Diseases caused by Fusarium (seedling diseases, root rot and Sudden death syndrome) are yield limiting in the major U.S. soybean production regions. In North Dakota, sudden death syndrome (SDS), caused by F. virguliforme, was confirmed in 2020, and two other Fusarium, F. solani and F. tricinctum, have been implicated to cause root rot. The management tools available to soybean farmers are limited to the use of few seed treatments, and varieties with tolerance to only F. virguliforme. Currently, the distribution of the SDS fungus and other Fusarium species throughout ND is not well understood. However, SDS is appearing in pockets within few ND fields, and we hypothesize that there are several factors that have contributed to the emergence of this disease. As examples, soybeans may be used as a continuous crop in commercial fields because of market prices. Farmers may implement no-till or reduced tillage practices leaving crop debris infested with Fusarium near the soil surface which can infect soybean and rotational crops. Also, it is possible that SDS is showing up in fields where soybean cyst nematode (SCN) may be present. With SCN known to exist in North Dakota’s major soybean producing areas, there is a need to thoroughly understand F. virguliforme and other Fusarium in terms of their prevalence, pathogenicity, and how they interact with the nematode. The information obtained from this research will complement NDSU’s efforts to educate farmers on managing SCN, SDS and other Fusarium with currently available and new tools.

• Characterize the species distribution of Fusarium associated with soybean.
• Characterize the pathogenicity of Fusarium species.
• Determine how the presence of SCN can affect development of SDS.
• Determine the impact of seed treatment on Fusarium.

• A map describing the current geographic distribution of F. virguliforme and other species of Fusarium in ND that will be made available to soybean farmers.
• Information on the potential for soybean crop damage caused by Fusarium
• Identification of soybean lines with root resistance for use in the breeding program to develop and release SDS resistant varieties.
• Information on how SCN impacts development of SDS, and why it is critical to manage the two diseases.
• Information on if the use of seed treatments can maximize soybean’s yield potential, i.e., the maximum yield that can be obtained when stress is absent.

Update:
Midyear report for North Dakota Soybean Council (July 1, 2023 to November 30, 2023)

a. Research Project Title, Principal and Co-Investigators

Title: Understanding How Fusarium Affects Soybean in North Dakota and Development of Disease Management Strategies

Principal Investigator: Febina Mathew (North Dakota State University, Department of Plant Pathology

Co-Investigators: Richard Webster, Samuel Markell, Carrie Miranda, Guiping Yan, Thomas Baldwin, and Joao Paulo Flores

b. Research Overview and Objectives

Research Overview: Seedling diseases, caused by Fusarium, can be a major problem in U.S. soybean production. In North Dakota, sudden death syndrome (SDS), which is caused by Fusarium virguliforme, was confirmed in 2020, and two other species of Fusarium, F. solani, and F. tricinctum, have been implicated in causing root rot. The ND soybean farmers have limited options to manage Fusarium diseases, which include the use of a few fungicide seed treatments, and varieties with tolerance to the SDS fungus, F. virguliforme. Currently, the distribution of Fusarium species (including SDS) and other seedling organisms such as species of Rhizoctonia and Pythium in ND is not understood. However, these diseases are occurring in pockets in ND soybean fields, and there may be several reasons for their emergence. Soybeans may be planted year after year because of higher market prices. Farmers may adopt no-till or reduced tillage practices, which leaves crop residues infested with Fusarium on or near the soil surface. Additionally, it is possible that Fusarium diseases are showing up in fields where soybean cyst nematode (SCN) may be present. Thus, in the proposed study, we developed these objectives: (1) Characterize the species distribution of Fusarium associated with soybean; (2) Characterize the pathogenicity of Fusarium species; (3) Determine how the presence of SCN can affect the development of SDS; and (4) Determine the impact of seed treatment on Fusarium. The information obtained from this study will complement NDSU’s efforts to help and educate farmers to manage SCN, SDS, and other Fusarium diseases with effective disease management strategies.

c. Completed Work: Deliverables and/or Milestones.

