To evaluate the kinetics of serum and urinary hepcidin levels along with anemia-related parameters during the infection course of infants with febrile urinary tract infection (UTI), we enrolled febrile infants aged one to four months in this prospective study. Febrile patients with UTI were allocated intoEscherichia coli(E. coli) or non-E. coligroups according to urine culture results. Septic workup, blood hepcidin, iron profile, urinalysis, and urinary hepcidin–creatinine ratio were collected upon admission and 3 days after antibiotic treatment. In total, 118 infants were included. On admission, the febrile UTI group showed a significant reduction in serum iron level and a significant elevation of urinary hepcidin–creatinine ratio compared to the febrile control counterpart. Moreover, urinary hepcidin–creatinine ratio had the highest odds ratio, 2.01, in logistics regression analysis. After 3 days of antibiotic treatment, hemoglobin and the urinary hepcidin–creatinine ratio were significantly decreased. Patients with anE. coliUTI had a significantly decreased urinary hepcidin–creatinine ratio after 3 days of antibiotics treatment, whereas the non-E. coligroup showed insignificant changes. Our study suggested that the urinary hepcidin–creatinine ratio elevated during acute febrile urinary tract infection and significantly decreased after 3 days of antibiotics treatment, especially inE. coliUTI.