To control the waterborne transmission of enterovirus 71(EV71), which is associated with hand foot and mouth disease (HFMD), it is essential to know the inactivation effectiveness of disinfectants on EV71 in water. In this article, we present a comparative analysis of the effects on EV71 following exposure to chlorine dioxide (ClO2) under different doses, pH, and temperature conditions. We show that the EV71 exhibited strong resistance to ClO2 (more than the MS2 standard) and that Ct value ranges required for a 4-log reduction of EV71 in buffered, disinfectant demand-free water at pH 7.2 and 20 °C by ClO2 were 4.24-6.62 mg/L·min according to the efficiency factor Hom model. ClO2 inactivation of the virus was temperature- and pH-dependent. The virucidal efficiency was higher at pH 8.2 than at pH 5.6 and pH 7.2 and higher at 36 °C than at 4 and 20 °C. In addition, we also observed the impact of ClO2 on the entire viral genome using RT-PCR, which indicated that the 5' noncoding region (5'-NCR) within the EV71 genome, specifically the 1-118 nt region, was the most easily damaged by ClO2 and correlated with viral infectivity. Our study has not only provided guidelines for EV71 disinfection strategies of waste and drinking water, but also confirmed the importance of the 5'-NCR for EV71 infectivity and may demonstrate a general inactivation by ClO2 of enteric virus by damaging the 5'-NCR. Furthermore, 5'-NCR can be used as a target region for PCR to investigate infectious virus contamination in environmental water and evaluate the inactivation effects of ClO2.