Contents lists available at ScienceDirect International Biodeterioration & Biodegradation journal homepage: www.elsevier.com/locate/ibiod In situ long-term monitoring of recolonization by fungi and lichens after innovative and traditional conservative treatments of archaeological stones in Fiesole (Italy) Daniela Pinna a,* , Monica Galeotti b , Brunella Perito c , Giulia Daly c , Barbara Salvadori d a University of Bologna, Ravenna Campus, Via Guaccimanni, 42, 48121 Ravenna, Italy b Opificio delle Pietre Dure, viale Filippo Strozzi 1, 50129 Firenze, Italy c Department of Biology, University of Florence, Via Madonna del Piano 6, 50019 Sesto Fiorentino, Firenze, Italy d Institute for the Conservation and Valorization of Cultural Heritage, National Research Council, Via Madonna del Piano 10, Sesto Fiorentino I-50019, Italy ARTICLE INFO Keywords: Untreated and treated stones Copper nanoparticles Recolonization Fungi Lichens Bioreceptivity ABSTRACT The research complements a study (Pinna et al., 2012) carried out to evaluate the effectiveness of mixtures of consolidants and water-repellent products (tetraethylorthosilicate, methylethoxy polysiloxane, Paraloid B72), with biocides (tributyltin oxide, dibutyltin dilaurate, copper nanoparticles) applied in situ to prevent biological growth on stones. The mixtures were tested over time on trial areas of three substrates - marble, sandstone, and plaster – in the archaeological site of Fiesole (Firenze, Italy). The 8-year-long study showed that the re- colonization of the three substrates after the conservation treatment related mainly to their bioreceptivity and to the climatic conditions. Although the mixtures of water repellents and consolidants with biocides and copper nanoparticles were effective in reducing the recolonization, they did not play a crucial role in preventing bio- films and lichens growth. This study demonstrated that it was not possible to draw common conclusions re- garding the products’ performance on the examined stones. Copper nanoparticles proved to be a suitable al- ternative to traditional biocides because they did not alter stones colour and contributed to the prevention of recolonization. The study provided information on the succession of fungi and lichens on untreated and treated stones, as well as on the variations of water repellency of treated stones. 1. Introduction A growing number of studies has considered the recolonization after treatment as one of the most challenging aspects nowadays in con- servation of cultural heritage objects. Particularly demanding are the following issues: the recolonization of bare surfaces after the removal of biofilms and lichens, and the recolonization of surfaces treated with consolidants and/or water repellents that can be useful in preventing biofouling (see review in Pinna, 2017). These substances can inhibit microbial growth by reducing the degree of saturation of the stone and limiting the amount of water available to organisms. As consolidants and water-repellents are currently applied in the final steps of re- storation/conservation processes, the development of microorganisms on treated surfaces is worthy of investigation. Two water-repellents, the products Rhodorsil H224 and Protectosil SC Concentrate, efficiently prevented the biological colonization for 2 years on limestone samples located outdoors in a park in Belgium, while tetraethoxysilane (TEOS, a consolidant) mixed with the antimicrobial agent Chitosan was not effective (Eyssautier-Chuine et al., 2014). A 160-day-long laboratory test evaluated the ability of water-repellents (silane and fluorinated compounds) to prevent colonization of the alga Graesiella emersonii on mortar samples (Martinez et al., 2014). The treatment notably reduced the progression of colonization compared with that obtained with the control specimens. Drawbacks can occur after the application of these products, how- ever. A study observed that the hydrophobicity provided by water-re- pellents on marble statues inhibited the spreading of raindrops over the surfaces (Charola et al., 2007) and created preferential water paths that favoured the accumulation of biomass and eventually the development of localized biofilms. The formation of these preferential water routes depended strongly on the structural characteristics of the stone sur- faces. In fact, autoclaved aerated concrete samples treated with alkyl- alkoxy silane showed streaking patterns of colonization when tested with an accelerated fouling experiment, while white concrete speci- mens treated with the same water-repellent did not show any streaking (De Muynck et al., 2009). According to these authors, the low https://doi.org/10.1016/j.ibiod.2018.05.003 Received 31 January 2018; Received in revised form 20 April 2018; Accepted 5 May 2018 * Corresponding author. E-mail address: daniela.pinna@unibo.it (D. Pinna). International Biodeterioration & Biodegradation 132 (2018) 49–58 0964-8305/ © 2018 Elsevier Ltd. All rights reserved. T