Differential effects of tree species identity on rhizospheric bacterial and fungal community richness and composition across multiple trace element-contaminated sites.

Soil microbial communities play a key role in plant nutrition and stress tolerance. This is particularly true in sites contaminated by trace metals, which often have low fertility and stressful conditions for woody plants in particular. However, we have limited knowledge of the abiotic and biotic factors affecting the richness and composition of microbial communities inhabiting the rhizosphere of plants in contaminated sites. Using high-throughput amplicon sequencing, we studied the rhizospheric bacterial and fungal community structures of 14 woody plant families planted in three contrasting sites contaminated by metals (Pb, Cd, Zn, Mn, Fe, S). The rhizospheric bacterial communities in the given sites showed no significant difference between the various woody species but did differ significantly between sites. The Proteobacteria phylum was dominant, accounting for over 25 % of the overall relative abundance, followed by Actinobacteria, Bacteroidetes and Gemmatimonadetes. Site was also the main driver of fungal community composition, yet unlike bacteria, tree species identity significantly affected fungal communities. The Betulaceae, Salicaceae and Fagaceae families had a high proportion of Basidiomycota, particularly ectomycorrhizal fungi, and the lowest diversity and richness. The other tree families and the unplanted soil harboured a greater abundance of Ascomycota and Mucoromycota. Consequently, for both bacteria and fungi, the site effect significantly impacted their community richness and composition, while the influence of plants on the richness and composition of rhizospheric microbial communities stayed consistent across sites and was dependent on the microbial kingdom. Finally, we highlighted the importance of considering this contrasting response of plant rhizospheric microbial communities in relation to their host identity, particularly to improve assisted revegetation efforts at contaminated sites.

Mycorrhizal inoculation effects on growth and the mycobiome of poplar on two phytomanaged sites after 7-year-short rotation coppicing.

Aims: Afforestation of trace-element contaminated soils, notably with fast growing trees, has been demonstrated to be an attractive option for bioremediation due to the lower costs and dispersion of contaminants than conventional cleanup methods. Mycorrhizal fungi form symbiotic associations with plants, contributing to their tolerance towards toxic elements and actively participating to the biorestoration processes. The aim of this study was to deepen our understanding on the effects of mycorrhizal inoculation on plant development and fungal community at two trace-element contaminated sites (Pierrelaye and Fresnes-sur-Escaut, France) planted with poplar (Populus trichocarpa x Populus maximowiczii). Methods: The 2 sites were divided into 4 replicated field blocks with a final plant density of 2200 tree h-1. Half of the trees were inoculated with a commercial inoculum made of a mix of mycorrhizal species. The sites presented different physico-chemical characteristics (e.g., texture: sandy soil versus silty-loam soil and organic matter: 5.7% versus 3.4% for Pierrelaye and Fresnes-sur-Escaut, respectively) and various trace element contamination levels. Results: After 7 years of plantation, inoculation showed a significant positive effect on poplar biomass production at the two sites. Fungal composition study demonstrated a predominance of the phylum Ascomycota at both sites, with a dominance of Geopora Arenicola and Mortierella elongata, and a higher proportion of ectomycorrhizal and endophytic fungi (with the highest values observed in Fresnes-sur-Escaut: 45% and 28% for ECM and endophytic fungi, respectively), well known for their capacity to have positive effects on plant development in stressful conditions. Furthermore, Pierrelaye site showed higher frequency (%) of mycorrhizal tips for ectomycorrhizal fungi (ECM) and higher intensity (%) of mycorrhizal root cortex colonization for arbuscular mycorrhizal fungi (AMF) than Fresnes-sur-Escaut site, which translates in a higher level of diversity. Conclusions: Finally, this study demonstrated that this biofertilization approach could be recommended as an appropriate phytomanagement strategy, due to its capacity to significantly improve poplar productivity without any perturbations in soil mycobiomes.

Factors inducing bryophyte growth on prehistoric pigments and effect of UV-C treatment.

In La Glacière cave (France), the touristic activity has been conducted to an environmental parameter change that has led to photosynthetic 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 the colonization 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, bone char). Microorganisms and bryophyte growth were monitored during 1 year, and then, the block samples were treated using UV-C light (254 nm). Thus, obtained results were compared with in situ treatment in La Glacière cave. Results have showed dense bryophyte propagation on the several blocks. However, the growth rate was correlated with the chemical composition of the pigment. 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 has been observed for samples maintained in laboratory. This difference was explained by the high bryophyte density under laboratory conditions that make UV-C light penetration difficult.

Bacterial diversity on an abandoned, industrial wasteland contaminated by polychlorinated biphenyls, dioxins, furans and trace metals.

