Apical periodontitis is an inflammatory disease of periradicular tissues caused by microbial infection of the endodontic space, after necrosis of the dental pulp. Necrotic pulp is colonized by microbial agents, most commonly bacteria, which exist as planktonic cells in the fluid phase, as aggregates and coaggregates, and in the form of biofilms. Inflammation in the periapex is a defense of the body against the penetration of pathogens and toxins into the body. Understanding the causative factors (microorganisms) and the host is very important for quality treatment in the clinical care of patients with apical periodontitis. The classical methods of cultivation of microorganisms as well as modern methods of molecular medicine have enabled the identification of numerous bacterial species associated with the development of the clinical picture of apical periodontitis. Primary apical periodontitis (unless caused by non-infectious irritation) is characterized by polymicrobial flora in the pulp space, while persistent apical periodontitis is characterized by infection with fewer microorganisms. The most common pathogens associated with primary periodontitis are Prevotella and Porphyromonas genera, Tannerella forsythia, Dialister species, Fusobacterium, Spirochetes, and gram positive anaerobic rods such as Pseudoramibacter alactolyticus, Filifactor alocis, Actinomyces spp., Propionibacterium propionicum. Microorganisms in the necrotic pulp habitat most often create biofilms, which enables their synergistic activity and requires a thorough approach in the chemomechanical treatment of the infected area with the aim of successful endodontic therapy and healing of the periapical lesion.