A single laccase acts as a key component of environmental sensing in a broad host range fungal pathogen.

Secreted laccases are important enzymes on a broad ecological scale for their role in mediating plant-microbe interactions, but within ascomycete fungi these enzymes have been primarily associated with melanin biosynthesis. In this study, a putatively secreted laccase, Sslac2, was characterized from the broad-host-range plant pathogen Sclerotinia sclerotiorum, which is largely unpigmented and is not dependent on melanogenesis for plant infection. Gene knockouts of Sslac2 demonstrate wide ranging developmental phenotypes and are functionally non-pathogenic. These mutants also displayed indiscriminate growth behaviors and enhanced biomass formation, seemingly as a result of their inability to respond to canonical environmental growth cues, a phenomenon further confirmed through chemical stress, physiological, and transcriptomic analyses. Transmission and scanning electron microscopy demonstrate apparent differences in extracellular matrix structure between WT and mutant strains that likely explain the inability of the mutants to respond to their environment. Targeting Sslac2 using host-induced gene silencing significantly improved resistance to S. sclerotiorum, suggesting that fungal laccases could be a valuable target of disease control. Collectively, we identified a laccase critical to the development and virulence of the broad-host-range pathogen S. sclerotiorum and propose a potentially novel role for fungal laccases in modulating environmental sensing.

Effect of Fungicide Application and Corn Hybrid Class on the Presence of Fusarium graminearum and the Concentration of Deoxynivalenol in Ear and Stalk Parts of Corn (Zea mays) Used for Silage.

In Wisconsin, the use of brown midrib (BMR) corn (Zea mays) hybrids for ensiling and subsequent feeding to dairy cows is quite common. The overall milk production from cows fed silage from BMR hybrids is typically higher than those fed silage made from dual-purpose hybrids. Gibberella diseases (ear and stalk rot) caused by Gibberella zeae (anamorph; Fusarium graminearum) and the accompanying accumulation of the mycotoxin deoxynivalenol (DON) can be significant issues during the field production of BMR hybrids. The work presented here aimed to understand the role of hybrid class on the distribution of F. graminearum DNA and DON in the ear and stalk parts of corn for silage. An ear and stalk partitioned sample experiment was conducted on silage corn from field trials in Arlington, Wisconsin, in 2020 and 2021. The trials were arranged in a randomized complete block design in both years, including one BMR hybrid, one dual-purpose hybrid, and seven fungicide application regimes. Paired ear and stalk samples were physically separated, dried, and ground at harvest before determining the concentration of F. graminearum DNA and DON in each sample. Across both years, the main effects of hybrid, treatment, and plant part were not significant (P > 0.1) on DON concentration. However, the hybrid-by-plant part interaction effect was significant (P < 0.01). Ears of the BMR hybrid accumulated the most DON, whereas the dual-purpose hybrid ears had the lowest DON concentration. The concentrations of DON and F. graminearum DNA were significantly (P < 0.01) and highly correlated in the ear (r = 0.73) but not in the stalk (r = 0.09, P = 0.33). These findings suggest that DON accumulation in the corn ear is a major contributor in the difference observed in the total DON between the hybrid classes. Therefore, growers and researchers are encouraged to focus production and breeding on hybrids in both classes that accumulate less DON in ears, resulting in lower total DON in corn chopped for silage.

Management of Soybean Cyst Nematode and Sudden Death Syndrome with Nematode-Protectant Seed Treatments Across Multiple Environments in Soybean.

