Abstract A total of 207 specimens were collected from different sources including patients, health care staff and hospital environment in Ibb city, Yemen. The study used the bacteriocin produced from active producer strains in typing of Staphylococcus aureus. Depending on the morphological, cultural and biochemical characteristics, 54 (26.09%) isolates of Staphylococcus aureus were identified. An antibiotic sensitivity test was done for the bacterial isolates, and the results showed that there were multiple resistant antibiotics. The Staphylococcin production of these isolates has been detected by using wells assay. Fifty one isolates were Staphylococcin producer. Four isolates (staph19, staph25, staph28 and staph43) were chosen as good Staphylococcin producers, and used locally as indicators in bacteriocin typing. Depending on S. aureus typing, the isolates fell into (9) groups. The most numerous group was characterized by susceptibility to all four staphylococcin and comprised 61.11% isolates of S. aureus, while the lowest numerous were found in three groups with a ratio of 1.85%; the remaining groups had little percentages ranging from 3.70% to 11.11%. We observed that about (94.44%) of the isolates were bacteriocin producers, and among them, four isolates had a strong bacteriocin production. Based on typing, most isolates had one pattern.