Aflatoxin, the most potent carcinogen found in nature, is produced by the fungus Aspergillus flavus and occurs naturally in maize, Zea mays L. Growing maize hybrids with genetic resistance to aflatoxin contamination are generally considered a highly desirable way to reduce losses to aflatoxin. Developing resistant hybrids requires reliable inoculation methods for screening maize germplasm for resistance to A. flavus infection and aflatoxin accumulation. The side-needle technique is a widely used inoculation technique: an A. flavus conidial suspension is injected underneath the husks into the side of the ear. This wounds the ear and limits expression of resistance associated with husk coverage, pericarp thickness, and seed coat integrity. In this investigation, the side-needle technique was compared with a second inoculation method that involved dispensing wheat kernels infected with A. flavus into plant whorls at 35 and 49 days after planting. Results showed that although the side-needle technique produced higher levels of aflatoxin accumulation, differences in A. flavus biomass produced by the two inoculation techniques were not significant. Both inoculation techniques were effective in differentiating resistant and susceptible single cross hybrids irrespective of the use of A. flavus infection or aflatoxin accumulation as a basis to define resistance. 1. Introduction Aflatoxin is produced by the fungus Aspergillus flavus and occurs naturally in maize, Zea mays L. Aflatoxin, the most potent carcinogen found in nature, is toxic to both humans and animals [1–3]. Dietary exposure to aflatoxin is one of the major causes of hepatocellular carcinoma, the fifth most common cancer in humans worldwide [4]. Although aflatoxin was first identified and recognized as a threat to animals after 100,000 turkeys died in England in 1961 [5–7], it is now known to be a threat to other livestock, pets, and wildlife [8–10]. The U. S. Food and Drug Administration restricts the sale of grain with aflatoxin levels exceeding 20？ng/g [2]. Aflatoxin was first recognized as a major problem for maize production in the southeastern United States in the 1970s. In 1977, over 90% of maize was contaminated with aflatoxin, and aflatoxin levels exceeded 20？ng/g in 90% of samples evaluated in Georgia [11, 12]. Aflatoxin contamination has remained a chronic problem in the Southeast where it reached devastating proportions in 1998 [13, 14]. Losses to aflatoxin-contaminated corn in Arkansas, Louisiana, Mississippi, and Texas were estimated at $85,000,000 to $100,000,000 [15]. An increase in
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