Genetic diversity and aggressiveness of Fusarium virguliforme isolates across the Midwestern United States

Sudden death syndrome (SDS) of soybean is a damaging disease caused by the fungus Fusarium virguliforme. Since this pathogen was first reported in the southern US state of Arkansas in 1971, it has spread throughout the Midwestern U.S. The SDS pathogen primarily colonizes roots but also produces toxins that translocate to and damage leaves. Previous studies detected little to no genetic differentiation among isolates, suggesting F. virguliforme in North America has limited genetic diversity and a clonal population structure. Yet, isolates vary in virulence to roots and leaves. We characterized a set of F. virguliforme isolates from the Midwestern U.S. representing a south to north latitudinal gradient from Arkansas to Minnesota. Ten previously tested microsatellite loci were used to genotype isolates and plant assays were conducted to assess virulence. Three distinct population clusters were differentiated across isolates. Although isolates ranged in virulence classes from low to very high, little correlation was found between virulence phenotype and cluster membership. Similarly, population structure and geographic location were not highly correlated. However, the earliest diverging cluster had the lowest genetic diversity and was detected only in southern states, while the other two clusters were distributed across the Midwest and were predominant in Minnesota. One of the Midwestern clusters had the greatest genetic diversity and was found along the northern edge of the known distribution. The results support three genetically distinct population clusters of F. virguliforme in the U.S., with two clusters contributing most to spread of this fungus across the Midwest.

Influence of Fusarium virguliforme Temporal Colonization of Corn, Tillage, and Residue Management on Soybean Sudden Death Syndrome and Soybean Yield

The asymptomatic host range of Fusarium virguliforme includes corn, a common crop rotated with soybean that we hypothesize may alter F. virguliforme population dynamics and disease management. A field-based approach explored the temporal dynamics of F. virguliforme colonization of corn and soybean roots under different tillage and residue managements. Experiments were conducted in IA, IN, MI, WI, and Ontario, Canada from 2016 to 2018. Corn and soybean roots were sampled at consecutive time points between 1 and 16 weeks after planting (WAP). DNA was extracted from all roots and analyzed by real-time qPCR for F. virguliforme quantification. Trials were rotated between corn and soybean, containing a two x two factorial of tillage (no-tilled or tilled) and corn residue (with or without) in several experimental designs. In 2016, low (ca. 100 fg/10 mg root tissue) F. virguliforme was detected in the inoculated IA, IN and MI locations, and non-inoculated WI corn fields. However, in 2017 greater levels of F. virguliforme DNA were detected in IA, IN and MI across sampling time points. Tillage practices showed inconsistent effects on F. virguliforme root colonization and SDS foliar symptoms among trials and locations. Yet, residue management did not alter root colonization of corn or soybean by F. virguliforme. Plots with corn residue had greater SDS foliar disease index in Iowa in 2016. However, this trend was not observed across the site-years, indicating corn residue may occasionally increase SDS foliar symptoms depending on the disease level, soil and weather factors.

Trichoderma Isolates Inhibit Fusarium virguliforme Growth, Reduce Root Rot, and Induce Defense-Related Genes on Soybean Seedlings

Sudden death syndrome (SDS) caused by Fusarium virguliforme is among the most important diseases affecting soybean in the United States. The use of biological control agents (BCAs) such as Trichoderma spp. can be a valuable resource to suppress F. virguliforme populations. Therefore, this research focused on screening possible BCAs against F. virguliforme and evaluating mycoparasitism and the induction of systemic resistance as mechanisms underlying the antagonistic activity of selected BCAs against F. virguliforme. In total, 47 potential BCAs, including 41 Trichoderma isolates and 6 Mortierella isolates, were screened in a dual-plate assay. The most effective isolates belonged to the Trichoderma harzianum species and were able to inhibit F. virguliforme radial growth by up to 92%. Selected Trichoderma isolates were tested in the greenhouse and in a microplot study. They reduced root rot caused by F. virguliforme when the plants were coinoculated with the pathogen and the BCA. The tested BCA’s ability to reduce F. virguliforme growth may be related to several mechanisms of action, including mycoparasitism and induction of defense-related genes in plants, as revealed by monitoring the expression of defense-related genes in soybean. Our results highlight the potential of native Trichoderma isolates to inhibit F. virguliforme growth and reduce SDS severity, providing the basis for future implementation of biological control in soybean production. More efforts are needed to implement the use of these approaches in production fields, and to deepen the current knowledge on the biology of these highly antagonistic isolates.

Reduction of Sudden Death Syndrome Foliar Symptoms and Fusarium virguliforme DNA in Roots Inoculated With Rhizophagus intraradices

There is increasing interest in incorporating arbuscular mycorrhizal fungi (AMF) into agricultural production because of the benefits they provide, including protection against pathogens and pests. Sudden death syndrome (SDS) of soybean is a devastating disease caused by the soilborne pathogen Fusarium virguliforme. Multiple management methods are needed to control SDS. The relationship between F. virguliforme and AMF is not well documented. The goal of this study was to determine whether soybean plants co-inoculated with F. virguliforme and the AMF species Rhizophagus intraradices showed reduced SDS foliar symptom severity and reduced relative F. virguliforme DNA quantities in soybean roots. Six soybean genotypes were inoculated with F. virguliforme alone or with R. intraradices in a greenhouse experiment. Averaged over the six soybean genotypes, area under the disease progress curve values and relative F. virguliforme DNA quantities were 45 and 28% lower (P < 0.05), respectively, in roots co-inoculated with R. intraradices compared with roots of control plants inoculated with F. virguliforme only. Weight of roots co-inoculated with R. intraradices were 58% higher (P < 0.05) compared with roots of plants not inoculated with R. intraradices. Nutrient analysis showed higher boron, phosphorus, potassium, sodium, and sulfur concentrations in root tissues of plants co-inoculated with R. intraradices compared with plants inoculated with F. virguliforme (P < 0.05). Overall, this study showed that R. intraradices reduced SDS severity and relative F. virguliforme DNA quantities while simultaneously increasing growth and nutrient uptake of plants. Further testing of AMF inoculants in the field will indicate whether incorporating them into soybean SDS management practices will reduce the impact of SDS on soybean production.

Lysobacter enzymogenes Employs Diverse Genes for Inhibiting Hypha Growth and Spore Germination of Soybean Fungal Pathogens

Lysobacter enzymogenes strain C3 (LeC3) is a potential biocontrol agent for plant diseases caused by fungi and oomycetes. Understanding the interaction between LeC3 and soybean pathogens at the molecular level could help improve its biocontrol efficacy. In this study, we obtained mutants with decreased abilities in inhibiting hypha growth of the white mold pathogen Sclerotinia sclerotiorum. Insertion sites for 50 mutants, which no longer inhibited S. sclerotiorum hypha growth in dual cultural assay, were determined and seven mutants were selected for further characterization. These seven mutants also completely lost their abilities in suppressing spore germination of Fusarium virguliforme, the causal agent of soybean sudden death syndrome. Furthermore, mutation of the seven genes, which encode diguanylate cyclase, transcriptional regulators from the TetR family, hemolysin III family channel protein, type IV secretion system VirB10 protein, phenol hydroxylase, and phosphoadenosine phosphosulfate reductase, respectively, led to reduced production or secretion of four extracellular enzymes and heat-stable antifungal factor (HSAF). These results suggest that these seven genes play important roles in L. enzymogenes in suppressing hypha growth and spore germination of fungal pathogens, probably by influencing production or secretion of extracellular enzymes and HSAF.