Dot-Immunobinding Assay for Detecting Xanthomonas campestris pv. holcicola in Sorghum. J. E. Leach, Assistant Professor, Department of Plant Pathology, Kansas State University, Manhattan 66506. B. A. Ramundo, D. L. Pearson, and L. E. Claflin. Research Assistant, Research Assistant, and Associate Professor, Department of Plant Pathology, Kansas State University, Manhattan 66506. Plant Dis. 71:30-33. Accepted for publication 4 September 1986. Copyright 1987 The American Phytopathological Society. DOI: 10.1094/PD-71-0030.

The application of a dot-immunobinding assay (DIA) for detecting Xanthomonas campestris pv. holcicola (X. c. pv. holcicola), the bacterial streak pathogen of sorghum (Sorghum bicolor), is described. Leaf sections with symptoms were soaked in sterile distilled water, and the soaking solution containing the bacteria was spotted onto nitrocellulose membranes. Membranes containing the absorbed antigens were incubated in antiserum to glutaraldehyde-fixed cells of X. c. pv. holcicola and then in protein A-alkaline phosphatase conjugate. To visualize the antigen-antibody-protein A complexes, membranes were exposed to naphthol AS-MX phosphate substrate with fast-violet B stain; a violet precipitate indicated a positive response. The assay was rapid (about 4 hr) and sensitive, i.e., 400 bacterial cells per 4-µl sample could be detected (equivalent to 10⁵ cfu/ml). Of 101 plant samples assayed, 46 of 48 known to be infected with X. c. pv. holcicola were detected by the DIA.