• Eight species of Fusarium (F. solani, F. acuminatum, F. caucasicum, F. oxysporum, F. curvatum, F. serpentinum, F. clavus, F. communae) have been isolated from soil samples collected from 75 fields in addition to Rhizoctonia and Pythium spp., which have been reported in the U.S. to cause seedling diseases of soybean.
• Among the species of Fusarium, F. caucasicum, F. curvatum, and F. clavus have not been reported on soybean previously in the U.S.
• For the first time, Phytopythium species were identified in ND.
• Results from the seed treatment trials show greater soybean yield with products such as ILEVO, Saltro, and CeraMax (11 to 17% yield increase) when compared to non-treated control.

d. Progress of Work and Results to Date

Objective 1. Characterize the species distribution of Fusarium associated with soybean (Objective in progress)

Mathew’s lab and Webster’s lab conducted a soybean disease survey across 30 counties (3-5 fields per county) at the vegetative (May – June) or reproductive growth stages (August – September) of soybeans to collect soil samples and infected plant samples. For fungal isolation, about 20 grams of homogenized soil from each field was moistened to 60 to 70 percent and baited with a single 7-day-old Barnes seedling. For each field, six replications were maintained. After 10 days of incubation, the infected roots were surface sterilized in 0.5 % bleach solution and 70 % ethanol for 30 sec, rinsed 2 times in sterile distilled water, and blotted dry. Root pieces were placed in antibiotic-amended (0.06% streptomycin sulfate) half-strength potato dextrose agar (PDA) media at 24± 2°C for 14 days under diffused light conditions. The fungal isolates were purified by hyphal tipping and morphological characterization was done based on colony appearance and spore production on PDA. Based on morphology, isolates were grouped, and representative samples were subjected to molecular characterization and identification using Internal Transcribed Spacer (ITS) gene sequencing for the Rhizoctonia or Pythium genus and Translation elongation factor 1 (EF1) gene sequencing for the Fusarium genus and the identity of the fungal organisms was confirmed using the type sequences available in the Mycobank database. About 461 isolates of seedling pathogens were recovered from 75 fields where Fusarium sp. and Rhizoctonia sp. added up to 58% of the total pathogen population, whereas the frequency of Pythium sp. was close to 20% . At least eight species of Fusarium (accounting for 50 % of the total pathogens) were obtained, which include F. solani species complex, Neocosmospora solani (syn. the F. solani species complex, FSSC), F. acuminatum (syn. F. tricinctum species complex), F. caucasicum, F. oxysporum, F. curvatum (syn. F. oxysporum species complex), F. serpentinum, F. clavus (syn. F. incarnatum-equiseti species complex), and F. communae. In addition, a difference in species distribution was observed across counties .

Objective 2. Characterize the pathogenicity of Fusarium species. (Objective in progress)

This objective was split into two sub-objectives: (1) Screen soybean accessions for root infection of F. virguliforme using an inoculum layer technique, and (2) Study cross-pathogenicity of isolates of Fusarium from soybean and wheat on both crops in the greenhouse using the layer (soybean) and point inoculation (wheat) techniques.

For sub-objective 1, Mathew’s lab set up a preliminary experiment with a single isolate of Fusarium virguliforme (obtained from Iowa State University). The experiment was set up in a completely randomized design using two varieties (‘Spencer’ SDS susceptible and ‘Ripley’ SDS resistant) in the greenhouse, and the layer inoculation method was used. Plastic cups were used which were filled with 80 grams of potting mix, 12 grams of fungal inoculum, followed by 20 grams of potting mix, into which three seeds of Spencer or Ripley were planted and covered with 20 grams of potting mix (inoculum density was 1:10). Throughout the experiment, the temperature was maintained at 23 + 2°C and humidity of 60 to 70% in the greenhouse, and the seedlings were watered every alternate day (water temperature at watering was 17°C and the potting mix was moistened to either 80% or 100% water holding capacity). After 14 days of incubations, seedlings were harvested, roots were washed, and ratings were taken to observe disease symptoms. It was observed that for the SDS-susceptible ‘Spencer’, more than 50% of the seeds decayed from fungal infection, regardless of whether the potting mix was moistened to 80 or 100 percent water holding capacity. In contrast, for the SDS-resistant ‘Ripley’, the seeds decayed when the potting mix was maintained at 100% water holding capacity, and the plants developed root rot infection when 80% water holding capacity was used (Fig. 4).

