Abstract: In order to understand the pathogen of the clinical mastitis in dairy cows in a dairy farm and the in vitro bactericidal effects of acid electrolytic water on the pathogen of mastitis in dairy cows, in this experiment, milk samples were collected from cows with clinical mastitis in a dairy farm, and the pathogenic bacteria were isolated from the milk samples; the species of the pathogenic bacteria were identified by Gram staining, microscopic examination, biochemical test, and 16S rDNA gene sequence analysis. The strain was also subjected to drug sensitivity test and animal pathogenicity test, and the bactericidal effect of acid electrolytic water on the strain was analyzed by in vitro bactericidal test. The results showed that the strain grew well on Baird-Parker（BP） medium, and the surface of the colonies was smooth, gray-black and opaque without hyaline ring after 24 h of incubation. Gram staining and microscopic examination showed that the strain was bluish-purple in color and arranged in the shape of grape bunches, and the strain was named QSPT2023. The biochemical characteristics of isolate QSPT2023 were basically consistent with those of Staphylococcus spp. The 16S rDNA gene sequence showed the highest similarity of 99.93% with that of Staphylococcus chromogenes OP068076.1 in GenBank. It was located in the same branch of the phylogenetic tree with Staphylococcus chromogenes OP068076.1 and had the closest affinity; it was closer to Staphylococcus chromogenes KJ783397.1, LC317309.1 and ON715882.1, and was more distant from Staphylococcus aureus, and Staphylococcus saprophyticus, Staphylococcus epidermidis. It was sensitive to chloramphenicol, cefotaxime, cefadroxil, ceftazidime, cefazolin, kanamycin, minocycline, oxfloxacin, doxycycline, florfenicol, and cotrimoxazole; and was resistant to daunorubicin, enrofloxacin, roxithromycin, ampicillin, amoxicillin, amphotericin B, and sulfadiazine. After the mice were injected intraperitoneally with 0.2 mL of bacterial solution of the isolate strain for 5 h, the symptoms such as tightly closed eyes, ooze around the eyes, and chilling and cuddling appeared; there were dead mice within 3 d, and the dead mice had obvious pathological characteristics after autopsy. The bactericidal rates of Staphylococcus chromogenes isolate treated with acid electrolytic water were 99.21% after 5 s, 99.99% after 10 s, and 100% after 15 s. The bactericidal effect was better than that of the traditional disinfectant 5% povidone-iodine. The results suggested that a strain of Staphylococcus chromogenes was successfully isolated from the milk samples of cows suffering from mastitis in a dairy farm, and that acid electrolytic water had a good bactericidal effect on this isolated strain.