The phytotoxic effects of aflatoxin B1(AFB1) onin vitrocultures of differentiating calli and regenerating plantlets ofNicotiana tabacumwere assessed. Callus appeared more sensitive to the effects of AFB1, with fresh mass accumulation and callus chlorophyll levels affected at low (approximately 0.5μg/ml) aflatoxin concentrations. Transmission electron microscopy revealed early deteriorative alterations in chloroplast morphology. Inhibitory effects of the toxin (up to and including 10 μg/ml) on callus fresh mass accumulation were reversed following a 3 week toxin‐free recovery period. In tobacco plantlets, root and leaf development, and root and leaf mass were significantly inhibited in a dose‐dependent fashion with increasing AFB1concentration above 0.5μg/ml. Inhibitory effects on plantlet root development were more pronounced that on leaf development.