A methodology aiming at identifying and removing biofilms from cultural heritage was applied to stones from tuff walls in historical sites. Identification of phototrophic encrusting microorganisms was carried out by optical and electron microscopy, as well as by molecular techniques (DNA analyses and denaturing gradient gel electrophoresis (DGGE)). In all sites, the examination of microbial components of biofilms resulted in the identification of 17 species belonging to Cyanobacteria, Rhodophyta, Bacillariophyta and Chlorophyta, with Cyanobacteria being the dominant components in all biofilms. In order to remove the biofilms, an innovative technique based on the use of nonthermal effects of radiofrequencies was adopted. The source of the electromagnetic fields was a signal generator connected to a horn antenna through an amplifier to provide the power boost required to generate the target field amplitude. Seven days after exposure to radiofrequency electromagnetic field, about 50 % reduction of biofilm was observed; after 14 days, biofilm extension was reduced by about 90 %. DGGE analyses performed after 14 days confirmed these visual inspections. Also, DGGE analyses carried out before and 14 days after treatments showed that 12 out of 17 identified species disappeared. A complete visual disappearance of biofilms was observed a month after the beginning of treatments. DGGE repeated at this time confirmed the total disappearance of biofilm-forming species. Treated stones, when transferred back to their original sites, did not show any microorganism re-growing after 6 months. No alteration in the color and structural consistency of tuff substrata was observed after radiofrequency treatments.
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