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The outstanding preservation of Paleolithic decorated caves is related to the buffering properties of their karstic environment. However, long-term monitoring of air/wall temperatures and gas compositions has recently revealed disruption signs in cave microclimates that had been maintained stable for hundreds of centuries. High precision and continuous temperature data records are currently monitored in various prehistoric caves in the South of France. Such operations have been promoted since the late 1990s by the French government for risk assessment and conservation. The most striking feature is the positive drift of underground temperatures (air and wall) which is now obvious in most sites except for Niaux Cave (> 300 m undersurface) and in the deepest parts of Mas-d’Azil and Chauvet Caves (> 50 m undersurface). In tourist caves (Pech-Merle, Mas-d’Azil, Gargas, Villars), the positive thermal trends could not be related to the energy increase brought by visitors which number is now stable, nor to the lighting systems whose energy demand was strongly reduced. In addition, the underground thermal drift nearly starts at the same time in many caves with uncertainties of +/- 1 year: 2012 for Chauvet with +0.4 °C/decade, 2011 for Pech Merle with +0.32 °C/decade, 2011 for Marsoulas Cave with +1.09 to +0.36 °C/decade from the entrance to the deep gallery, 2011 for Gargas with +0.69 °C, +0.54 °C and +0.36 °C for the deeper station. It is worth noting that a 0.3-0.4 °C thermal drift is consistent with that predicted from global warming in these regions. The thermal drifts were already in progress when monitoring began in Villars Cave in 1996 (+0.17 °C to +0.39 °C/decade), in Mas-d’Azil in 2012 and in Bruniquel in 2015. Marsoulas (+1.09 °C/decade) and Mas-d’Azil (more than +1 °C/decade in 6 of the 16 stations) present a much higher drift rate compared with that of surface, which suggests a thermal amplification process. As measurements are performed in heterothermal zones, the long-term thermal drifts are modulated by persisting smoothed and out-of-phase yearly variations. A notable exception is the case of Bruniquel main gallery where the temperature records show a quasi-linear increase. In that case, the decadal evolutions of temperature +0.31 °C, +0.175 °C, and +0.24 °C, are not related to the depth of monitoring stations (32 m, 55 m, and 38 m, respectively) nor to their distances from the entrance. In 2018, those drift rates induced a permanent inversion of thermal gradient in the main gallery. In Gargas, the drift rate is more pronounced in the outer parts of the karst body, thus inducing a continuous evolution of the thermal gradients within the galleries. Such underground microclimate disruption of patrimonial caves is a warning signal of direct threat on the preservation of remains. Karst physical organization and its related underground environment are themselves legacies of past climates; the current functioning of transfer zones of karst aquifers which includes the caves, are directly dependent on the outside climate. A more comprehensive approach and modelling of possible tipping points are urgently needed for conservation issues.

International audience; In La Glacière cave (France), the touristic activity has been conducted to an environmental parameter change that has led tophotosynthetic organism proliferation (microalgae, diatoms, cyanobacteria, bryophytes). The present study is focused on bryophyte development occurring in the show cave that was responsible of limestone biodeterioration. In order to understand thecolonization process of limestone, we have maintained limestone blocks under optimal Lampenflora growth conditions.Moreover, some limestone blocks were painted with several pigments that were used in the prehistory (e.g., red ocher, bonechar). Microorganisms and bryophyte growth were monitored during 1 year, and then, the block samples were treated using UVC light (254 nm). Thus, obtained results were compared with in situ treatment in La Glacière cave. Results have showed densebryophyte propagation on the several blocks. However, the growth rate was correlated with the chemical composition of thepigment. In fact, the presence of some chemical elements such as As, Cr, Ti, and Co contributed to reduce bryophyte growth.Finally, moss treatment using UV-C light has demonstrated high efficiency under in situ condition, while a fast recolonization hasbeen observed for samples maintained in laboratory. This difference was explained by the high bryophyte density underlaboratory conditions that make UV-C light penetration difficult.

Microbial organisms can cause huge crises in decorated caves, as seen in emblematic sites such as Lascaux cave. The preservation of such sites involves understanding the healthy microbial behavior of caves before the damage occurs. Indeed, knowledge of normal cave behavior is a prerequisite to identifying potential imbalance. This study seeks to determine whether models of aerobiological behavior could be identified in several caves of different sizes, ranging from rock shelters to large caves that are open or closed to the public. Aerial rates of fungi and bacteria were monitored over 3 years in nine sites in Dordogne (France). This study revealed that in a context of caves where public visits were carefully managed, fungal and bacterial rates were more affected by the size of the caves than by the opening of sites to the visitors. The study confirmed that large caves can generally be described as “self-purifying caves” as they were strongly affected by the exterior environment at their entry but much less so at locations further inside the cave, while small caves can be described as “non-self-purifying caves” since they were strongly affected by the exterior environment throughout their whole length. The results also highlighted the difficulty to determine a limit value of microbial rates valid for all caves because of the specificities of each one.

