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A 15-week experiment was performed in a riverside laboratory flume (with diverted river water) to check variations of river biofilm structure (biomass, algal and bacterial compositions) and function (community gross primary production GPP and respiration) under constant flow while water quality went through natural temporal variations. One major suspended matter pulse coinciding with one river flood was recorded after 10 weeks of experiment. Epilithic biofilm first exhibited a 10-week typical pattern of biomass accrual reaching 33 g ash-free dry matter (AFDM) m–2 and 487 mg chlorophyll-a m–2 and then, experienced a shift to dominance of loss processes (loss of 60% AFDM and 80% chlorophyll-a) coinciding with the main suspended matter pulse. Algal diversity remained low and constant during the experiment: Fragilaria capucina and Encyonema minutum always contribute over 80% of cell counts. DGGE banding patterns discriminated between two groups that corresponded to samples before and after biomass loss, indicating major changes in the bacterial community composition. GPP/R remained high during the experiment, suggesting that photoautotrophic metabolism prevailed and detachment was not autogenic (i.e., due to algal senescence or driven by heterotrophic processes within the biofilm). Observational results suggested that silt deposition into the biofilm matrix could have triggered biomass loss.