Cultural heritage objects constitute a very diverse environment, inhabited by various bacteria and fungi. The impact of these microorganisms on the degradation of artworks is undeniable, but at the same time, some of them may be applied for the efficient biotreatment of cultural heritage assets. Interventions with microorganisms have been proven to be useful in restoration of artworks, when classical chemical and mechanical methods fail or produce poor or short-term effects. The path to understanding the impact of microbes on historical objects relies mostly on multidisciplinary approaches, combining novel meta-omic technologies with classical cultivation experiments, and physico-chemical characterization of artworks. In particular, the development of metabolomic- and metatranscriptomic-based analyses associated with metagenomic studies may significantly increase our understanding of the microbial processes occurring on different materials and under various environmental conditions. Moreover, the progress in environmental microbiology and biotechnology may enable more effective application of microorganisms in the biotreatment of historical objects, creating an alternative to highly invasive chemical and mechanical methods.

The Lubin underground mine, is one of three mining divisions in the Lubin-Glogow Copper District in Lower Silesia province (Poland). It is the source of polymetallic ore that is rich in copper, silver and several heavy metals. Black shale is also significantly enriched in fossil organic matter in the form of long-chain hydrocarbons, polycyclic aromatic hydrocarbons, organic acids, esters, thiophenes and metalloporphyrins. Biological analyses have revealed that this environment is inhabited by extremophilic bacteria and fungi. Kupfershiefer black shale and samples of water, bottom and mineral sediments from the underground (below 600 m) Lubin mine were taken and twenty bacterial strains were isolated and characterized. All exhibited multi-resistant and hypertolerant phenotypes to heavy metals. We analyzed the plasmidome of these strains in order to evaluate the diversity and role of mobile DNA in adaptation to the harsh conditions of the mine environment. Experimental and bioinformatic analyses of 11 extrachromosomal replicons were performed. Three plasmids, including a broad-host-range replicon containing a Tn3 family transposon, carried genes conferring resistance to arsenic, cadmium, cobalt, mercury and zinc. Functional analysis revealed that the resistance modules exhibit host specificity, i.e. they may increase or decrease tolerance to toxic ions depending on the host strain. The other identified replicons showed diverse features. Among them we identified a catabolic plasmid encoding enzymes involved in the utilization of histidine and vanillate, a putative plasmid-like prophage carrying genes responsible for NAD biosynthesis, and two repABC-type plasmids containing virulence-associated genes. These findings provide an unique molecular insight into the pool of extrachromosomal replicons and highlight their role in the biology and adaptation of extremophilic bacteria inhabiting terrestrial deep subsurface.

Although interactions between microorganisms involved in biogas production are largely uncharted, it is commonly accepted that methanogenic Archaea are essential for the process. Methanogens thrive in various environments, but the most extensively studied communities come from biogas plants. In this study, we employed a metagenomic analysis of deeply sequenced methanogenic communities, which allowed for comparison of taxonomic and functional diversity as well as identification of microorganisms directly involved in various stages of methanogenesis pathways. A comprehensive metagenomic approach was used to compare seven environmental communities, originating from an agricultural biogas plant, cattle-associated samples, a lowland bog, sewage sludge from a wastewater treatment plant and sediments from an ancient gold mine. In addition to the native consortia, two laboratory communities cultivated on maize silage as the sole substrate were also analyzed. Results showed that all anaerobic communities harbored genes of all known methanogenesis pathways, but their abundance varied greatly between environments and that genes were encoded by different methanogens. Identification of microorganisms directly involved in different stages of methane production revealed that hydrogenotrophic methanogens, such as Methanoculleus, Methanobacterium, Methanobrevibacter, Methanocorpusculum or Methanoregula, predominated in most native communities, whereas acetoclastic Methanosaeta seemed to be the key methanogen in the wastewater treatment plant. Furthermore, in many environments, the methylotrophic pathway carried out by representatives of Methanomassiliicoccales, such as Candidatus Methanomethylophilus and Candidatus Methanoplasma, seemed to play an important role in methane production. In contrast, in stable laboratory reactors substrate versatile Methanosarcina predominated. The metagenomic approach presented in this study allowed for deep exploration and comparison of nine environments in which methane production occurs. Different abundance of methanogenesis-related functions was observed and the functions were analyzed in the phylogenetic context in order to identify microbes directly involved in methane production. In addition, a comparison of two metagenomic analytical tools, MG-RAST and MetAnnotate, revealed that combination of both allows for a precise characterization of methanogenic communities.

