As the human population increases, the landscape is altered to provide housing, food, and industry. Human activity poses a risk to the health of natural habitats that, in turn, affect biodiversity. Biodiversity is necessary for a functioning ecosystem, as species work synergistically to create a livable environment. It is, therefore, important to know how human practices and natural events threaten these habitats and the species living in them. A universal method of modeling habitat threats does not exist. This paper details the use of a literature review to formulate a new framework called Define–Investigate–Estimate–Map (DIEM). This framework is a process of defining threats, investigating an area to discover what threats are present, estimating the severity of those threats, and mapping the threats. Analysis of 62 studies was conducted to determine how different authors define and characterize threats in various contexts. The results of this analysis were then applied to a case study to evaluate the Choctawhatchee River and Bay Watershed. Results suggest that the most abundant threat in the watershed is agricultural development, and the most destructive threat is urban development. These two threats have the greatest impact on the total threat level of the watershed. Applying the DIEM framework demonstrates its helpfulness in regional analysis, watershed modeling, and land development planning.