Although infection by Corynebacterium diphtheriae is a model of extracellular mucosal pathogenesis, and diphtheria is one of the most worried diseases, this microorganism can be associated also with invasive infections such as endocarditis, septic arthritis, and osteomyelitis. Invasive infections are usually caused by non-toxigenic C. diphtheriae strains. Over the last years severe pharyngitis/tonsillitis associated with the isolation of non-toxigenic C. diphtheriae have been described. Penicillin treatment failure of these infections could only partially be explained by penicillin tolerance of the causing strain. Thus, we examined the in vitro ability of non-toxigenic C. diphtheriae throat clinical isolates to adhere to, and enter human respiratory epithelial cells. Trasmission and scanning electron microscopy demonstrated intracellular C. diphtheriae in laryngeal (HEp-2 cells) and pharyngeal (Detroit D562 cells) tissue culture. Live intracellular bacteria were detectable up to 48 h post-infection. Using a variety of compound that act on eukariotic cell structures, the internalization of C. diphtheriae seems to occur via a zipper-like mechanism. It is likely that internalization of C. diphtheriae can be involved in throat colonization contributing to bacterial eradication failure and asymptomatic carriage.