DETECTION OF PENICILLIUM-AURANTIOGRISEUM BY ELISA UTILIZING ANTIBODIES PRODUCED AGAINST ITS EXOANTIGENS

An indirect competitive ELISA was developed using rabbit antiserum aga inst the exoantigens (ExAgs) of Penicillium aurantiogriseum, and cross -reactivities were determined with 15 ExAgs from other species and gen era of fungi and with the water-soluble extracts from four grains. The assay had good sensitivity to P. aurantiogriseum ExAgs (1 mu g per ml serum), with generally low or no cross-reactivity with grain extracts and the other fungi, including five species of Penicillium, four spec ies of Aspergillus, three species of Fusarium, two species of Mucor an d one species of Alternaria. Immunoblotting analysis of the ExAgs from P. aurantiogriseum yielded at least 20 distinct antigenic bands, with about 3 or 4 being dark, 10-12 light and 5-8 faint in colour. Most of the bands had molecular masses of about 70-90 kDa. In contrast to P. aurantiogriseum, the ExAgs from the other fungi either did not react i n the immunoblotting assay with the antiserum, or reacted only weakly with it. One antigenic band from P. roqueforti, however, reacted moder ately strongly with the antiserum. The ELISA and immunoblotting analys is were used to detect P. aurantiogriseum in naturally contaminated wh eat samples, and in wheat samples spiked with a second common storage fungus, Aspergillus ochraceus. The results demonstrated that the antis erum was able to detect P. aurantiogriseum in a background of other fu ngal species, and that these did not produce false positives. There wa s also a positive association between the concentration of P. aurantio griseum ExAgs, as measured by ELISA, and the amount of mould as determ ined by the number of colony-forming units. Immunoblotting qualitative ly confirmed the ELISA results obtained with fungi when cultured on li quid or on solid (wheat) media. The data suggest that the immunoassays developed for P. aurantiogriseum are useful for the detection and ide ntification of P. aurantiogriseum on the basis of its ExAgs, with the advantages of being more efficient, simple and reliable than conventio nal techniques.