# Biological Risk for Hypogea: Shared Data from
## 3 Results
### The Etruscan Tombs  
#### 3.1.1 Recurrent Biodeterioration Phenomena  
Research which commenced in the 1980s indicate that the Etruscan tombs are mainly populated by _Actinobacteria_ (belonging mainly to the order Actinomycetesales) which give rise to biodeterioration phenomena of the painted layers. Firmicutes, in particular Bacillaceae, and Alphaproteobacteria (order Rhizales) also seem to play an important role. Modern biomolecular tools utilised for the Tarquinia and Chiusi tombs have permitted a more precise identification of these microbiomes, even if some species do not seem to play a part in the biodeterioration processes. In the case of the Chiusi tombs, microbial colonisation was observed on both painted and unpainted wall surfaces, and a total of 456 non-chimeric bacterial sequences were obtained from different areas and pigments, mostly belonging to the Alphaproteobacteria (order Rhizales) and Firmicutes groups, such as _Rhodococcus_ sp., _Streptomyces_ spp., _Bacillus_ spp., _Paenibacillus_ sp., and also of _Nocardi_ and _Pseudonocardi_ genera [46; 47; 48]. In the Mercareccia tombs (Tarquinia), 142 different heterotrophic colony morphotypes were identified, 16% of which had not been described in relation to artistic heritage up to now. The most representative bacteria belonged to Actinomycetes and Bacillales, and the most common genera were _Bacillus_ (24 strains) and _Streptomyces_ (18 strains), together with _Rhodococcus_ (seven strains), _Paenibacillus_ (two strains) and _Pseudomonas_ (one strain) [39]. In one Tarquinia tomb (Shields) [42], were also detected moonmilk deposits, which consist of very fine crystals of calcium carbonate, made up of calcite needle fibres (1-2 \(\upmu\)m) and nanofibres (less than 1 \(\upmu\)m). This phenomenology arises from the progressive accumulation of calcite which also traps microorganisms in the crystal matrix, until they are completely encompassed in the crystals [88; 89]. The patina is similar to talcum or chalk powder when dry, and the white layers having different consistencies. One is softer while the other is thick and hard, thus more difficult to remove. The microbial community was made up of Proteobacteria, representing approximately 50% of the population, followed by the phyla _Acidobacteria_ and Actinobacteria [42]. More precisely, the class Alphaproteobacteria predominated the community (25% of the population), followed by Betaproteobacteria, Gammaproteobacteria and Actinobacteria. The most representative genera were _Pseudonocardi_ and _Hypomicrobium_. In this case, the high affinity in composition highlighted how representative they were of microbiota contained in the tomb [42]. A comparison of the taxa using biomolecular techniques often shows that microbial communities can vary greatly, in fact, the two tombs of Chiusi have very few in common [47; 48]. The presence of fungi on the mural paintings displayed a variable, though not negligible, aspect. In the Chiusi tombs, amplification of archaea and fungi was unsuccessful from all samples taken from the whitish alterations [47; 48]. On the contrary, in the Tarquinia, Cerveteri and Veio tombs, a certain amount of fungi was recorded. For example, in the Mercareccia Tomb (Monterozzi Necropolis), fungi occurred in both the dromos and inner chamber. The taxa identified were Sordariomycetes, Eurotiomycetes, and Dothideomycetes, along with several species of _Penicillium_ (_P. rugulosum, P. communis, P. brevicompactum_) and _Aspergillus_ (_A. candidum, A. versicolor, A. flavus_). Other species that were detected included _Cladosporium cladosporioides, Preussia terricola, Trichocladium asperum, Scopulariopsis brevicailis, Engyodontium album_, _Eurotium herbiorum_, and _Isaria farinosa_[39]. Our present study on the Moretti and Blue Demons Tombs in Tarquinia highlighted melanised fungi as responsible for the dark pattern deterioration found in both tombs (Figure 3). _Acremonium_-like fungi (three strains), _Cladosporium_ sp.