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Aspergillus flavus is a ubiquitous facultative pathogen that routinely infects important crops leading to formation of aflatoxins during crop development and after harvest. Corn and peanuts in warm and/or drought-prone regions are highly susceptible to aflatoxin contamination. Controlling aflatoxin using atoxigenic A. flavus is a widely adopted strategy. However, no A. flavus genotypes are currently approved for use in China. The current study aimed to select atoxigenic A. flavus endemic to Guangxi Zhuang Autonomous Region with potential as active ingredients of aflatoxin biocontrol products. A total of 204 A. flavus isolates from corn, peanuts, and field soil were evaluated for ability to produce the targeted mycotoxins. Overall, 57.3% could not produce aflatoxins while 17.15% were incapable of producing both aflatoxins and CPA. Atoxigenic germplasm endemic to Guangxi was highly diverse, yielding 8 different gene deletion patterns in the aflatoxin and CPA biosynthesis gene clusters ranging from no deletion to deletion of both clusters. Inoculation of corn and peanuts with both an aflatoxin producer and selected atoxigenic genotypes showed significant reduction (74 to 99%) in aflatoxin B 1 (AFB 1) formation compared with inoculation with the aflatoxin producer alone. Atoxigenic genotypes also efficiently degraded AFB 1 (61%). Furthermore, atoxigenic isolates were also highly efficient at reducing aflatoxin concentrations even when present at lower concentrations than aflatoxin producers. The use of multiple atoxigenics was not always as effective as the use of a single atoxigenic. Effective atoxigenic genotypes of A. flavus with known mechanisms of atoxigenicity are demonstrated to be endemic to Southern China. These A. flavus may be utilized as active ingredients of biocontrol products without concern for detrimental impacts that may result from introduction of exotic fungi. Field efficacy trials in the agroecosystems of Southern China are needed to determine the extent to which such products may allow the production of safer food and feed. Atoxigenic A. flavus isolates obtained from naturally contaminated sources, screened for aflatoxin production potential and molecular basis of atoxigenecity. Selected atoxigenic isolates effectively reduced aflatoxin formation in corn and peanuts when co-inoculated with toxigenic A. flavus. [Display omitted] • Atoxigenic biocontrol using native strains is a sustainable approach for aflatoxins mitigation. • The population of atoxigenic A. flavus endemic to Southern China and their genetic basis of atoxigenecity were explored. • Southern China atoxigenic A. flavus exhibited 8 gene deletion patterns in aflatoxin and CPA biosynthesis gene clusters. • Selected isolates were highly efficient in inhibiting aflatoxin formation in corn (up to 95.85%) and peanuts (99.49%). • AFB 1 degradation by atoxigenic isolates proved the second mechanism of aflatoxin mitigation. [ABSTRACT FROM AUTHOR]