Microscopic and biomolecular complementary approaches to characterize bioweathering processes at petroglyph sites from the Negev Desert, Israel

Abstract

Throughout the Negev Desert highlands, thousands of ancient petroglyphs sites are susceptible to deterioration processes that may result in the loss of this unique rock art. Therefore, the overarching goal of the current study was to characterize the composition, diversity and effects of microbial colonization of the rocks to find ways of protecting these unique treasures. The spatial organization of the microbial colonizers and their relationships with the lithic substrate were analysed using scanning electron microscopy. This approach revealed extensive epilithic and endolithic colonization and close microbial-mineral interactions. Shotgun sequencing analysis revealed various taxa from the archaea, bacteria and some eukaryotes. Metagenomic coding sequences (CDS) of these microbial lithobionts exhibited specific metabolic pathways involved in the rock elements' cycles and uptake processes. Thus, our results provide evidence for the potential participation of the microorganisms colonizing these rocks during different solubilization and mineralization processes. These damaging actions may contribute to the deterioration of this extraordinary rock art and thus threaten this valuable heritage. Shotgun metagenomic sequencing, in conjunction with the in situ scanning electron microscopy study, can thus be considered an effective strategy to understand the complexity of the weathering processes occurring at petroglyph sites and other cultural heritage assets.

A Multi-Analytical Approach to Infer Mineral–Microbial Interactions Applied to Petroglyph Sites in the Negev Desert of Israel

“…The microorganism communities associated with rock varnish have been previously studied to determine whether or not there is a connection between the varnish and the microorganisms inhabiting the stone. The results of our metagenomic study are consistent with the findings reported in the literature [25,33], revealing a microbiome dominated by bacteria, with lower proportions of eukaryotes, and archaea representing less than 1% of the total assigned reads. Within the bacteria, Actinobacteria and Cyanobacteria were the most abundant phyla, followed by Proteobacteria and Bacteroidetes, as found also by other authors in metagenomics studies on microbial communities of stones in desert environments [16,27,30].…”

“…Previous studies mentioned the presence of fungi and lichens, such as Caloplaca sp., as a weathering agent of rock substrates due to the production of oxalic acid and consequent formation of calcium oxalates that can facilitate the disintegration of the calcareous matrix [33]. In this study, however, we could not detect the presence of calcium oxalates.…”

“…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].…”

“…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 Nir 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.…”

“…A few studies report serious damage in numerous rock art sites due to lichen colonization [31,32]. Nir et al [33], with a study combining scanning electron microscopy (SEM) and metagenomic sequencing methods, suggest that the stone surface of petroglyphs in the Negev desert is colonized by a complex microbial and lichen community with the biochemical potential to induce biodeterioration. Despite these studies, comprehensive research on deterioration of rock art panels, particularly by microorganisms, is still needed.…”