Objective 4. Determine the impact of seed treatment on Fusarium. (Objective completed)

The trial was planted at the NDSU’s Main Research Station in Fargo, ND by Dr. Markell’s crew and Dr. Webster. The trial was established on a randomized complete block design with seven treatments (Base, Base + ILEVO, Base + Saltro, Base + TBZ + Headsup + Biost + SAR, Base + Saltro + Ataplan, Base + CeraMax, Base + ILEVO + CeraMax, and a non-treated control) on a susceptible soybean variety and there were four replications per seed treatment. Each plot (replication) was about 20 ft. long, and 10 ft wide (>100,000 plants/A). The trial was not inoculated. Plant emergence was evaluated three weeks after planting, and the number of emerged plants was statistically compared with the non-treated control plots. We did not observe any significant differences in the number of plants per acre. We used drone sensors (cameras available at NDSU) in collaboration with Flores’s lab to assess crop response to seed treatments and calculated normalized difference vegetation index (NDVI) from the sensor data to assess the density of vegetation. We took the drone readings three times during the season but did not observe significant differences in NDVI among treatments. However, we noticed that the vegetation was denser in August when compared to July (NDVI values close to 1 indicate dense vegetation) and certain products such as ILEVO, Saltro, and CeraMax provided greater NDVI values when compared to the non-treated control plots. Yield data was obtained at the end of the growing season (October), and we observed greater yield with products such as ILEVO, Saltro, and CeraMax (11 to 17% yield increase) when compared to non-treated control plots).

e. Work to be Completed.

Objective 1. Characterize the species distribution of Fusarium associated with soybean.
• Mathew’s lab will complete the processing of soil samples collected from the remaining 25 fields to make it to a total of 100 fields.

Objective 2. Characterize the pathogenicity of Fusarium species.
• Mathew’s lab is screening 50 soybean accessions for resistance to root rot caused by Fusarium virguliforme (This experiment is in progress)
• Baldwin’s lab will be performing cross-pathogenicity of Fusarium graminearum isolates obtained from soybean roots on wheat to understand the underlying pathogenicity genes associated with the fungus. Mathew’s lab has provided isolates to Baldwin’s lab for the study. (This experiment is in progress).

Objective 3. Determine how the presence of SCN can affect the development of SDS.
• Mathew’s lab in collaboration with Yan’s lab will set up a greenhouse experiment to determine how the presence of SCN can affect the root rot caused by Fusarium graminearum. (This experiment will be set up in January)

f. Other relevant information: potential barriers to achieving objectives, risk mitigation strategies, or breakthroughs.

Fusarium virguliforme was not successfully isolated from the soil samples collected from the fields surveyed in 2023, and this may be because the fields that were surveyed may not have been at a possible risk of SDS development. This was communicated to NDSC, and we will continue the research by using either the most prevalent or a virulent species of Fusarium (F. graminearum) in place of the SDS fungus.

