeds, which makes wheat less attractive for the plant breeding industry and leads to lower investments and progress compared to other crops where the hybrid technology is established. Hybrid breeding is a worldwide success story in many crops but is not yet established in wheat. Hybrid wheat promises increased yield gain due to the exploitation of heterosis, higher yield stability and stabilized return on investments for breeding companies which warrants further investment and breeding progress in this important stable crop. The self-pollinating nature of wheat is a major bottleneck for hybrid seed production and efficient hybrid wheat breeding requires the redesign of the wheat floral architecture to enhance cross-pollination. Furthermore, the longterm success of hybrid wheat is crucially dependent on the establishment of heterotic groups, on the identification of a high yielding heterotic pattern, and finally, on the realized amount of heterosis and hybrid performance. Therefore, the main objectives of my thesis research were to: (i) analyze the genetic diversity and adaptation in a global winter wheat collection and evaluate how diversity trends could be used to support the development of heterotic groups in wheat; (ii) assess the relationship between heterosis and genetic distance under maximized diversity and evaluate the usefulness of exotic germplasm for hybrid wheat breeding; (iii) dissect the genetic architecture underlying male floral traits in wheat to enable genomics-assisted breeding approaches and investigate the trait seed set which is most crucial for an efficient hybrid seed production. The analyses of genetic diversity in a large worldwide panel of 1,110 winter wheat varieties released during the past decades showed no major population structure but revealed genetically distinct subgroups. Most of the global diversity trends could be explained by breeding history and were associated with geographical origin and long-term domestication. We found that the frequency of the copy number variants at the Photoperiod-B1 (Ppd-B1) and the Vernalization-A1 (Vrn-A1) loci reflect wheat adaptation to the environmental conditions of the different regions of origin. Thus, adaptation issues add an additional layer of complexity and hamper the direct introgression of genetic diversity to support the genetic divergence between heterotic pools. Based on all these analyses, we proposed HyBFrame, a unified framework illustrating how global wheat genetic diversity can be used to support and accelerate reciprocal recurrent selection for the development of genetically distinct heterotic groups in wheat. In a second experiment, we produced 2,046 wheat hybrids by crossing elite with elite lines as well as elite with exotic lines and performed multi-environmental field trials. Interestingly, we found an average midparent heterosis of about 10% in elite crosses as well as in exotic crosses and observed no evidence for a breakdown of heterosis under maximized genetic distance among the hybrid parents. Genetic distance based on genome-wide molecular markers revealed only a very weak association with midparent heterosis for grain yield. Here, we elaborated a functional Rogers’ distance giving weight to heterosis loci and observed a strong positive association between heterosis and this novel distance measure. Hence, considering the genetic architecture of heterosis revealed a more accurate picture of the relationship between heterosis and genetic distance. In addition, the genetic architecture of heterosis in wheat is crucially dependent on the genetic background. We found that a higher number of negative dominance and dominance-by-dominance epistatic effects can reduce the level of absolute heterosis in wide crosses between exotic lines and elite testers. Moreover, hybrid performance in wheat is mainly driven by parental per se performance. Thus, elite lines are favorable for hybrid breeding and should be employed as the starting material for heterotic grouping. Hybrid seed production is the major bottleneck for hybrid wheat breeding and explains the low market share of hybrid wheat varieties. Seed set on the female plants in crossing blocks is the most crucial trait for hybrid seed production in wheat. We tested 31 male lines and evaluated the hybrid seed set on two female tester lines in crossing blocks. Seed set showed a large genotypic variation and a high heritability suggesting that recurrent selection for increased seed set is feasible. The major problem is the synchronized flowering between male and female lines, making the evaluation of seed set in large panels very complex and difficult. Hence, indirect male floral traits with high correlation to the trait seed set would be promising to breed for improved hybrid seed production. We found a strong association between seed set and visual anther extrusion, underscoring that indirect male floral traits have a high potential for preliminary male screenings. We also dissected the genetic architecture underlying promising male floral traits and assessed the potential of genomics-assisted approaches for their improvement. We employed a panel of 209 diverse wheat lines and found a complex genetic architecture underlying all male floral traits. The Reduced height gene Rht-D1 was identified as the only major QTL, for which the commonly used height-reducing allele showed negative effects on male floral traits. This genetic architecture with many moderate- or small-effect QTL limits classical marker-assisted selection. In contrast, genomic prediction yielded moderate to high prediction abilities for anther extrusion. Finally, we proposed a breeding scheme to increase cross-pollination in wheat based on a combination of phenotypic and genomics-assisted selection. Taken together, hybrid breeding in wheat is a very promising approach and the next years will show if all of the current issues can be solved. This thesis research contributed to breeding strategies for hybrid wheat breeding and to the general understanding of heterosis in crops.
