Anthropogenic effects including river regulation, watershed development, contamination, and fish introductions have substantially affected the majority of freshwater habitats in Europe and North America. This pattern of resource development and degradation is widespread in the tropics, and often little is known about the resources before they are lost. This article describes the freshwater resources of Puerto Rico and identifies factors that threaten conservation of native fishes. The fishes found in freshwater habitats of Puerto Rico represent a moderately diverse assemblage composed of 14 orders, 29 families, and 82 species. There are fewer than 10 species of native peripherally-freshwater fish that require a link to marine systems. Introductions of nonindigenous species have greatly expanded fish diversity in freshwater systems, and native estuarine and marine species (18 families) also commonly enter lowland rivers and brackish lagoons. Environmental alterations, including land use and development, stream channelization, pollution, and the impoundment of rivers, combined with nonnative species introductions threaten the health and sustainability of aquatic resources in Puerto Rico. Six principal areas for attention that are important influences on the current and future status of the freshwater fish resources of Puerto Rico are identified and discussed.