The dry rot fungus Serpula lacrymans is the most destructive fungal agent of wood building materials in Europe, Russia, North America, and Japan. The identification of this wood-deteriorating agent and the discrimination of different fungal isolates is very important for the control of buildings in general and for the preservation of cultural heritage in particular. The objective of the study was to develop a Fourier transform infrared (FTIR) method coupled with a partial least square discriminant analysis (PLS-DA) for the sample preparation and identification of S. lacrymans. Five distinct S. lacrymans strains were analysed and compared to two strains of unrelated fungal species. Different methods of mycelial growth, sample preparation, and FTIR spectral data normalisation were compared. FTIR analysis of a harvested mycelium grown on the surface of a polyether sulfone microfiltration membrane deposited on a malt extract agar medium, followed by vector normalization and PLS-DA statistical analysis, resulted in 100% correct attribution at phylum, species, and strain level, regardless of the type of standardization used. This study confirms the applicability of FTIR spectroscopy for the identification of S. lacrymans and the discrimination of different strains belonging to this species.

This study focuses on analysis of the archaeal and fungal diversity associated with gypsum efflorescences damaging the walls of the Sorcerer’s prehistoric cave registered as a world cultural heritage site. Archaeal 16S rDNA and fungal internal transcribed spacer (ITS) clone libraries were constructed and analysed. Two thaumarchaeotal OTUs belonging to the Nitrososphaeraceae family dominated the archaeal community (100% of clones). Nitrososphaeraceae are obligate aerobic, chemolithoautotrophic organisms that derive their energy from the oxidation of ammonia and may contribute to primary productivity in the cave. Seven fungal OTUs belonging to Ascomycota and one belonging to Basidiomycota were present. The Cordycipitaceae family, mainly represented by entomophilous fungi, dominated the analysis (66.7% of clones). We show that archaeal and fungal OTUs are associated with gypsum efflorescences damaging the walls of the Sorcerer’s cave. The role of these microorganisms in the deterioration of the walls of the cave remains to be determined.

L’etude et la restauration de la sculpture de la Vierge a l’Enfant du musee Sainte-Croix de Poitiers a ete l’occasion d’appliquer une methode innovante de traitement des micro- organismes chlorophylliens : l’exposition aux rayons UV-C. L’œuvre en pierre calcaire, possedant encore des traces de polychromie, avait ete conservee en exterieur et etait colonisee par les algues, mousses et lichens. La problematique du nettoyage de ces micro-organismes a entraine une etude technico-scientifique menee au Laboratoire de Recherche des Monuments Historiques. L’exposition sous rayons UV-C, nocifs pour les organismes vivants, presentait l’interet d’eviter l’utilisation d’un produit biocide. Les bons resultats de l’etude ont permis d’appliquer cette technique sur la Vierge a l’Enfant, une premiere dans le domaine de la restauration de sculpture en pierre.

In recent decades, the Azé cave has begun to suffer from microorganism proliferation due to artificial lighting installations for touristic activity. The present work aims at understanding the origin of the Lampenflora and bacterial community populating inside the cave. We performed high throughput sequencing of the communities populating the outside, the entrance and the inside of the Azé cave. Our results indicated that 68.2–74.5% of the phototrophic community was represented by Eukaryotes, and 25.5 to 31.8% by cyanobacteria regardless of the sampled area. We observed a decrease of bacterial and phototroph species richness from the outside to the bottom of the cave. The phototrophic communities were significantly different in term of specific OTU richness but not in term of composition. For photosynthetic organisms, only 4 OTU were specific to the inside of the cave, representing less than 0.1% of the total OTUs. On the contrary, more than 400 bacterial OTUs were specific to the inside of the cave. These metabarcoding-based findings in the Azé cave revealed a complex community structure that is similar but less diverse in comparison to the outside.

1. Problematiques et bilan des principaux resultats La tombe de Padiamenope (TT 33) est la plus grande tombe d’Egypte mais sa structure est particulierement atypique. Si elle a en commun avec les autres grandes tombes de l’Assassif les trois premieres salles avec une chapelle de culte, elle possede aussi un ensemble cultuel central (le cenotaphe) avec un couloir de circumambulation. Claude Traunecker a montre que le ...

