Bacterial colony enumeration is an essential tool for many widely used biomedical assays. This chapter introduces a cost-effective and fully automatic bacterial colony counter which accepts digital images as its input. The proposed counter can handle variously shaped dishes/plates, recognize chromatic and achromatic images, and process both color and clear medium. In particular, the counter can detect dish/ plate regions, identify colonies, separate aggregated colonies, and finally report consistent and accurate counting result. The authors hope that understanding the complicated and labor-intensive nature of colony counting will assist researchers in a better understanding of the problems posed and the need to automate this process from a software point of view, without relying too much on specific hardware. © 2009, IGI Global.