Autophagy protects against many infections by inducing the lysosomal-mediated degradation of invading pathogens. However, previous in vitro studies suggest that some enteroviruses not only evade these protective effects but also exploit autophagy to facilitate their replication. We generated Atg5^f/f/Cre⁺ mice, in which the essential autophagy gene Atg5 is specifically deleted in pancreatic acinar cells, and show that coxsackievirus B3 (CVB3) requires autophagy for optimal infection and pathogenesis. Compared to Cre⁻ littermates, Atg5^f/f/Cre⁺ mice had an ∼2,000-fold lower CVB3 titer in the pancreas, and pancreatic pathology was greatly diminished. Both in vivo and in vitro, Atg5^f/f/Cre⁺ acinar cells had reduced intracellular viral RNA and proteins. Furthermore, intracellular structural elements induced upon CVB3 infection, such as compound membrane vesicles and highly geometric paracrystalline arrays, which may represent viral replication platforms, were infrequently observed in infected Atg5^f/f/Cre⁺ cells. Thus, CVB3-induced subversion of autophagy not only benefits the virus but also exacerbates pancreatic pathology.