International audience; Carbon dioxide (CO 2) concentration (CDC) is an essential parameter of underground atmospheres for safety and cave heritage preservation. In the Chauvet cave (South France), a world heritage site hosting unique paintings dated 36,000 years BP, a high-sensitivity monitoring, ongoing since 1997, revealed: 1) two compartments with a spatially uniform CDC, a large volume (A) (40,000 to 80,000 m 3) with a mean value of 2.20 ± 0.01% vol. in 2016, and a smaller remote room (B) (2000 m 3), with a higher mean value of 3.42 ± 0.01%; 2) large CDC annual variations with peak-to-peak amplitude of 2% and 1.6% in A and B, respectively; 3) long-term changes, with an increase of CDC and of its annual amplitude since 1997, then faster since 2013, reaching a maximum of 4.4% in B in 2017, decreasing afterwards. While a large effect of seasonal ventilation is ruled out, monitoring of seepage at two dripping points indicated that the main control of CDC seasonal reduction was transient infiltration. During periods of water deficit, calculated from surface temperature and rainfall, CDC systematically increased. The carbon isotopic composition of CO 2 , correlated with water excess, is consistent with a time-varying component of CO 2 seeping from above. The CO 2 flux, which is the primary driver of CDC in A and B, inferred using box modelling, was found to confirm the relationship between water excess and reduced CO 2 flux into A, compatible with a more constant flux into B. A buoyancy-driven horizontal CO 2 flow model in the vadose zone, hindered by water infiltration, is proposed. Similarly, pluri-annual and long-term CDC changes can likely be attributed to variations of water excess, but also to increasing vegetation density above the cave. As CDC controls the carbonate geochemistry, an increased variability of CDC raises concern for the preservation of the Chauvet cave paintings.

“La Muette” is an eighteenth-century hunting lodge located in the forest of Saint-Germain-en-Laye, France. This study consisted in the implementation of multidisciplinary analytical tools to characterize the fungal biocontamination of woods inside the monument. The fungal colonization was first mapped by macroscopic visual observation of all the rooms. Brown rot and white rot present in different rooms and fruiting bodies of Serpula lacrymans were observed on the first and second floors in localized areas. At the same time, air sampling was carried out, and the volatile organic compounds (VOCs) specific to the activity of fungi were analysed by gas chromatography. The analysis of the VOCs confirmed the presence of active developments of moulds and S. lacrymans in the pavilion. Wood samples were collected from areas with visible fungal colonization and subjected to culturing, extraction of DNA and analysis of the ITS sequences after PCR amplification. Culturing and microscopic observation revealed the presence of several moulds: Penicillium sp., Cladosporium sp., Acremonium sp., Humicola sp., Rhizopus sp. and Mucor sp. Isolates not identified by this approach were identified by analysing ITS sequences as Trichoderma atroviride, Engyodontium album and Umbelopsis isabellina. The presence of additional Ascomycota (Acremonium charticola, Aspergillus conicus, Chaetomium elatum, Lecanicillium sp.) and of the Basidiomycota Coprinellus aff. radians was revealed after DNA extraction, amplification and analysis of ITS sequences directly on wood samples. In conclusion, a high fungal diversity including moulds, S. lacrymans and C. aff. radians was demonstrated in association with altered wood in the monument.

UV-C and biocide treatments are frequently used to treat historical monuments contaminated by autotrophic biofilms. In this study, the authors compare for the first time the efficacy of these treatments against microorganisms such as cyanobacteria and algae proliferating in the Vicherey church (Vicherey, Vosges Department, France). To identify the most environmentally friendly and also efficient method, an allelopathic treatment was also tested. Colorimetric and physiological measurements of treated biofilms were thus monitored for 6 months. Fungi growing on necrotic matter from treated biofilms were sampled and sequenced. With biocides, results showed incomplete eradication of biofilms, even after two treatments. Biofilm color loss was delayed in comparison to UV-C treatment, which appeared more efficient after just one treatment. Moreover, quantum yield (Fv/Fm) decreased immediately after UV-C treatment, indicating inhibition of algae and cyanobacteria photosynthesis. However, two species of fungi colonized the cyanobacteria biofilms treated with UV-C. Allelopathic treatment was not efficient on biofilm and showed no deleterious effect on photosynthesis. The present findings demonstrate that the UV-C method, coupled with a cleaning phase for necrotic organic matter, may be considered environmentally friendly and the best alternative to chemicals.

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The crypt of Saint-Savin-sur-Gartempe Abbey church contains a notable example of Romanesque wall paintings. A recurrent white fungal covering has affected these wall paintings since the late 1970s. Its composition and changes were studied from 2005 to 2015 through microbial isolations, ATP activity measurements and antimicrobial susceptibility testing. ATP monitoring showed that the fungal activity level remained high throughout the ten years, with spatial and temporal variations over time. Engyodontium album was the main agent isolated from the fungal covering, showing change in fungal strains compared to those isolated before 2005. This change in the fungal community could result from the previous application of econazole nitrate as a biocidal product. However, multiple susceptibility tests showed a possible variation of active ingredients that could lead to the successful elimination of the fungal contamination. Alternative solutions for the preservation of the wall paintings against fungal covering are discussed.

