Background: Buruli ulcer is a chronic, indolent, necrotizing infectious disease of the skin and soft tissues characterized by the formation of large ulcers, often in the arms or legs. Nigeria is a Buruli ulcer disease (BUD)endemic country with its control programme still in infancy. As a result, samples are sent to laboratories outside the country. Some patients go to neighbouring countries with more established programmes for polymerase chain reaction (PCR) as a basis of diagnosis and treatment. Hence, this study was embarked upon to assist the national control programme in overcoming the PCR diagnosis test challenge. Method: This was a cross-sectional and community-based type of study of Buruli ulcer patients from 15 states,mostly from southern Nigeria; Abia, Akwa Ibom, Anambra, Bayelsa, Cross Rivers, Delta, Ebonyi, Ekiti, Enugu, Imo, Lagos, Ogun, Ondo, Osun, Rivers and the Federal Capital Territory (FCT) Abuja. Swab and Fine Needle Aspiration (FNA) samples were received from January 2016 to June 2018. DNA was extracted, and each sample was subjected to IS2404-based nested PCR. Results: Out of 920 samples received, 427 (46.4% ) were IS2404-based nested PCR positive. Of which 204were males, and 223 were females. The patients’ mean age was 35.7 ± 19.94, with 172 (18.7%) being children ≤15 years, while 748 (81.3%) were ≥ 15 years. During the study period, the highest number of samples, 171(18.6%) and 170 (18.5%) were received from Cross Rivers and Delta states, respectively. In contrast, the least number of samples, 2 (0.2%) and 3 (0.3%), came from Ekiti and Lagos states, respectively. Conclusion: This is the first in-country PCR confirmed diagnosis of a large cohort of BU patients. The results show a high prevalence in southern Nigeria, which is an indicator of high transmission. Our findings suggest the need for a prompt intervention by the government by providing the needed health facilities and education for the communities.