# Comparison of Methods to Identify and Monitor Mold Damages in Buildings
## 4 Discussion
### Pros and Cons of Different Methods: Microscopy vs. DNA Metabarcoding  
All fungal detection techniques have specific benefits and drawbacks, providing a fragmented picture of the mycobiota present in a given sample. This has been demonstrated by diverse comparative studies on indoor dust and/or material samples, especially focused on the comparison between culture-dependent and -independent methods [39; 40; 41]. In the present study, we compared two contrasting techniques (microscopy vs. DNA metabarcoding) for the characterization of indoor surfaces with visible mold growth. The microscopy of tape lifts is a very simple and cheap method to document which fungi are actively growing on building materials, as its results are based on the observation of fungal reproductive structures [12]. Similar to the morphological identification of cultivated isolates, this microscopy technique sometimes reaches taxonomic identification down to the species level, when the key structures are present and the observer is skilled enough to classify them. However, microscopy is not able to identify many of the spores and hyphal fragments that settle on the sampled areas, which are especially abundant in dust samples collected from dry surfaces. Based on the microscopy results, we were able to identify the main indicator fungi associated with the studied mold damages, where 39.6% of the morphological identifications were at the species level. Many identifications could only reach the genus level, e.g., species of _Aspergillus_, _Penicillium_, _Cladosporium_, _Acremonium_, and _Trichoderma_, due to the lack of the species-specific features on the tapes. Species identification within these genera often requires further analyses, such as cultivation using special media and incubation conditions, metabolic characterization, and/or the DNA sequencing of multiple markers [29; 42]. Our microscopy results also showed some uncertainty at the genus level. For example, _Ullocladium_ sp. was found growing in four mold samples (M4, M24, M37, and M60; Tables S2 and S3) where the DNA-based relative abundances of _Alternaria_ were considerably high. In view of the morphological similarity between these two taxa, whose only distinctive feature is the type of spore ontogenesis on the conidiophore [29], both of them most likely correspond to the same indicator fungus. It is difficult to determine the most accurate identification for this fungus because these contradictory results may be due to (i) the arguable taxonomic weight of this morphological trait and/or (ii) the difficulty of distinguishing between the genera _Alternaria_ and _Ullocladium_ based on the ITS2 sequence divergence. For instance, the representative ITS2 sequence of the most abundant OTU assigned to _Alternaria_ showed 100% similarity against members of both genera. Previous DNA-based studies revealed multiple non-monophyletic genera, including _Alternaria_ and _Ullocladium_, within the so-called _Alternaria_ complex [43]. The main advantages of DNA metabarcoding are (i) the high sensitivity based on both the PCR and HTS and (ii) the quantitative potential providing relative abundances of comprehensive lists of taxa [21; 41]. In contrast, the microscopy of tape lifts can only detect a few dominant taxa with some qualitative information about how abundant they are growing on the studied surface area. In this study, considering the small tape area observed and the lack of homogeneity between the samples of these qualitative data, we decided to exclude them from the analyses. Comparing the richness in the mold samples, the DNA metabarcoding reported a mean of 16 OTUs per sample, while the microscopy of the tape lifts only identified two taxa per sample on average. It is worthwhile to mention that DNA metabarcoding is a semi-quantitative technique, as relative quantities are not always correlated to absolute measurements. [PERSON] et al.