## 4 Discussion
The lack of Ca in the superficial layers of rocks coated by the desert varnish was previously observed by other authors [33, 46], who attribute the calcite matrix deficiency to dissolution actions by microorganisms. Biodeterioration of calcareous rocks due to microbial colonization is a well-studied phenomenon [56, 75, 76, 77], and acidification due to the production of carbonic, sulfuric and nitric acids as byproducts of cell metabolisms has been proposed as a possible cause of calcium carbonate dissolution [29, 78]. In particular [PERSON] et al. [79] mention cyanobacteria genera such as _Gloecocapsa_, _Nostoc_ and _Oscillatoria_ as able to degrade calcareous rocks. These cyanobacteria genera were all detected as part of the bacterial communities by the metagenomic analysis carried out in this study. The presence of microorganisms in the microcavities below the crust was detected by SEM analysis; further, by means of Raman spectroscopy, we were able to identify carotenoid pigments in the reddish layer below the dark coating, indicating biological colonization. Moreover, extensive microbial colonization in the cavities and discontinuities of the rocks associated with petroglyph sites through SEM-BSE was previously broadly studied by [PERSON] et al. [33], who identified a complex microbial community (cyanobacteria, heterotrophic bacteria, free-living fungi and lichens) inhabiting the superficial layers of the rocks in which the Ca matrix is no longer present. [PERSON] and [PERSON] [46] state that cyanobacteria and fungi dissolve the carbonates in limestone rocks to reach optimal living conditions such as temperature, humidity and nutrient supply. Indeed, the interior of the rocks can be a suitable niche for microorganisms to survive harsh condition due to better nutrient and water availability [54, 55]. In the case of stones covered with desert varnish, this might act as an extra shelter from the extreme environmental conditions [16]. Moreover, [PERSON] et al. [25] hypothesized that some species might take advantage of the rich transition-metal coating, being able to utilize the varnish minerals; they give as an example the use of Mn by cyanobacteria in their photosynthetic apparatus. Moreover, the authors of this study report that they detect enrichment in the metabolic process genes of microorganisms that relay on varnish-enriched transition metals. It is known that a wide variety of endolithic microorganism has the ability to mobilize metal cations [55] and in particular iron [72] and manganese, which are key metals used by microorganism for protection against UV radiation. [PERSON] and [PERSON] [60] state that the cellular accumulation of Mn by microorganisms might be a widespread mechanism to survive oxidative stress, and they investigated Mn\({}^{2+}\) function in _Rubrobacter_ spp., suggesting that Mn-antioxidant complexes are involved in radiation resistance, while a recent study [80] on the radiation resistance of _Deinococcus Radiodurans_ established the key role played by intracellular Fe and Mn in oxidative protection of this microorganism. In this study, _Rubrobacter_ spp. were dominant in all of our samples, and especially in sample EZ4, and might uptake both iron and manganese from the black crust. Moreover, the genus _Rubrobacter_, as mentioned previously, is known to grow on stone monuments with the ability to form biofilms and penetrate the porous rock, enhancing mechanical damage through the detachment of mineral grains [62]. These observations support the hypothesis that in sample EZ4, the black varnish might have been degraded by microorganism, including the genus Acinetobacter (18% of total reads of sample EZ4), which is reported to be able to solubilize Al, Si and Fe from rocks [58]. More and more recent studies seem to support the hypothesis that the black crust is of biological origin [25; 81; 82; 83; 84; 85; 86; 87; 88; 89; 90; 91; 92; 93; 94; 95; 96; 97; 98; 99; 100  
**Author Contributions:** Conceptualization, K.S., G.P. and A.K. ; analytical measurements, L.R., I.N., E.E. and M.J.P.