In an effort to uncover the role of the high affinity Zn(II) uptake system in uropathogenic Escherichia coli CFT073, we deleted the znuB gene, which encodes for the transmembrane component of the ZnuABC transporter system. The null mutant for znuB did not grow on minimal medium unless supplemented with excess Zn(II) (50 µM ZnCl2). In contrast, the E. coli K-12 ΔznuB cell line grew well on minimal medium that was not supplemented with Zn(II). The ΔznuB mutant was significantly deficient in formation of biofilm under static conditions and also showed a substantially-reduced migration front of swarm cells. Because motility and biofilm formation are important for E. coli CFT073 pathogenicity, we propose that the high affinity Zn(II) uptake system may contribute to the virulence of this pathogen in the urinary tract.