Caves are considered oligotrophic habitats exhibiting constant temperature and relative humidity throughout the year. While darkness inhibits photosynthetic microorganism growth, introducing artificial lights to promote touristic activity can induce algae and cyanobacteria proliferation. Besides the aesthetic problem, microorganisms are responsible of physical and chemical degradation of limestone wall with possibly a degradation of prehistoric painting of cultural value. In our studies, we identified lampenflora with new-generation sequencing (NGS) in five French show caves and also on a 180, 000 years old contaminated bear bones (Ursus deningeroides). Afterward, we attempted to find an ecological and efficient method to eradicate photosynthetic biofilms. Our previous works showed the good efficiency of UV-C against biofilms. To ensure the safety of UV-C treatment, pigments used for prehistoric parietal painting were strongly irradiated around 4800 kJ m−2. Afterward, we treated all biofilms in La Glaciere show cave. Following the treatment, irradiated biofilm color evolution was monitored. Results of sequencing showed that biofilms were mainly composed of bacteria, fungi, cyanobacteria, chlorophyceae, and diatoms. Multicellular mosses were also present in each biofilm. Furthermore, X-ray crystallography, infrared spectroscopy, and colorimetric measurement showed no change in pigment color nor in their chemical structure. Finally, UV-C treatment results showed that biofilms were completely eradicated with no algae proliferation observed after 21 months. Moreover, a complete bleaching is obtained with UV-C, which is not always the case with chemical conventional methods. In conclusion, the use of UV-C could be considered as an environmentally friendly good alternative to chemical products. Moreover, microorganisms are killed by radiation, while no change in pigment color nor in their chemical structure has been observed.

International audience; Fungi are present in natural and non-touristic caves due to the presence of organic matter provided mainly by insects or animals such as bats. In show caves, however, tourist infrastructure and the visitors themselves are an important source of organic matter. In addition, photosynthetic biofilms provide a high amount of carbon and nitrogen sources for fungi. This study was conducted to identify the fungal communities present in caves along with the potential use of UV-C treatment against their proliferation. Thus, fungal communities proliferating in biofilms in six French and Swiss show caves were analyzed using high throughput sequencing. The results show 385 species recorded, some of them previously described in cases of fungal outbreak. This preliminary study also aimed to test the use of UV-C light as an environmentally friendly method to treat fungal proliferation. Six fungal strains, from three different sources (Lascaux cave, La Glaciere cave, a church in Vicherey, France), were cultivated in an agar dish. Spores, mycelia and the entire colony were irradiated using several UV-C intensities. Results showed that four of the six fungi spores and mycelium died following a low-intensity UV-C treatment (2 kJ m(-2), 160 s), though Ochroconis lascauxensis and Penicillium bilaiae spores showed higher resistance. Finally, it was demonstrated that the fungal colony could resist the UV-C light due to a shadow effect. The structure of the fungal colony was affected from the periphery to its inner part. However, after four 30 kJ m(-2) treatments (39 min irradiation) all strains there definitively eradicated. Further studies will be necessary to examine the potential of UV-C light under cave conditions as a preventive and curative treatment.

This study focused on a French prehistoric decorated cave which suffers from the development of saline efflorescences. The goal was to determine the ecology of microorganisms associated with efflorescences, and determine if the sessile microbiota could have a role in the deterioration process. Samples from 2 sampling campaigns were analyzed. Elemental, mineralogical and isotopic analyses showed gypsum containing sulfur originating mainly from the host-rock. Microscopic observations, direct cell counting, culture-dependent assays and high-throughput sequencing (HTS) of bacterial 16S rDNA sequences indicated an abundant sessile microbiota, where Actinobacteria, Alphaprotebacteria, Gammaproteobacteria, Bacteroidetes and Planctomycetes were the main phylogenetic groups. Although a relatively low abundance of genera associated with sulfur cycling were identified, the presence of dense biofilms at the interface rock/efflorescences composed of a high relative abundance of Actinobacteria may reflect previously described indirect biomineralization. An increase of microbial abundance and bacterial richness between October 2014 and April 2015 might be attributed to fluctuations in environmental factors. This study gives insight into origin and bacterial composition of efflorescences that threatened Sorcerer's cave engravings. HTS has rarely been used in the field of cultural heritage and thus represents one of the originalities of this work.

Photosynthetic biofilms proliferating on heritage monuments represent a major threat for curators leading to biodegradation and esthetic issues. Previous studies demonstrated that UV-C, used as a tool for biofilm eradication, is a promising avenue to combat microbial proliferation. In this study, this environmentally friendly method was tested on biofilm-forming Chlorella vulgaris suspension. Algal physiological response to UV-C was then assessed. Results showed that > 10 kJ m−2 UV-C exposure was enough to directly kill cells whereas low UV-C exposure reduced quantum yield of photosystem II and inhibited both respiration and photosynthesis. Clear relationships between UV-C exposure times and physiological responses were found. In addition, the use of VIS-light after UV-C treatment enhances chlorophyll bleaching. Our findings contribute to a better understanding of the physiological responses of Chlorella vulgaris to UV-C radiation allowing thus an optimization of the UV-C treatment reported in our previous studies.

