Deployment of a Natural Biofumigant for Cyst Nematode and White Mold Mitigation in Soybean Crops
Sustainable Production
Crop protectionDiseaseField management
Parent Project:
This is the first year of this project.
Lead Principal Investigator:
Zhengrong Gu, South Dakota State University
Co-Principal Investigators:
Project Code:
Contributing Organization (Checkoff):
Institution Funded:
Brief Project Summary:
The purpose of this project is to utilize a natural biofumigant from oilseeds to mitigate cyst nematodes and white mold in soybean crops. The secondary metabolite, glucosinolates, has fungicidal and nematocidal properties that make it an excellent candidate as a biofumigant in soybeans. Once recovered from oilseed meals, the glucosinolates can be applied in fields, be incorporated into the soil, and have a controlled release during the growing season to mitigate the negative impacts of soybean cyst nematodes and white mold. Research will evaluate the overall effectiveness of the glucosinolates on SCN and white mold.
Key Beneficiaries:
#agronomists, #extension specialists, #farmers
Unique Keywords:
#biofumigants, #scn, #soybean cyst nematodes, #soybean diseases, #white mold
Information And Results
Project Summary

The purpose of this project is to utilize a natural biofumigant from oilseeds to mitigate cyst nematodes and white mold in soybean crops. The secondary metabolite, glucosinolates (GSL), has fungicidal and nematocidal properties that make it an excellent candidate as a biofumigant in soybean crops. Once recovered from oilseed meals, the glucosinolates can be applied in fields, be incorporated into the soil, and have a controlled release during the growing season to mitigate the negative impacts of soybean cyst nematodes and white mold. The expected project duration is one year with an estimated budget of $38,688.

Project Objectives

The main objective of this project will be to deploy glucosinolates as a biofumigant to treat SCN and white mold that may cause yield loss This will be the general scope of the project, and some goals that are included, but not limited to, are:

• Evaluate the effectiveness of GSL inhibition in vitro:
o Soybean Cyst Nematode.
o White mold.
• Evaluate the effectiveness of GSL inhibition in situ:
o Soybean Cyst Nematode.
o White mold.
• Evaluate the overall impact of GSL on soybean yield.

Project Deliverables

Professors and students in ABE and AHPS will collaborate to deliver a GSL derived product that can mitigate the growth and spread of SCN and white mold. The estimate time to completion to deliver results on a GSL biofumigant will be one year. The breakdown of the timeline is as follows:

Month 1: Isolate GSL from canola meal. Grow SCN juveniles and white mold cultures in incubator. Prepare soybean plots in a biocontainment greenhouse.
Month 2 – 10: Evaluate GSL inhibition of SCN and white mold in Petri dishes. Plant and grow soybeans, inoculate with juvenile SCNs early in growing, then with white mold when flower buds appear. These trials will have several replications.
Month 11: Analyze data, formulate conclusions and next steps.
Month 12: Prepare publications and final report of findings.

Progress Of Work

Updated October 27, 2022:
The first three months of the project went well overall with surprising discoveries. Extract was obtained from oilseed meals successfully and early trials of soybean cyst nematode treatment were conducted. Soybeans were planted with control and 3 treatment levels of glucosinolate extract, all infected with 300 juvenile nematodes per plant. The early trials were repeated twice, and in both cases cyst nematodes were not significantly affected without adding glucosinolate hydrolysis enzyme. However, soybean root mass and plant height increased with increasing treatment dosage levels in both experiments. This is a surprising discovery, as the extract was only expected to treat the cyst nematodes and was not hypothesized to behave as a biofertilizer. The reason this is surprising is due to the fact that the meal was not used in any of the trials and that was believed to contain the nutrients that allowed oilseed meals to act as a “green manure” from the earlier reviewed literature. Since cyst nematodes were not significantly treated in the early trials, some changes will be made to the downstream processing of glucosinolate recovery to make the treatment more effective (i.e. concentration, purification, etc.). Hopefully, this can make the treatment effective at treating cyst nematodes while maintaining the biofertilizer effect.
The graduate research assistant position was not filled for the first three months and is not expected to be filled in the next three months for the FA2022 semester. Due to this, and the surprising discovery, it is requested that half of the funds for GRA salary and tuition remission be moved to laboratory supplies. The supplies that we plan to acquire will be for identifying and analyzing the nutrients that are coming with the glucosinolates causing the biofertilizer effect. The other half of the GRA funds is requested to be left as originally budgeted, so if a student can be recruited in time for the SP2023 semester the support is still available to them.
There is no request for assistance from the farmers at this time. Trials will continue to be conducted with increased attention to the biofertilizer properties of the proposed treatment.

