Revelation of antifungal susceptibility of botanicals and genetic diversity of aflatoxin-producing Aspergillus flavus in chili (Capsicum annuum)

Aflatoxins associated with Aspergillus flavus pose a significant risk to food safety and public health. This study determined the minimum inhibitory concentration (MIC) of five medicinal plant extracts against A. flavus. Molecular identification, performed by amplifying the ITS rDNA region, confirmed the isolate as A. flavus (99.80% similarity to the GenBank sequence OP480002.1). The antifungal activities of botanical extracts (Piper betle, Centella asiatica, Azadirachta indica, Mentha spicata, and Ocimum tenuiflorum) at different concentrations (600 mg/ml, 300 mg/ml, and 100 mg/ml) were evaluated using disc diffusion and broth dilution methods, with Fluconazole as a reference. Except for M. spicata, all extracts exhibited inhibitory activity, with A. indica showing the highest inhibition (19.83 mm at 600 mg/ml), surpassing Fluconazole (15.67 mm). The MIC values were lowest for Fluconazole (4.68 mg/ml), followed by A. indica (9.375 mg/ml). RAPD-PCR analysis revealed significant genetic diversity among isolates, with 12 primers generating 50 polymorphic bands. Genetic similarity ranged from 73% to 99.2%, with the highest diversity observed between isolates AS3 and AS4. These findings highlight the potential of these medicinal plant extracts as natural alternatives to synthetic fungicides at specific concentrations for reducing aflatoxin contamination in agricultural products. Furthermore, the observed genetic diversity may help in assessing the aflatoxin production capacity of A. flavus in different chili cultivars.