g. Summary

Seedling pathogens (Fusarium, Rhizoctonia, Pythium, etc.) are a major yield-limiting factor in the U.S. soybean production regions (loss of $9.84/A). In North Dakota, the survey conducted in 2023 (75 fields across 30 counties) with funding from the North Dakota Soybean Council indicated that the major seedling pathogens are the species Fusarium, Rhizoctonia, Pythium, and possibly Macrophomina. Among species of Fusarium, F. solani, F. acuminatum, F. caucasicum, F. oxysporum, F. curvatum, F. serpentinum, F. clavus, and F. communae were identified. In addition, for the first time, a species of Phytopythium was identified in ND, and three species of Fusarium, F. caucasicum, F. curvatum, and F. clavus were not previously reported in the U.S. We hypothesize that factors such as heavy rains (above-normal precipitation) across ND in 2023, tillage practices, and limited host resistance options may have contributed to the prevalence of seedling diseases. Considering management options for farmers are limited, fungicide seed treatments were evaluated, and the results show greater soybean yield with products such as ILEVO, Saltro, and CeraMax (11 to 17% yield increase) when compared to non-treated control. Our results highlight the role of Fusarium in causing soybean seedling disease and root rot in ND, and the information obtained from the seed treatment trials and screening soybean germplasm for disease resistance will be used for developing and refining disease management programs targeting Fusarium.

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Updated June 28, 2024:
Final report for North Dakota Soybean Council (July 1, 2023 to June 30, 2024)

a. Research Project Title, Principal and Co-Investigators

Title: Understanding How Fusarium Affects Soybean in North Dakota and Development of Disease Management Strategies

Principal Investigator: Febina Mathew

Co-Investigators: Richard Webster, Samuel Markell, Carrie Miranda, Guiping Yan, Thomas Baldwin, and Joao Paulo Flores

b. Research Overview and Objectives

Seedling diseases, caused by Fusarium, can be a major problem in U.S. soybean production. In North Dakota, sudden death syndrome (SDS), which is caused by Fusarium virguliforme, was confirmed in 2020, and two other species of Fusarium, F. solani, and F. tricinctum, have been implicated in causing root rot. The ND soybean farmers have limited options to manage Fusarium diseases, e.g. varieties with tolerance to F. virguliforme. Currently, the distribution of Fusarium species (including SDS) and other seedling organisms such as species of Rhizoctonia and Pythium in ND is not understood. Additionally, Fusarium diseases may be showing up in fields where soybean cyst nematode (SCN) may be present. Thus, in the proposed study, we developed these objectives: (1) Characterize the species distribution of Fusarium associated with soybean; (2) Characterize the pathogenicity of Fusarium species; (3) Determine how the presence of SCN can affect the development of SDS; and (4) Determine the impact of seed treatment on Fusarium. The information obtained from this study will complement NDSU’s efforts to help and educate farmers to manage SCN, SDS, and other Fusarium diseases with effective disease management strategies.

c. Materials and Methods

Objective 1. Characterize the species distribution of Fusarium associated with soybean (Mathew, Webster)

A soybean disease survey was conducted across 30 counties (3-5 fields per county) at the vegetative (May – June) or reproductive growth stages (August – September) of soybeans to collect soil samples and infected plant samples. For fungal isolation, about 20 grams of homogenized soil from each field was moistened to 60 to 70 percent and baited with a single 7-day-old Barnes seedling. For each field, six replications were maintained. After 10 days of incubation, the infected roots were surface sterilized in 0.5 % bleach solution and 70 % ethanol for 30 sec, rinsed 2 times in sterile distilled water, and blotted dry. Root pieces were placed in antibiotic-amended (0.06% streptomycin sulfate) half-strength potato dextrose agar (PDA) media at 24± 2°C for 14 days under diffused light conditions. The fungal isolates were purified by hyphal tipping and morphological characterization was done based on colony appearance and spore production on PDA. Based on morphology, isolates were grouped, and representative samples were subjected to molecular characterization and identification using Internal Transcribed Spacer (ITS) gene sequencing for the Rhizoctonia or Pythium genus and Translation elongation factor 1 (EF1) gene sequencing for the Fusarium genus and the identity of the fungal organisms was confirmed using the type sequences available in the Mycobank database and Fusarium ID. About 600 isolates of seedling pathogens were recovered from 100 fields where Fusarium sp. and Rhizoctonia sp. added up to 44% of the total pathogen population, whereas the frequency of Pythium sp. was close to 15% (Fig. 2). At least nine species of Fusarium (accounting for 38% of the total pathogens) were obtained, which include F. acuminatum (syn. F. tricinctum species complex), F. armeniacum (syn. F. sambucinum species complex), F. clavum (syn. F. incarnatum-equiseti species complex), F. oxysporum (syn. F. oxysporum species complex), F. sambucinum (syn. F. sambucinum species complex), F. scirpi (syn. F. incarnatum-equiseti species complex), F. solani (syn. F. solani species complex), F. sporotrichioides (syn. F. sambucinum species complex) and F. vanettenii (syn. F. solani species complex). In addition, a difference in species distribution was observed across counties.

