## 3 Results and Discussion  
### Fungal Isolation and Identification  
From the sampling performed, a total of 43 fungal isolates were obtained across all five samples. The leather component of ANT.Moc.490 was the sample with the highest number of isolates (11), followed by ANT.Moc.349 (10), ANT.Moc.344 (9) and the wooden component of ANT.Moc.490 (7), with MCUC.2013.1.3 showing the lowest number of isolates (6). Molecular analysis of the several sequences obtained indicated that these 43 isolates belonged to 15 different previously described species (Table A1). The majority (approximately 79%) of identified species belonged to the _Aspergillus_ genus, with _A. pseudoglaucus_ representing the majority of isolates (Figure 2). Other _Aspergillus_ species included _A. caesiellus_, _A. candidus_, _A. chevalieri_, _A. conicus_, _A. creher_, _A. proliferans_ and _A. ruber_. _Penicillium_ was the second most common genus (7%), with two different species (_P. brevicompactum_ and _P. phoenicium_), followed by _Talaromyces_ (_T. amestolkiae_ and _T. coalescens_) and _Wallenia_ (_W. sebi_), with both genera representing 5% each of the total number of isolates. The two other genera identified were _Epicoccum_ and _Trichoderma_, with each representing 2% of all recovered isolates, and both represented by only one species (_E. nigrum_ and _T. longibrachiatum_, respectively). Figure 2: Taxa identified through culture-dependent methods: (**a**) relative abundance of genera across all samples; (**b**) species identification and relative abundance per sample. Of the 15 different species isolated, _A. pseudoglaucus_ was the only one present in all samples, thus suggesting it was the main fungus responsible for the visible contamination observed in the artifacts. The _Aspergillus_ genus is of particular significance in the context of biodeterioration, being associated with the production of organic acids, calcium carbonate dissolution and mineralization, and the production of enzymes with ligninolytic, fibrinolytic, keratinocyte, estereolytic, lipolytic, proteolytic and cellulolytic activity [50; 51], while also producing mycotoxins that can lead to health issues for those that come in contact with them [50]. _Aspergillus pseudoglaucus_, a xerophilic filamentous fungus [52], has been previously isolated in other cultural heritage artifacts, such as leather luggage from the Tianjin Museum in China [18], the tomb of the first Portuguese king [53], cathedral wall paintings, paper, textiles, wooden sculptures [54] and easel paintings [55]. And while previous studies reported a negative reaction to in vitro biodeteriorative potential tests evaluating cellulolytic and proteolytic activity, the ability to perform calcium carbonate dissolution and acid and pigment production [56], a more recent study indicates it is a strong lipase and lecithinase producer [57]. During our own present study, strong yellow pigment production was observed on PDA culture media, thus posing a potential threat to artifacts from the MCUC collection susceptible to these biodeterioration phenomena. Moreover, _A. pseudoglaucus_ has been associated with maxillary sinusitis and superficial infections [58], making it a health concern for museum staff and visitors. Among the other _Aspergillus_ isolates identified, _A. caesellus_, a labophilic and thermotolerant fungus [59; 60], has well-established lignocellulosic activity [60], thus posing a significant threat to lignocellulosic cultural heritage objects, such as the one it was isolated from in this study (ANT.Moc.349), where it represented 30% of recovered isolates. _Aspergillus candidus_ has a strong biodeteriorative potential, with recorded proteolytic, lipolytic, cellulolytic, pectinolytic and estereolytic abilities [51; 54], with previous records of its occurrence in cultural heritage objects, including canvas and wooden artworks, wall paintings, books, manuscripts and other paper-based objects, photographs and even mummified human remains [54].