The structure of complex biological systems reflects not only their function but also the environments in which they evolved and are adapted to. Reverse Ecology-an emerging new frontier in Evolutionary Systems Biology-aims to extract this information and to obtain novel insights into an organism's ecology. The Reverse Ecology framework facilitates the translation of high-throughput genomic data into large-scale ecological data, and has the potential to transform ecology into a high-throughput field. In this chapter, we describe some of the pioneering work in Reverse Ecology, demonstrating how system-level analysis of complex biological networks can be used to predict the natural habitats of poorly characterized microbial species, their interactions with other species, and universal patterns governing the adaptation of organisms to their environments. We further present several studies that applied Reverse Ecology to elucidate various aspects of microbial ecology, and lay out exciting future directions and potential future applications in biotechnology, biomedicine, and ecological engineering.