The ability of microbes to grow as surface-associated communities, or biofilms, is seen as a lifestyle widely spread in all biomes on planet earth, including extremely acidic ecosystems. Within these harsh and constantly changing habitats, the biofilm mode of growth is thought to offer the microbial community a regular condition so as to cope with the adverse scenery. Though acidophilic biofilm communities are mainly dominated by bacterial species, archaeal and eukaryotic populations appear to play crucial roles, broadening metabolic diversity and maintaining the community structure. Environmental signals and molecular mechanisms that underlay the biofilm lifestyle have been largely characterized for bacteria that colonize human cavities. In contrast, the genetic basis that enable environmental acidophiles to form and develop biofilms are far from comprehended. Nonetheless, the molecular mechanisms that govern the biofilm developmental processes in a few model acidophiles have begun to be elucidated. This chapter describes what is currently known on the cellular and molecular mechanisms that promote biofilm formation of bacteria and archaea inhabiting extremely acidic ecosystems.