Green microalgae colonizing stone surfaces represent a major problem for the conservation of heritage monuments, since they lead to biodegradation and aesthetic issues. Previous studies in La Glaciere show cave (France) have demonstrated that UV-C may have a strong effect on microalgae, thus leading to chlorophyll bleaching, which was increased when biofilms were maintained under VIS-light condition unlike to those maintained in the dark. To understand the physiological mechanisms underlying this response and in order to optimize in situ treatment, 30 kJ m−2 UV-C exposure times were applied to Chlorophyta Chlorella sp. and chlorophyll degradation kinetics were then monitored. UV-C irradiation was enough to inhibit photosynthesis and to directly kill all algal cells. Results also showed that chlorophyll a was degraded faster than chlorophyll b and that 14 h were necessary for complete degradation of all the present chlorophyll. In addition, our results highlighted the importance of visible light exposition after UV-C treatment which leading to chlorophyll bleaching. Irradiated algae cultivated in the dark were still green 5 days after treatment while cultivated samples in the light lost their green color after 14 h. An efficient UV-C treatment applicable to show caves and other heritage monuments was proposed.

International audience; In recent decades, show caves have begun to suffer from microorganism proliferation due to artificial lighting installations for touristic activity. In addition to the aesthetic problem, light encourages microorganisms that are responsible for physical and chemical degradation of limestone walls, speleothems and prehistoric paintings of cultural value. Microorganisms have previously been described by microscopy or culture-dependent methods, but data provided by new generation sequencing are rare. The authors identified, for the first time, microorganisms proliferating in one Swiss and in four French show caves using three different primers. The results showed that both photosynthetic and non-photosynthetic bacteria were the dominant taxa present in biofilms. Microalgae were heavily represented by the Trebouxiophyceae, Eustigmatophyceae and Chlorophyceae groups. Twelve diatoms were also recorded, with dominance of Syntrichia sp. (96.1%). Fungi were predominantly represented by Ascomycota, Zygomycota and Basidiomycota, fully half of the sampled biofilms where Fungi were detected. Comparing microbial communities from bleach-treated caves to those in untreated caves showed no significant difference except for a low-level change in the abundance of certain taxa. These findings provided by Illumina sequencing reveal a complex community structure in the 5 caves based on the assembly of bacteria, cyanobacteria, algae, diatoms, fungi and mosses.

The microbial diversity on the walls of the UNESCO World cultural Heritage site Laugerie-Haute east shelter, France, was surveyed in a context of degradation by formation of salt efflorescences. Samples of salt efflorescence and underlying rock were collected in autumn and spring. Sessile microorganisms were studied by cultivation, microscopy, and molecular methods. Concentrations of the order of 10 3 fungal and 10 7 bacterial cells per gram of sample were quantified by culturing at both seasons. Higher concentrations (10 7 to 10 9 cells per gram) of sessile microorganisms were determined by microscopic counting after DAPI staining with a significant increase in sessile cells in spring. The presence of biofilms was confirmed by electron microscopy with the observation of microorganisms embedded in an abundant matrix in association with rock. Members of Firmicutes, Actinobacteria, and Ascomycota were identified among the cultivable sessile microflora. Bacillus herbersteinensis and Engyodontium album were the only microorganisms to be isolated in both seasons. Sequence analysis of 16S rRNA genes and internal transcribed spacer clone libraries showed dominance of Bacteroidetes, Proteobacteria, and Ascomycota. Bacteroidetes and Actinobacteria were more abundant in autumn, and in spring, respectively. Cyanobacteria were present at both seasons. Bacterial richness increased during spring. Whatever the season, clones affiliated to the 4 bacterial genera Aureimonas, Sphingopyxis , Halomonas , and Crossiella , and to the two fungal genera Engyodontium and Teratosphaeria were present. In conclusion, we demonstrated the presence of abundant biofilm at efflorescence salt on the walls of the shelter; that was increased in quantity and diversity in spring.

The aim of the present study was to analyze the epilithic bacterial diversity of the ruins of the Chaalis Abbey, France. To do this, stone samples were collected at the surface of three areas presenting different macroscopic characteristics: a Discolored Area (DCA), a Damaged Area (DMA) and an Undamaged Area (UDMA). The bacterial diversity investigation after DNA extraction and amplified ribosomal DNA restriction analysis targeting the 16S rRNA genes revealed that clone libraries were dominated by Alphaproteobacteria and Actinobacteria . The main part of recovered phylotypes belonged to genera previously associated with stone colonization and/or biodeterioration. Indeed most abundant Alphaproteobacteria phylotypes belonged to the genus Sphingomonas and to the species Bosea thiooxidans , and most abundant Actinobacteria phylotypes belonged to the genera Rubrobacter and Arthrobacter . Members of Cyanobacteria , Bacteroidetes , TM7, Betaproteobacteria , Deinococcus , Acidobacteria and Chloroflexi were also recovered but to a lesser extent. In conclusion, these results suggest that the ruins of the Chaalis Abbey are covered by bacterial communities mainly composed of Alphaproteobacteria and Actinobacteria able to colonize and, for some of them, to deteriorate stone.