Updated January 27, 2023:
The greenhouse had a pipe freeze and burst in December resulting in our room losing heat. Thus, our plants froze and died before conclusive data was gathered for our latest fall trials. We replanted 1.5x the plants that we normally do for the next trial to try and catch back up on the research. If all goes well, we should have a large report in the spring.

Updated May 1, 2023:
Currently analyzing data from soybean trials, more students will be assisting on both the FY23 and recently funded FY24 project (continuation of FY23). So far, developing the enzyme to function properly is the largest technical risk for nematicidal effect, and the biostimulant effects for improve growth, stress resistance, etc. is easier and carries a lower technical risk. The SDSU Chem. Dept. will be consulted to help analyze and identify the biocompounds in our treatment that are causing the surprising biostimulant effects for better development. We may be requesting the Final Report for FY23 have a later due date (~Sept. 30) so we can include data and reports from the summer student research fellows in the final report. This will make the research around the FY23 more robust and thorough. Kayla Christopherson, presented the continuation of this project, is currently putting an application proposal together for an NSF iCorps program. This program inspires entrepreneurship in scientists through customer discovery activities. The main technology proposed for her program will be a seed treatment from the technology being developed in this project. This will allow Kayla and the team to connect directly with farmers, seed dealers, ag-chemical dealers, etc. to develop a Business Model Canvas to allow smooth implementation of the technology from a minimum viable product. This will lower the overall risk of technology commercialization and development to directly address farmers needs (create a pull for the tech, rather than a push into the market). These activities will allow us to identify methods for application, that were discussed at the meeting, like dry box spreading, seed coating, no-till applications, etc.

Updated July 30, 2023:
As part of previous research, we simply tried spraying antimicrobial compounds on topsoil. However, this did not yield the results we were hoping for. We believe this is because the antimicrobial compounds become "lost" and separated when simply sprayed on the soil. For this reason, we are trying a new delivery technique: encapsulation. To encapsulate the antimicrobial compounds, we opted for a hydrogel approach. Hydrogels are composed of polymer chains that have been cross-linked to form a 3D network structure. This structure allows hydrogels to swell and absorb large amounts of water, or in our case, GSL and myrosinase. As previously discussed with the Soybean Council, glucosinolates from the Brassicaceae plant family are a form of metabolites that contain many nitrogen and sulfur groups. When hydrolyzed by myrosinase, a catalyst, the glucosinolates break down to form bioactive compounds such as isothiocyanates, nitriles, and thiocyanates. These bioactive compounds possess anti-inflammatory, antioxidant, and antimicrobial capabilities.

For the first few weeks of this summer, we searched for the most effective way to develop a hydrogel. Initially, we used starch as the main structural element and citric acid as the crosslinker. However, this method proved ineffective, as the crosslinker did not strengthen the hydrogel and instead left it as a pile of goop that was very sticky (Image 1). Our next hydrogel utilized pectin and chitosan, cross-linked with sodium hydroxide. Although it took a long time to cure, it eventually formed a nice thin sheet. However, an issue arose. The hydrogel would initially absorb water, but if left to dry a second time, the thin sheet would become brittle and crumble to the touch (Image 2). We attempted to compensate for this issue by adjusting the pectin-to-chitosan ratio, but ultimately, this led to a decrease in the mechanical properties, making it unsuitable for long storage periods. Finally, at the start of July, we used sodium alginate and cross-linked it with a calcium cation either through calcium chloride or calcium carbonate (Image 3). This hydrogel seemed to possess ideal stability.

