The contamination of corn (maize) by fungi and the accumulation of mycotoxins are a serious agricultural problem for human and animal health. One particular devastating group of mycotoxins, called aflatoxins, has been intensely studied since the 1960s. Studies of Aspergillus flavus, the agriculturally relevant producer of aflatoxins, have led to a well-characterized biosynthetic pathway for aflatoxin production, as well as a basic understanding of the organism’s life cycle. Unfortunately, these efforts have not resulted in corn production practices that substantially reduce aflatoxin contamination. Similarly, the use of agrochemicals (e.g., fungicides) results in very limited reduction of the fungus or the toxin. Thus, cultural management (fertility and irrigation) coupled with aggressive insect management is current recommendation for integrated aflatoxin management. The development of resistant hybrids appears to be a very promising technology, but commercial hybrids are still not available. Thus, biocontrol appears to be the most promising available avenue of reducing aflatoxin accumulation. Biocontrol utilizes nontoxigenic strains of Aspergillus to reduce the incidence of toxin-producing isolates through competitive displacement. To maximize the effectiveness of biocontrol, a thorough knowledge of the environmental factors influencing colonization and growth of Aspergillus is needed. A. flavus not only colonizes living plant tissue, but it also grows saprophytically on plant tissue in the soil. These residues serve as a reservoir for the fungus, allowing it to overwinter, and under favorable conditions it will resume growth and release new conidia. The conidia can be transmitted by air or insects to serve as new inoculum on host plants or debris in the field. This complex ecology of Aspergilli has been studied, but our understanding lags behind what is known about biosynthesis of the toxin itself. Our limited understanding of Aspergilli soil ecology is in part due to limitations in evaluating Aspergilli, aflatoxin, and the biosynthetic genes in the varying aspects of the environment. Current methods for assessing Aspergillus and aflatoxin accumulation rely heavily on cultural and analytical methods that are low throughput and technically challenging. Thus to understand Aspergillus ecology and environmental effects in contamination to maximize biocontrol efforts, it is necessary to understand current treatment effects and to develop methodologies capable of assessing the fungal populations present. In this manuscript we discuss the current knowledge of A. flavus ecology, the application of selected molecular techniques to field assessments, and crop practices used to reduce aflatoxin contamination, focusing on chemical treatments (fungicides and herbicides), insect management, and crop management.

This study compared cultural and analytical methods for detecting aflatoxin production by Aspergillus species. Aspergillus isolates were obtained from various Mississippi Delta crops (corn, peanut, rice, cotton) and soils. Most of the isolates (99%) were A. flavus and the remainder comprised A. parasiticus and A. nomius. The following three cultural methods were evaluated on potato dextrose agar: fluorescence (FL) on β-cyclodextrin-containing media (CD), yellow pigment (YP) formation in mycelium and medium, and color change after ammonium hydroxide vapor exposure (AV). Aflatoxins in culture extracts were confirmed by thin-layer chromatography (TLC) and quantified by enzyme-linked immunosorbent assay (ELISA). Of the 517 isolates, 314 produced greater than 20 ng/g of total aflatoxin based on ELISA, and 180 produced greater than 10 000 ng/g of aflatoxin in the medium. Almost all the toxigenic isolates (97%) were confirmed by TLC as producers. Of the toxigenic isolates, as determined by ELISA, 93%, 73%, and 70% gave positive FL, YP, and AV responses, respectively. Of the 203 isolates producing less than 20 ng/g of aflatoxin, 20%, 6%, and 0% of respective FL, YP, and AV methods gave false-positive responses. The 9% false-positive results from TLC fall within this range. This study showed good agreement among all tested cultural methods. However, these cultural techniques did not detect aflatoxin in all cultures that were found to produce aflatoxins by ELISA, LC/MS, and TLC. The best results were obtained when the AV color change and CD fluorescence methods were used together, yielding an overall success rate comparable to TLC but without the need for chemical extraction and the time and expense of TLC.Key words: aflatoxins, analytical methods, Aspergillus flavus, Aspergillus parasiticus, cultural methods, β-cyclodextrin, fluorescence enhancers, mycotoxins, yellow pigment.

The fungus Aspergillus flavus is responsible for producing carcinogenic mycotoxins, the aflatoxins, on corn (maize) and other crops. An additional harmful toxin, cyclopiazonic acid, is produced by some isolates of A. flavus. Several A. flavus strains that do not produce one or both of these mycotoxins are being used in biological control to competitively exclude the toxin-producing strains from the agroecosystem, particularly from seeds, grain and other marketable commodities. Three well-studied non-aflatoxigenic strains, including two that are commercially available, have been compared in side-by-side field trials. The results of that study, together with a growing understanding of A. flavus ecology and new genetic insights, are guiding the selection of biocontrol strains and influencing crop management decisions for safe and sustainable production.