Diagnosis of Soybean Diseases Caused by Fungal and Oomycete Pathogens: Existing Methods and New Developments

Soybean (Glycine max) acreage is increasing dramatically, together with the use of soybean as a source of vegetable protein and oil. However, soybean production is affected by several diseases, especially diseases caused by fungal seed-borne pathogens. As infected seeds often appear symptomless, diagnosis by applying accurate detection techniques is essential to prevent propagation of pathogens. Seed incubation on culture media is the traditional method to detect such pathogens. This method is simple, but fungi have to develop axenically and expert mycologists are required for species identification. Even experts may not be able to provide reliable type level identification because of close similarities between species. Other pathogens are soil-borne. Here, traditional methods for detection and identification pose even greater problems. Recently, molecular methods, based on analyzing DNA, have been developed for sensitive and specific identification. Here, we provide an overview of available molecular assays to identify species of the genera Diaporthe, Sclerotinia, Colletotrichum, Fusarium, Cercospora, Septoria, Macrophomina, Phialophora, Rhizoctonia, Phakopsora, Phytophthora, and Pythium, causing soybean diseases. We also describe the basic steps in establishing PCR-based detection methods, and we discuss potentials and challenges in using such assays

Pushing forward white lupin as a local source for protein and nitrogen in Central Europe

White lupin (Lupinus albus L.) is a promising leguminous crop. Europe is fully dependent on protein and nitrogen fertilizer imports. This has tremendous negative effects both in Europe and the producing countries, such as loss of terrestrial biodiversity, pollution of freshwater, increase of greenhouse gases and soil acidification. Diverse crop-rotations with a substantial amount of pulses are a proven solution. The protein composition and yield potential of white lupin suggest that it could become the ‘Soy of the North’. Currently, the seed-borne pathogen Colletotrichum lupini is substantially impeding the cultivation of white lupin in Central Europe. We developed a DNA-based diagnostic test to identify and quantify the fungal pathogen in plants and seeds. This technique will allow us to improve our understanding of the Colletotrichum lupini life cycle and, thereby, lay the basis for an advanced resistance breeding approach

Genetic mapping of anthracnose resistance in white lupin

White lupin (Lupinus albus) is a valuable grain legume with a high protein content and quality, contributing to soil fertility (Monteiro et al., 2014, Lambers et al., 2013). Its high yield potential could make it a sustainable alternative for imported soybean in Europe (Lucas et al., 2015). However, lupin anthracnose, caused by the air- and soilborne fungus Colletotrichum lupini severely limits cultivation as low levels of seed infestation can already cause total yield loss (Talhinhas et al., 2016). Host resistance is crucial for managing anthracnose but a better insight into the genetic basis is required. We developed a high-throughput phenotyping tool that identifies field-relevant anthracnose resistance under controlled conditions. For inoculation, we identified a local, highly virulent C. lupini strain. Phylogenetic analyses revealed that the strain belongs to a globally dispersed genetic group corresponding to Dubrulle et al.’s (2020) C. lupini group II. Using the developed tool we phenotyped a diverse collection of 200 white lupin accessions, revealing a strong segregation between susceptible and resistant plants, potentially holding novel sources of resistance. Genotyping-bysequencing was performed and the generated single-nucleotide polymorphic markers (SNPs) are currently being used for genetic mapping. Quantitative trait loci (QTLs) for anthracnose resistance will be presented aiding to improve and speed up white lupin breeding programs

Genome‑wide association study reveals white lupin candidate gene involved in anthracnose resistance

White lupin (Lupinus albus L.) is a re-emerging protein crop and promising alternative to soybean. Its cultivation, however, is severely threatened by anthracnose disease caused by the fungal pathogen Colletotrichum lupini. To dissect the genetic architecture for anthracnose resistance, genotyping-by-sequencing (GBS) was performed on white lupin accessions collected from the center of domestication and traditional cultivation regions. GBS resulted in 4,611 high-quality single-nucleotide polymorphisms (SNPs) for 181 accessions, which were combined with resistance data observed under controlled conditions to perform a genome-wide association study (GWAS). Obtained disease phenotypes were shown to highly correlate to overall three-year disease assessments under Swiss field conditions (r > 0.8). GWAS results identified two significant SNPs associated with anthracnose resistance on gene Lalb_Chr05_g0216161 encoding a RING zinc-finger E3 ubiquitin ligase which is potentially involved in plant immunity. 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 scarcity of modern breeding efforts in white lupin. Together with 15 highly resistant accessions identified in the resistance assay, our findings show promise for further crop improvement. This study provides the basis for marker-assisted selection, genomic prediction and studies aimed at understanding anthracnose resistance mechanisms in white lupin and contributes to improving breeding programs worldwide

Uromyces fabae: development, metabolism, and interactions with its host Vicia faba

This MiniReview is intended to provide an overview of the current knowledge regarding cytological, physiological, and molecular aspects of Uromyces fabae. For almost five decades this rust fungus has served as a model system to gain insight into the features characterizing an obligate biotrophic parasite. While earlier studies mostly focused on cytological aspects, later studies were concerned with biochemical and molecular characteristics. Despite the fact that there is still no stable transformation system available for any obligate biotroph, recent molecular analyses have provided new insights into this highly sophisticated interaction of a fungus with its host

