Soybean is one of the most economically important crops impacted by Sclerotinia sclerotiorum, a broad host range fungal pathogen, also called white mold. White mold is a recurring problem for Minnesota soybean growers. Cultural management of white mold, such as crop rotation and row spacing modification, has limited economic success. One of the most sustainable and economical ways to control crop diseases is to identify and generate resistant varieties of plants. In the Minnesota soybean breeding program, several varieties of soybeans have been bred. White mold disease is a significant issue in soybean production areas of Minnesota. The disease is caused by a broad host range fungal pathogen Sclerotinia sclerotiorum that causes disease on more than 400 different plant species. Cultural management of white mold, such as crop rotation and row spacing modification, has limited success, and hence white mold management is largely dependent on fungicide treatment. S. sclerotiorum forms a black survival structure called sclerotia at the end of the disease cycle. These sclerotia can survive up to 7 years in the soil, contributing to long-term presence in the field and making it difficult to manage with traditional management strategies. Planting a susceptible variety of soybean can increase the soil's sclerotia load, making it economically unviable for growing the same variety in the following years. One of the most sustainable and economical ways to control crop diseases is to generate resistant varieties. Unfortunately, no soybean lines are known to have complete resistance against white mold.
The breeding effort at UMN has led to the generation of hundreds of lines. However, they have not been adequately screened in the absence of a standardized resistance screening method. Our lab has standardized a petiole inoculation method to perform high throughput screening of soybean lines against S. sclerotiorum (Figure 1). Our preliminary study assessed ten soybean lines for S.sclerotiorum infection responses. We have found consistent and reproducible results and identified six susceptible, one moderately resistant, and three highly resistant lines (Figure 2). These lines can serve as check panels, help accurately assess the white mold resistance in soybean germplasm, and aid in breeding and commercial soybean development.


Figure 1. Symptom development in highly susceptible (Bret MN01), moderately resistant (MN091), and highly resistant soybean lines (MIN Soy). Disease symptoms were observed following petiole inoculation with an agar plug containing actively growing mycelia of S. sclerotiorum at 1,2,3,4,5 and 6 days post-inoculation (dpi). At 6dpi, there is a significant difference in lesion length shown in red boxes.



Figure 2. Lesion length development in highly susceptible (light red), moderately Resistant (light blue), and highly resistant soybeans lines (light green) following S. sclerotiorum 6 days post- inoculation (dpi). Error bars represent the standard deviation of the mean (SDM) across two experimental runs with two replications (n=12).

S. sclerotiorum isolates from different regions can have a range of aggressiveness, causing a wide range of severity for different soybean genotypes. We initially performed an aggressiveness check for eight isolates of S. sclerotiorum collected from different regions of Minnesota to represent different geographical distributions. These isolates were used to challenge the most susceptible variety of Soybean (Bret MN 01, MN0083, MN0095, and William 82). We found one isolate, S.S. Sibley, isolated from Sibley county to be highly aggressive. Greenhouse inoculations using a soybean check panel and one isolate would increase the effectiveness of the breeding and selection process for selecting the resistance lines. Once we zero in on a few resistant/elite soybean varieties, these can be further challenged with the eight-isolate to confirm their disease response. This strategy will help us save our time and money to screen all the lines against all the isolates.
One of the important aspects of the study is screening the soybean lines in a naturally infested field. We have identified a highly naturally infected plot in Albert Lea, Faribault county, Mn County managed by Johnson farms of Wells. They will be our cooperator in facilitating the proposed research. Greenhouse study and Field trial of the soybean variety will give us comprehensive knowledge about their resistance phenotype elite germplasm targets for future crosses and variety development. As co-investigators on this project, Drs. Ranjan, Lorenz, and Seth will continue to work together to discover new sources and integrate them into the breeding program.
To answer the above-discussed problems, we have divided our proposal into two major objectives/tasks, which are discussed below -

Objective 1. Conduct white mold disease screening on the soybean varieties produced through the UMN soybean breeding program and commercial varieties in the naturally infested field and controlled environment, greenhouse/growth chamber.
Goal 1. The study aims to identify white mold resistant lines/cultivars for growers and use best lines as parental lines for future breeding programs for sustained breeding effort towards white mold-resistant cultivars productions. The study results will be published through the MN Ag Experiment Station publications.
Goal 2. Provide an unbiased source of information to growers on the level of white mold resistance in commercially available varieties and their performance at S. sclerotiorum infested sites.
Objective 2. Evaluate the yield impact of white mold in treated and non-treated soybean varieties and quantify their yield potential under standard farm management practices.
Goals 1. Provide a yield potential of soybean lines under S. sclerotiorum disease pressure.

