Shredder organisms play a key role in rivers by feeding and fragmenting coarse organic matter that will then be exploited by other consumers. The effects of microplastics (MPs) on Gammarus sp., an ubiquitous genus of freshwater amphipods, and its shredding activity have been broadly investigated. However, the potential behavioral and physiological effects of different sizes of MPs on Gammarus sp. remain overlooked despite the recognized influence of MP size on MP toxicity. This study investigated the effects of a 28-day exposure to four different concentrations of two size fractions of PVC-microplastics (PVC-MPs), on Gammarus fossarum mortality rate, feeding rate, assimilation efficiency, and expression of proteins involved in key processes. Increased mortality was observed for all treatments exposed to PVC-MPs, with higher mortality in the presence of smaller PVC-MPs at the highest concentration. No protein biomarker modulation was observed in presence of PVC-MPs, suggesting that no metabolic stress but direct physical damages of PVC-MPs might have led to the observed mortalities. No difference was observed for feeding rates, but a higher assimilation efficiency was measured for individuals exposed to PVC-MPs, regardless of the concentration. This could be due to energy reallocation towards defense mechanisms or indicate a potential shift in digestive microbiota. This study highlighted the toxicity of PVC-MPs, particularly of smaller sizes and even at relatively low concentration, for Gammarus fossarum. PVC-MP pollution may therefore alter the functional integrity of river ecosystems by reducing the abundance of shredder organisms and, subsequently, the process of leaf litter decomposition.