The present study aimed at reviewing the literature on extraradicular infections of endodontically treated teeth, summarizing the main hypotheses on etiopathogenesis and describing the most suitable techniques to identify the composition of pathogenic extraradicular microorganisms. Medline database was searched using the keywords “Apical biofilm,” “extraradicular infection,” “secondary endodontic lesion,” “endodontic retreatment,” “biofilm” either alone or combined with AND. A further hand search was performed on the main endodontic journals. The most frequent bacterial species identified in different studies and with different techniques may vary considerably. Although the presence of some species of microorganisms seems to be determinant, the true origin of extraradicular infection is still undetermined. The literature analysis showed marked differences in methodology, materials, aims, and techniques adopted, which led to highly heterogeneous outcomes. The picture emerging from this review is that extraradicular infection is likely a multifactorial disease that requires further systematic investigation using standardized techniques.