The abundance and distribution of New England cottontails (NEC; Sylvilagus transitionalis) have been declining for several decades. Remnant populations in some regions are known to be vulnerable to extirpation but little is known about the status of populations in most areas. We conducted a survey of the historic range (ca. 1960) of NEC to determine the current distribution and relative status of extant populations. Because NEC were sympatric with eastern cottontails (S. floridanus) and snowshoe hares (Lepus americanus) in much of their historic range, identity of resident lagomorphs was based on DNA extracted either from tissue of captured cottontails or from fecal pellets of free-ranging lagomorphs. We searched 2,301 patches of suitable habitat and detected NEC in 162. We identified 5 disjunct populations in approximately 14% of the historic range. Forest maturation and fragmentation are the most plausible explanations for the widespread decline of NEC. Contraction of the historic distribution was toward eastern and southern edges where a variety of anthropogenic disturbances (e.g., brushy edges of highways and railroad corridors and idle portions of agricultural fields) provided habitat. Land-ownership patterns (dominated by small acreages) and land-use activities (expanding development and limited forest management) within the currently occupied range of NEC suggest a continued decline of suitable habitats. As a result, we recommend efforts to enhance remaining populations of NEC that include responses at 2 spatial scales. At the population or landscape scale, current land uses should guide habitat manipulations that expand existing populations. At the regional scale, we recommend consideration should be given to increasing dispersal among remnant populations, possibly by generating “stepping stones” of suitable habitat. In addition to improving long-term viability of NEC, other species of conservation concern that are dependent on early successional habitats will benefit from these efforts.