Aims To explore antibacterial activities of coagulase-negative staphylococci (CoNS) from teat apices of dairy cows towards mastitis-causing pathogens. Methods and Results Of 254 CoNS, 38 displayed bacteriocin-like activity after a first screening. Seven of these strains displayed activity against at least one mastitis-related pathogen (Streptococcus uberis, Streptococcus dysgalactiae and Staphylococcus aureus). Staphylococcus chromogenes L217 displayed the strongest inhibitory effect, being active against all tested mastitis-related pathogens and most tested CoNS. Based on cation exchange and reversed-phase chromatography, in addition to N-terminal Edman degradation and PCR, the antibacterial peptide was identified as a nukacin-type bacteriocin and named nukacin L217. Although staphylococcal bacteriocins are generally found in the cell-free supernatants of liquid cultures, Staph.chromogenes L217 only led to detectable activity when grown on agar medium. Conclusions Bacteriocin-like activities are not uncommon among CoNS from teat apices and may inhibit mastitis-causing pathogens, as found for nukacin L217 production by Staph.chromogenes L217. Significance and Impact of the Study Nukacin L217 is the first identified bacteriocin of the species Staph.chromogenes and displays unusual production kinetics, that is, requiring surface growth of its producer. The fact that nukacins are produced by different CoNS species suggests a role in the teat skin ecosystem.