Results from the proposed research will be presented at farmer field days held by both the Minnesota Agricultural Experiment Station and Minnesota Soybean Research and Promotion Council, including the Minnesota Ag Expo. This is a new proposal, and I hope to continue for a few more years to give a comprehensive result of the study. The study will lead to the identification of new genetic resources for breeding new soybean varieties for better yield and white mold resistance. The research will also be published in extension articles.

Update:
The project commenced in May 2022. The primary objective of the proposal is to screen the soybean varieties for white mold disease resistance/susceptibility in the naturally infested field and controlled environment, greenhouse/growth chamber. I will be discussing and updating our progress in two sub-sections –
1. Screening soybean varieties in the naturally infested white mold field
2. Screening soybean varieties in growth chamber experiments.
1. Screening soybean varieties in naturally infested white mold field -
For the field resistance study of 48 soybean varieties to white mold, plots were arranged in a randomized complete block design with four replications at a Johnson Farm's (farmer cooperator's) field near Albert Lea, Faribault county, MN. Fourty eight soybean seeds were selected from UMN breeding program and few commercial varieties (provided Dr. Lorenz and Dr. Seth). They were planted by Dr. Seth's group on 31st May 2022 using a custom plot planter, with four-row plots 12 feet in length and rows spaced 30 inches apart. The maturity group of the selected soybean varieties varies between 1.5 to 2.0.
We counted the germinated seeds and stalked the plots, as shown in Figure 1 (uploaded document as pdf) on 17th June. We also performed the first round of manual weeding. All soybean varieties germinated well and had consistent count numbers. Our next visit to the plot was on the 15th July to stage the plant growth; they were in the R1-R2 stages of growth. We also performed a second round of manual weeding. In between, we also got help from our collaborator Johnson's farm for weeding. On 5th August, we scouted for the disease symptom. We saw only a few incidences of early disease symptoms. This was followed by 25th August scouting, and we could see a significant level of disease in soybean. We scouted all 192 plots and scored them for disease incidences and severity. We could see all the stages of disease progression (Figure 2). We counted 157 total incidences of disease in the plots. We also noted that a few plots had been damaged due to herbicide drift from adjacent farm plots and mechanized weeding. Overall, good disease pressure in the field was observed. We will also be performing a few more data collections in the coming weeks.
2. Screening soybean varieties in growth chamber experiments.
S. sclerotiorum isolates from different regions can have a range of aggressiveness, causing a wide range of severity for different soybean genotypes. We initially performed an aggressiveness check for eight isolates of S. sclerotiorum collected from different regions of Minnesota to represent different geographical distributions. But this collection did not include Johnson's farm S. sclerotiorum isolate (Faribault). Therefore, we planned to check the aggressiveness of the isolate to already established three different check lines (William 82 and Bert, susceptible check line and Noir, highly tolerant check line) in our lab and compared them to highly virulent Sibley and WT1980 (standard lab isolate). We concluded that Johnson's farm isolate (Faribault strain) is highly virulent (Figure 3).
We decided to use the Faribault strain to screen the soybean varieties in the growth chamber study. As discussed in our proposal, we will perform three biological replicates of each variety. We have completed the first biological replicates of 12 soybean varieties (Figure 4) and currently performing our second biological replicates. For the study's statistical significance, we will perform another biological replicate. Similarly, we will be screening the rest of the 36 soybean varieties.

