Soybean cyst nematode (SCN) is a major yield-limiting factor of soybean. This disease has spread to at least 24 soybean-producing counties in North Dakota (ND). Cultivar resistance is the primary management tactic. SCN is known to be genetically diverse populations and can develop new virulent forms over time due to continuous use of the same resistance. The shift in SCN populations has led to a decrease in resistance in soybean cultivars derived from PI 88788. Hence, it is imperative to screen soybean cultivars and germplasm to identify resistant soybeans against SCN. In 2019, we screened soybean cultivars, germplasm, and breeding lines for SCN. We targeted on early maturity groups in which resistance can be potentially transferred and used in ND. From the one-time experiment, we found seven soybeans showed resistance to both HG types 2.5.7 and 0, 12 were resistant to HG 2.5.7 only, and 17 were resistant to HG 0 only. However, these results were from a one-time experiment and need to be repeated to confirm the resistance reactions. Additionally, some soybean seed samples have been tested for SCN resistance requested from growers. New seed samples from growers need to be tested to help them select resistant cultivars before planting in infested fields. Sudden death syndrome (SDS) is an important fungal disease of soybean. This disease was first reported in ND in 2020. However, the soybeans used in our SCN resistance screening have not been tested for resistance to SDS.

1. Evaluate 80 soybean cultivars, germplasm and breeding lines for their resistance reactions to two prevalent SCN populations detected in North Dakota.
2. Evaluate 20 soybean cultivars, germplasm and breeding lines for their resistance reactions to SDS detected in North Dakota.

1. Resistance reactions of 80 soybean cultivars, germplasm and breeding lines to SCN will be identified or confirmed.
2. Resistance levels of 20 soybeans will be disclosed.
3. The results will be summarized and made available to soybean farmers and breeders.

Updated November 30, 2021:
Resistance of Soybean Cultivars and Germplasm to Soybean Cyst Nematode

PI: Guiping Yan, Ph.D.
Collaborators: Greg LaPlante, Drs. Carrie Miranda, Sam Markell, and Berlin Nelson

Objectives of the research

1. Evaluate 80 soybean cultivars, germplasm and breeding lines for their resistance reactions to two prevalent SCN populations detected in North Dakota.
2. Evaluate 20 soybean cultivars, germplasm and breeding lines for their resistance reactions to SDS detected in North Dakota.

Completed work

Forty-seven soybean seed samples (commercial cultivars) obtained from growers were tested against HG type 0, and 44 seed samples from growers were assessed against HG type 2.5.7. Fourteen breeding lines from the soybean breeding program at NDSU were tested with HG type 0. A total of 149 early maturity soybean accessions originated from different countries (South Korea, North Korea, Canada, China, Japan, Russia, Ukraine, Vietnam, and United States) were screened against the HG type 2.5.7. These soybean accessions were acquired from the USDA-ARS Soybean Germplasm Collection in Illinois. The soybean cultivar Barnes from North Dakota was included as a susceptible check. Two common SCN populations, HG type 2.5.7. and HG type 0 were collected from two soybean fields in Traill and Richland counties, ND.

Pre-germinated seeds from each of the soybean cultivars and lines and the susceptible check were planted in small cone-tainers each containing 100 cc of sterilized sandy soil in four replicates. Each plant was inoculated with 2,000 eggs of each of SCN HG type 2.5.7 and SCN HG type 0. The plants were kept in a growth chamber maintained at a constant temperature of 27o C with a daylight period of 16 hours (Figure 1). After 35 days of growth, SCN white females (cysts) from plant roots and soil in each cone-tainer were extracted and then counted under a microscope. The mean number of white females produced on the roots of each cultivar and line was used for calculating the female index (FI) by comparing with the susceptible check Barnes using the formula (FI = mean no. of white females produced on a tested line/mean no. of white females in the susceptible check x 100%). Resistance response of each of the lines was categorized based on the female index and the resistance rating system described by Schmitt and Shannon (1992) (resistant: FI < 10%; moderately resistant: FI = 10-30%; moderately susceptible: FI = 31-60%; susceptible: FI > 60%).

The inoculum for SDS resistance evaluation was provided by Dr. Berlin Nelson. Fungal isolate of Fusarium virguliforme was sub-cultured in eleven petri plates with autoclaved PDA media (Figure 1). The inoculated petri plates were covered by lid and wrapped by parafilm tape before incubation. Plugs of actively growing fungal hyphae will be used to inoculate sorghum seeds. The sorghum seeds were provided by Dr. Marisol Berti (NDSU) upon request. The original SDS isolate was kept in daylight to enhance conidia production. The original isolate of F. virguliforme was also cryopreserved in 50% glycerol solution at -80°C for future use. Twenty soybean germplasm lines showing least susceptibility to both SCN populations were selected and will be tested with the fungal pathogen.

