Immunodetection and quantification of Botrytis cinerea on harvested wine grapes.
1991
Ricker R.W. | Marois J.J. | Dlott J.W. | Bostock R.M. | Morrison J.C.
Water-soluble antigens produced by Botrytis cinerea were detected in spiked and naturally infected grape juice by using an enzyme immunoassay with an indirect format of antibody horseradish-peroxidase conjugates bound to polyclonal rabbit antibodies directed against B. cinerea (anti-Bc IgG). Protein A purified gamma globulin from an early-bled antiserum (803-7), which reacted primarily with low molecular weight compounds present only in extracts of B. cinerea, was used to specifically detect B. cinerea and quantify levels of infection in juice from infected grape berries. Late-bled, higher titer antiserum (803-19), which cross-reacted with proteins and carbohydrates present in extracts from species of Botrytis, Aspergillus, Penicillium, and Uncinula, was used to quantify levels of rot caused by the presence of multiple fungi. Minimum detectable levels of infection, based on mixtures of clean and infected juice, were 0.25-0.5% with 803-7 IgG, and 0.02% with 803-19 IgG. Affinity purification of 803-19 IgG by using antigens from Aspergillus niger coupled to Sepharose beads improved specificity of anti-Bc IgG to B. cinerea but decreased detection sensitivity to approximately 0.5% infection. Cross-reactivity of all anti-Bc IgG collections was consistently low with juice extracted from uninfected grape berries. In contrast, cross-reactivity of anti-Bc IgG with water-soluble antigens extracted from sterile and reproductive structures of several fungi was negligible in early-bled antiserum and increased in subsequent collections. The increase in cross-reactivity in late-bled antisera corresponded with an increase in the overall serum titers for anti-Bc IgG to antigens from B. cinerea. Nonspecific binding of 803-19 IgG was high with extracts from A. niger and an unidentified species of Penicillium, suggesting numerous epitopes common to antigens from these fungi.
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