The long-term survival of three human enterovirus serotypes, coxsackievirus B3, echovirus 7, and poliovirus 1 was examined in samples of surface freshwater collected from five sites of physically different character. These were an artificial lake created by damming a creek, a small groundwater outlet pond, both a large and a medium sized river, and a small suburban creek. Survival was studied at temperatures of -20, +01 and +23 celsius. The average amount of viral inactivation was 6.5 to 7.0 log base 10 units over 8 weeks at +23C, 4 to 5 log base 10 units over 12 weeks at +01C, and 0.4 to 0.8 log base 10 units over 12 weeks at -20C. The effect of incubation temperature upon viral inactivation rate was statistically significant (P <0.00001). As determined by pairing tests, survival was also significantly related to both viral serotype and water source at each of the three incubation temperatures (P < or = 0.05). Efforts were made to determine whether the rate of viral inactivation observed at the different incubation temperatures was related to characteristics inherent to the water that was collected from the different locations.