Colletotrichum acutatum is an important phytopathogenic fungus causing anthracnose in commercially important fruit crops, such as strawberry. The conidia produced by the fungus are survival structures which play a key role in host infection and fungal propagation. Despite its relevance to the fungal life cycle, conidial biology has not been extensively investigated. Here, we provide the first proteomic description of the conidial germination in C. acutatum by comparing the proteomic profiles of ungerminated and germinated conidia. Using two-dimensional electrophoresis combined with MALDI-TOF/TOF mass spectrometry, we have identified 365 proteins in 354 spots, which represent 245 unique proteins, including some proteins with key functions in pathogenesis. All these proteins have been classified according to their molecular function and their involvement in biological processes, including cellular energy production, oxidative metabolism, stress, fatty acid synthesis, protein synthesis, and folding. This report constitutes the first comprehensive study of protein expression during the early stage of the C. acutatum conidial germination. It advances our understanding of the molecular mechanisms involved in the conidial germination process, and provides a useful basis for the further characterization of proteins involved in fungal biology and fungus life cycles.