The dataset includes 151 clinical isolates from different sources, collected from the Microbiology Laboratory of the First Hospital of Jilin University, covering 19 clinically significant bacterial species. Data collection was conducted under controlled conditions using professional imaging equipment, including a closed background, stable top and bottom light sources, and consistent imaging angles, following standardized procedures. The final dataset consists of 950 original colony images, with 59,221 colony targets manually annotated. Data augmentation was performed through random horizontal mirroring or vertical flipping, expanding the dataset to 1,900 images, resulting in a total of 118,442 colony targets.