International audience; A laboratory investigation of UV-C effects was conducted over a 62-h period: a much higher dose than in classic UV-C treatment was applied to five pigments and two painting binders used by prehistoric humans. Colorimetric parameters were compared to a control to see if UV-C can change pigment and binder color. Infrared spectroscopy, scanning electron microscopy, inductively coupled plasma and X-ray crystallography were also carried out to confirm colorimetric measurement. In order to understand how microorganism may physically deteriorate paintings, limestone blocks were painted and monitored until their complete colonization by algae, cyanobacteria, fungi and/or mosses. The results show that UV-C has no effect on mineral compounds. Conversely, it is noteworthy that binder color changed under both UV-C light conditions as well as in visible light. Concerning painted blocks, a fast proliferation has been observed with deterioration of the paintings. These results show the high importance of treating biofilm as soon as possible. Moreover, these findings may be a promising avenue inducing cave managers to use friendly UV-C light to treat contaminated cave paintings and also in the prevention of biodeterioration by lampenflora.

EA BIOmE; International audience; Ultra-violet C (UV-C) treatment is commonly used in sterilization processes in industry, laboratories, and hospitals, showing its efficacy against microorganisms such as bacteria, algae, or fungi. In this study, we have eradicated for the first time all proliferating biofilms present in a show cave (theLa Glacière Cave, Chaux-lès-Passavant, France).Colorimetric measurements of irradiated biofilms were then monitored for 21 months. To understand the importance of exposition of algae to light just after UVradiation, similar tests were carried out in laboratory conditions. Since UV-C can be deleterious for biofilm support, especially parietal painting,we investigated their effects on prehistoric pigment. Results showed complete eradication of cave biofilms with no algae proliferation observed after 21 months. Moreover, quantum yield results showed a decrease directly after UV-C treatment, indicating inhibition of algae photosynthesis. Furthermore, nochanges in pigment color nor in chemical and crystalline properties has been demonstrated. The present findings demonstrate that the UV-C method can be considered environmentally friendly and the best alternative to chemicals. This inexpensive and easily implemented method is advantageousfor cave owners and managers.

International audience; Algal biofilms in caves visited by tourists are a significant threat to karstic formations. They cause a dirty greenish appearance on cave formations and provoke biophysical and biochemical erosion. Seeking an alternative to the use of chemicals, we investigated the effects of high doses of UV-C irradiation on the green algae Chlorella minutissima-dominating biofilms in the Moidons Cave (Jura, France). The algae were harvested and subjected to laboratory subculturing under conditions similar to the conditions in the cave (low temperatures and a short photoperiod with weak light intensities). After one or two doses of UV-C of 150 or 300 kJ m−2, the samples were incubated for 21 days, and the physiological parameters (pigment concentrations and photosynthetic activity) were monitored every seven days. Hydrogen peroxide (H2O2) and malondialdehyde (MDA) concentrations were also measured to detect oxidative stress, and analyses of DNA integrity were performed. The results showed that all UV-C treatments caused chlorophyll bleaching with completely inhibited metabolic activity. In addition, high UV-C doses caused oxidative stress and programmed cell death. Irradiation was tested on a natural biofilm that proliferated in the Moidons Cave. Colorimetric measurements of the treated biofilm indicated that UV-C treatment caused significant chlorophyll bleaching.

Algal growth is responsible for aesthetic defects on roofing tiles. Accelerated water-streaming tests were done on different building materials. The results establish the ranges of porosity and roughness that can initiate the colonization under humid saturation. A smooth surface is then recommended to reduce the settlement. Experiments done on limestone and clay tiles demonstrate that coupling high porosity and rough surface should be banished to avoid any rapid algal colonization. However, the alkaline composition of concrete tiles can strongly affect the algal settlement, whatever their intrinsic porosity and roughness. Observations by Environmental Scanning Electron Microscopy show that calcite crystals could promote the settlement of algae on the surface.

A major fungal invasion was discovered in the prehistoric painted cave of Lascaux in France in Sep 2001. At least three species of the Fusarium solani complex were isolated and identified with a portion of the translation elongation factor 1alpha gene (EF-1alpha), a portion of the nuclear large subunit rDNA (LSU) and nuclear ribosomal intergenic spacer region (ITS). This study represents the first time that Fusarium species have been reported from a cave containing prehistoric paintings. Significant interspecific molecular variability was observed, suggesting that there might have been repeated introduction of the species, possibly carried by water from soils above the cave.

