Roads built through or near wetlands cause significant mortality of reptiles and amphibians and create barriers to migration and dispersal. I investigated the number of times turtles and other herpetofauna attempted to cross a 4-lane highway at Lake Jackson, Florida, USA, during a period of severe drought (Feb–Apr 2000). Levels of road mortality were so high that I designed and installed a temporary drift fence system to work with an existing drainage culvert and for the next 2.5 years I evaluated its effectiveness at reducing road mortality and facilitating migration. I monitored roads and fences several times per day for 44 months, during both drought and non-drought conditions. A total of 10,229 reptiles and amphibians of 44 species were found either road killed or alive behind drift fences: 8,842 turtles, 838 frogs, 363 snakes, 152 lizards, 32 alligators, and 2 salamanders. Drift fences combined with intensive monitoring greatly reduced turtle road kills and facilitated the use of an under-highway culvert. Along a 0.7-km section of the highway, turtle mortality before installation of the fence (11.9/km/day) was significantly greater than post-fence mortality (0.09/km/day) and only 84 of 8,475 turtles climbed or penetrated the drift fences. Pre-fence data provided strong evidence that turtles cannot successfully cross all 4 lanes of U.S. Highway 27, as 95% of 343 turtles were killed as they first entered the highway adjacent to the shoulder and the remaining 5% were killed in the first two traffic lanes. According to a probability model, the likelihood of a turtle successfully crossing U.S. Highway 27 decreased from 32% in 1977 to only 2% in 2001 due to a 162% increase in traffic volume. Therefore, at least 98% of turtles diverted by the fences probably would have been killed if fences were not in place. The results of this study represent the highest attempted road-crossing rate ever published for turtles (1,263/km/year). Because of demographic and life history constraints, turtle populations may incur irreversible declines in areas where road mortality is high, especially when mass migrations are triggered by periods of drought.