Objective 2. Characterize the pathogenicity of Fusarium species (Mathew, Baldwin, Miranda).

This objective was split into two sub-objectives: (1) Screen soybean accessions for root infection of F. proliferatum using an inoculum layer technique, and (2) Study cross-pathogenicity of isolates of Fusarium graminearum from soybean and barley on both crops in the greenhouse using the layer (soybean) and point inoculation (barley) techniques.

Sub-objective (1): For the screening study, seeds of 53 accessions were obtained from Dr. Carrie Miranda’s breeding program at NDSU. One F. proliferatum isolate FUS026 (McCook County, SD) was arbitrarily selected from 2 isolates that showed the same median disease severity of 4 (= germination and greater than 75% of the roots having lesions) in a study by Okello et al. (2020) on the susceptible check ‘Asgrow 1835’. The fungus was grown on potato dextrose agar (PDA; Oxoid, Basingstoke, UK) amended with streptomycin sulfate (0.3 g/L) to prevent bacterial contamination. The Petri plates were then incubated at 23±2°C for 14 days under a 12-hour light/dark cycle. To prepare the inoculum, a sand and corn meal mixture (3:1 ratio) was autoclaved twice and inoculated with five mycelial plugs of 15 mm square size were added. The inoculated mixture was incubated at 23±2°C for 14 days and thoroughly mixed. At planting, 40 grams of potting mix was added to 475 ml transparent plastic cups, followed by adding 20 grams of inoculum and 20 grams of potting mix. Over this layer, three 3-day pre-germinated seeds were planted and covered with 20 grams of potting mix. The experiment was performed in the greenhouse (24 ±2°C; 12-hour light and dark conditions) and laid out in a completely randomized design. Six replicates were maintained for each accession. The plants were watered at 80 % of the water-holding capacity every other day. The experiment was performed twice. The plants were pulled out 21 days after planting and gently cleaned under running tap water. The root rot severity was determined immediately after washing and rated on a disease scale of 1-5 (Acharya et al.2015, Okello et al. 2020) where, 1 = seedlings germinated with no visible root discoloration, 2 = seedlings germinated with1 to 19% root lesions, 3 = seedlings germinated with 20 to 74% root lesions, 4 = seedlings germinated with roots having 75% or greater percentage lesions; and 5 = No seeds germination and complete colonization by the fungus. Using non-parametric statistics (Shah and Madden 2004), a significant effect of the RTEs caused by F. proliferatum isolate FUS026 was observed on the accessions (ATS = 6.81; df = 5.87; P = <0.0001). Based on 95 % confidence intervals, none of the accessions tested were significantly less susceptible to the fungus compared to the control ‘RG200RR’.

Sub-objective (2): Two barley isolates (322, 354) and three soybean isolates (357, 362, 364) were grown in PDA for seven days, and DNA was extracted using a modified CTAB method. Isolates were grown in mungbean agar for seven days before harvesting. Spores were standardized to 500,000 per ml, and 10 ul was inoculated to 2nd or 3rd full kernel of the wheat head. After point inoculation, the whole head was covered with small plastic bags. After 72 hours, the plastic bags were replaced with glassine bags. Heads were evaluated after 14 days for present disease severity calculating infected/total kernel*100. Among the isolates, only isolate 322 was not Fusarium graminearum. The disease severity rating showed that this caused minimal damage to the barley heads, giving statistically comparable results to isolate 357. Both of these isolates had <25% severity. Isolate 354 and 362 have statistically similar severity (>25%, <50%). Among the isolates, 364 obtained the highest severity (>50%, <100%). It is also worth noting that only isolate 322 did not spread in the entire barley heads. Overall, the F. graminearum isolates from soybeans exhibited a wide range of virulence on wheat, including an isolate that was highly virulent and an isolate that did not spread through the rachis and might be incapable of producing DON.