Rust haustoria : nutrient uptake and beyond

Haustoria are morphological features of an extremely successful class of plant parasites, the obligate biotrophs. The broad phylogenetic spectrum of organisms producing haustoria suggests that these structures have arisen many times in the course of evolution and represent specific adaptations of these organisms to the close interaction with their respective host plants. This close interaction and the fact that these structures cannot be produced in vitro have hampered an analysis of the roles of haustoria in biotrophy for many decades. Only recently has it become possible to analyse haustorial function at a molecular level. A picture is beginning to emerge indicating that haustoria do not only serve in nutrient uptake a task postulated for these elements ever since their discovery. Moreover, they seem to perform enormous biosynthetic duties. They also seem to be engaged in the suppression of host defense responses and in redirecting or reprogramming the host s metabolic flow. This review intends to summarize current knowledge about the structure and function especially of rust haustoria

Impact of genomics on fungal biology

Fungi represent an extremely diverse and complex class of organisms, and their categorization as lower eukaryotes should by no means be mistaken as meaning low-end. At present, fungi serve as model systems for various aspects of molecular and cellular biology, for example cell cycle regulation, intracellular signaling, metabolic pathway analysis and transcriptional regulation (Feldbrügge et al., 2004; Jiang, 2006; Oliver, 2006). They are also increasingly used on an industrial scale in the production of chemical compounds or in bioremediation (Grimm et al., 2005; Tortella et al., 2005). Some of the most recent and exciting advances within the field of fungal biology have been linked with genomic studies. To explore these, the IXth International Fungal Biology Conference & 16th New Phytologist Symposium entitled Impact of Genomics in Fungal Biology was held in Nancy, France (http://www.newphytologist. org/fungal-genomics/default.htm). The meeting brought together nearly 100 scientists, from all areas of fungal research, and highlighted a wide range of impacts that genome sequencing has and will have on our understanding of fungal biology

A novel beta-glucosidase in Uromyces fabae : feast or fight?

Efficient nutrient mobilization is a key element for biotrophic plant parasites such as the rust fungi. In the course of a cDNA library screen for elements involved in sugar utilization in Uromyces fabae, we identified a sequence with homology to beta-glucosidases. Full-length genomic and cDNA clones of the gene, termed BGL1, were isolated and sequenced. The BGL1 gene comprises 3,372 nucleotides, including nine introns. The open reading frame encompasses 2,532 bases and codes for a polypeptide of 843 amino acids with an apparent molecular mass of 92.4 kDa. Analysis of the polypeptide revealed a potential secretion signal, indicating an extracellular localization of mature BGL1p (89.8 kDa). BGL1 seems to be expressed in all stages of growth, including haustoria, the feeding structures of rust fungi. In the course of immunolocalization studies, the gene product BGL1p was localized in the periphery of intercellular hyphae and haustoria. On the basis of sequence homology, the BGL1 gene was identified as a fungal beta-glucosidase

The Uredinales : Cytology, Biochemistry, and Molecular Biology

Fungi belonging to the order Uredinales are commonly referred to as rust fungi. All members of the Uredinales are parasitic on plants, often causing dramatic losses in various important crop plants (Alexopoulos et al. 1996). Together with the powdery mildew fungi and the downy mildew-causing oomycetes, rust fungi form an extremely successful group of parasites, the obligate biotrophs. The term obligate biotrophic characterizes a specific lifestyle in which the pathogen is absolutely dependent on a living host to complete its life cycle. In turn, the host plant as a whole usually suffers only limited damage over an extended period of time (Staples 2000). By contrast, necrotrophic parasites kill their hosts quickly after infection and subsequently thrive on the dead plant material (Staples 2001). Hemibiotrophic fungi, such as Colletotrichum spp., are characterized by a more or less extended biotrophic phase before switching to necrotrophic growth and killing their host (Perfect and Green 2001). In order to separate the true obligate biotrophic pathogens from hemibiotrophs and necrotrophs we suggest the following six criteria:1. Obligate biotrophs are not culturable in vitro (at least not to a point representing the parasitic phase)2. They form highly differentiated infection structures (variations of the normally tubular cell shape, which are necessary for pathogenesis)3. They have limited secretory activity4. They establish a narrow contact zone separating fungal and plant plasma membranes5. They engage in a long-term suppression of host defense responses6. They form haustoria (specialized hyphae that penetrate host cells).The peculiarities of the lifestyle of obligate biotrophs, paired with their huge economic impact, make rust fungi a versatile field of study at both the fundamental and the applied level. This chapter on Uredinales can by no means cover the complete literature on rust fungi. It is intended to summarize key references, review articles, and books to provide the interested reader witha gateway to more specialized literature on most aspects of research involving rust fungi. Readers new to the field are encouraged to consult the excellent textbooks by Alexopoulos et al. (1996) and Webster and Weber (2007) to gain easier access into the exciting field of mycology in general and obligate biotrophic plant parasites like the rust fungi in particular