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This review is focused on the biogeochemistry of arsenic in freshwaters and, especially, on the key role that benthic microalgae and prokaryotic communities from biofilms play together in through speciation, distribution, and cycling. These microorganisms incorporate the dominant iAs (inorganic arsenic) form and may transform it to other arsenic forms through metabolic or detoxifying processes. These transformations have a big impact on the environmental behavior of arsenic because different chemical forms exhibit differences in mobility and toxicity. Moreover, exposure to toxicants may alter the physiology and structure of biofilms, leading to changes in ecosystem function and trophic relations. In this review we also explain how microorganisms (i.e., biofilms) can influence the effects of arsenic exposure on other key constituents of aquatic ecosystems such as fish. At the end, we present two real cases of fluvial systems with different origins of arsenic exposure (natural vs. anthropogenic) that have improved our comprehension of arsenic biogeochemistry and toxicity in freshwaters, the Pampean streams (Argentina) and the Anllóns River (Galicia, Spain). We finish with a briefly discussion of what we consider as future research needs on this topic. This work especially contributes to the general understanding of biofilms influencing arsenic biogeochemistry and highlights the strong impact of nutrient availability on arsenic toxicity for freshwater (micro) organisms.