Abstract The honeybee disease American foulbrood (AFB) is a serious problem since its causative agent (Paenibacillus larvae) has become increasingly resistant to conventional antibiotics. One of the feasible alternative treatments being used for control of this disease are plants extracts. The aim of the present work was to evaluate the effect of crude extract and fractions of Scutia buxifolia against six Paenibacillus species, including P. larvae, and its potential use for the control of AFB. In vitro activity of S. buxifolia samples against Paenibacillus species were evaluated by the disk diffusion and microdilution methods, and the minimal inhibitory concentration (MIC) were also determined. All Paenibacillus species were sensitive to crude extract and fractions of S. buxifolia. The dichloromethane (DC) fraction showed the better MIC (1.56mg/mL), followed by ethyl acetate (EtAc) (6.25mg/mL), n-butanol (BuOH) (25mg/mL) and Crude extract (CE) (50mg/mL). Toxic effect of S. buxifolia crude extracts and fractions against bees were also evaluated by the spraying application method of the same concentrations of MICs. The samples tested showed no toxic effects for the bees after 15days of observation. These results are first time described for this species and showed that S. buxifolia presented a important activity against Paenibacillus species and proved to be a natural alternative for the prevention/control of AFB.