In the Saint-Marcel cave (France), wood barrels and thousands of bottles containing red wine were stored for vinification. After storage began, a fungal and bacterial outbreak occurred, and the area was invaded by numerous types of mold colonizing the cave ceilings and all objects related to human activities (the stairwell and oenological materials). In this study, using the metabarcoding approach, we have studied the microbial outbreak and have linked the identified microorganisms to oenological activity. Both 16S and ITS primers were used to sequence the samples collected from the cave. The results showed that the dominant microorganisms proliferating in the cave were related to wine vinification. For instance, Zasmidium cellare, a strain known for living in dark and ethanol-rich environments, was the dominant fungus on the cave stairwell. Furthermore, Guehomyces pullulans, a cold-adapted yeast used for juice clarification, was recorded as the major species on the blackened limestone ceilings. These findings reveal a complex community structure in the studied cave based on the assembly of bacteria and fungi. Finally, our results demonstrate that oenological activities could seriously affect cave preservation, changing the natural microbial communities populating cave environments.