Diversity in the tropical multipurpose shrub legumes Cratylia argentea (Desv.) O. Kuntze and Flemingia macrophylla (Willd.) Merrill.
(2006) Andersson, Meike S.; Schultze-Kraft, Rainer
Cratylia argentea (Desv.) O. Kuntze and Flemingia macrophylla (Willd.) Merrill are promising tropical multipurpose shrub legumes. Both are drought-tolerant, well adapted to low-fertility, acid soils, and especially suited for low-input smallholder production systems in the sub-humid and humid tropics. They can be used e.g. as dry season forage supplementation, live soil cover or mulch, erosion barrier hedges, and shade-providing shrubs in young coffee and cocoa plantations. Germplasm collections were assembled from the wild-legume flora in Brazil (C. argentea) and Southeast Asia (F. macrophylla), but research and development are so far based on only a few accessions. Knowledge about the extent of genetic diversity within these collections is very limited. In addition, the potential utilization of F. macrophylla is so far limited by poor forage quality and acceptability of the few evaluated accessions. The objective of the present study, conducted in a research cooperation with the International Centre for Tropical Agriculture (CIAT), Cali, Colombia, was to assess the diversity in the germplasm collections of C. argentea (38 accessions) and F. macrophylla (69 accessions) in terms of morphological and phenological traits, agronomic and forage quality traits, and molecular markers, and to identify superior genotyes. Based on these different characterization approaches, the objective was furthermore to establish core collections for F. macrophylla, and to compare and validate the different strategies, giving particular consideration to their practical implications (time and cost efficiency) for the application to small collections of perennial wild tropical legumes. Cratylia argentea High diversity in terms of phenological and agronomic as well as forage quality traits was detected in the collection, with scope for plant improvement in terms of higher dry season DM production. Accessions CIAT 18674 and 22406 were identified as promising for further evaluation since they were similar to the commercial cultivar "Veraniega" in terms of forage quality, and superior in terms of DM production, particularly in the dry season. Molecular marker analysis with RAPDs showed that the genetic diversity in the collection was relatively low and fairly homogeneously distributed. Accessions CIAT 22373, 22378, 22380, 22381 and 22411 were identified as possible duplicates. Flemingia macrophylla High diversity in terms of morphological and agronomic as well as forage quality traits was detected among the 69 accessions. The identification of four morphotypes in the collection probably has taxonomic implications. Scope for plant improvement was identified with respect to forage quality - one of the species´ main limitations. Accessions CIAT 18437, 21083 and 21090 had similar DM production and higher digestibility than the control accession, and were virtually free of extractable condensed tannins. Problems with low palatability and low seed production of these promising accessions need to be further studied. Genetic diversity in F. macrophylla was higher than in C. argentea, and corresponded closely to the four morphotypes revealed by conventional characterization. Various duplicate accessions were identified, and evidence was provided that the non-Asian accessions are not native to their collection site regions, but rather introduced from Southeast Asia. The results have direct applications for plant improvement of these promising multipurpose legumes. The superior genotypes selected in this study will be used in work with farmers in CIAT-research sites in Central America and distributed to partners. It must be recognized, however, that the diversity assessed is influenced by the climatic and edaphic conditions at the site where the studies were conducted. Therefore, multilocational trials should be considered with a selected subset (including the promising accessions) of C. argentea and F. macrophylla i) to assess the extent of genotype x environment interaction, and ii) to identify genotypes with consistently high performance in a range of distinct environments. Research on the reproduction system of both species is urgently required to determine the potential extent and impact of outcrossing. Beyond the immediate application of these species for farmer utilization, the results of the use and comparison of different approaches to assess diversity and to establish core collections can help to improve germplasm management and characterization of wild tropical legume species in general. Random sampling has been identified as a valuable and resource-efficient strategy for the creation of core collections when no additional information about accessions is available, and in the absence of adequate funds. The validation of the findings of this study with a broader range of perennial tropical wild legumes is necessary to assess their applicability to other species.