View uploaded report

Update:
The project commenced in May 2022. The primary objective of the proposal is to screen the soybean varieties for white mold disease resistance/susceptibility in the naturally infested field and controlled environment, greenhouse/growth chamber. I will be discussing and updating our progress on three research goals -
1. Screening soybean varieties in the naturally infested white mold field
2. Screening soybean varieties in growth chamber experiments.
3. Soybean yield estimation in the naturally infested white mold field
1. Screening soybean varieties in naturally infested white mold field -
For the field resistance study of 48 soybean varieties to white mold, plots were arranged in a randomized complete block design with four replications at a Johnson Farm's (farmer cooperator's) field in wells, Faribault county, MN. Forty-eight soybean variety seeds were selected from the UMN breeding program and a few commercial varieties (provided Dr. Lorenz and Dr. Seth). They were planted by Dr. Seth's group on 31st May 2022 using a custom plot planter, with four-row plots 12 feet in length and rows spaced 30 inches apart. The maturity group of the selected soybean varieties varies between 1.5 to 2.0.
We counted the germinated seeds and stalked the plots, as shown in Figure 1 (uploaded document as pdf) on 17th June. We also performed the first round of manual weeding. All soybean varieties germinated well and had consistent count numbers. Our next visit to the plot was on 15th July to stage the plant growth; they were in the R1-R2 stages of growth. We performed a second round of manual weeding. In between, we got help from our collaborator Johnson's farm for weeding. On 5th August, we scouted for the disease symptom. We saw only a few incidences of early disease symptoms. This was followed by 25th August scouting, and we could see a significant level of disease in soybean. We scouted all 192 plots and scored them for disease incidences and severity. We could see all the stages of disease progression (Figure 2). We counted 157 total incidences of disease in the plots. We also noted that a few plots were damaged due to herbicide drift from adjacent farm plots and mechanized weeding. Overall, good disease pressure in the field was observed. We performed our last white mold disease data collection on 8th September (diseased soybean plots, Figure 3) and scored them for disease severity and incidences. The disease incidence (DI) was assessed by taking three separate 1-yard (3 feet) sections within the inner two center rows of the plot by counting the cumulative number of diseased plants. The disease incidence was calculated by dividing the total number of infected plants by the number of plants. Disease severity (DS) was measured by taking five separate one-foot-long sections of the inner rows of a plot and determining the highest severity within each length. Severity ratings were scored on a scale of 0 to 3 (Figure 2). DS is calculated by taking the sum of all severity scores and dividing by the number of one-foot sections (=5) where the disease is present. The disease severity index (DIX) is calculated by dividing the DS by three and multiplying by the DI (DIX = DS/3 x DI) (Figure 4). As can be expected with field trials, a number of plots suffered from herbicide drift, mechanical tractor damage, and/or herbivory, leaving 27 varieties with three or more replicates (Figure 4).
2. Screening soybean varieties in growth chamber experiments.
S. sclerotiorum isolates from different regions can have a range of aggressiveness, causing a wide range of severity for different soybean genotypes. We initially performed an aggressiveness check for eight isolates of S. sclerotiorum collected from different regions of Minnesota to represent different geographical distributions. But this collection did not include Johnson's farm S. sclerotiorum isolate (Faribault). Therefore, we planned to check the aggressiveness of the isolate to already established three different check lines (William 82 and Bert, susceptible check line and Noir, highly tolerant check line) in our lab and compared them to highly virulent Sibley and WT1980 (standard lab isolate). We concluded that Johnson's farm isolate (Faribault strain) is highly virulent (Figure 5).
We decided to use the Faribault strain to screen the soybean varieties in the growth chamber study. The study helped us to identify highly resistant and susceptible soybean varieties to white mold. As discussed in our proposal, we will perform three biological replicates of each variety. We have completed the two biological replicates of 32 soybean varieties (Figure 6) and currently performing our second biological replicates for the rest of them. Figure 6 represents the average disease lesion length (in mm) of 32 soybean varieties.
3. Soybean yield estimation in the naturally infested white mold field – The plots for yield estimation were harvested on 5th October by Dr. Seth's group. Yield data from the field trial has been calculated, but trends and significant interactions have yet to be assessed.
Conclusion and discussion (project deliverables) – These forty-eight soybean varieties are being screened for white mold in growth chambers. At the same time, we have completed our first year of the field screening study. As we performed a simultaneous evaluation of soybean varieties in a controlled environment and a field study, it gave us a comprehensive understanding of their differential responses to white mold disease. The initial results from the simultaneous evaluation of soybean varieties in a controlled environment and field study suggest that soybean lines 1940KN, AG17XF2, and 2022 N are highly resistant, while lines O.1718N, A2121XF, and MN1901CN are highly susceptible to white mold. Interestingly, MN1904HOLL displays the highest mean lesion length score from the growth chamber experiments while falling in the bottom six for DIX scores from the field. This seemingly conflicted observation could be due to the uneven distribution of S. sclerotiorum at some patches in the plots or as a result of making comparisons before three biological replicates of the growth chamber experiment and only one year of field disease screening study. Continuation of our experiments for the next two years (especially the field screening study) will increase our sample size and give a comprehensive understanding and conclusive results.
Results from the proposed research will be presented at farmer field days held by both the Minnesota Agricultural Experiment Station and Minnesota Soybean Research and Promotion Council, including the Minnesota Ag Expo. The study is in its first year, and we hope to continue for a few more years to give a comprehensive study result.