As of today, we have begun conducting mechanical property testing on various concentrations of sodium alginate to determine the strongest concentration. We have currently sent the hydrogels to a team in the engineering department and have yet to receive the results. We have also initiated microbial inhibition testing, in which we are mixing ampicillin within the hydrogel before it is cured. Subsequently, we will use the Kirby-Bauer inhibition test, except instead of placing an antibiotic disk, we will place a small section of the hydrogel containing ampicillin (Image 4). This test will provide a baseline for when GSL and myrosinase are added, as we will have a better understanding of the diffusion rates of the various concentrations of sodium alginate.

Moving forward, we will perform biodegradability tests. However, due to limited space and time, this aspect of the project has been put on the back burner. Additionally, we will begin incorporating GSL and myrosinase into the hydrogels and conduct inhibition testing.

Final Project Results

Benefit To Soybean Farmers

The soybean cyst nematode and white mold are a hinderance on the yield and thus profitability of South Dakota soybean farmers. There are currently several solutions to managing white mold in soybeans but mitigating the effects of the soybean cyst nematode continues to be a challenge. The soybean cyst nematode (SCN), while currently not a prevalent issue in South Dakota is becoming a growing concern for farmers and processors. Current methods for mitigating the effects of SCN can be expensive, labor intensive, and time consuming. One example is that farmers need to clean equipment when moving between fields to attempt to mitigate the spread. This however can cost valuable time during the planting season and may not always be effective, as SCN eggs are microscopic, so is difficult to determine if the equipment is effectively cleaned. Another technique would be fumigation; however, this can be expensive and in some cases the field may need to lay barren for several years. There is a clear need for a more effective SCN mitigation strategy as the concern grows.

Utilization of biofumigants to manage molds and nematodes has been tested before as a “green manure” approach. Potato farmers in Idaho utilized Brassicaceous biomass in soils contaminated with potato cyst nematodes (PCN). This technique has been proven to be successful but has its drawbacks. First, was a large amount of biomass required to be incorporated into the soil in order to be effective at treating PCN. Second, the meal has valuable use as a feedstock in agriculture and aquaculture due to its protein and amino acid profile. It would be more feasible to reduce the mass and volume needed to be applied in fields to achieve the same outcome. Also, recovering the meal after isolating the active component can prove to be economically feasible. The active component that treated the PCN but is detrimental to the value of canola meal as a feedstock is glucosinolates but has strong potential as a biofumigant.

Isolating the glucosinolates while keeping valuable proteins and amino acids in canola meal can prove to be effective and economical. The recovered glucosinolates can be applied in fields to treat the issue of SCN and white mold. When applied, the soil can be cultivated to incorporate the GSL deeper into the ground. This then can expose the SCN to GSL thus killing and treating the pest. As the GSL breaks down in the soil via microflora, a bioactive and volatile allyl isothiocyanate (A.ITC) is released. A.ITC is known to have potent effects on many types of molds and fungi. As the A.ITC is released from the soil as the GSL is broken down during the growing season, this can provide a secondary effect to protect budding soybeans from white mold. This is a two-in-one benefit from applying a single natural, biofumigant into the soil. However, this will require precision, as the right amounts, at the right times will be crucial for the GSL biofumigant to be effective. Overall, our central hypothesis is that glucosinolates can be an effective treatment to SCN and white mold in South Dakota soybean crops.

The United Soybean Research Retention policy will display final reports with the project once completed but working files will be purged after three years. And financial information after seven years. All pertinent information is in the final report or if you want more information, please contact the project lead at your state soybean organization or principal investigator listed on the project.