Rotavirus (at 1 × 10 7.5 /mL TCID 50 ) was incubated with T. esculentum water and ethanolic extracts at room temperature for 1 h. Following the incubation, dilutions of the incubated virus were added to CLAB, CIEB, and H4 cell monolayers in 96well plates and then incubated for 1 h at 37 • C in a humidified atmosphere of 5% CO 2 and 95% air, after which the unattached virus and the extracts were washed off with PBS. DMEM (Sigma-Aldrich, Grand Island, USA), supplemented with 10% foetal calf serum (Cambrex, Verviers, Belgium), L-glutamine (2 mmol/L, Sigma), penicillin (100 units/mL, Sigma), and streptomycin (1 mg/mL, Fluka, Buchs, Switzerland), was then introduced on the cells and then incubated for 24 h at 37 • C in atmosphere of 5% CO 2 until viral cytopathic effects were observed in control wells. Cell viability was then determined with quantification of crystal violet as described previously. As control, viral only, T. esculentum only and cell culture medium only treatments were plated onto the epithelial cells. Relative protection by T. esculentum extracts against RV infection was determined by crystal Tests were done in triplicate wells. Tylosema esculentum extracts used were as follows: cotyledon ethanolic extract (MCE), seed coat ethanolic extract (MSCE), tuber water extract (MTW), cotyledon water extract (MCW), and seed coat water extract (MSCW). Tylosema esculentum extracts were introduced at and below IC 50 or at 1 and 0.1 mg/mL of MCE (the highest concentration of MCE was not cytotoxic).