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

Candidate Effectors From Uromyces appendiculatus, the Causal Agent of Rust on Common Bean, Can Be Discriminated Based on Suppression of Immune Responses

Rust fungi are devastating pathogens for several important crop plants. The biotrophic lifestyle of rust fungi requires that they influence their host plants to create a favorable environment for growth and reproduction. Rust fungi secrete a variety of effector proteins that manipulate host target proteins to alter plant metabolism and suppress defense responses. Because of the obligate biotrophic lifestyle of rust fungi, direct evidence for effector function is difficult to obtain, and so suites of experiments utilizing expression in heterologous systems are necessary. Here, we present results from a yeast cell death suppression assay and assays for suppression of PAMP-triggered immunity (PTI) and effector triggered immunity (ETI) based on delivery of effectors through the bacterial type III secretion system. In addition, subcellular localization was tested using transient expression of GFP fusion proteins in Nicotiana benthamiana through Agrobacterium infiltration. We tested 31 representative effector candidates from the devastating common bean rust pathogen Uromyces appendiculatus. These effector candidates were selected based on features of their gene families, most important lineage specificity. We show that several of our effector candidates suppress plant defense. Some of them also belong to families of effector candidates that are present in multiple rust species where their homologs probably also have effector functions. In our analysis of candidate effector mRNA expression, some of those effector candidates that gave positive results in the other assays were not up-regulated during plant infection, indicating that either these proteins have functions at multiple life stages or that strong up-regulation of RNA level in planta may not be as important a criterion for identifying effectors as previously thought. Overall, our pipeline for selecting effector candidates based on sequence features followed by screening assays using heterologous expression systems was successful in discriminating effector candidates. This work lays the foundation for functional characterization of U. appendiculatus effectors, the identification of effector targets, and identification of novel sources for resistance in common bean

Suppression or Activation of Immune Responses by Predicted Secreted Proteins of the Soybean Rust Pathogen Phakopsora pachyrhizi

Rust fungi, such as the soybean rust pathogen Phakopsora pachyrhizi, are major threats to crop production. They form specialized haustoria that are hyphal structures intimately associated with host-plant cell membranes. These haustoria have roles in acquiring nutrients and secreting effector proteins that manipulate host immune systems. Functional characterization of effector proteins of rust fungi is important for understanding mechanisms that underlie their virulence and pathogenicity. Hundreds of candidate effector proteins have been predicted for rust pathogens, but it is not clear how to prioritize these effector candidates for further characterization. There is a need for high-throughput approaches for screening effector candidates to obtain experimental evidence for effector-like functions, such as the manipulation of host immune systems. We have focused on identifying effector candidates with immune-related functions in the soybean rust fungus P. pachyrhizi. To facilitate the screening of many P. pachyrhizi effector candidates (named PpECs), we used heterologous expression systems, including the bacterial type III secretion system, Agrobacterium infiltration, a plant virus, and a yeast strain, to establish an experimental pipeline for identifying PpECs with immune-related functions and establishing their subcellular localizations. Several PpECs were identified that could suppress or activate immune responses in nonhost Nicotiana benthamiana, N. tabacum, Arabidopsis, tomato, or pepper plants

The haustorial transcriptomes of Uromyces appendiculatus and Phakopsora pachyrhizi and their candidate effector families

Haustoria of biotrophic rust fungi are responsible for the uptake of nutrients from their hosts and for the production of secreted proteins, known as effectors, which modulate the host immune system. The identification of the transcriptome of haustoria and an understanding of the functions of expressed genes therefore hold essential keys for the elucidation of fungus–plant interactions and the development of novel fungal control strategies. Here, we purified haustoria from infected leaves and used 454 sequencing to examine the haustorial transcriptomes of Phakopsora pachyrhizi and Uromyces appendiculatus, the causal agents of soybean rust and common bean rust, respectively. These pathogens cause extensive yield losses in their respective legume crop hosts. A series of analyses were used to annotate expressed sequences, including transposable elements and viruses, to predict secreted proteins from the assembled sequences and to identify families of candidate effectors. This work provides a foundation for the comparative analysis of haustorial gene expression with further insights into physiology and effector evolution

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

Food losses from farm to retail operations: agricultural produces supply chain of Baja Peninsula, México

Objective: To evaluate food losses (FL) volumes generated by farms in Baja California Peninsula, México, of five agricultural commodities. Design/methodology/approach: Baja California Sur (BCS) state was the study area. Information was gathered from a total of 380 sampled chain actors in asparagus, mango, strawberry, orange and tomato by survey and personal interviews. Tobit technique was applied to identify factors that influence FL percentage. Results: Data shows about 11.8% of asparagus is lost during harvesting and distribution, as well as 8.5% of strawberry, 26% of mango, 17.8% of oranges and 3.5% of tomatoes, representing 29.9% loss rate of marketed yield. Limitations on study/implications: This study did not classify commodities in the last steps of the supply chain. The five commodities used in the current study correspond to the more important agricultural produces in BCS, but given changing market, harvesting time and produce availability did not consider the waste of the supply chain. Findings/conclusions: Commodity, type of transportation and distribution, education, and human resources has been identified as influence factors in the volume of FL. This exploratory study fills the void in information in terms of its geographic scope and food group number, and farm owners willing to manage food losses for the purpose of obtaining bioactive compound. Keywords: Food loss, agrifood, desert agriculture, food security, retailObjective: To evaluate food losses (FL) volumes generated by farms in Baja California Peninsula, México, of five agricultural commodities. Design/methodology/approach: Baja California Sur (BCS) state was the study area. Information was gathered from a total of 380 sampled chain actors in asparagus, mango, strawberry, orange and tomato by survey and personal interviews. Tobit technique was applied to identify factors that influence FL percentage. Results: Data shows about 11.8% of asparagus is lost during harvesting and distribution, as well as 8.5% of strawberry, 26% of mango, 17.8% of oranges and 3.5% of tomatoes, representing 29.9% loss rate of marketed yield. Limitations on study/implications: This study did not classify commodities in the last steps of the supply chain. The five commodities used in the current study correspond to the more important agricultural produces in BCS, but given changing market, harvesting time and produce availability did not consider the waste of the supply chain. Findings/conclusions: Commodity, type of transportation and distribution, education, and human resources has been identified as influence factors in the volume of FL. This exploratory study fills the void in information in terms of its geographic scope and food group number, and farm owners willing to manage food losses for the purpose of obtaining bioactive compound