Serological characterization of Pythium ultimum

An ELISA protocol was developed for rapid identification of P. ultimum using a species-specific monoclonal antibody, MAb E5. Microscopic identification was compared with ELISA for identification of 246 cultures of Pythium spp. P. ultimum isolates were identified by ELISA with {dollar}>{dollar}99% accuracy.; Culture factors influenced the detection of P. ultimum by MAb E5 in ELISA. Antigen production was greatest in Czapek-Dox broth. Sucrose, glucose, maltose, fructose, and mannose stimulated antigen production; whereas xylose, ribose, and galactose suppressed production. Seed exudates also stimulated antigen production. These results indicate that antigen production depends on substrate composition.; The antigen that binds to Mab E5 is a 46 kDa glycoprotein. The epitope is in the glycosidic moiety and could be composed of {dollar}beta{dollar}-(1-3)-linked glucose residues. The antigen was thermostable and reactivity with Mab E5 increased after P. ultimum mycelia were boiled. The Mab E5 antigen is localized in the cell wall. The antigen is produced only by young actively-growing mycelia and is then released into the culture media. Antigen concentrations decreased in mycelia and sporangia as they aged, but the antigen reappeared in hyphal tips after aged mycelia were exposed to fresh nutrients or in sporangia when they were induced to germinate by nutrients.; Other MAbs to P. ultimum, X4, A3, and A4 also recognized a 46 kDa protein in all Pythium isolates tested by Western blot. They did not recognize any protein in P. irregulare isolates.