Objective 3. Determine how the presence of soybean cyst nematode can affect the development of root rot caused by Fusarium graminearum (Mathew, Yan)

The study was conducted in a growth chamber using the SCN/F. graminearum-susceptible soybean cultivar 'Williams 82'. The F. graminearum isolate FUS052 used in this study was recovered from diseased soybean roots sampled from McCook County in South Dakota in 2014. To prepare the inoculum, the fungus was grown on potato dextrose agar (PDA) and a sand-cornmeal mixture was inoculated as described previously. For SCN inoculum, an HG type 2.5.7 isolate of SCN provided by Dr. Guiping Yan's lab was used. To prepare the inoculum, nematode cysts were collected from the soil using a decanting and sieving protocol (Krusberg et al. 1994). A 100-cc soil sample with SCN was collected in a bucket filled three-fourths with water. The soil was mixed thoroughly by stirring to suspend it in water. The suspension was then poured through a 710-µm-pore sieve (No. 25) placed above a 250-µm-pore sieve (No. 60). The SCN cysts collected on the No. 60 sieve were washed into a beaker using water gently sprayed from a spray bottle to avoid harming the SCN cysts. The suspension was brought to 50 ml, and the eggs and juveniles were extracted from the cysts by rupturing them with a rotating rubber stopper (Faghihi and Ferris 2000). The eggs and juveniles were collected in a 75-µm-pore sieve (No. 200) placed above a 20-µm-pore sieve (No. 635). The SCN eggs were collected and suspended in 100 ml of distilled water. The number of eggs present per ml of suspension was counted under a dissecting microscope at 40X magnification using a nematode counting slide. For inoculation, the total number of nematode eggs was adjusted to approximately 600 eggs (600 ± 100) per ml.

The experiment was performed in a growth chamber, and the treatments were F. graminearum only, SCN only, F. graminearum + SCN, and control. Each treatment had five replications, and plastic cone-tainers filled with river sand with or without inocula and one 4-day pregerminated seed served as a replication. The cones were filled with river sand following the inoculum layer inoculation method modified by Bilgi et al. (2008). For F. graminearum only treatment, each cone was filled initially with 60 grams of pasteurized river sand, and then 20 grams of fungal inoculum was added, followed by another 20 grams of river sand. A pre-germinated seed of 'Williams 82' was planted on top of this layer and covered with 20 grams of river sand. For F. graminearum + SCN treatment, a small hole was made using a pipette tip in the river sand layer above the fungal inoculum layer, and a 3.1 ml suspension of approximately 600 eggs/ml was added to the hole using a pipette. The pre-germinated seed was sown near this hole and covered with river sand. For SCN-only treatments, the cones were filled with pasteurized river sand and inoculated as described for the F. graminearum + SCN treatment. For all treatments containing SCN, efforts were made to ensure that the primary roots (radicle) of the seedlings were in contact with the nematode egg suspension. The cones for the non-inoculated control treatment were filled with 100 grams of river sand, and a pre-germinated seed was sown on top, which was then covered with 20 grams of river sand. To prevent soil loss while watering and to restrict the roots within the cone-tainers, the undersides of the cones were covered with a black cloth held with rubber bands. Each cone was watered every other day with 7-8 ml of water, approximately 70% of the water holding capacity. The temperature in the growth chamber was maintained at 23±2°C with a relative humidity of 70-80% and 12-hour light and dark conditions. The experiment was performed three times. After 42 days of inoculation, each experiment was terminated, and the roots were gently pulled out from each cone and washed under running tap water. The shoot length and root length (tap root) of each plant were measured using a ruler. The washed roots were rated for root rot severity on a disease scale of 1-5 (Okello et al. 2020). To count the nematode cysts, the plants in the cone-tainers were gently removed and placed in a beaker along with the infested soil. Using the decanting and sieving protocol (Krusberg et al. 1994), SCN cysts were collected, and the eggs were extracted and counted following the protocol described by Faghihi and Ferris (2000). The total number of SCN eggs per ml was calculated from all the cones inoculated with the nematode. The non-inoculated control and the F. graminearum-only treatments were also examined for the presence of nematodes. Since the disease scoring data was ordinal and the data for root length, shoot length, and SCN egg count, non-parametric statistics (Shah and Madden 2004) was used. After 42 days of inoculation, approximately 90% of the plants inoculated with F. graminearum and a combination of F. graminearum and SCN showed complete seed infection. A significant effect of treatments was observed on the root rot severity (ATS = 230.16; df = 1.72; P = 1.60 x 10^-87) at 95% confidence intervals.

