Although shale drilling operations for oil and natural gas have increased greatly in the past decade, few studies directly quantify the impacts of shale development on plants and wildlife. We evaluate knowledge gaps related to shale development and prioritize research needs using a quantitative framework that includes spatial and temporal extent, mitigation difficulty, and current level of understanding. Identified threats to biota from shale development include: surface and groundwater contamination; diminished stream flow; stream siltation; habitat loss and fragmentation; localized air, noise, and light pollution; climate change; and cumulative impacts. We find the highest research priorities to be probabilistic threats (underground chemical migration; contaminant release during storage, during disposal, or from accidents; and cumulative impacts), the study of which will require major scientific coordination among researchers, industry, and government decision makers. Taken together, our research prioritization outlines a way forward to better understand how energy development affects the natural world.