Landscape genetics has seen tremendous advances since its introduction, but parameterization and optimization of resistance surfaces still poses significant challenges. Despite increased availability and resolution of spatial data, few studies have integrated empirical data to directly represent ecological processes as genetic resistance surfaces. In our study, we determine the landscape and ecological factors affecting gene flow in the western slimy salamander (Plethodon albagula). We used field data to derive resistance surfaces representing salamander abundance and rate of water loss through combinations of canopy cover, topographic wetness, topographic position, solar exposure and distance from ravine. These ecologically explicit composite surfaces directly represent an ecological process or physiological limitation of our organism. Using generalized linear mixed-effects models, we optimized resistance surfaces using a nonlinear optimization algorithm to minimize model AIC. We found clear support for the resistance surface representing the rate of water loss experienced by adult salamanders in the summer. Resistance was lowest at intermediate levels of water loss and higher when the rate of water loss was predicted to be low or high. This pattern may arise from the compensatory movement behaviour of salamanders through suboptimal habitat, but also reflects the physiological limitations of salamanders and their sensitivity to extreme environmental conditions. Our study demonstrates that composite representations of ecologically explicit processes can provide novel insight and can better explain genetic differentiation than ecologically implicit landscape resistance surfaces. Additionally, our study underscores the fact that spatial estimates of habitat suitability or abundance may not serve as adequate proxies for describing gene flow, as predicted abundance was a poor predictor of genetic differentiation.