A significant effect of RTE caused by F. graminearum was observed for root length (ATS = 60.68; df = 2.05; P = 9.68 x 10-28) shoot length (ATS = 111.52; df = 2.48; P = 2.18 x 10-60) and SCN egg counts (ATS = 66.16; df = 1.82; P = 4.14 x 10-27) at 42 days after inoculation. The presence of SCN didn’t significantly increase the severity of root rot caused by the fungus. A significant reduction in the root length and shoot length was observed for plants with F. graminearum only and F. graminearum + SCN treatments compared to the control. Additionally, the root and shoot lengths of plants inoculated only with SCN were significantly less compared to the control (Fig. 5). The SCN-only treatment showed a significant increase in the SCN egg count compared to the control. The SCN egg count was reduced in the treatment that had a combination of F. graminearum and SCN compared to the initial inoculum amount. The results from this study show that the presence of SCN didn’t increase the severity of root rot caused by F. graminearum.

Objective 4. Determine the impact of seed treatment on Fusarium. (Mathew, Markell, Webster, Flores)

The trial was planted at the NDSU’s Main Research Station in Fargo, ND by Dr. Markell’s crew and Dr. Webster. The trial was established on a randomized complete block design with seven treatments (Base, Base + ILEVO, Base + Saltro, Base + TBZ + Headsup + Biost + SAR, Base + Saltro + Ataplan, Base + CeraMax, Base + ILEVO + CeraMax, and a non-treated control) on a susceptible soybean variety and there were four replications per seed treatment. Each plot (replication) was about 20 ft. long, and 10 ft wide (>100,000 plants/A). The trial was not inoculated. Plant emergence was evaluated three weeks after planting, and the number of emerged plants was statistically compared with the non-treated control plots. We did not observe any significant differences in the number of plants per acre as indicated in Table 1. We used drone sensors (cameras available at NDSU) in collaboration with Flores’s lab to assess crop response to seed treatments and calculated normalized difference vegetation index (NDVI) from the sensor data to assess the density of vegetation. We took the drone readings three times during the season but did not observe significant differences in NDVI among treatments. However, we noticed that the vegetation was denser in August when compared to July (NDVI values close to 1 indicate dense vegetation) and certain products such as ILEVO, Saltro, and CeraMax provided greater NDVI values when compared to the non-treated control plots. Yield data was obtained at the end of the growing season (October), and we observed greater yield with products such as ILEVO, Saltro, and CeraMax (11 to 17% yield increase) when compared to non-treated control plots.