View uploaded report

Update:
The project commenced in May 2022. The primary objective of the proposal is to screen the soybean varieties for white mold disease resistance/susceptibility in the naturally infested field and controlled environment, greenhouse/growth chamber. I will be discussing and updating our progress on three research goals -
1. Screening soybean varieties in the naturally infested white mold field
2. Screening soybean varieties in growth chamber experiments.
3. Soybean yield estimation in the naturally infested white mold field
1. Screening soybean varieties in naturally infested white mold field – We had updated our soybean screening study on 30th November 2022 update (Figure related to this study is shown in uploaded Figure 1-4).
2. Screening soybean varieties in growth chamber experiments.
S. sclerotiorum isolates from different regions can have a range of aggressiveness, causing a wide range of severity for different soybean genotypes. We initially performed an aggressiveness check for eight isolates of S. sclerotiorum collected from different regions of Minnesota to represent different geographical distributions. But this collection did not include Johnson's farm S. sclerotiorum isolate (Faribault). Therefore, we planned to check the aggressiveness of the isolate to already established three different check lines (William 82 and Bert, susceptible check line and Noir, highly tolerant check line) in our lab and compared them to highly virulent Sibley and WT1980 (standard lab isolate). We concluded that Johnson's farm isolate (Faribault strain) is highly virulent (Figure 5).
We decided to use the Faribault strain to screen the soybean varieties in the growth chamber study. The study helped us to identify highly resistant and susceptible soybean varieties to white mold. As discussed in our proposal, we will perform three biological replicates of each variety. We have completed one biological replicate of 50 soybean varieties (Figure 6), including two susceptible soybean check lines “Bert” and “sturdy”. Figure 6 represents the area under disease progress curve of 50 soybean varieties. Currently, we are performing our second biological replicates screening and have completed screening for 39 soybean varities.
3. Soybean yield estimation in the naturally infested white mold field – The plots for yield estimation were harvested on 5th October by Dr. Seth's group. Yield data from the field trial has been calculated, but trends and significant interactions have yet to be assessed.
Conclusion and discussion (project deliverables) – Fifty soybean varieties, including two susceptible soybean check lines, are being screened for white mold in growth chambers. We performed a simultaneous evaluation of soybean varieties in a controlled environment and a field study, which gave us a comprehensive understanding of their differential responses to white mold disease. At the same time, we have completed our first year of the field screening study. The initial results from the simultaneous evaluation of soybean varieties in a controlled environment and field study suggest that soybean lines 1940KN, MP152074, and 2022 N are highly resistant, while lines O.1718N, A2121XF, and MN1901CN are highly susceptible to white mold. Interestingly, MN1904HOLL displays the highest mean lesion length score from the growth chamber experiments while falling in the bottom six for DIX scores from the field. This seemingly conflicted observation could be due to the uneven distribution of S. sclerotiorum at some patches in the plots or as a result of making comparisons before three biological replicates of the growth chamber experiment and only one year of field disease screening study. Continuation of our experiments for the next two years (especially the field screening study) will increase our sample size and give a comprehensive understanding and conclusive results.
Results from the proposed research will be presented at farmer field days held by both the Minnesota Agricultural Experiment Station and Minnesota Soybean Research and Promotion Council, including the Minnesota Ag Expo. The study is in its first year, and we hope to continue for a few more years to give a comprehensive study result.