As soybean (Glycine max) production continues to expand in the United States and Canada, so do pathogens and pests that directly threaten soybean yield potential and economic returns for farmers. One such pathogen is the soybean cyst nematode (SCN; Heterodera glycines). SCN has traditionally been managed using SCN-resistant cultivars and rotation with nonhost crops, but the interaction of SCN with sudden death syndrome (SDS; caused by Fusarium virguliforme) in the field makes management more difficult. Nematode-protectant seed treatments have become options for SCN and SDS management. The objectives of this study were to evaluate nematode-protectant seed treatments for their effects on (i) early and full season SCN reproduction, (ii) foliar symptoms and root-rot caused by SDS, and (iii) soybean yield across environments accounting for the above factors. Using a standard protocol, field trials were implemented in 13 states and one Canadian province from 2019 to 2021 constituting 51 site-years. Six nematode-protectant seed treatment products were compared with a fungicide + insecticide base treatment and a nontreated check. Initial (at soybean planting) and final (at soybean harvest) SCN egg populations were enumerated, and SCN females were extracted from roots and counted at 30 to 35 days postplanting. Foliar disease index (FDX) and root rot caused by the SDS pathogen were evaluated, and yield data were collected for each plot. No seed treatment offered significant nematode control versus the nontreated check for in-season and full-season nematode response, no matter the initial SCN population or FDX level. Of all treatments, ILEVO (fluopyram) and Saltro (pydiflumetofen) provided more consistent increases in yield over the nontreated check in a broader range of SCN environments, even when FDX level was high.

Impact of Foliar Fungicide Application on the Culturable Fungal Endophyte Community of Soybean Seed in the Midwest United States.

The purpose of our study was to determine whether the application of quinone outside inhibitor (QoI) and pyrazole-carboxamide fungicides as a tank mix would impact the endophyte community of soybean seed. Field trials during 2018 in Iowa, South Dakota, and Wisconsin, U.S.A., investigated the impact of a single combination fungicide spray at early pod set in soybeans. The composition of culturable endophytic fungi in mature soybean seed was assessed on three cultivars per state, with maturity groups (MGs) ranging from 1.1 to 4.7. An unusually wet 2018 season delayed harvest, which led to a high level of fungal growth in grain. The survey included 1,080 asymptomatic seeds that were disinfested and individually placed on 5-cm-diameter Petri plates of acidified water agar. The survey yielded 721 fungal isolates belonging to 24 putative species in seven genera; taxa were grouped into genera based on a combination of morphological and molecular evidence. The dominant genera encountered in the survey were Alternaria, Diaporthe, and Fusarium. The study showed that the fungicide treatment reduced the incidence of Fusarium in Wisconsin seed, increased the incidence of Diaporthe in seed from all states, and had no impact on the incidence of Alternaria. This is one of the first attempts to characterize the diversity of seed endophytes in soybean and the first to characterize the impacts of fungicide spraying on these endophyte communities across three states. Our study provides evidence that the impact of a fungicide spray on soybean seed endophyte communities may be influenced by site, weather, and cultivar maturity group.

Uncovering the environmental conditions required for Phyllachora maydis infection and tar spot development on corn in the United States for use as predictive models for future epidemics.

Phyllachora maydis is a fungal pathogen causing tar spot of corn (Zea mays L.), a new and emerging, yield-limiting disease in the United States. Since being first reported in Illinois and Indiana in 2015, P. maydis can now be found across much of the corn growing regions of the United States. Knowledge of the epidemiology of P. maydis is limited but could be useful in developing tar spot prediction tools. The research presented here aims to elucidate the environmental conditions necessary for the development of tar spot in the field and the creation of predictive models to anticipate future tar spot epidemics. Extended periods (30-day windowpanes) of moderate mean ambient temperature (18-23 °C) were most significant for explaining the development of tar spot. Shorter periods (14- to 21-day windowpanes) of moisture (relative humidity, dew point, number of hours with predicted leaf wetness) were negatively correlated with tar spot development. These weather variables were used to develop multiple logistic regression models, an ensembled model, and two machine learning models for the prediction of tar spot development. This work has improved the understanding of P. maydis epidemiology and provided the foundation for the development of a predictive tool for anticipating future tar spot epidemics.

A global-temporal analysis on Phytophthora sojae resistance-gene efficacy.

Plant disease resistance genes are widely used in agriculture to reduce disease outbreaks and epidemics and ensure global food security. In soybean, Rps (Resistance to Phytophthora sojae) genes are used to manage Phytophthora sojae, a major oomycete pathogen that causes Phytophthora stem and root rot (PRR) worldwide. This study aims to identify temporal changes in P. sojae pathotype complexity, diversity, and Rps gene efficacy. Pathotype data was collected from 5121 isolates of P. sojae, derived from 29 surveys conducted between 1990 and 2019 across the United States, Argentina, Canada, and China. This systematic review shows a loss of efficacy of specific Rps genes utilized for disease management and a significant increase in the pathotype diversity of isolates over time. This study finds that the most widely deployed Rps genes used to manage PRR globally, Rps1a, Rps1c and Rps1k, are no longer effective for PRR management in the United States, Argentina, and Canada. This systematic review emphasizes the need to widely introduce new sources of resistance to P. sojae, such as Rps3a, Rps6, or Rps11, into commercial cultivars to effectively manage PRR going forward.

