The molecular evolution of antibiotic resistance in clinical isolates of Escherichia coli has been studied. The isolates came from a patient who was on long-term antibiotic therapy for multiple infected liver cysts. Over a period of two years seven E. coli strains were isolated from his blood during periods of septicaemia and directly from liver cysts during exploratory surgery. The liver cysts have provided a protected environmental from the antibiotic therapy the patient was receiving. The low antibiotic concentrations inside the liver cysts have allowed the bacteria to develop multiple antibiotic resistance mechanisms over time. Pulsed field gel electrophoresis patterns show that the seven isolates are closely related and are not a series of successive competitive infections. In addition, each isolate has the same basic ompC sequence that is distinct from other E. coli isolates, which suggests that they derive from the same founder population. However, the isolates differ in many characteristics including their MIC values for a range of antibiotics, auxotrophic markers, -lactamase activities, mutations in the ampC promoter sequence, mutations in gyrA and parC genes and their outer membrane permeability. The data provide strong evidence for a single focal infection expanding via parallel pathways of evolution to give a range of antibiotic resistant isolates. It is probable that the infected cysts are providing numerous protected environments that are the foci for the separate development of distinct variants.