Genetic diversity in germplasm of cornsalad (Valerianella locusta L.), radish (Raphanus sativus L.), and celeriac (Apium graveolens L. var. rapaceum), investigated with PCR-based molecular markers
(2004) Muminovic, Jasmina; Melchinger, Albrecht E.
During the last couple of decades, production and economic importance of cornsalad (Valerianella locusta L.; fam. Valerianaceae), radish (Raphanus sativus L. var. sativus convar. radicula; fam. Brassicaceae), and celeriac (Apium graveolens L. var. rapaceum; fam. Apiaceae) have been considerably increasing in Europe. Nevertheless, genetic diversity currently utilized for breeding cornsalad, radish, and celeriac is narrow, whereas their germplasm collections in gene banks are relatively poor. Assessment of genetic diversity among breeding materials and genetic resources is an important consideration for the optimal design of further breeding programs. The major objective of this study was to investigate genetic diversity in germplasm of cornsalad, radish, and celeriac, applying amplified fragment length polymorphisms (AFLPs) and inter simple sequence repeats (ISSRs) molecular markers. In particular, the objectives were to (i) analyze relationships among breeding materials of the three vegetable crops (referred to as elite germplasm), as well as among their formerly grown varieties, gene bank and botanical garden accessions (referred to as exotic germplasm), (ii) reveal genetic structure of radish germplasm to establish heterotic pools for hybrid breeding, and (iii) evaluate the usefulness of introducing exotic materials for broadening of the elite germplasm in cornsalad, radish, and celeriac. Average genetic similarity in 34 elite varieties of cornsalad was very high (GS = 0.90), which is comparable with other autogamous crops. The majority of elite varieties clustered closely applying the UPGMA analysis because of a narrow-based germplasm in cornsalad breeding. A substantial level of genetic diversity (GS = 0.47) was detected in 30 cornsalad varieties representing exotic germplasm. Exotic varieties that interspersed the sub-clusters of the elite may serve as a direct genetic resource for broadening the elite cornsalad germplasm base, whereas Valerianella locusta-related species that were distinct from cultivated germplasm can contribute to the introgression of new (resistance) genes. Sixty-eight varieties of cultivated radish (garden and Black radish) created sub-clusters with GS estimates higher than 0.70, thus supporting the assumption that the currently used radish germplasm in Europe relies on a narrow genetic base. Owing to a high degree of heterogeneity and heterozygosity within radish varieties, the detected between-variety diversity was low, but there still was a substantial overall diversity in available radish germplasm. Applying both UPGMA and principal coordinate analyses, Black radish varieties were distinct from garden radish. A further unambiguous division within garden radish germplasm was revealed with the model-based clustering approach. These sub-groups can be employed for establishment of heterotic pools within European modern cultivars of garden radish. In addition, ISSRs can substantially reduce hybrid radish production costs by an early detection of two closely related weed species (R. raphanistrum and R. sativus L. var. sativus convar. sinensis). AFLPs and the evaluation of morphological traits were used to investigate genetic diversity in 34 varieties of elite celeriac germplasm and 35 accessions of exotic germplasm. Only two morphological traits supported the clustering pattern obtained with UPGMA analysis of morphological distance estimates. AFLP-based GS estimates offered a clearer view of diversity present in elite (GS = 0.68-0.95) and exotic germplasm (GS = 0.05-0.95), and clustered the two sets in distinct UPGMA-based sub-clusters. This indicated that only a small fraction of available genetic diversity is exploited for current breeding of celeriac. Exotic celeriac germplasm as well as varieties of celery and leaf celery might substantially improve commercial celeriac breeding. Wild relatives of Apium graveolens are valuable resources for the introgression of resistance genes. Regarding the generally high level of GS in celeriac germplasm conserved in the German gene bank, a broadening of the germplasm collection was suggested. This study demonstrated the capacity of molecular markers to be highly discriminating among varieties of cornsalad, radish, and celeriac. AFLP-based genetic similarity estimates in the three vegetable crops (i) allowed the first insight into the genetic diversity and structure present in the germplasm, (ii) offered suggestions for germplasm broadening, and (iii) proposed a way of rationalization and utilization of available germplasm resources.