A well-balanced microbial consortium is crucial for efficient biogas production. In turn, one of a major factor that influence on the structure of anaerobic digestion consortium is a source of microorganisms which are used as an inoculum. This study evaluated the influence of inoculum sources (with various origin) on adaptation of a biogas community and the efficiency of the biomethanization of maize silage. As initial inocula for anaerobic digestion of maize silage the samples from: (i) an agricultural biogas plant (ABP) which utilizes maize silage as a main substrate, (ii) cattle slurry (CS), which contain elevated levels of lignocelluloses materials, and (iii) raw sewage sludge (RSS) with low content of plant origin materials were used. The adaptation of methanogenic consortia was monitored during a series of passages, and the functionality of the adapted consortia was verified through start-up operation of anaerobic digestion in two-stage reactors. During the first stages of the adaptation phase, methanogenic consortia occurred very slowly, and only after several passages did the microbial community adapts to allow production of biogas with high methane content. The ABP consortium revealed highest biogas production in the adaptation and in the start-up process. The biodiversity dynamics monitored during adaptation and start-up process showed that community profile changed in a similar direction in three studied consortia. Native communities were very distinct to each other, while at the end of the Phase II of the start-up process microbial diversity profile was similar in all consortia. All adopted bacterial communities were dominated by representatives of Porphyromonadaceae, Rikenellaceae, Ruminococcaceae and Synergistaceae. A shift from low acetate-preferring acetoclastic Methanosaetaceae (ABP and RSS) and/or hydrogenotrophic Archaeae, e.g. Methanomicrobiaceae (CS) prevailing in the inoculum samples to larger populations of high acetate-preferring acetoclastic Methanosarcinaceae was observed by the end of the experiment. As a result, three independent, functional communities that syntrophically produced methane from acetate (primarily) and H2/CO2, methanol and methylamines were adapted. This study provides new insights into the specific process by which different inocula sampled from typical methanogenic environments that are commonly used to initiate industrial installations gradually adapted to allow biogas production from maize silage.

Background: Halomonas sp. ZM3 was isolated from Zelazny Most post-flotation mineral waste repository (Poland), which is highly contaminated with heavy metals and various organic compounds. Mobile DNA of the strain (i.e. plasmids and transposons) were analyzed in order to identify genetic information enabling adaptation of the bacterium to the harsh environmental conditions. Results: The analysis revealed that ZM3 carries plasmid pZM3H1 (31,370 bp), whose replication system may be considered as an archetype of a novel subgroup of IncU-like replicons. pZM3H1 is a narrow host range, mobilizable plasmid (encodes a relaxase of the MOB_V family) containing mercury resistance operon (mer) and czcD genes (mediate resistance to zinc and cobalt), which are part of a large truncated Tn3 family transposon. Further analysis demonstrated that the phenotypes determined by the pZM3H1 resistance cassette are highly dependent on the host strain. In another strand of the study, the trap plasmid pMAT1 was employed to identify functional transposable elements of Halomonas sp. ZM3. Using the sacB positive selection strategy two insertion sequences were identified: ISHsp1 - representing IS5 group of IS5 family and ISHsp2 - a distinct member of the IS630 family. Conclusions: This study provides the first detailed description of mobile DNA in a member of the family Halomonadaceae. The identified IncU plasmid pZM3H1 confers resistance phenotypes enabling adaptation of the host strain to the Zelazny Most environment. The extended comparative analysis has shed light on the distribution of related IncU plasmids among bacteria, which, in many cases, reflects the frequency and direction of horizontal gene transfer events. Our results also identify plasmid-encoded modules, which may form the basis of novel shuttle vectors, specific for this group of halophilic bacteria.