Agricultural expansion and intensification are major threats to tropical biodiversity. In addition to the direct removal of native vegetation, agricultural expansion often elicits other human-induced disturbances, many of which are poorly addressed by existing environmental legislation and conservation programmes. This is particularly true for tropical freshwater systems, where there is considerable uncertainty about whether a legislative focus on protecting riparian vegetation is sufficient to conserve stream fauna. To assess the extent to which stream fish are being effectively conserved in agricultural landscapes, we examined the spatial distribution of assemblages in river basins to identify the relative importance of human impacts at instream, riparian and catchment scales, in shaping observed patterns. We used an extensive dataset on the ecological condition of 83 low-order streams distributed in three river basins in the eastern Brazilian Amazon. We collected and identified 24,420 individual fish from 134 species. Multiplicative diversity partitioning revealed high levels of compositional dissimilarity (DS) among stream sites (DS = 0.74 to 0.83) and river basins (DS = 0.82), due mainly to turnover (77.8% to 81.8%) rather than nestedness. The highly heterogeneous fish faunas in small Amazonian streams underscore the vital importance of enacting measures to protect forests on private lands outside of public protected areas. Instream habitat features explained more variability in fish assemblages (15%-19%) than riparian (2%-12%), catchment (4%-13%) or natural covariates (4%-11%). Although grouping species into functional guilds allowed us to explain up to 31% of their abundance (i.e. for nektonic herbivores), individual riparian - and catchment - scale predictor variables that are commonly a focus of environmental legislation explained very little of the observed variation (partial R2 values mostly <5%). Policy implications. Current rates of agricultural intensification and mechanization in tropical landscapes are unprecedented, yet the existing legislative frameworks focusing on protecting riparian vegetation seem insufficient to conserve stream environments and their fish assemblages. To safeguard the species-rich freshwater biota of small Amazonian streams, conservation actions must shift towards managing whole basins and drainage networks, as well as agricultural practices in already-cleared land. © 2017 British Ecological Society.