The study investigated the mycological safety of Bambara groundnut (Vigna subterranea (L.) Verdc) from Southern Africa. Conventional and molecular methods identified aflatoxin-producing Aspergillus flavus in Bambara seed while a modified QuEChERS method for mycotoxin detection confirmed the presence of aflatoxin in the flour. Flour samples obtained by milling, roasting and milling, and spontaneous fermentation stored for 30 days at simulated tropical conditions (30¿±¿1¿°C, 85¿±¿2% relative humidity) were analyzed for changes in fungal counts, water activity and aflatoxin production. A co-occurrence of Aflatoxin (AF) B1 (0.13–6.90), AFB2 (0.14–2.90), AFG1 (1.38–4.60), and AFG2 (0.15–1.00) was recorded in the flour samples. Over the storage period, there was significant (p¿=¿0.05) increase in the water activity of roasted Bambara groundnut flour (0.09–0.95), fermented Bambara groundnut flour (0.34–1.02), and Bambara groundnut flour (0.42–0.89); and in the fungal counts irrespective of the processing method. Roasting at 140¿°C for 20¿min completely degraded aflatoxin in the seeds. Fungal and mycotoxin contamination of Bambara groundnut raises concerns on health of consumers.