The frequency of, risk factors for, and outcome of polyclonal gram-negative bacteremia are still unknown. We investigated them in a prospective cohort study of patients for whom a blood culture yielded >/=1 species of gram-negative aerobic rod. For each patient, pulsed field gel electrophoresis (PFGE) was performed on 4 colonies of each morphologic type. Episodes of bacteremia were considered polyclonal if caused by >1 PFGE type of the same species. Ten (6.5%) of 153 investigated patients had polyclonal bacteremia. Bacteremia due to nonfermenting rods was the single significant risk factor for polyclonal bacteremia. Complications were equally frequent in all patient groups. However, patients with polyclonal bacteremia received more extensive antibiotic therapy than did patients with monoclonal bacteremia. Nearly 20% of episodes of bacteremia due to nonfermenting rods were polyclonal, but it remains unclear why nonfermenting rods were more likely to cause polyclonal bacteremia than were other gram-negative rods.