Integration of Soybean (Glycine max) Resistance Levels to Sclerotinia Stem Rot into Predictive Sclerotinia sclerotiorum Apothecial Models.

Sclerotinia stem rot (SSR) is a major disease of soybean across the Upper Midwest region of the United States. Management of this disease has relied on fungicide applications, but due to the environmental conditions necessary for SSR to develop, many of these applications are unnecessary. To mitigate this, predictive models have been developed using localized weather data for predicting the formation of Sclerotinia sclerotiorum apothecia, the inoculum source of SSR, and these models were integrated into a decision support system called Sporecaster. However, these models do not account for the soybean resistance levels to SSR. In this study, fungicide trials were performed across seven site-years in Wisconsin between 2020 and 2022 examining fungicide applications applied at one of three action thresholds (low, moderate, and high) following Sporecaster recommendations in combination with four soybean varieties representing three SSR resistance levels (susceptible, moderately resistant, and resistant). From these trials, the low and moderate action thresholds resulted in similarly low disease severity index (DIX) levels comparable to the standard across all varieties. However, the low action threshold was most accurate for predicting SSR development in the susceptible variety, and the high action threshold was most accurate for predicting SSR development for the three resistant varieties. Both the susceptible soybean and a moderately resistant line yielded similarly high results. Additionally, the use of all fungicide applications led to similar partial profits at grain sale prices of either $0.44 or $0.55 kg-1. Overall, this study uncovered relationships between soybean resistance levels to SSR and Sporecaster, allowing for improved recommendations for fungicide applications.

Fungicide Sensitivity of Sclerotinia sclerotiorum from U.S. Soybean and Dry Bean, Compared to Different Regions and Climates.

Fungicide use is integral to reduce yield loss from Sclerotinia sclerotiorum on dry bean and soybean. Increasing fungicide use against this fungus may lead to resistance to the most common fungicides. Resistance has been reported in Brazil (Glycine max) and China (Brassica napus subsp. napus), however, few studies have investigated fungicide sensitivity of S. sclerotiorum in the United States. This work was conducted to determine if there was a difference in fungicide sensitivity of S. sclerotiorum isolates in the United States from: (i) dry bean versus soybean and (ii) fields with different frequencies of fungicide application. We further hypothesized that isolates with fungicide applications of a single active ingredient from tropical Brazil and subtropical Mexico were less sensitive than temperate U.S. isolates due to different management practices and climates. The EC50(D) fungicide sensitivity of 512 S. sclerotiorum isolates from the United States (443), Brazil (36), and Mexico (33) was determined using a discriminatory concentration (DC) previously identified for tetraconazole (2.0 ppm; EC50(D) range of 0.197 to 2.27 ppm), boscalid (0.2; 0.042 to 0.222), picoxystrobin (0.01; 0.006 to 0.027), and thiophanate-methyl, which had a qualitative DC of 10 ppm. Among the 10 least sensitive isolates to boscalid and picoxystrobin, 2 presented mutations known to confer resistance in the SdhB (qualitative) and SdhC (quantitative) genes; however, no strong resistance was found. This study established novel DCs that can be used for further resistance monitoring and baseline sensitivity of S. sclerotiorum to tetraconazole worldwide plus baseline sensitivity to boscalid in the United States.

A broadly conserved fungal alcohol oxidase (AOX) facilitates fungal invasion of plants.

