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The efficiency and feasibility of geothermal utilisation depends strongly on the characteristics and behaviour of the fluids that transfer heat between the geosphere and the engineered components of a power plant. Chemical and physical processes such as precipitation, corrosion, or degassing are induced by pressure and temperature changes, with potentially serious consequences for power plant operation and project economics. The EU Horizon 2020-funded project REFLECT aims to avoid such problems by collecting high-quality chemical, physical, and microbiological data at extreme salinities, pressures or temperatures and improving the understanding of kinetic processes through laboratory experiments. These data are presented in a European geothermal fluid atlas and implemented in predictive models in order to provide recommendations on how to best operate geothermal systems for a sustainable future.

Evolutionary game theory has provided various models to explain the coexistence of competing strategies, one of which is the rock-paper-scissors (RPS) game. A system of threeEscherichia colistrains—a toxin-producer, a resistant, and a sensitive—has become a classic experimental model for studying RPS games. Previous experimental and theoretical studies, however, often ignored the influence of ecological factors such as nutrients and toxin dynamics on the evolutionary game dynamics. In this work, we combine experiments and modeling to study how these factors affect competition dynamics. Using 3D-printed mini-bioreactors, we tracked the frequency of the three strains in different culturing media and under different flow regimes. Although our experimental system fulfilled the requirements of cyclic dominance, we did not observe clear cycles or long-term coexistence between strains. We found that both nutrients and flow rates strongly impacted population dynamics. In our simulations, we explicitly modeled the release, removal and diffusion of toxin. We showed that the amount of toxin that is retained in the system is a simple indicator that can predict competition outcomes across broad parameter space. Moreover, our simulation results suggest that high rates of toxin diffusion might have prevented cyclic patterns from emerging in our experimental system.

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The coexistence of competing species is a long-lasting puzzle in evolutionary ecology research. Despite abundant experimental evidence showing that the opportunity for coexistence decreases as niche overlap increases between species, bacterial species and strains competing for the same resources are commonly found across diverse spatially heterogeneous habitats. We thus hypothesized that the spatial scale of competition may play a key role in determining bacterial coexistence, and interact with other mechanisms that promote coexistence, including a growth-motility tradeoff. To test this hypothesis, we let two Pseudomonas putida strains compete at local and regional scales by inoculating them either in a mixed droplet or in separate droplets in the same Petri dish, respectively. We also created conditions that allow the bacterial strains to disperse across abiotic or fungal hyphae networks. We found that competition at the local scale led to competitive exclusion while regional competition promoted coexistence. When competing in the presence of dispersal networks, the growth-motility tradeoff promoted coexistence only when the strains were inoculated in separate droplets. Our results provide a mechanism by which existing laboratory data suggesting competitive exclusion at a local scale is reconciled with the widespread coexistence of competing bacterial strains in complex natural environments with dispersal.

Bacterial endospores are highly specialized cellular forms that allow Endospore forming Firmicutes (EFF) to tolerate harsh environmental conditions. EFF are considered ubiquitous in natural environments in particular those subjected to stress conditions. In addition to natural habitats EFF are often the cause of contamination problems in anthropogenic environments such as industrial production plants or hospitals. It is therefore desirable to assess their prevalence in environmental and industrial fields. To this end a high sensitivity detection method is still needed. The aim of this study was to develop and evaluate an approach based on quantitative PCR. For this the suitability of functional genes specific for and common to all EFF were evaluated. Seven genes were considered but only spo0A was retained to identify conserved regions for qPCR primer design. An approach based on multivariate analysis was developed for primer design. Two primer sets were obtained and evaluated with 16 pure cultures including representatives of the genera Bacillus Paenibacillus Brevibacillus Geobacillus Alicyclobacillus Sulfobacillus Clostridium and Desulfotomaculum as well as with environmental samples. The primer sets developed gave a reliable quantification when tested on laboratory strains with the exception of Sulfobacillus and Desulfotomaculum. A test using sediment samples with a diverse EFF community also gave a reliable quantification compared to 16S rRNA gene pyrosequencing. A detection limit of about 104 cells (or spores) per gram of initial material was calculated given this method a promising potential for the detection of EFF in a wide range of applications.