In this work, three pieces of historical on-site glass windows dated from the 13th to 16th century and one archeological sample (8th century) showing Mn-rich brown spots at their surface or subsurface have been characterized by optical microscopy and Scanning Electron Microscopy coupled with Energy Dispersive X-ray spectroscopy. The oxidation state of Mn as well as the Mn environment in the alteration phase have been characterized by X-ray absorption spectroscopy at the Mn K-edge. Results show that the oxidation state of Mn and therefore the nature of the alteration phase varies according to the sample considered and is correlated with the extent of the brown alteration. The larger the brown areas the more oxidized the Mn. However, by contrast with literature, the samples presenting the more extended brown areas are not similar to pyrolusite and contain Mn mainly under a (+III) oxidation state.

Biodeterioration can be defined as any undesired change of the properties of a material caused by biological activity of living organisms. The biodeterioration of stone materials is related to the production of pigments (aesthetic action), to cell metabolism (biochemical action) and to the mechanical action of the biomass colonizing the material during its growth (physical action). Quantification of the sessile biomass and characterization of microbial communities colonizing stone are essential first steps to ensure the diagnosis of biodeterioration processes and to implement control strategies and appropriate treatment. Different destructive and non-destructive approaches can be used to sample stone specimens from monuments: scraping, swab using, and cutting. Different analytical methods can be used depending on the type of microorganism sought: determination of chlorophyll content and color analysis for pigmented microorganisms; measurement of in situ physiological activity of surface microcolonies by applying fluorogenic substrate analogues or confocal laser scanning microscopy observations after CTC staining for active biomass; scanning or transmission electron microscopy observation for biofilm visualization; enzyme-linked immunosorbent assay for the investigation of both microorganisms that can and cannot be cultured; classical microbiological methods, which consist in cultivation of microorganisms on synthetic media; and molecular methods for the study of microbial biodiversity based on the polymorphism of molecular markers using PCR, hybridization, classical or high throughput sequencing. The aim of this review is to present basics of the different biodeterioration mechanisms and to focus on the main techniques that can be used to characterize and quantify the biodeterioration biomass.

International audience; The proliferation of epilithic algae that form biofilms in subterranean environments, such as show caves, is a major problem for conservators. In an effort to reduce the use of chemical cleansers when addressing this problem, we proposed investigating the effects of UV-C on combating algal biofilm expansion in a cave located in northeastern France (Moidons Cave). First, the biofilms and cavity were studied in terms of their algal growth-influencing factors to understand the dynamics of colonization in these very harsh environments. Next, colorimetric measurements were used both to diagnose the initial colonization state and monitor the UV-C-treated biofilms for several months after irradiation. The results indicated that passive dispersal vectors of the viable spores and cells were the primary factors involved in the cave's algae repartition. The illumination time during visits appeared to be responsible for greater colonization in some parts of the cave. We also showed that colorimetric measurements could be used for the detection of both thin and thick biofilms, regardless of the type of colonized surface. Finally, our results showed that UV-C treatment led to bleaching of the treated biofilm due to chlorophyll degradation even one year after UV-C treatment. However, a re-colonization phenomenon was colorimetrically and visually detected 16 months later, suggesting that the colonization dynamics had not been fully halted.

International audience; Biological aerosols from air constitute a significant source of exposure to microorganisms in public places. Airborne microorganisms are involved in the development of certain respiratory symptoms, allergies, or infections among users and occupants. Various sampling instruments have commonly been used in aerobiology to collect bacteria and fungi suspended in the air. The objective of this study was to develop a reliable procedure for sampling in indoor public environments presenting different levels of occupancy, airborne bacteria and fungi to be subjected to molecular analysis (bacteria and fungi quantitative PCR, capillary electrophoresis single strand conformation polymorphism fingerprinting). Four different sampling devices were tested in situ in an office building (open-plan type) and the sampling strategy chosen was tested in two museum contexts. In accordance with the drawbacks involved to our study (quantitative and qualitative aspects, cost, and overcrowding), cyclone device appeared to be most suitable. The results underline the effectiveness of this high-volume aerosol sampling device for both qualitative and quantitative molecular analysis. Four in situ sampling collections were carried out in 1 day in the Louvre Museum to study quantitative and qualitative variations of airborne bacterial and fungal diversity. The quantitative results revealed a similar order of magnitude for the numbers of both bacteria and fungi. In the Louvre Museum, the samples yielded between 3.7x104 and 4.1x104 genome equivalent (GE) bacteria/m3 air and between 5.0x104 and 5.9x104 GE fungi/m3 air and in the Decorative Arts Museum between, 2.1x104 and 2.5x104 GE bacteria/m3 air and between 1.4x104 and 1.7x104 GE fungi/m3 air. The results also indicate that the dominant bacterial community displayed a stable structure over a short period of time whereas dominant eukaryotic airborne community appeared more variable.