Alcohol oxidases (AOXs) are ecologically important enzymes that facilitate a number of plant-fungal interactions. Within Ascomycota they are primarily associated with methylotrophy, as a peroxisomal AOX catalysing the conversion of methanol to formaldehyde in methylotrophic yeast. In this study we demonstrate that AOX orthologues are phylogenetically conserved proteins that are common in the genomes of nonmethylotrophic, plant-associating fungi. Additionally, AOX orthologues are highly expressed during infection in a range of diverse pathosystems. To study the role of AOX in plant colonization, AOX knockout mutants were generated in the broad host range pathogen Sclerotinia sclerotiorum. Disease assays in soybean showed that these mutants had a significant virulence defect as evidenced by markedly reduced stem lesions and mortality rates. Chemical genomics suggested that SsAOX may function as an aromatic AOX, and growth assays demonstrated that ΔSsAOX is incapable of properly utilizing plant extract as a nutrient source. Profiling of known aromatic alcohols pointed towards the monolignol coniferyl alcohol (CA) as a possible substrate for SsAOX. As CA and other monolignols are ubiquitous among land plants, the presence of highly conserved AOX orthologues throughout Ascomycota implies that this is a broadly conserved protein used by ascomycete fungi during plant colonization.

Field and Greenhouse Assessment of Seed Treatment Fungicides for Management of Sudden Death Syndrome and Yield Response of Soybean.

Seed treatments for the management of sudden death syndrome (SDS) caused by Fusarium virguliforme are available in the United States and Canada; however, side-by-side comparisons of these seed treatments are lacking. Sixteen field experiments were established in Illinois, Indiana, Iowa, Michigan, and Wisconsin, United States, and Ontario, Canada, in 2019 and 2020 to evaluate seed treatment combinations. Treatments included a nontreated check (NTC), fungicide and insecticide base seed treatments (base), fluopyram, base + fluopyram, base + saponin extracts from Chenopodium quinoa, base + fluopyram + heat-killed Burkholderia rinojenses, base + pydiflumetofen, base + thiabendazole + heat-killed B. rinojenses, and base + thiabendazole + C. quinoa extracts + heat-killed B. rinojenses. Treatments were tested on SDS moderately resistant and susceptible soybean cultivars at each location. Overall, NTC and base had the most root rot, most foliar disease index (FDX), and lowest yield. Base + fluopyram and base + pydiflumetofen were most effective for managing SDS. Moderately resistant cultivars reduced FDX in both years but visual root rot was greater on the moderately resistant than the susceptible cultivars in 2020. Yield response to cultivar was also inconsistent between the 2 years. In 2020, the susceptible cultivar provided significantly more yield than the moderately resistant cultivar. Treatment effect for root rot and FDX was similar in field and greenhouse evaluations. These results reinforce the need to include root rot evaluations in addition to foliar disease evaluations in the breeding process for resistance to F. virguliforme and highlights the importance of an integrated SDS management plan because not a single management tactic alone provides adequate control of the disease.

Detection and discovery of plant viruses in soybean by metagenomic sequencing.

BACKGROUND: Viruses negatively impact soybean production by causing diseases that affect yield and seed quality. Newly emerging or re-emerging viruses can also threaten soybean production because current control measures may not be effective against them. Furthermore, detection and characterization of new plant viruses requires major efforts when no sequence or antibody-based resources are available. METHODS: In this study, soybean fields were scouted for virus-like disease symptoms during the 2016-2019 growing seasons. Total RNA was extracted from symptomatic soybean parts, cDNA libraries were prepared, and RNA sequencing was performed using high-throughput sequencing (HTS). A custom bioinformatic workflow was used to identify and assemble known and unknown virus genomes. RESULTS: Several viruses were identified in single or mixed infections. Full- or nearly full-length genomes were generated for tobacco streak virus (TSV), alfalfa mosaic virus (AMV), tobacco ringspot virus (TRSV), soybean dwarf virus (SbDV), bean pod mottle virus (BPMV), soybean vein necrosis virus (SVNV), clover yellow vein virus (ClYVV), and a novel virus named soybean ilarvirus 1 (SIlV1). Two distinct ClYVV isolates were recovered, and their biological properties were investigated in Nicotiana benthamiana, broad bean, and soybean. In addition to infections by individual viruses, we also found that mixed viral infections in various combinations were quite common. CONCLUSIONS: Taken together, the results of this study showed that HTS-based technology is a valuable diagnostic tool for the identification of several viruses in field-grown soybean and can provide rapid information about expected viruses as well as viruses that were previously not detected in soybean.