View uploaded report

Update:
The project commenced in May 2022. The primary objective of the proposal is to screen the soybean varieties for white mold disease resistance/susceptibility in the naturally infested field and controlled environment, greenhouse/growth chamber. I will be discussing and updating our progress on three research goals -
1. Screening soybean varieties in the naturally infested white mold field
2. Screening soybean varieties in growth chamber experiments.
3. Soybean yield estimation in the naturally infested white mold field
1. Screening soybean varieties in naturally infested white mold field – We had updated our soybean screening study on 30th November 2022 update (Figure related to this study is shown in uploaded Figure 1-4).
2. Screening soybean varieties in growth chamber experiments.
S. sclerotiorum isolates from different regions can have a range of aggressiveness, causing a wide range of severity for different soybean genotypes. We initially performed an aggressiveness check for eight isolates of S. sclerotiorum collected from different regions of Minnesota to represent different geographical distributions. But this collection did not include Johnson's farm S. sclerotiorum isolate (Faribault). Therefore, we planned to check the aggressiveness of the isolate to already established three different check lines (William 82 and Bert, susceptible check line and Noir, highly tolerant check line) in our lab and compared them to highly virulent Sibley and WT1980 (standard lab isolate). We concluded that Johnson's farm isolate (Faribault strain) is highly virulent (Figure 5).
We decided to use the Faribault strain to screen the soybean varieties in the growth chamber study. The study helped us to identify highly resistant and susceptible soybean varieties to white mold. As discussed in our proposal, we will perform three biological replicates of each variety. We have completed two biological replicates of 50 soybean varieties (Figure 6), including two susceptible soybean check lines “Bert” and “sturdy”. Figure 6 represents the area under disease progress curve of 50 soybean varieties. Currently, we are performing our third biological replicates screening and have completed screening for 21 soybean varieties.
3. Soybean yield estimation in the naturally infested white mold field – The plots for yield estimation were harvested on 5th October by Dr. Seth's group. Yield data from the field trial has been calculated, but trends and significant interactions have yet to be assessed.
Conclusion and discussion (project deliverables) – Fifty soybean varieties, including two susceptible soybean check lines, are being screened for white mold in growth chambers. We performed a simultaneous evaluation of soybean varieties in a controlled environment and a field study, which gave us a comprehensive understanding of their differential responses to white mold disease. At the same time, we have completed our first year of the field screening study. The initial results from the simultaneous evaluation of soybean varieties in a controlled environment and field study suggest that soybean lines 1940KN, MP152074, and 2022 N are highly resistant, while lines O.1718N, A2121XF, and MN1901CN are highly susceptible to white mold. Interestingly, MN1904HOLL and M14-250018 display the higher mean lesion length score from the growth chamber experiments while falling in the bottom six for DIX scores in the field trial. This seemingly conflicted observation could be due to the uneven distribution of S. sclerotiorum at some patches in the plots, environmental conditions or as a result of making comparisons before three biological replicates of the growth chamber experiment and only one year of field disease screening study. Continuation of our experiments for the next two years (especially the field screening study) will increase our sample size and give a comprehensive understanding and conclusive results.
Results from the proposed research will be presented at farmer field days held by both the Minnesota Agricultural Experiment Station and Minnesota Soybean Research and Promotion Council, including the Minnesota Ag Expo. The study is in its first year, and we hope to continue for a few more years to give a comprehensive study result.

View uploaded report

The proposed research addresses three areas targeted by the Production Action Team's mission statement: (1) Pest management; (2) Plant breeding and functional genomics; (3) White mold (Sclerotinia stem rot). The benefit of this research is three pronged and will positively impact MN soybean farmers. These are –
1. The study will identify resistant lines out of 50 soybean lines, including some commercial varieties. This will help Soybean growers to make an informed decision about planting white mold resistant varieties.
2. The identified white mold resistant and higher yield cultivars can become important genetic resources for future breeding program decisions and local growers' use.