The antibacterial activity of Korean honey bee venom (KBV) was examined against seven major bacterial mastitis pathogens, Enterococcus faecium, Escherichia coli, Methicillin resistant Staphylococcus aureus (MRSA), Staphylococcus aureus, Streptococcus intermedius, Streptococcus oralis and Streptococcus uberis isolated from infected mammary quarters of cows. Seven bacterial mastitis pathogens were studied for antibacterial activity of the KBV by disc diffusion assay, minimal inhibitory concentrations (MIC) and bacterial count in milk samples. The KBV showed activity against Ent. faecium, E. coli, MRSA, Staph. aureus, Strep. intermedius. The order of susceptibility of the bacteria against the KBV was Staph. aureus>MRSA>E. coli>Strep. intermedius>Ent. faecium>Strep. oralis>Strep. uberis. The MIC against Staph. aureus, MRSA and E. coli were stronger effect as compared with standard drug. The effect of the KBV ($100{mu}g;ml^{-1}$) on the viability of Ent. faecium, E. coli, MRSA, Staph. aureus, Strep. intermedius, Strep. oralis and Strep. uberis in milk differed significantly with each other within 12 h incubation period. The results indicate that KBV has significant antibacterial effects against major bacterial mastitis bacteria, Ent. faecium, E.coli, MRSA, Staph. aureus, Strep. intermedius. Results of the study indicate the potential use of KBV as alternative to antibiotic therapy. Further investigations are needed though to confirm its efficacy and its effects on the animals.