Active microbial ecosystem in Iron-Age tombs of the Etruscan civilization.

Earth's microbial biosphere extends down through the crust and much of the subsurface, including those microbial ecosystems located within cave systems. Here, we elucidate the microbial ecosystems within anthropogenic 'caves'; the Iron-Age, subterranean tombs of central Italy. The interior walls of the rock (calcium-rich macco) were painted ~2500 years ago and are covered with CaCO3 needles (known as moonmilk). The aims of the current study were to: identify biological/geochemical/biophysical determinants of and characterize bacterial communities involved in CaCO3 precipitation; challenge the maxim that biogenic activity necessarily degrades surfaces; locate the bacterial cells that are the source of the CaCO3 precipitate; and gain insight into the kinetics of moonmilk formation. We reveal that this environment hosts communities that consist primarily of bacteria that are mesophilic for temperature and xerotolerance (including Actinobacteria, Bacteroidetes and Proteobacteria); is populated by photosynthetic Cyanobacteria exhibiting heterotrophic nutrition (Calothrix and Chroococcidiopsis); and has CaCO3 precipitating on the rock surfaces (confirmation that this process is biogenic) that acts to preserve rather than damage the painted surface. We also identified that some community members are psychrotolerant (Polaromonas), acidotolerant or acidophilic (members of the Acidobacteria), or resistant to ionizing radiation (Brevundimonas and Truepera); elucidate the ways in which microbiology impacts mineralogy and vice versa; and reveal that biogenic formation of moonmilk can occur rapidly, that is, over a period of 10 to 56 years. We discuss the paradox that these ecosystems, that are for the most part in the dark and lack primary production, are apparently highly active, biodiverse and biomass-rich.

Calcite moonmilk of microbial origin in the Etruscan Tomba degli Scudi in Tarquinia, Italy.

A white deposit covering the walls in the Stanza degli Scudi of the Tomba degli Scudi, Tarquinia, Italy, has been investigated. In this chamber, which is still preserved from any kind of intervention such as cleaning and sanitization, ancient Etruscans painted shields to celebrate the military power of the Velcha family. Scanning electron microscopy analysis has revealed the presence of characteristic nanostructures corresponding to a calcite secondary mineral deposit called moonmilk. Analysis of the microbial community identified Proteobacteria, Acidobacteria and Actinobacteria as the most common phyla in strong association with the moonmilk needle fibre calcite and nanofibers of calcium carbonate. Employing classical microbiological analysis, we isolated from moonmilk a Streptomyces strain able to deposit gypsum and calcium carbonate on plates, supporting the hypothesis of an essential contribution of microorganisms to the formation of moonmilk.

A role for microbial selection in frescoes' deterioration in Tomba degli Scudi in Tarquinia, Italy.

Mural paintings in the hypogeal environment of the Tomba degli Scudi in Tarquinia, Italy, show a quite dramatic condition: the plaster mortar lost his cohesion and a white layer coating is spread over almost all the wall surfaces. The aim of this research is to verify if the activity of microorganisms could be one of the main causes of deterioration and if the adopted countermeasures (conventional biocide treatments) are sufficient to stop it. A biocide treatment of the whole environment has been carried out before the conservative intervention and the tomb has been closed for one month. When the tomb was opened again, we sampled the microorganisms present on the frescoes and we identified four Bacillus species and one mould survived to the biocide treatment. These organisms are able to produce spores, a highly resistant biological form, which has permitted the survival despite the biocide treatment. We show that these Bacillus strains are able to produce calcium carbonate and could be responsible for the white deposition that was damaging and covering the entire surface of the frescoes. Our results confirm that the sanitation intervention is non always resolutive and could even be deleterious in selecting harmful microbial communities.