Artefact conditions need to be continuously monitored to avoid degradation effects naturally caused by time and public exploitation in order to increase the value of cultural assets. In this way, the atmospheric analysis of both biological and chemical pollutants potentially present inside conservation environments represents valid support for the adoption of preventive conservation actions by evaluating periodically the presence of risk for the same artefacts. The aim of the present study was to analyze the fungal particles, potentially biodeteriogen, through aerobiological volumetric monitoring, particularly inside valuable historical, artistic, and cultural sites. Different exposition and conservation typologies of the artefacts with different flows of visitors were considered. The applied methodologies have furnished a reliable description of biological air pollution due to the presence of fungal spores—moreover, they have allowed for the prevention of risk situations and the measurement of their evolution in order to limit degradation processes. Through aerobiological monitoring, it was possible to provide important indications for interventions of prevention, conservation and restoration of cultural heritage in indoor environments.