Presence and Correlation of Fusarium graminearum and Deoxynivalenol Accumulation in Silage Corn Plant Parts.

Corn silage, made from Zea mays, is a high-energy feed that is important for feeding dairy cows. Plant diseases, such as those caused by Fusarium graminearum, can decrease silage corn yields and quality. F. graminearum (teleomorph Gibberella zeae) is an ascomycete fungus that causes Gibberella ear and stalk rot in corn. F. graminearum produces deoxynivalenol (DON), a secondary metabolite toxic to humans and animals. An understanding of the distribution of DON and F. graminearum throughout the corn plant is important for determining the quality of corn silage. A partitioned sample experiment that included two brown midrib silage hybrids and three fungicide treatments was conducted in research plots located in Arlington, WI, U.S.A., in 2018 and 2019. At harvest, stalk and ear parts were physically separated, dried, and ground for analysis. DON concentration (in parts per million) was determined using an enzyme-linked immunosorbent assay, and F. graminearum DNA concentration (in picograms per nanogram) was determined using quantitative PCR. DON and F. graminearum DNA were detected in all samples, demonstrating accumulation of the fungus in stalks and ears of the plant. In 2018, DON contamination was as high as 30 ppm and varied drastically between stalks and ears. In 2019, DON concentrations were much lower (<5 ppm), but were consistently higher in stalk samples than ear samples. Across all samples, DON concentrations and F. graminearum accumulation were highly correlated within the separated stalk (r = 0.78) and ear portions (r = 0.87) but were not correlated between ears and stalks. Depending on the weather and planting conditions in a given year, stalk infections or ear infections may occur by F. graminearum, leading to subsequent DON increases in those respective parts that are independent of each other.

Integration of Row Spacing, Seeding Rates, and Fungicide Applications for Control of Sclerotinia Stem Rot in Glycine max.

Soybean (Glycine max) farmers in the Upper Midwest region of the United States often experience severe yield losses due to Sclerotinia stem rot (SSR). Previous studies have revealed benefits of individual management practices for SSR. This study examined the integration of multiple control practices on the development of SSR, yield, and the economic implications of these practices. Combinations of row spacings, seeding rates, and fungicide applications were examined in multisite field trials across the Upper Midwest from 2017 to 2019. These trials revealed that wide row spacing and low seeding rates individually reduced SSR levels but also reduced yields. Yields were similar across the three highest seeding rates examined. However, site-years where SSR developed showed the highest partial profits at the intermediate seeding rates. This finding indicates that partial profits in diseased fields were reduced by high seeding rates, but this trend was not observed when SSR did not develop. Fungicides strongly reduced the development of SSR while also increasing yields. However, there was a reduction in partial profits due to their use at a low soybean sale price, but at higher sale prices fungicide use was similar to not treating. Additionally, the production of new inoculum was predicted from disease incidence, serving as an indicator of increased risk for SSR development in future years. Overall, this study suggests using wide rows and low seeding rates in fields with a history of SSR while reserving narrow rows and higher seeding rates for fields without a history of SSR.

Host-Induced Gene Silencing of a Sclerotinia sclerotiorum oxaloacetate acetylhydrolase Using Bean Pod Mottle Virus as a Vehicle Reduces Disease on Soybean.

A lack of complete resistance in the current germplasm complicates the management of Sclerotinia stem rot (SSR) caused by Sclerotinia sclerotiorum in soybean. In this study, we used bean pod mottle virus (BPMV) as a vehicle to down-regulate expression of a key enzyme in the production of an important virulence factor in S. sclerotiorum, oxalic acid (OA). Specifically, we targeted a gene encoding oxaloacetate acetylhydrolase (Ssoah1), because Ssoah1 deletion mutants are OA deficient and non-pathogenic on soybean. We first established that S. sclerotiorum can uptake environmental RNAs by monitoring the translocation of Cy3-labeled double-stranded and small interfering RNA (ds/siRNAs) into fungal hyphae using fluorescent confocal microscopy. This translocation led to a significant decrease in Ssoah1 transcript levels in vitro. Inoculation of soybean plants with BPMV vectors targeting Ssoah1 (pBPMV-OA) also led to decreased expression of Ssoah1. Importantly, pBPMV-OA inoculated plants showed enhanced resistance to S. sclerotiorum compared to empty-vector control plants. Our combined results provide evidence supporting the use of HIGS and exogenous applications of ds/siRNAs targeting virulence factors such as OA as viable strategies for the control of SSR in soybean and as discovery tools that can be used to identify previously unknown virulence factors.