Preliminary results

Out of the 47 soybean seed samples obtained from growers tested for SCN HG type 0, 13% of them showed resistant reaction, 19% had moderately resistant reaction, 45% had moderately susceptible reaction, and 23% of them had susceptible reaction. Out of the 44 soybean seed samples from growers tested for SCN HG type 2.5.7, 9% of them showed resistant reaction, 11% of them showed moderately resistant reaction, 30% had moderately susceptible reaction, and 50% had susceptible reaction (Figure 2).

Among the 14 breeding lines obtained from the NDSU soybean breeding program tested for SCN HG type 0, one of them showed resistant reaction, eight of them had moderately resistant reaction, and five of them had moderately susceptible reaction based on the resistance classification described by Schmitt and Shannon (1992).

Out of the 149 soybean germplasm lines from the USDA-ARS Soybean Germplasm Collection tested for SCN HG type 2.5.7, 13 accessions were resistant, 17 accessions were moderately resistant, 81 accessions were moderately susceptible, and 38 accessions were susceptible. Out of the 13 resistant accessions, three originated from China, four from Canada, four from Japan, one from North Korea, and one from Ukraine. These results have been summarized and presented in a manuscript which is now being published in the journal, Plant Health Progress.

Work to be completed

During the rest of the funding cycle, 20 soybean germplasm lines from the USDA-ARS Soybean Germplasm Collection, which showed least susceptibility to SCN, will be tested with the fungal pathogen (F. virguliforme) to determine the levels of resistance or susceptibility to SDS. Twenty-five new soybean seed samples obtained from growers will be tested with both SCN populations, HG type 0 and HG type 2.5.7, upon request. An experiment with 20 soybean lines showing resistant or moderately resistant to HG type 2.5.7 in our previous experiment is being conducted to confirm their resistance reactions. These lines will be tested with another SCN population HG type 0 to further confirm their resistance reactions and to select soybean lines with high resistance to both HG type 0 and 2.5.7. The majority of screened soybean cultivars and lines in our experiments were not resistant to SCN. The valuable resistant soybean lines identified in this study have the potential to be used in breeding SCN-resistant cultivars. The information obtained from this research will be useful to identity resistance genes for developing soybean cultivars with improved genetic resistance to SCN.

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Updated June 30, 2022:
RESISTANCE OF SOYBEAN CULTIVARS AND GERMPLASM TO SOYBEAN CYST NEMATODE

TECHNICAL REPORT

NORTH DAKOTA SOYBEAN COUNCIL
JUNE 2020

Dr. Guiping Yan, Principal Investigator, Dept. Plant Pathology, NDSU
Co-investigators: Dr. Carrie Miranda, Dept. Plant Sciences, NDSU and Dr. Sam Markell, Dept. Plant Pathology, NDSU

Soybean cyst nematode (SCN; Heterodera glycines) and sudden death syndrome (SDS; Fusarium virguliforme) are two economically important disease affecting soybean production worldwide. SCN was first found in North Dakota (ND) in Richland county in 2003 and has spread to more than 26 soybean-producing counties, while SDS was first reported in the same county in 2018 and have been reported in two counties in ND so far. Planting resistant cultivars is often considered the best management strategy to manage SCN and SDS. But only few cultivars are found to be completely resistant to SCN and SDS. The continued use of the limited source of resistance has resulted in high selection pressure and a shift in the SCN virulence populations is capable of breaking the resistance. As a result, the need to find novel sources of resistance and broaden the genetic basis of resistance is increasing.

SDS has a synergistic relationship with SCN. SDS is most likely to occur in areas where SCN is a problem and the disease complex of SCN and SDS causes the most yield losses of soybean. Therefore, soybean lines that have resistance to both SCN and SDS should be identified to manage the disease complex. The major goal of this research is to manage SCN and SDS by identifying new resistance sources for breeding programs and by identifying and utilizing resistant soybean cultivars. The specific objectives of this research were to screen 80 soybean cultivars, germplasms and breeding lines for resistance to HG type 0 and HG type 2.5.7 detected in North Dakota, and to screen 20 soybean cultivars, germplasm and breeding lines for their resistance reactions to SDS detected in North Dakota.