Genetische Diversität von Echinococcus granulosus sensu stricto
(2017) Ebi, Dennis; Mackenstedt, Ute
The dog tapeworm Echinococcus granulosus sensu stricto, distributed worldwide, is the most important agent of human Cystic Echinococcosis, a zoonotic disease which is considered as one of the priority ‘neglected zoonotic diseases’ by the World Health Organisation. The parasite is known from a large number of host species and is conventionally subdivided into several genotypes. The epidemiological consequences of the variation, however, are still unknown. In the last years several studies on the intraspecific variation of this parasite were published. Most of these studies were based on small numbers of isolates from geographically restricted regions, so that the relevance of that diversity concerning pathogenicity, host specificity, transmission cycles and biogeography is still little understood. To achieve a comprehensive estimate of its genetic diversity, 1085 isolates of E. granulosus s.s. from humans and animal hosts originating from widely different regions of the world were analyzed for the sequence polymorphism of a 1609 bp fragment of the mitochondrial cox1 gene. For analysis haplotype networks, diversity and neutrality indices of subpopulations of different geographic origin were compared. To improve the analytical efficiency of the sequence-data, a LabVIEW application was written. It could be shown that most diverse sub-population is found in the Middle Eastern region and that the genetic diversity is lower in the other analyzed regions, which confirms a previous tentative hypothesis on the origin of the domestic cycle of E. granulosus s.s. in that region. Records of identical haplotypes in different regions, inclusion of published data, calculation of parameters of population genetics and haplotype network analysis led to hypotheses explaining the distribution history of E. granulosus s.s.. This in line with fossil and molecular data on domestic sheep as the principal final host. A set of new and easily calculable genetic diversity indices were developed and their added value was explained. Additionally, a statistical resampling experiment was programmed which showed that at least 40 well distributed samples are necessary to provide a reliable estimate of the genetic diversity of the parasite in a region. Using an extension for that software, it was shown that the newly developed diversity indices do not strongly depend on the number of analyzed samples, and provide appropriate tools to assess the genetic diversity of parasite populations.
High-throughput field phenotyping reveals genetic variation in photosynthetic traits in durum wheat under drought
(2021) Zendonadi dos Santos, Nícolas; Piepho, Hans‐Peter; Condorelli, Giuseppe Emanuele; Licieri Groli, Eder; Newcomb, Maria; Ward, Richard; Tuberosa, Roberto; Maccaferri, Marco; Fiorani, Fabio; Rascher, Uwe; Muller, Onno
Chlorophyll fluorescence (ChlF) is a powerful non‐invasive technique for probing photosynthesis. Although proposed as a method for drought tolerance screening, ChlF has not yet been fully adopted in physiological breeding, mainly due to limitations in high‐throughput field phenotyping capabilities. The light‐induced fluorescence transient (LIFT) sensor has recently been shown to reliably provide active ChlF data for rapid and remote characterisation of plant photosynthetic performance. We used the LIFT sensor to quantify photosynthesis traits across time in a large panel of durum wheat genotypes subjected to a progressive drought in replicated field trials over two growing seasons. The photosynthetic performance was measured at the canopy level by means of the operating efficiency of Photosystem II (Fq′/Fm′) and the kinetics of electron transport measured by reoxidation rates (Fr1′ and Fr2′). Short‐ and long‐term changes in ChlF traits were found in response to soil water availability and due to interactions with weather fluctuations. In mild drought, Fq′/Fm′ and Fr2′ were little affected, while Fr1′ was consistently accelerated in water‐limited compared to well‐watered plants, increasingly so with rising vapour pressure deficit. This high‐throughput approach allowed assessment of the native genetic diversity in ChlF traits while considering the diurnal dynamics of photosynthesis.