This study used subcellular fractionation combined with proteomics for a comprehensive overview of the protein localizations of R. salmoninarum proteins within enriched subcellular fractions of cytosolic, membrane, extracellular and membrane vesicle (MV) proteins. With this approach, 1,674 protein groups were label-free quantified and the abundance of proteins involved in virulence processes were compared between the subcellular fractions.