Objectives: Several occupations in developing countries lag behind in ensuring the safety of their workers in occupational settings. Lack of implementation of safety guidelines at workplaces can expose workers to health risks. In Pakistan, barbers are one of the un-regulated occupational groups. Low literacy, increased frequency of direct skin contact and blade/razors use can expose barbers to body fluids including blood of the customers. We conducted this study in order to determine hepatitis B virus (HBV) prevalence among barbers and their knowledge, attitude and practices in a peri-urban district of Sindh. Material and Methods: Three hundred eighty-five barbers from the Sukkur district were interviewed using a structured questionnaire. Blood samples were collected and tested for HBsAg. A scale was built to determine the proportions of responses to knowledge, attitude and practice items. Results: The prevalence of HBV among barbers was 2.1%. The barbers’ knowledge on HBV and its transmission routes was poor. The response to attitude items was good, except that only 35.1% of the participants agreed to have vaccination against HBV. The overall performance on the knowledge and practice scales was poor compared to the attitude scale on which 80% of the barbers performed well. Conclusion: The prevalence of HBV among barbers was lower compared to the available national figures for the prevalence among the general population.