Abstract Numerous freshwater species are consumed or exploited through artisanal fisheries in the rivers of the islands of Guadeloupe, French West Indies. Autochthonous production of organic matter is limited in these flowing rivers and is mainly represented by scarce filamentous green algae and an abundant epilithic biofilm growing on wet stones in the river bed. Stable isotope analysis was used to quantify the relative importance of biofilm and other riverine allochthonous and autochthonous food sources in the diet of tropical shrimps (Palaemonidae, Atyidae and Xiphocarididae) and fishes (Gobiidae and Eleotridae) consumed by the local people. The epilithic biofilm was exploited by most species, constituted an important source of autochthonous carbon and was an important source of organic matter production at the base of freshwater food webs in Caribbean rivers. Biofilm percentages in the diet reached 32% for molluscs, 85% for atyid shrimps, 29% for xiphocaridid shrimps, 14% for palaemonid shrimps and 13% for fish. Assessment of biofilm in nine rivers showed that blue and red cyanobacteria were quantitatively dominant with a moderately rich diatom flora. These results address the interactions between river biofilm and Caribbean freshwater fauna where trophic links between consumers and their potential resources have poorly been documented.