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The aim of this study was evaluate the treatment of Bubalus bubalis (dairy buffaloes) affected with clinical mastitis, through the use of Mimosa tenuiflora botanical extract, using as parameters for this study: the semiotic examination and strip cup test, number of colonies count by using Petrifilm plates; isolation and identification of microorganisms found in the animal’s milk before and after treatment, toxicological testing and bactericidal kinetics with strains of Staphylococcus aureus collected from buffalo milk. We observed that the extract of M. tenuiflora exerts bactericidal effect on S. aureus, at the concentration of 107.5 mg/mL as low acute toxicity. In this context, it can be concluded that M. tenuiflora extract presents an antimicrobial activity about S. aureus, with relatively low toxicity. Thus, these outcomes suggested further studies about alternative economically viable for control and prevention of infections in veterinary medicine.