The clearing and fragmentation of terrestrial ecosystems is commonly acknowledged as a major cause of the decline of biodiversity. These and other predicted responses to habitat fragmentation are derived from theory, which ecologists have tested with empirical approaches ranging from observations to experimental manipulations. These empirical approaches have also identified areas of theory in need of additional development. For example, experimental studies of fragmentation have provided insights such as the key role played by the matrix surrounding fragments, the importance of edge effects, and the impacts of corridors linking fragments with one another. Much less clear, however, is the extent to which these theoretical and empirical studies – while advancing our conceptual understanding of ecological responses to landscape change – help guide management and conservation practice. We review lessons learned from landscape-scale fragmentation experiments and observational studies, present the results of a survey of fragmentation and conservation experts which probed for links and mismatches between fragmentation studies and conservation practice, and discuss how future studies can contribute to conservation practice. Our survey showed that respondents consider fragmentation theory and empirical studies and their findings important for guiding conservation and management practices. The survey also identified that there are disconnects between what is typically studied by fragmentation ecologists and factors that are central to the practice of biodiversity conservation, notably, community-based human dimensions (e.g. economic, social, health issues), policy and governance, ecosystem services, eco-evolutionary responses of species, and interaction of multiple threats to biodiversity and ecosystem processes. We discuss how these disconnects can present opportunities for experiments to continue to provide valuable recommendations for conservation practice in fragmented landscapes.