Effectors of Plant Necrotrophic Fungi.

Plant diseases caused by necrotrophic fungal pathogens result in large economic losses in field crop production worldwide. Effectors are important players of plant-pathogen interaction and deployed by pathogens to facilitate plant colonization and nutrient acquisition. Compared to biotrophic and hemibiotrophic fungal pathogens, effector biology is poorly understood for necrotrophic fungal pathogens. Recent bioinformatics advances have accelerated the prediction and discovery of effectors from necrotrophic fungi, and their functional context is currently being clarified. In this review we examine effectors utilized by necrotrophic fungi and hemibiotrophic fungi in the latter stages of disease development, including plant cell death manipulation. We define "effectors" as secreted proteins and other molecules that affect plant physiology in ways that contribute to disease establishment and progression. Studying and understanding the mechanisms of necrotrophic effectors is critical for identifying avenues of genetic intervention that could lead to improved resistance to these pathogens in plants.

Disarming the Host: Detoxification of Plant Defense Compounds During Fungal Necrotrophy.

While fungal biotrophs are dependent on successfully suppressing/subverting host defenses during their interaction with live cells, necrotrophs, due to their lifestyle are often confronted with a suite of toxic metabolites. These include an assortment of plant defense compounds (PDCs) which can demonstrate broad antifungal activity. These PDCs can be either constitutively present in plant tissue or induced in response to infection, but are nevertheless an important obstacle which needs to be overcome for successful pathogenesis. Fungal necrotrophs have developed a number of strategies to achieve this goal, from the direct detoxification of these compounds through enzymatic catalysis and modification, to the active transport of various PDCs to achieve toxin sequestration and efflux. Studies have shown across multiple pathogens that the efficient detoxification of host PDCs is both critical for successful infection and often a determinant factor in pathogen host range. Here, we provide a broad and comparative overview of the various mechanisms for PDC detoxification which have been identified in both fungal necrotrophs and fungal pathogens which depend on detoxification during a necrotrophic phase of infection. Furthermore, the effect that these mechanisms have on fungal host range, metabolism, and disease control will be discussed.

Chemotype and Aggressiveness Evaluation of Fusarium graminearum and Fusarium culmorum Isolates from Wheat Fields in Wisconsin.

Fusarium graminearum commonly causes Fusarium head blight on wheat, barley, rice, and oats. F. graminearum produces nivalenol (NIV) and deoxynivalenol (DON) and forms derivatives of DON based on its acetylation sites. The fungus is profiled into chemotypes based on DON derivative chemotypes (3 acetyldeoxynivalenol [3ADON] chemotype; 15 acetyldeoxynivalenol [15ADON] chemotype), and/or the NIV chemotype. This study assessed the Fusarium population found on wheat and the chemotype profile of the isolates collected from 2016 and 2017 in Wisconsin. F. graminearum was isolated from all locations sampled in both 2016 and 2017. Fusarium culmorum was isolated only from Door County in 2016. Over both growing seasons, 91% of isolates were identified as the 15ADON chemotype, while 9% of isolates were identified as the 3ADON chemotype. Aggressiveness was quantified by area under the disease progress curve (AUDPC). The isolates with the highest AUDPC values were from the highest wheat-producing cropping districts in the state. Deoxynivalenol production in grain and sporulation and growth rate in vitro were compared with aggressiveness in the greenhouse. Our results showed that 3ADON isolates in Wisconsin were among the highest in sporulation capacity, growth rate, and DON production in grain. However, there were no significant differences in aggressiveness between the 3ADON and 15ADON isolates. The results of this research detail the baseline frequency and distribution of 3ADON and 15ADON chemotypes observed in Wisconsin. Chemotype distributions within populations of F. graminearum in Wisconsin should continue to be monitored in the future.

