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Peanuts (Arachis hypogaea) are of significant agricultural importance due to their versatile uses, providing a valuable source of edible oil and protein-rich food products and serving in industrial applications. Peanuts are susceptible to various diseases, one of them being net syndrome, a poorly understood threat triggered by an undefined causal agent, with symptoms including black coloring and lesions of the peanut pod that affect product quality. In this study, amplicon sequencing was harnessed to explore the fungal and bacterial populations associated with the syndrome. Using this approach, we identified a specific Fusarium sp. amplicon sequence variant that demonstrated a significant correlation with the presence of net syndrome symptoms in both young and mature peanuts. This finding was further validated by Koch's postulates testing, including isolation, cultivation, and application of this strain to corroborate its involvement in net syndrome in peanuts. Our findings contribute to a better understanding of the interactions between peanuts and their microbial communities and to identification of the previously undetermined agent involved in net syndrome and the etiology of the disease.