DEVELOPMENT OF SOLID-PHASE IMMUNOASSAYS FOR THE DETECTION AND QUANTIFICATION OF ASPERGILLUS-OCHRACEUS IN WHEAT-GRAIN

Aspergillus ochraceus Wilhelm is a common contaminant of stored foods, especially cereal grains, which produces the mycotoxin, ochratoxin A (OA). Solid-phase immunoassays (enzyme-linked immunosorbent assay [ELI SA] and immunoblotting), utilizing antibodies raised in rabbits to A. ochraceus exoantigens (ExAgs, a mixture of extracellular macromolecule s), were evaluated for their ability to provide an index of the degree of A. ochraceus contamination in cereals. A. ochraceus could be readi ly detected with a minimal degree of interference or cross-reactivity from the wheat matrix or from other fungal species naturally present i n wheat. The ELISA was sensitive, with the limits of detection for ExA gs being as low as 50 ng ml(-1) . The amounts of A. ochraceus ExAgs de tected in wheat that was naturally molded and inoculated with A. ochra ceus correlated favorably with other parameters indicative of the pres ence of the fungus. These parameters included the amount of OA (r = 0. 93, P < 0.05), the percentage of A. ochraceus infection (r = 0.89, P < 0.05), the number of colony-forming units (r = 0.68, P < 0.05) and th e glucosamine content (r = 0.64, P < 0.05). Immunoblotting patterns of ExAgs extracted from liquid and wheat cultures of A. ochraceus demons trated that ExAgs consisted of several antigenic components, with the immunodominant ones having molecular weights of approximately 20 or 30 kDa. The concentration of particular components appeared to be influe nced by the media on which A. ochraceus was cultivated. The data sugge st that the immunoassays developed for A. ochraceus ExAgs can be used for the identification and quantitation of this fungus in grain.