International audience; Plus d’un tiers du parc immobilier français est contaminé par des moisissures [1]. Or cette croissance microbienne présente un risque avéré tant pour les occupants que pour les matériaux. En effet, ces biocontaminants et leurs métabolites peuvent être à l’origine de symptômes voire de pathologies (irritations, infections, toxi-infections, allergies) chez les personnes exposées. Concernant l’aspect biodétérioration des matériaux, de nombreux travaux ont mis en évidence le rôle des produits de construction et de décoration comme « niche écologique » fournissant, par leur composition même ou leur encrassement, les nutriments nécessaires au développement des moisissures. Au niveau microbiologique, après l’implantation ou bioadhésion de la particule biologique sur un substrat, phénomène de nature purement physico-chimique, le micromycète se développe au détriment du support qu’il dégrade grâce à son arsenal enzymatique constitutif et/ou induit. Dans ce contexte, l’objet de cette étude consiste à´etudier la dynamique de colonisation fongique sur ces supports afin d’identifier des leviers d’action permettant de ralentir voire limiter la croissance des moisissures sur les produits de construction et de décoration. À cet effet, la colonisation de produits, a été´etudiée en caractérisant les communautés microbiennes au cours du temps. Cette étude a permis de caractériser le processus de colonisation microbienne de deux produits, dans un contexte thermohydrique simulant une infiltration discontinue. Cette approche vient compléter la littérature axée essentiellement sur la description de la flore fongique d’un support au terme d’une infestation.

International audience; Current techniques are unable to detect either moulds at an early stage in their development or hidden contaminants. Thus most cases of fungal development are only detected once major damage to colonised substrates has been observed. In this context, the aim of the present study was to develop a methodology based on measuring the Volatile Organic Compounds (VOC) emitted by these micro-organisms and adapted to monitoring works of art in terms of the risks of mould development. This article describes the development of four specific indices. One of these indices was tested in situ during measurements performed in the prehistoric Lascaux caves contaminated by Fusarium solani, before and after the site was treated with biocidal agents. Preliminary tests were also carried out using polymer sensors which, in the laboratory, were able to discriminate between mould-affected and sterile environments.

International audience

Development of biofilm on monument heritage are often considered as agents leading to aesthetic issues and biodeterioration of their support. The aim of this study is the use of UV-C light as alternative treatment against Lampenflora. Bacterial, algal and fungal communities were studied using new generation sequencing approach. Following UV-C treatments, physiological responses of Chlorella sp. were studied in laboratory (cell death, chlorophyll degradation, photosystem II status, etc.). Damages at cellular and molecular levels (cell death, bleaching of the chlorophyll) were taken into consideration to optimize the UV-C treatment. Then, to ensure the safety of UV-C for biofilms support (parietal paintings), several pigments and binders, used by prehistoric human, were highly irradiated. The results showed that UV-C radiations were not deleterious for inorganic compounds, while a color change was observed on organic binders. Finally, UV-C treatment has proven its effectiveness in La Glacière show cave by eradicating all biofilms. In addition, no recolonization was observed two years after treatment, showing that this treatment, which is the most environmental friendly to date, is effective over time.; Dans le cadre de la conservation du patrimoine, les micro-organismes se développant sous forme de biofilm sont souvent considérés comme des agents entraînant des problèmes esthétiques et de détérioration du support colonisé. L’objectif général de cette thèse est l’utilisation de la lumière UV-C comme méthode de traitement des biofilms. Dans le but de comprendre la diversité des biofilms, les communautés bactériennes, fongiques et des protistes autotrophes ont été étudiées à l’aide d’une technique de séquençage haut débit. Puis, afin de comprendre les effets des UV-C sur les micro-algales (Chlorella sp.), des essais en laboratoire ont permis de caractériser les réponses physiologiques algales (mort cellulaire, dégradation de la chlorophylle, état de photosystème II, etc.) suite à différents traitements aux UV-C. Tous les dommages observés au niveau cellulaire et moléculaire (dont la mort cellulaire et le « bleaching » de la chlorophylle) ont permis d’optimiser la méthode de traitement. Dans le but de s’assurer de l’innocuité du traitement aux UV-C sur le support des biofilms (peintures pariétales), des pigments et des liants, utilisés à la préhistoire, ont été fortement irradiés. Les résultats ont montré que les composés inorganiques sont insensibles aux UV-C au contraire des molécules organiques. Enfin, le traitement UV-C a démontré son efficacité dans une grotte touristique en permettant d’éradiquer tous les biofilms. De plus, aucune recolonisation n’a été observée deux ans après le traitement, démontrant que la lumière UV-C est le traitement le plus écologique à ce jour.