Most former industrial sites are contaminated by mixtures of trace elements and organic pollutants. Levels of pollutants do not provide information regarding their biological impact, bioavailability and possible interactions between substances. There is genuine interest in combining chemical analyses with biological investigations. We studied a brownfield where several industrial activities were carried out starting in the 1970s, (incineration of pyralene transformers, recovery of copper by burning cables in the open air). Four representative plots showing different levels of polychlorobiphenyls were selected. Organic and trace metal levels were measured together with soil pedological characteristics. The bacterial community structure and functional diversity were assessed by 16S metagenomics with deep sequencing and community-level physiological profiling. Additionally, a vegetation survey was performed. Polychlorobiphenyls (8 mg.kg-1 to 1500 mg.kg-1) were from 2.4 × 103-fold to 6 × 105-fold higher than the European background level of 2.5 µg.kg-1. Polychlorinated dibenzo-p-dioxins and dibenzofurans ranged from 0.5 to 8.0 µg.kg-1. The soil was also contaminated with trace metals, i.e., Cu > 187, Zn > 217 and Pb > 372 mg.kg-1. Location within the study area, trace metal content and soil humidity were stronger determinants than organic pollutants of bacterial community structures and activities. Thus, the highest biological activity and the greatest bacteriological richness were observed in the plot that was less contaminated with trace metals, despite the high level of organic pollutants in the plot. Moreover, trace element pollution was associated with a relatively low presence of Actinobacteria and Rhizobia. The plot with the highest metal contamination was rich in metal-resistant bacteria such as Sphingomonadales, Geodermatophilaceae and KD4-96 (Chloroflexi phylum). Acidobacteria and Sphingomonadales, capable of resisting trace metals and degrading persistent organic pollutants, were dominant in the plots that had accumulated metal and organic contamination, but bacterial activity was lower in these plots than in the other plots.

Fungal and bacterial outbreak in the wine vinification area in the Saint-Marcel show cave.

In the Saint-Marcel cave (France), wood barrels and thousands of bottles containing red wine were stored for vinification. After storage began, a fungal and bacterial outbreak occurred, and the area was invaded by numerous types of mold colonizing the cave ceilings and all objects related to human activities (the stairwell and oenological materials). In this study, using the metabarcoding approach, we have studied the microbial outbreak and have linked the identified microorganisms to oenological activity. Both 16S and ITS primers were used to sequence the samples collected from the cave. The results showed that the dominant microorganisms proliferating in the cave were related to wine vinification. For instance, Zasmidium cellare, a strain known for living in dark and ethanol-rich environments, was the dominant fungus on the cave stairwell. Furthermore, Guehomyces pullulans, a cold-adapted yeast used for juice clarification, was recorded as the major species on the blackened limestone ceilings. These findings reveal a complex community structure in the studied cave based on the assembly of bacteria and fungi. Finally, our results demonstrate that oenological activities could seriously affect cave preservation, changing the natural microbial communities populating cave environments.

Poplar rotation coppice at a trace element-contaminated phytomanagement site: A 10-year study revealing biomass production, element export and impact on extractable elements.

Growing lignocellulosic crops on marginal lands could compose a substantial proportion of future energy resources. The potential of poplar was explored, by devising a field trial of two hectares in 2007 in a metal-contaminated site to quantify the genotypic variation in the growth traits of 14 poplar genotypes grown in short-rotation coppice and to assess element transfer and export by individual genotypes. Our data led us to conclusions about the genotypic variations in poplar growth on a moderately contaminated site, with the Vesten genotype being the most productive. This genotype also accumulated the least amounts of trace elements, whereas the Trichobel genotype accumulated up to 170 mg Zn kg-1 DW in the branches, with large variation being exhibited among the genotypes for trace element (TE) accumulation. Soil element depletion occurred for a range of TEs, whereas the soil content of major nutrients and the pH remained unchanged or slightly increased after 10 years of poplar growth. The higher TE content of bark tissues compared with the wood and the higher proportion of bark in branches compared with the wood led us to recommend that only stem wood be harvested, instead of the whole tree, which will enable a reduction in the risks encountered with TE-enriched biomass in the valorization process.

Assessment of fungi proliferation and diversity in cultural heritage: Reactions to UV-C treatment.

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 Glacière 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.

Biofilm biodiversity in French and Swiss show caves using the metabarcoding approach: First data.

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.

Bleaching of biofilm-forming algae induced by UV-C treatment: a preliminary study on chlorophyll degradation and its optimization for an application on cultural heritage.

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 Glacière 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.

UV-C as an efficient means to combat biofilm formation in show caves: evidence from the La Glacière Cave (France) and laboratory experiments.

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 (the La 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 UV radiation, 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, no changes 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 advantageous for cave owners and managers.

UV-C as a means to combat biofilm proliferation on prehistoric paintings: evidence from laboratory experiments.

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.