101 soybean lines (29 soybean accessions of early maturity group ranging from 0-I from USDA-ARS Soybean Germplasm Collection in Urbana, IL and 72 commercial cultivars from growers) were screened for their resistance reactions to HG type 2.5.7. 110 soybean lines (21 soybean accessions of early maturity group ranging from 0-I from USDA-ARS Soybean Germplasm Collection in Urbana, IL, 75 commercial cultivars from growers, and 14 breeding lines from the NDSU soybean breeding program) were screened for their resistance responses to HG type 0.

Pregerminated seedlings of each soybean line were planted into a cone container each containing 100 cm3 of pasteurized river sand soil replicated four times. Each plant was inoculated with 2000-2500 eggs at the time of planting and kept in a controlled growth chamber maintained at 27 °C and daylight of 16 hours for 30 days. After 30 days, the plants were harvested and SCN white females were extracted from the roots and soil of individual plants and counted on a lined petri-plate under a microscope. The number of white females in the four replicates was averaged to calculate the mean number of white females which was used to calculate the female index (FI) as, FI = mean no. of white females produced on a tested soybean line/mean no. of white females on the susceptible check Barnes x 100 %. Soybean lines were classified for their resistance reactions based on the female index as described by Schmitt and Shannon (1992) as resistant (R) (FI < 10%), moderately resistant (MR) (10% = FI < 30%), moderately susceptible (MS) (30% = FI < 60%), and susceptible (S) (FI = 60%).

Based on the previous SCN screening experiments, 21 soybean accessions from USDA-ARS Soybean Germplasm Collection in Urbana, IL, that were resistant to either HG type 0 or HG type 2.5.7 were selected and tested for their resistance reactions to the SDS pathogen found in North Dakota. Fusarium virguliforme, the causal agent of SDS, was grown in half strength PDA media and used to infect autoclaved sorghum seed in a conical flask with 10 plugs (8mm diameter) of actively growing fungal hyphae. Sorghum seeds were incubated at room temperature for 2 weeks and air dried before use.

Two experiments were conducted to screen the soybeans for resistance to the SDS pathogen (Figure 1). 500 ml plastic cups were filled with 350 gram of pasteurized river sand soil and inoculated by a layered method with 3-5 gm of fungus-infested sorghum seeds. Six seeds of each line were planted per pot and covered with a top layer of soil. Each line was replicated 5 times. Cultivar Barnes with autoclaved sorghum seeds was used as a control. Only four healthy plants were allowed to grow per pot after germination, and kept in the greenhouse for 40 days. The disease was evaluated based on foliar disease severity and root rot severity. Foliar disease severity was estimated based on the percentage of chlorotic or necrotic leaf area in total leaf area and root rot severity was estimated according to the percentage of root area showing brown lesions or discoloration in total area of tap root.

Among the 110 soybean cultivars, breeding lines and germplasm tested for HG type 0, 16% were resistant, 29 % were moderately resistant, 32% were moderately susceptible, and 23% were susceptible. Among the 101 soybean cultivars and germplasm tested for HG type 2.5.7, 15% were resistant, 5% were moderately resistant, 24% were moderately susceptible, and 56% were susceptible. Multiple lines were resistant to both HG type 0 and 2.5.7. The testing results for selected soybean cultivars, breeding lines and germplasm are shown in Tables 1 and 2.

In addition, 149 soybean germplasm lines from the USDA-ARS Soybean Germplasm Collection were tested for SCN HG type 2.5.7. The results have been summarized and presented in a manuscript which is now published in the journal, Plant Health Progress (Acharya, K. and Yan, G. P. 2022. Screening of early maturing soybean accessions for resistance against HG Type 2.5.7 of soybean cyst nematode, Heterodera glycines. Plant Health Progress 23:166-173). Briefly, 13 of the accessions were resistant, 17 of the accessions were moderately resistant, 81 were moderately susceptible, and 38 were susceptible. Out of the 13 resistant accessions, three originated from China, four from Canada, four from Japan, one from North Korea, and one from Ukraine.

