Viral hepatitides are satisfactorily diagnosed in the laboratory by immunoassays for either antigen or antibody detection in serum samples. However, the early detection of acute infections during the window period, investigation of occult infections, and issues related to the establishment and follow-up of antiviral therapy in chronic infections pose new challenges that only molecular methods can meet. In addition, full characterization of epidemic outbreaks and surveillance of the emergence of viral variants able to escape from vaccine protection are major public health objectives that can only be achieved through the use of these techniques. As a further attempt to improve the viral safety of blood transfusions, the incorporation of molecular biology techniques into the routine work of transfusion centers has generated new technical and scientific demands on microbiology laboratories, which must in turn incorporate these methods to respond to the challenge. After more than a decade, automatic methods for the detection, quantification, characterization and sequencing of the genomes of these viruses have become a reality for the clinical laboratory, reaffirming the essential role of the microbiologist in the hospital setting.