The rise of multidrug-resistant bacterial infections necessitates the discovery of novel antimicrobial strategies. Here, we show that protein design provides a generalizable means of generating new antimicrobials by neutralizing the function of bacterial adhesins, which are virulence factors critical in host-pathogen interactions. We designed high-affinity miniprotein binders to FimH and Abp1D/Abp2D chaperone usher pili adhesins from uropathogenic and , respectively, which are implicated in mediating both uncomplicated and catheter-associated urinary tract infections (UTI) responsible for significant morbidity and mortality worldwide. The designed antagonists have high specificity and stability, disrupt bacterial recognition of host receptors, block biofilm formation, and are effective in treating and preventing uncomplicated and catheter-associated UTIs . Targeting virulence factors outside the cell membrane with protein design provides a rapid route to next-generation therapeutics that can disrupt pathogenesis and thus are capable of treating and preventing disease in an antibiotic-sparing manner.