While often considered as harmful for cultural heritage microorganisms can also be used for its safeguarding. Indeed there is a growing interest for the development of biological technologies that are environmental friendly (close to ambient temperature and pressure neutral pH) and do not require the use of toxic materials. A real progress could be expected in terms of durability effectiveness and toxicity. Over the last decades the development of biological methods and materials became a significant alternative for the conservation restoration of cultural artefacts. Hence biotechnology has been applied with success in different domains such as bioremediation or corrosion control. In particular different soil bacteria have already been reported as a conservation treatment for ornamental stone demonstrating the great opportunities offered by using such alternative treatments. This research topic will focus on two different areas: the microorganisms involved in biodegradation and control methods used to reduce the deterioration of cultural artefacts and the preservation and protection of cultural artefacts using fungi and bacteria. The aim is to achieve a comprehensive view on the role and potential of microorganisms pro and against degradation of cultural materials (stones metals graphic documents textiles paintings glasses…). Research papers case studies reviews perspectives and opinion papers on these themes are welcome.

Klebsiella oxytoca was stressed by incubation in seawater microcosms for ten years. Despite being introduced into sterile seawater, the added Klebsiella oxytoca declined to levels undetectable by plate counts on nutrient agar. After two years of starvation, no culturable strain was observed. Incubation of samples in nutrient-rich broth medium not supplemented with growth factors, however, allowed resuscitation of viable but non culturable (VBNC) cells so that subsequent plating yielded observable colonies for significantly extended periods of time. Recovery of VBNC Klebsiella oxytoca was obtained by incubation in nutrient broth even after ten years of starvation. To see whether the samples contain the same strain of Klebsiella oxytoca inoculated 10 years ago, the complete 16S rDNA gene was PCR amplified and sequenced from initial, stressed and revived strains of Klebsiella oxytoca. The 16S rDNA gene sequences from ten-year stressed strains were homologous in a high degree of similarity to the GenBank reference strain and were identical to each other.

Can microbes be used to preserve archaeological iron objects? Lucrezia Comensolia, J. Maillardb, P. Juniera and E. Josepha,c a Laboratory of Microbiology, Institute of Biology University of Neuchâtel, Switzerland b Laboratory for Environmental Biotechnology, Ecole Polytechnique Fédérale de Lausanne (EPFL), Switzerland c Haute Ecole Arc Conservation-Restauration, Neuchâtel, Switzerland lucrezia.comensoli@unine.ch Without any conservation-restauration intervention archaeological iron objects are affected by a dramatic corrosion that leads to an irreversible loss of shape. The main issue with this metallic substrate is the chlorine content of the corrosion layers, which, by reaction with H2O and O2, causes an irreversible deterioration of the objects once excavated. Conservation-restauration technics available nowadays are ineffective, too expensive, time consuming and employ toxic substances. Therefore, the “MAIA” project (Microbe for Archaeological Iron Artworks) aims to develop new conservation-restauration methods based on microorganisms. For this purpose, iron reduction and chlorine uptake/volatilization capacities have been studied in selected microorganisms. These two metabolic abilities will allow to actively remove chlorine from the chlorinated iron compounds present in the corrosion layer and also to produce more stable iron minerals. For this aim, two anaerobic bacterial strains of Desulfitobacterium hafniense (strains LBE and TCE1, reported for dehalorespiration) and several fungal strains were studied. Different iron sources were tested either as soluble (iron citrate) or solid-phase iron compounds (powdered iron compounds from real archaeological artefacts). Spectrophotometric analyses were carried out to ascertain iron reduction and the evolution of the bacterial growth was quantified by quantitative Polymerase Chain Reaction (qPCR). Scanning Electronic Microscopy allows studying the fungal absorption of iron and chlorine, Raman Spectroscopy was used to identify the bio-mineral produced and finally we performed the first bacterial treatment of real archaeological iron objects. The results obtained show that microbes have the potential to stabilize iron objects by both reduction into more stable minerals and by removing chlorine from the object.

Fungi and bacteria form various associations that are central to numerous environmental processes. In the so called fungal highway bacteria disperse along fungal mycelium. We developed a novel tool for the in situ isolation of bacteria moving along fungal hyphae as well as for the recovery of fungi potentially involved in dispersal both of which are attracted towards a target culture medium. We present the validation and the results of the first in situ test. Couples of fungi and bacteria were isolated from soil. Amongst the enriched organisms we identified several species of fast growing fungi (Fusarium sp. and Chaetomium sp.) as well as various potentially associated bacterial groups including Variovorax soli Olivibacter soli Acinetobacter calcoaceticus and several species of the genera Stenotrophomonas Achromobacter and Ochrobactrum. Migration of bacteria along fungal hyphae across a discontinuous medium was confirmed in most of the cases. Although the majority of the bacteria for which migration was confirmed were also positive for flagellar motility not all motile bacteria dispersed using their potential fungal partner. In addition the importance of hydrophobicity of the fungal mycelial surface was confirmed. Future applications of the columns include targeting different types of microorganisms and their interactions either by enrichment or by state of the art molecular biological methods.

