A concept for international societally relevant microbiology education and microbiology knowledge promulgation in society.

EXECUTIVE SUMMARY: Microbes are all pervasive in their distribution and influence on the functioning and well-being of humans, life in general and the planet. Microbially-based technologies contribute hugely to the supply of important goods and services we depend upon, such as the provision of food, medicines and clean water. They also offer mechanisms and strategies to mitigate and solve a wide range of problems and crises facing humanity at all levels, including those encapsulated in the sustainable development goals (SDGs) formulated by the United Nations. For example, microbial technologies can contribute in multiple ways to decarbonisation and hence confronting global warming, provide sanitation and clean water to the billions of people lacking them, improve soil fertility and hence food production and develop vaccines and other medicines to reduce and in some cases eliminate deadly infections. They are the foundation of biotechnology, an increasingly important and growing business sector and source of employment, and the centre of the bioeconomy, Green Deal, etc. But, because microbes are largely invisible, they are not familiar to most people, so opportunities they offer to effectively prevent and solve problems are often missed by decision-makers, with the negative consequences this entrains. To correct this lack of vital knowledge, the International Microbiology Literacy Initiative-the IMiLI-is recruiting from the global microbiology community and making freely available, teaching resources for a curriculum in societally relevant microbiology that can be used at all levels of learning. Its goal is the development of a society that is literate in relevant microbiology and, as a consequence, able to take full advantage of the potential of microbes and minimise the consequences of their negative activities. In addition to teaching about microbes, almost every lesson discusses the influence they have on sustainability and the SDGs and their ability to solve pressing problems of societal inequalities. The curriculum thus teaches about sustainability, societal needs and global citizenship. The lessons also reveal the impacts microbes and their activities have on our daily lives at the personal, family, community, national and global levels and their relevance for decisions at all levels. And, because effective, evidence-based decisions require not only relevant information but also critical and systems thinking, the resources also teach about these key generic aspects of deliberation. The IMiLI teaching resources are learner-centric, not academic microbiology-centric and deal with the microbiology of everyday issues. These span topics as diverse as owning and caring for a companion animal, the vast range of everyday foods that are produced via microbial processes, impressive geological formations created by microbes, childhood illnesses and how they are managed and how to reduce waste and pollution. They also leverage the exceptional excitement of exploration and discovery that typifies much progress in microbiology to capture the interest, inspire and motivate educators and learners alike. The IMiLI is establishing Regional Centres to translate the teaching resources into regional languages and adapt them to regional cultures, and to promote their use and assist educators employing them. Two of these are now operational. The Regional Centres constitute the interface between resource creators and educators-learners. As such, they will collect and analyse feedback from the end-users and transmit this to the resource creators so that teaching materials can be improved and refined, and new resources added in response to demand: educators and learners will thereby be directly involved in evolution of the teaching resources. The interactions between educators-learners and resource creators mediated by the Regional Centres will establish dynamic and synergistic relationships-a global societally relevant microbiology education ecosystem-in which creators also become learners, teaching resources are optimised and all players/stakeholders are empowered and their motivation increased. The IMiLI concept thus embraces the principle of teaching societally relevant microbiology embedded in the wider context of societal, biosphere and planetary needs, inequalities, the range of crises that confront us and the need for improved decisioning, which should ultimately lead to better citizenship and a humanity that is more sustainable and resilient. ABSTRACT: The biosphere of planet Earth is a microbial world: a vast reactor of countless microbially driven chemical transformations and energy transfers that push and pull many planetary geochemical processes, including the cycling of the elements of life, mitigate or amplify climate change (e.g., Nature Reviews Microbiology, 2019, 17, 569) and impact the well-being and activities of all organisms, including humans. Microbes are both our ancestors and creators of the planetary chemistry that allowed us to evolve (e.g., Life's engines: How microbes made earth habitable, 2023). To understand how the biosphere functions, how humans can influence its development and live more sustainably with the other organisms sharing it, we need to understand the microbes. In a recent editorial (Environmental Microbiology, 2019, 21, 1513), we advocated for improved microbiology literacy in society. Our concept of microbiology literacy is not based on knowledge of the academic subject of microbiology, with its multitude of component topics, plus the growing number of additional topics from other disciplines that become vitally important elements of current microbiology. Rather it is focused on microbial activities that impact us-individuals/communities/nations/the human world-and the biosphere and that are key to reaching informed decisions on a multitude of issues that regularly confront us, ranging from personal issues to crises of global importance. In other words, it is knowledge and understanding essential for adulthood and the transition to it, knowledge and understanding that must be acquired early in life in school. The 2019 Editorial marked the launch of the International Microbiology Literacy Initiative, the IMiLI. HERE, WE PRESENT: our concept of how microbiology literacy may be achieved and the rationale underpinning it; the type of teaching resources being created to realise the concept and the framing of microbial activities treated in these resources in the context of sustainability, societal needs and responsibilities and decision-making; and the key role of Regional Centres that will translate the teaching resources into local languages, adapt them according to local cultural needs, interface with regional educators and develop and serve as hubs of microbiology literacy education networks. The topics featuring in teaching resources are learner-centric and have been selected for their inherent relevance, interest and ability to excite and engage. Importantly, the resources coherently integrate and emphasise the overarching issues of sustainability, stewardship and critical thinking and the pervasive interdependencies of processes. More broadly, the concept emphasises how the multifarious applications of microbial activities can be leveraged to promote human/animal, plant, environmental and planetary health, improve social equity, alleviate humanitarian deficits and causes of conflicts among peoples and increase understanding between peoples (Microbial Biotechnology, 2023, 16(6), 1091-1111). Importantly, although the primary target of the freely available (CC BY-NC 4.0) IMiLI teaching resources is schoolchildren and their educators, they and the teaching philosophy are intended for all ages, abilities and cultural spectra of learners worldwide: in university education, lifelong learning, curiosity-driven, web-based knowledge acquisition and public outreach. The IMiLI teaching resources aim to promote development of a global microbiology education ecosystem that democratises microbiology knowledge.

