DEVELOPMENT OF METHODOLOGY FOR NONDESTRUCTIVE ASSAY OF BACTERIA, FUNGI AND VIRUSES IN SEEDS OF LARGE-SEEDED FIELD CROPS

Germplasm collections are a major source of potentially harmful seedbo rne pathogens. Conventional procedures to assay for pathogens in seeds generally result in destruction of the seeds, which is undesirable fo r germplasm accessions containing small numbers of high-value seeds. M ethods to assay seeds nondestructively for pathogens were investigated in a series of pathogen/host combinations: Pseudomonas syringae pv. s yringae van Hall, P. syringae pv. phaseolicola (Burkh.) Dowson, and Xa nthomonas campestris pv. phaseoli (E. F. Smith) Dowson in bean (Phaseo lus vulgaris L.); soybean mosaic virus in soybean (Glycine max L. [Mer r.]); and seedborne fungi in com (Zea mays L.), including Helminthospo rium carbonum Ullstrup, Diplodia maydis (Burkh.) Sacc., Penicillium, a nd Fusarium spp. Excising cores of seed tissues proved to be the most successful approach over the three major steps in the process: extract ion of the pathogen; assay of the extract according to pre-existing me thods such as incubation on culture plates and blotters, or enzyme-lin ked immunosorbent assay (ELISA); and restoration of seeds to a safe ph ysiological state. Short-term incubation of seeds on a culture medium or leaching pathogens from seeds in a buffer solution or liquid medium showed promise as means of extracting and assaying for the pathogen, but resulted in severely reduced seed germination. The feasibility of nondestructively assaying seeds of large-seeded plant species for bact erial, viral, and fungal pathogens was demonstrated.