While often considered as a nuisance and potentially harmful for cultural heritage microorganisms can also be used for its safeguarding. Indeed there is a growing interest for the development of biological technologies that are environmentally friendly (close to ambient temperature and pressure neutral pH) and that do not require the use of toxic materials. A real progress could be expected in terms of durability effectiveness and toxicity. Over the last decades the development of biological methods and materials became a significant alternative for the preservation of ancient heritage. In particular microbial metabolism is exploited aiming to consolidate clean stabilize or even protect surfaces of cultural items. This contribution intends to present an overview of the current research projects of the Laboratory of Microbiology of the University of Neuchâtel in the field of cultural heritage. Using the capabilities of specific bacteria and fungi the projects MAIA and BIOPATINA combine innovative aspects in biogeochemistry of microorganisms and conservation science. The MAIA project is dedicated to the development of a microbial desalination protocol for archaeological iron while the BIOPATINA project is directed towards proposing an alternative stabilization treatment for copper based surfaces. Achieved results and ongoing experiences will be presented here.

It has generally been assumed that the bioreduction of hexavalent uranium in groundwater systems will result in the precipitation of immobile uraninite (UO2). In order to explore the form and stability of uranium immobilized under these conditions, we introduced lactate (15 mM for 3 months) into flow-through columns containing sediments derived from a former uranium-processing site at Old Rifle, CO. This resulted in metal-reducing conditions as evidenced by concurrent uranium uptake and iron release. Despite initial augmentation with Shewanella oneidensis, bacteria belonging to the phylum Firmicutes dominated the biostimulated columns. The immobilization of uranium (similar to 1 mmol U per kg sediment) enabled analysis by Xray absorption spectroscopy (XAS). Tetravalent uranium associated with these sediments did not have spectroscopic signatures representative of U-U shells or crystalline UO2. Analysis by microfocused XAS revealed concentrated micrometer regions of solid U(IV) that had spectroscopic signatures consistent with bulk analyses and a poor proximal correlation (mu m scale resolution) between U and Fe. A plausible explanation, supported by biogeochemical conditions and spectral interpretations, is uranium association with phosphoryl moieties found in biomass; hence implicating direct enzymatic uranium reduction. After the immobilization phase, two months of in situ exposure to oxic influent did not result in substantial uranium remobilization. Ex situ flow-through experiments demonstrated more rapid uranium mobilization than observed in column oxidation studies and indicated that sediment-associated U(IV) is more mobile than biogenic UO2. This work suggests that in situ uranium bioimmobilization studies and subsurface modeling parameters should be expanded to account for non-uraninite U(IV) species associated with biomass. (C) 2011 Elsevier Ltd. All rights reserved.

The functional gene amoA was used to compare the diversity of ammonia-oxidizing bacteria (AOB) in the water column and sediment–water interface of the two freshwater lakes Plußsee and Schöhsee and the Baltic Sea. Nested amplifications were used to increase the sensitivity of amoA detection, and to amplify a 789-bp fragment from which clone libraries were prepared. The larger part of the sequences was only distantly related to any of the cultured AOB and is considered to represent new clusters of AOB within the Nitrosomonas/Nitrosospira group. Almost all sequences from the water column of the Baltic Sea and from 1-m depth of Schöhsee were related to different Nitrosospira clusters 0 and 2, respectively. The majority of sequences from Plußsee and Schöhsee were associated with sequences from Chesapeake Bay, from a previous study of Plußsee and from rice roots in Nitrosospira -like cluster A, which lacks sequences from Baltic Sea. Two groups of sequences from Baltic Sea sediment were related to clonal sequences from other brackish/marine habitats in the purely environmental Nitrosospira-like cluster B and the Nitrosomonas-like cluster. This confirms previous results from 16S rRNA gene libraries that indicated the existence of hitherto uncultivated AOB in lake and Baltic Sea samples, and showed a differential distribution of AOB along the water column and sediment of these environments.

