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United States Department of Agriculture

Agricultural Research Service

Research Project: Developing Resistance to Aflatoxin through Seed-Based Technologies

Location: Food and Feed Safety Research

Title: Evaluation of resistance to aflatoxin contamination in kernels of maize genotypes using a GFP-expressing Aspergillus flavus strain

Authors
item Rajasekaran, Kanniah
item Sickler, Christine
item Brown, Robert
item Cary, Jeffrey
item Bhatnagar, Deepak

Submitted to: World Mycotoxin Journal
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 5, 2013
Publication Date: April 22, 2013
Citation: Rajasekaran, K., Sickler, C.M., Brown, R.L., Cary, J.W., Bhatnagar, D. 2013. Evaluation of resistance to aflatoxin contamination in kernels of maize genotypes using a GFP-expressing Aspergillus flavus strain. World Mycotoxin Journal. 6(2):151-158.

Interpretive Summary: Aflatoxins produced by Aspergillus spp. contaminate diverse food and feed crops including corn, cotton, peanuts and tree nuts. Identification of native resistant genes in germplasm is essential for conventional or molecular crop breeding programs. Unfortunately, resistance to this group of saprophytic fungi is very rare in cultivated crops due to lack of precise host-pathogen relationship. However, in corn, several genotypes have been identified as resistant to aflatoxin contamination due to several factors such as physical barriers (e.g., wax or tough seed coat) or the presence of antifungal proteins in corn kernels. To be successful in breeding programs, it is equally important to know how the fungus invades maize kernels so that objectives of conventional or molecular breeding for tissue or organ specific expression of resistant factors or genes. Recently we utilized an aflatoxigenic A. flavus strain, engineered to express the green fluorescent protein from the jelly fish, to study the site of entry, fungal growth and colonization in corn kernels. We observed that the predominant site of entry is through the stalk or pedicel unless the kernels are injured by insect or other damages. This method of assaying for resistance to the fungus is very sensitive, rapid and simple and can be used to track the fungal colonization in real time. This simple method will be very useful to breeders, geneticists, agronomists and biotechnologists alike who are interested in evaluating germplasm for resistance to Aspergillus.

Technical Abstract: Evaluation of resistance or susceptibility of corn inbreds to infection by Aspergillus flavus was evaluated by a kernel screening assay. A GFP-expressing strain of A. flavus was used to accomplish this study to measure fungal spread and aflatoxin levels in real time. Among the four inbreds tested, Mi82 showed the most resistance and Ga209 the least. TZAR101 was also resistant to fungal infection. Va 35 was susceptible to fungal infection; however, it produced less aflatoxin levels compared to the susceptible line Ga209. Fluorescence microscopy indicated that the site of entry was consistantly through the pedicel. Fungal entry through the pericarp was never observed. Once the fungus entered through the pedicel, it spread quickly through the open spaces between the pericarp and the aleurone layer ultimately colonizing the endosperm and scutellum and finally, the embryo. A clear correlation was established between fungal fluorescence and aflatoxin levels. This method provides a quick, reliable means of evaluating resistance to Aspergillus flavus in undamaged kernels.

Last Modified: 7/22/2014
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