With human activities increasingly impacting natural resources in relatively remote locations, there is a need for simple and efficient methods to explore the ecological consequences of these activities. Little is understood about the influences of off-highway vehicle (OHV) use on wildlife populations. We examined the effect of OHV activity on developmental instability in a phrynosomatid lizard (i.e., western fence lizard [Sceloporus occidentalis]) in the western Great Basin, USA. We measured fluctuating asymmetry (FA) of bilateral head-scale patterns in populations of lizards at 3 OHV and 3 non–OHV sites. Fluctuating asymmetry was higher at OHV sites relative to non-OHV sites, supporting the idea that OHV activity can stress wildlife populations. We found FA to be a good tool for uncovering responses to stress in natural populations, and we recommend exploring FA as a means of uncovering developmental instability in other systems that merit conservation interest