Development and Application of an In Vitro Method to Evaluate Anthracnose Resistance in Soybean Germplasm

Anthracnose caused by Colletotrichum truncatum is a major fungal disease of soybean, especially vegetable soybean (edamame). Studies of this disease have mainly focused on resistance evaluation, but the primary methods used-in vivo inoculation of pods or plants under greenhouse or field conditions-have limitations with respect to accuracy, stability, scale, and environmental safety. In this study, we developed a method for inoculating pods in vitro by soaking in a mycelial suspension. We optimized the crucial components, including the mycelial suspension concentration (40 to 60 mg mL^-1), the maturity of the sampled pods (15 days after flowering), and the post-inoculation incubation period (5 days). Application of the mycelial suspension by soaking rather than spraying improved the efficiency of inoculation and made large-scale evaluation possible. Using this method, we evaluated 589 soybean germplasm resources (275 cultivars, 233 landraces, and 81 wild accessions). We identified 25 highly resistant cultivars, 11 highly resistant landraces, but only one highly resistant wild accession. Our results will aid future research on soybean anthracnose resistance, including gene discovery, the elucidation of molecular mechanisms, and the breeding of resistant cultivars.