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 Iowa, Indiana, Michigan, and Wisconsin, United States and Ontario, Canada from 2016 to 2018. Corn and soybean roots were sampled at consecutive timepoints between 1 and 16 weeks after planting. DNA was extracted from all roots and analyzed by real-time quantitative PCR for F. virguliforme quantification. Trials were rotated between corn and soybean, containing a two-by-two factorial of tillage (no-tilled or tilled) and corn residue (with or without) in several experimental designs. In 2016, low amounts (approximately 100 fg per 10 mg of root tissue) of F. virguliforme were detected in the inoculated Iowa, Indiana, and Michigan locations and noninoculated Wisconsin corn fields. However, in 2017, greater levels of F. virguliforme DNA were detected in Iowa, Indiana, and Michigan across sampling timepoints. Tillage practices showed inconsistent effects on F. virguliforme root colonization and sudden death syndrome (SDS) foliar symptoms among trials and locations. However, 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 that corn residue may occasionally increase SDS foliar symptoms depending on the disease level and soil and weather factors.

Identification of Soybean (Glycine max) Check Lines for Evaluating Genetic Resistance to Sclerotinia Stem Rot.

Soybean production in the upper midwestern United States is affected by Sclerotinia stem rot (SSR) caused by the fungal pathogen Sclerotinia sclerotiorum. Genetic resistance is an important management strategy for this disease; however, assessing genetic resistance to S. sclerotiorum is challenging because a standardized method of examining resistance across genotypes is lacking. Using a panel of nine diverse S. sclerotiorum isolates, four soybean lines were assessed for reproducible responses to S. sclerotiorum infection. Significant differences in SSR severity were found across isolates (P < 0.01) and soybean lines (P < 0.01), including one susceptible, two moderately resistant, and one highly resistant line. These four validated lines were used to screen 11 other soybean genotypes to evaluate their resistance levels, and significant differences were found across genotypes (P < 0.01). Among these 11 genotypes, five commercial and public cultivars displayed high resistance and were assessed during field studies across the upper midwestern United States growing region to determine their response to SSR and yield. These five cultivars resulted in low disease levels (P < 0.01) in the field that were consistent with greenhouse experiment results. The yields were significantly different in fields with disease present (P < 0.01) and disease absent (P < 0.01), and the order of cultivar performance was consistent between environments where disease was present or absent, suggesting that resistance prevented yield loss to disease. This study suggests that the use of a soybean check panel can accurately assess SSR resistance in soybean germplasm and aid in breeding and commercial soybean development.

Diaporthe Seed Decay of Soybean [Glycine max (L.) Merr.] Is Endemic in the United States, But New Fungi Are Involved.

Diaporthe seed decay can compromise seed quality in soybean [Glycine max (L.) Merr.] in the warm and humid production areas of the United States during crop maturation. In the current study, 45 isolates of Diaporthe were recovered from seed sampled from soybean fields affected by Diaporthe-associated diseases in eight U.S. states in 2017. The isolates obtained belonged to 10 species of Diaporthe based on morphology and phylogenetic analyses of the internal transcribed spacer, partial translation elongation factor 1-α, and ß-tubulin gene sequences. The associated species included D. aspalathi, D. caulivora, D. kongii, D. longicolla, D. sojae, D. ueckerae, D. unshiuensis, and three novel fungi, D. bacilloides, D. flavescens, and D. insulistroma. One isolate each of the 10 species was examined for pathogenicity on seed of cultivar Sava under controlled conditions. Seven days postinoculation, significant differences in the percentages of decayed seeds and seedling necrosis were observed among the isolates and the noninoculated control (P < 0.0001). While the isolates of D. bacilloides, D. longicolla, and D. ueckerae caused a significantly greater percentage of decayed seeds (P < 0.0001), the isolate of D. aspalathi caused the greatest seedling necrosis (P < 0.0001). The observation of new fungi causing Diaporthe seed decay suggests the need for a more comprehensive survey in U.S. soybean producing areas since members of the genus Diaporthe appear to form a complex that causes seed decay.