Listeria monocytogenes is the etiological agent of listeriosis, a severe human foodborne infection characterized by gastroenteritis, meningitis, encephalitis, abortions, and perinatal infections. This gram-positive bacterium is a facultative intracellular pathogen that induces its own uptake into nonphagocytic cells and spreads from cell to cell using an actin-based motility process. This review covers both well-established and recent advances in the characterization of L. monocytogenes virulence determinants and their role in the pathophysiology of listeriosis.