Significant difference in root rot severity was observed among the 21 lines screened for SDS (Figure 2). PI 603151A and PI 603424A showed the maximum root rot severity (28.3%) and foliar disease severity (10.9%), respectively, in Experiment 1 while PI 6031148 showed the maximum root rot severity (38.5%) and foliar disease severity (8.4%) in Experiment 2. Among the lines that had root rot severity of less than 10%, PI 548642 had the lowest root rot severity (2.5%) followed by PI 603426B, PI 594314, and PI 603438A in Experiment 1. In Experiment 2, among the lines having root rot severity less than 10%, PI603169 and PI603426B had the lowest root rot severity (7.5%) followed by PI603153. In both trials, PI 603426B (7.8% and 7.5%) had root rot severity less than 10%. Foliar symptoms were not common in both the trials and therefore the disease was mainly evaluated based on the root rot severity.

The data have been communicated to the crop consultants, farmers, and breeder who provided the seed. The results will be further validated to confirm the resistance reactions of the soybean lines to SCN and SDS. Molecular techniques will be used to further elucidate the resistance genes or loci in the germplasm before using them in breeding programs to develop cultivars resistant to SCN. This research will be useful for the farmers to select resistant soybean cultivars in infested fields and for the breeder to develop new cultivars with improved resistance to manage SCN and SDS.

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RESISTANCE OF SOYBEAN CULTIVARS AND GERMPLASM TO SOYBEAN CYST NEMATODE

EXECUTIVE SUMMARY

NORTH DAKOTA SOYBEAN COUNCIL
JUNE 2022

Dr. Guiping Yan, Principal Investigator, Dept. Plant Pathology, NDSU
Co-investigators: Dr. Carrie Miranda, Dept. Plant Sciences, NDSU and Dr. Sam Markell, Dept. Plant Pathology, NDSU

Research Conducted
101 soybean cultivars, breeding lines and germplasm were screened for their resistance reactions to SCN HG type 2.5.7 and 110 were screened for HG type 0. Plants were inoculated with SCN eggs and kept in a growth chamber at 27 °C for 30 days (Figure 1). SCN white females were extracted and female index was calculated to categorize resistance phenotypes. 21 lines were further selected and tested for their reactions to Fusarium virguliforme, the fungal pathogen of sudden death syndrome (SDS). Root rot severity was evaluated for levels of resistance or susceptibility.

Why the research is important to ND soybean farmers
Both SCN and SDS are important diseases in soybean. SDS is most likely to occur in areas where SCN is a problem and SDS combined with SCN cause the most yield losses. Soybean cultivars with resistance to both SDS and SCN are desirable for managing this disease complex. This research is important to navigate the new resistance sources that should be introduced to breeding programs for developing new resistant cultivars and help growers select resistant cultivars.

Final findings of the research
Among the 101 soybeans tested for HG type 2.5.7, 15% were resistant, 5% were moderately resistant, 24% were moderately susceptible, and 56% were susceptible. Among the 110 soybeans for HG type 0, 16% were resistant, 29 % were moderately resistant, 32% were moderately susceptible, and 23% were susceptible. Multiple lines were resistant to both HG type 0 and 2.5.7.

Significant difference in root rot severity was observed among the 21 lines screened for SDS (Figure 2). PI 548642 had the lowest root rot severity (2.5%) followed by PI 603426B, PI 594314, and PI 603438A in Experiment 1. PI 548642 also showed resistance to HG type 2.5.7 and moderate resistance to HG type 0. In Experiment 2, PI603169 and PI603426B had the lowest root rot severity (7.5%) followed by PI603153. These lines had root rot severity less than 10%, indicating a good level of resistance. PI 603426B had root rot severity less than 10% in both trials.

Benefits/Recommendations to North Dakota soybean farmers and industry
The data have been communicated to the crop consultants, farmers, and breeder who provided the seed. The findings of this research will be useful for the farmers to select resistant soybean cultivars in infested fields and for the breeder to develop new cultivars with improved resistance to manage SCN and SDS.

Both SCN and SDS are imponant diseases in soybean. SDS is most likely to occur in areas where SCN is a problem and SDS combined with SCN cause the most yield losses of soybean. The resistant soybean cultivars identified in this proposed research will be recommended to farmers for suppressing SCN and SDS in infested fields. The soybean cultivars with dual resistance to both SDS and SCN are desirable for managing this disease complex. The highly resistant soybean germplasm and breeding lines confirmed in this proposed research will be provided to soybean breeding programs for transferring the resistance to locally adapted susceptible cultivars to develop new cultivars with improved genetic resistance. Thus, this research is important to navigate the new resistance sources that should be introduced to soybean breeding programs for developing new resistant cultivars and help growers select the resistant cultivars for controlling the nematode and fungal diseases to increase soybean yield.