AimThis study evaluated the binding capacity of aflatoxin B-1 (AFB(1)) by two Enterococcus faecium strains (MF4 and GJ40) isolated from faeces from healthy dogs. Materials and MethodsThe binding assay was performed using 50 and 100ppb of AFB(1) analysing the effects of the viability, incubation time and pH on AFB(1) binding(.) Binding stability was determined by washing three times the bacteria-AFB(1) complexes with phosphate buffer saline. ResultsBoth GJ40 and MF4 strains have the ability to remove AFB(1) from aqueous solution. Viable cells were slightly more effective in AFB(1) binding than nonviable ones for both strains. Enterococcus faeciumGJ40 removes 24-27% and 17-24%, and Ent.faeciumMF4 removes 36-42% and 27-32% of AFB(1) (50 and 100ppb, respectively) throughout a 48h incubation period. In general, the removal of AFB(1) was highest at pH 700 for both strains. The stability of the bacteria-AFB(1) complex formed was found to be high (up to 50% of AFB(1) remained bounded in bacterial cell after three washes with phosphate buffered saline). ConclusionThe Ent.faecium strains assayed are capable of removing AFB(1) under different conditions in vitro. Significance and Impact of the StudyThis is the first AFB(1) binding assay performed with Ent.faecium strains isolated from dog faeces, being an interesting strategy for AFB(1) decontamination of pet food.