The invasive capacity of streptococcal strains belonging to groups A and B was evaluated by infecting human epithelial and endothelial cells and monitoring the number of viable intracellular bacteria at different times postinfection. All strains tested entered eukaryotic cells (HeLa, HEp2 and HUVE), with Streptococcus pyogenes exhibiting a higher invasion efficiency than group B streptococci (GBS). No intracellular multiplication was observed, and GBS remained viable 24 h postinfection, whereas S. pyogenes were gradually killed. We found that cytochalasin D almost completely inhibited internalization of all bacterial strains, whereas colchicine had no effect, indicating that host microfilaments play a major role in bacterial internalization. Moreover, the use of the lysosomotropic agent ammonium chloride enabled us to demonstrate that a pH increase in the intracellular vesicles did not affect streptococcal entry. These results were documented by electron microscopic observations which revealed the different steps in the invasion pathway, including a fusion event between phagosomes containing S. pyogenes and lysosomes.