The cinematographic films are a reproduced version of reality and have become vital documents to study everything around us. For their preservation it is necessary to study the biodeterioration of these documents. The aims of this study were to characterize the biodeterioration caused mainly by microorganisms in two cinematographic films by applying molecular methodologies, electron microscopy and epifluorescence microscopy as well as to determine the enzymatic characterization of the fungal species isolated. From the Cuban Institute for Cinematographic Industry and Arts (ICAIC) the samples on two damaged cinematographic films were collected. The films were analyzed by different microscopic techniques, including the two types of electron microscopy. Also, the degradative potential of the isolated fungi was determined by qualitative evaluation of the enzymatic activities. A significant fungal colonization on both sides of the films and the damages caused by these microorganisms in the material were observed as well as the exoskeletons of dust mites of the families Tydeidae and Tarsonemidae as part of biofouling that were used as nutrients by the fungi. By epifluorescence microscopy was observed that some of the microorganisms were still viable and active. Using molecular biology techniques could be identified several species corresponding to the fungal genera Aspergillus, Cladosporium, Penicillium and Microascus as well as the bacteria genera Bacillus, Staphylococcus and Kocuria, which were responsible of the biodeterioration of these films. All isolated fungal species were capable to degrade the cellulose and gelatin as well as to excrete organic acids and pigments. Bacteria were detected in low concentrations as well as other biological agents, but it was observed that the films were being colonized by a high concentration of various fungal species with a significant biodeteriogenic potential, demonstrating that they were severely affecting the films.