d. Research results/outcomes

• Nine species of Fusarium have been isolated from soil samples collected from 100 fields in addition to Rhizoctonia and Pythium spp. Among the species of Fusarium, F. clavum, F. scirpi, and F. vanettenii have not been reported on soybeans previously in the U.S.
• Preliminary results suggested that the presence of SCN didn’t increase the severity of root rot caused by F. graminearum.
• Preliminary results also suggest that F. graminearum isolates from soybeans exhibited a wide range of virulence, including an isolate that was highly virulent and an isolate that did not spread through the wheat rachis and might be incapable of producing DON.
• Results from the seed treatment trials show greater yield with products such as ILEVO, Saltro, and CeraMax (11 to 17% yield increase) when compared to non-treated control.
• One manuscript published from Fusarium research in 2024 (Rafi et al. 2024 - https://doi.org/10.1094/PDIS-02-24-0477-RE ) in a peer-reviewed journal, Plant Disease

e. Listings of any disclosures of inventions or plant varieties

None

f. Discussion

In North Dakota, a survey conducted in 2023 across 100 fields in 30 counties, with funding from the North Dakota Soybean Council, revealed that the major seedling pathogens are Fusarium, Rhizoctonia, Pythium, and possibly Macrophomina. Among the Fusarium species identified were F. solani, F. acuminatum, F. caucasicum, F. oxysporum, F. curvatum, F. serpentinum, F. clavus, and F. communae. Furthermore, three species of Fusarium, namely F. caucasicum, F. curvatum, and F. clavus, were previously unreported in the United States. Considering the limited management options available to farmers, the survey evaluated fungicide seed treatments. The results demonstrated that products such as ILEVO, Saltro, and CeraMax led to higher soybean yields (11 to 17% increase) compared to the non-treated control. Moreover, our study highlighted that certain Fusarium species, such as F. graminearum, known to be pathogenic to rotational crops like wheat, also pose a threat to soybeans. This emphasizes the importance of developing resistant soybean cultivars to combat Fusarium infections and reduce inoculum for crop rotation.

g. Conclusion/Benefits to the North Dakota Soybean Farmers and the Industry

Our study highlights the importance of investigating seedling pathogens, such as Fusarium, Rhizoctonia, Pythium, and others, in North Dakota. These organisms are known to cause a yield loss of $9.84 per acre in soybean production regions throughout the United States. We propose that factors such as heavy rainfall (above-normal precipitation) in North Dakota in 2023, tillage practices, and limited options for host resistance may have contributed to the widespread occurrence of seedling diseases. Specifically, our findings emphasize the role of Fusarium in causing soybean seedling disease and root rot in North Dakota. The data gathered from our seed treatment trials will be used to develop and refine disease management programs that target Fusarium specifically. Additionally, we will continue collaborating with Dr. Miranda's breeding program to screen different varieties for resistance to Fusarium species, which can then be adopted by farmers in the future. Furthermore, our study on the Fusarium-SCN experiment highlights the importance of screening cultivars for resistance to both pathogens, ultimately leading to more effective disease control.

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We collected soil samples from 100 fields in North Dakota and isolated nine species of Fusarium, along with Rhizoctonia and Pythium spp. Out of these Fusarium species, F. clavum, F. scirpi, and F. vanettenii have not been previously reported on soybeans in the U.S. Our study suggest that the presence of soybean cyst nematode does not increase the severity of root rot caused by F. graminearum. Furthermore, our study indicates that F. graminearum isolates from soybeans vary in their virulence on wheat heads, with one highly virulent isolate and another isolate that does not spread through the wheat rachis and may be unable to produce DON. Seed treatment trials show that using products such as ILEVO, Saltro, and CeraMax results in higher yields (11 to 17% increase) compared to the non-treated control. In 2024, a manuscript on Fusarium research was published by Rafi et al. in the peer-reviewed journal Plant Disease. The manuscript can be accessed at https://doi.org/10.1094/PDIS-02-24-0477-RE.

Soybeans represent an economically important crop for ND farmers and Fusarium diseases (including SDS) can be yield-limiting particularly under cool, wet conditions (early in the season or during the growing season). Through this research, farmers will obtain information on how to minimize the impact of Fusarium and their association with SCN (such as use of SCN-resistant and SDS tolerant varieties, seed treatments) on soybean yield and maximize their return on investment.