Corynebacterium, Listeria and Bacillus are ubiquitous gram-positive aerobic bacilli, which colonize environment, animals and human body. Corynebacterium diphtheriae produces diphtheria either cutaneous or respiratory forms, a disease controlled by vaccination only in some countries and with increased incidence due to immigration and vaccine rejection. Listeria monocytgenes is associated with bacteremia and meningitis in pregnant women, neonates, elderly and immunosuppressed patients, although its incidence at a global level remains unknown. Bacillus anthracis is considered a potential bioterrorism agent because of its lethality. Considering that all cutaneous, gastrointestinal and respiratory symptoms of anthrax could be unspecific, diagnostic procedures that allow their detection with greater safety in case of suspicion have been developed. Toxin production, invasiveness and intracellular survival, resistance in the environment and lytic enzymes production are mechanisms of virulence of these species. Other Corynebacterium or Bacillus species are considered opportunistic pathogens and their clinical relevance has increased along the last decades, with the increase of immunosuppressed patients and the use of medical devices and prostheses susceptible of colonization.