Objective To establish a simple and reliable experimental rodent model sensitive to coxsackievirus A16 (CVA16).Methods Mongolian gerbils with different age were selected and inoculated intraperitoneally with live CVA16, and the gerbils were observed daily until 14 days postinoculation to screen for the most optimal ages sensitive to the virus. The dose-dependent symptoms were evaluated and the 50% lethal dose (LD50) was determined. The virus titers were measured in blood and various tissues of CVA16-infected Mongolian gerbils 3 days post-infecton. Finally, the gerbils were immunized twice with inactivated CVA16 vaccine at day 1 and day 11, respectively, followed by challenge with the virus with a dose of LD50 at day 14. The gerbils were then observed for another 2 weeks to record their body weight, symptom and mortality rate. Their blood samples were collected from the eyes, and CVA16-specific neutralizing antibody titers and total antibody titers was checked by microneutralization test and ELISA, respectively. Results Various clinical symptoms, such as inactivity, hind limb weakness, paralysis and even death occurred in gerbils following CAV16 infection. 7-day-old and 14-day-old gerbils are susceptible to CVA16 infection whereas 28-day-old gerbils are resistant. The most sensitive and appropriate age is 14-day-old. The 50% lethal dose was determined to be 1×104.5 CCID50. High titers of the virus were confirmed in blood and various tissues of Mongolian gerbils contracted CAV16 3 days post-infecton. The survival rate is 87.5% for 14-day-old gerbils preimmunized with two doses of inactivated CVA16 vaccine and challenged with the virus. The geometric mean titers (GMTs) of neutralizing antibody was 28.14, and the seroprevalence was 87.5%. Conclusions Mongolian gerbils is sensitive to CVA16 and the virus reproduces actively in Vivo. Thus, it can be used as a reliable small animal model for studies of CVA16 pathogenesis, vaccine development and drug evaluation.