A novel endospore forming bacterium designated strain GSsed3T was isolated from deposits clogging aboveground filters from the geothermal power plant of Gross Schoenebeck in Northern Germany. The novel isolate was Gram staining positive facultative anaerobe catalase positive and oxidase positive. Optimum growth occurred at 60 °C 0.5 (w/v) NaCl and pH 7 8. Analysis of the 16S rRNA sequence similarity indicated that strain GSsed3T belongs to the genus Anoxybacillus and showed 99.8 sequence similarity to Anoxybacillus rupiensis R270T 98.2 similarity to Anoxybacillus tepidamans GS5 97T 97.9 similarity to Anoxybacillus voinovskiensis TH13T 97.7 similarity to Anoxybacillus caldiproteolyticus DSM 15730T and 97.6 similarity to Anoxybacillus amylolyticus MR3CT. DNA DNA hybridization (DDH) indicated only 16 relatedness to A. rupiensis DSM 17127T. Furthermore DDH estimation based on genomes analysis indicated only 19.9 overall nucleotide similarity to A. amylolyticus DSM 15939T. The major respiratory menaquinone was MK 8. The polar lipid profile consisted of phosphatidylethanolamine phosphatidylglycerol diphosphatidylglycerol one unknown phosphoglycolipid and one unknown phospholipid. The predominant cellular fatty acids were iso C15:0 iso C17:0 C16:0 iso C16:0 and anteiso C17:0. The peptidoglycan type was A1? meso Dpm direct. The genomic DNA G+C content of the strain was 46.9 mol. The phenotypic genotypic and chemotaxonomic characterization indicated that strain GSsed3T differs from related species of the genus. Therefore strain GSsed3T is considered to be a novel species of the genus Anoxybacillus for which the name Anoxybacillus geothermalis sp. nov. is proposed. The type strain of Anoxybacillus geothermalis is GSsed3T ( = CCOS808T = ATCC BAA2555T).

Eukaryotic studies have shown considerable transcriptional variation among individuals from the same population. It has been suggested that natural variation in eukaryotic gene expression may have significant evolutionary consequences and may explain large-scale phenotypic divergence of closely related species, such as humans and chimpanzees (M.-C. King and A. C. Wilson, Science 188:107–116, 1975, http://dx.doi.org/10.1126/science.1090005; M. F. Oleksiak, G. A. Churchill, and D. L. Crawford, Nat Genet 32:261–266, 2002, http://dx.doi.org/10.1038/ng983). However, natural variation in gene expression is much less well understood in prokaryotic organisms. In this study, we used four sequenced strains of the marine bacterium Shewanella baltica to better understand the natural transcriptional divergence of a stratified prokaryotic population. We found substantial low-magnitude expressional variation among the four S. baltica strains cultivated under identical laboratory conditions. Collectively, our results indicate that transcriptional variation is an important factor for ecological speciation., In bacterial populations, subtle expressional differences may promote ecological specialization through the formation of distinct ecotypes. In a barrier-free habitat, this process most likely precedes population divergence and may predict speciation events. To examine this, we used four sequenced strains of the bacterium Shewanella baltica, OS155, OS185, OS195, and OS223, as models to assess transcriptional variation and ecotype formation within a prokaryotic population. All strains were isolated from different depths throughout a water column of the Baltic Sea, occupying different ecological niches characterized by various abiotic parameters. Although the genome sequences are nearly 100% conserved, when grown in the laboratory under standardized conditions, all strains exhibited different growth rates, suggesting significant expressional variation. Using the Ecotype Simulation algorithm, all strains were considered to be discrete ecotypes when compared to 32 other S. baltica strains isolated from the same water column, suggesting ecological divergence. Next, we employed custom microarray slides containing oligonucleotide probes representing the core genome of OS155, OS185, OS195, and OS223 to detect natural transcriptional variation among strains grown under identical conditions. Significant transcriptional variation was noticed among all four strains. Differentially expressed gene profiles seemed to coincide with the metabolic signatures of the environment at the original isolation depth. Transcriptional pattern variations such as the ones highlighted here may be used as indicators of short-term evolution emerging from the formation of bacterial ecotypes. IMPORTANCE Eukaryotic studies have shown considerable transcriptional variation among individuals from the same population. It has been suggested that natural variation in eukaryotic gene expression may have significant evolutionary consequences and may explain large-scale phenotypic divergence of closely related species, such as humans and chimpanzees (M.-C. King and A. C. Wilson, Science 188:107–116, 1975, http://dx.doi.org/10.1126/science.1090005; M. F. Oleksiak, G. A. Churchill, and D. L. Crawford, Nat Genet 32:261–266, 2002, http://dx.doi.org/10.1038/ng983). However, natural variation in gene expression is much less well understood in prokaryotic organisms. In this study, we used four sequenced strains of the marine bacterium Shewanella baltica to better understand the natural transcriptional divergence of a stratified prokaryotic population. We found substantial low-magnitude expressional variation among the four S. baltica strains cultivated under identical laboratory conditions. Collectively, our results indicate that transcriptional variation is an important factor for ecological speciation.