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Roads impact a disproportionately larger area than the land area that they occupy, often called the road effect zone. This effect extends to wildlife populations. Impacts on federally listed species include habitat loss and fragmentation, road mortality, the barrier effect, and the habituation effect. Per the Endangered Species Act, these impacts must be assessed before roadway impacts occur. This publication provides an overview of the potential impacts to 29 of Florida’s threatened and endangered animal species from roads. Potential mitigation options include road configuration, signage, speed limits, habitat protection, improved technology, wildlife fencing, and wildlife crossing structures. The goal is to educate transportation engineers and planners to allow for sustainable transportation infrastructure growth while protecting, conserving, and ultimately recovering Florida’s threatened and endangered species.