BACKGROUND: Intensive agriculture can disrupt adjacent stream ecosystems by increasing nutrient runoff, suspended and dissolved organic matter, and pesticide loads. Freshwater biofilms are surface-attached microbial communities that host complex interaction networks and play a key role in nutrient cycling and bioremediation. If adjacent land use drastically shifts microbial community composition and assembly, the functional resilience and adaptive capacity of biofilms under changing conditions may be impaired. In this study, we compared developing and mature biofilm samples from three sites along the Otterbach and two sites along the Perlenbach stream (Bavaria, Germany). The Otterbach sites, located in an area with agriculture and a nearby village, were adjacent to an intensively managed cropland, an extensively managed grassland, and a forest, while the Perlenbach flows through an area free of cropping and fertilization, with sites adjacent to an extensively managed grassland and a forest. Bacterial community composition was assessed through 16S rRNA gene amplicon sequencing. Bacterial diversity, differential abundance, community assembly and co-occurrence network analyses were performed. RESULTS: Adjacent intensive land use increased bacterial alpha diversity in both developing and mature biofilms. Moreover, adjacent intensive and extensive land use shaped bacterial community composition and increased the relevance of deterministic processes in bacterial community assembly, especially in developing biofilms, increasing the presence of key responding taxa such as Arenimonas, Blastocatella, Gemmatimonas, Flectobacillus, Leptothrix, Flavobacterium or Rhodoferax. These taxa were also detected in the co-occurrence networks of agriculturally influenced sites, displaying strong connectivity and centrality. These effects were limited to the Otterbach stream, which exhibited higher overall nutrient concentrations. CONCLUSIONS: Agricultural land use strongly influenced bacterial richness, composition, and assembly in biofilms from adjacent stream ecosystems, particularly in developing biofilms from the most anthropogenically impacted stream, driven by the proliferation of key responding taxa. This showcased how anthropogenic nutrient inputs can redirect biofilm development pathways and potentially alter the ecological role of biofilms in stream ecosystems.