Genome-wide transcriptional response to silver stress in extremely halophilic archaeon Haloferax alexandrinus DSM 27206 T.

BACKGROUND: The extremely halophilic archaeon Haloferax (Hfx.) alexandrinus DSM 27206 T was previously documented for the ability to biosynthesize silver nanoparticles while mechanisms underlying its silver tolerance were overlooked. In the current study, we aimed to assess the transcriptional response of this haloarchaeon to varying concentrations of silver, seeking a comprehensive understanding of the molecular determinants underpinning its heavy metal tolerance. RESULTS: The growth curves confirmed the capacity of Hfx. alexandrinus to surmount silver stress, while the SEM-EDS analysis illustrated the presence of silver nanoparticles in cultures exposed to 0.5 mM silver nitrate. The RNA-Seq based transcriptomic analysis of Hfx. alexandrinus cells exposed to 0.1, 0.25, and 0.5 mM silver nitrate revealed the differential expression of multiple sets of genes potentially employed in heavy-metal stress response, genes mostly related to metal transporters, basic metabolism, oxidative stress response and cellular motility. The RT-qPCR analysis of selected transcripts was conducted to verify and validate the generated RNA-Seq data. CONCLUSIONS: Our results indicated that copA, encoding the copper ATPase, is essential for the survival of Hfx. alexandrinus cells in silver-containing saline media. The silver-exposed cultures underwent several metabolic adjustments that enabled the activation of enzymes involved in the oxidative stress response and impairment of the cellular movement capacity. To our knowledge, this study represents the first comprehensive analysis of gene expression in halophillic archaea facing increased levels of heavy metals.

Karst spring microbiome: Diversity, core taxa, and community response to pathogens and antibiotic resistance gene contamination.

Karst aquifers are important water resources for drinking water supplies worldwide. Although they are susceptible to anthropogenic contamination due to their high permeability, there is a lack of detailed knowledge on the stable core microbiome and how contamination may affect these communities. In this study, eight karst springs (distributed across three different regions in Romania) were sampled seasonally for one year. The core microbiota was analysed by 16S rRNA gene amplicon sequencing. To identify bacteria carrying antibiotic resistance genes and mobile genetic elements, an innovative method was applied, consisting of high-throughput antibiotic resistance gene quantification performed on potential pathogen colonies cultivated on Compact Dry™ plates. A taxonomically stable bacterial community consisting of Pseudomonadota, Bacteroidota, and Actinomycetota was revealed. Core analysis reaffirmed these results and revealed primarily freshwater-dwelling, psychrophilic/psychrotolerant species affiliated to Rhodoferax, Flavobacterium, and Pseudomonas genera. Both sequencing and cultivation methods indicated that more than half of the springs were contaminated with faecal bacteria and pathogens. These samples contained high levels of sulfonamide, macrolide, lincosamide and streptogramins B, and trimethoprim resistance genes spread primarily by transposase and insertion sequences. Differential abundance analysis found Synergistota, Mycoplasmatota, and Chlamydiota as suitable candidates for pollution monitoring in karst springs. This is the first study highlighting the applicability of a combined approach based on high-throughput SmartChip™ antibiotic resistance gene quantification and Compact Dry™ pathogen cultivation for estimating microbial contaminants in karst springs and other challenging low biomass environments.

Description of two cultivated and two uncultivated new Salinibacter species, one named following the rules of the bacteriological code: Salinibacter grassmerensis sp. nov.; and three named following the rules of the SeqCode: Salinibacter pepae sp. nov., Salinibacter abyssi sp. nov., and Salinibacter pampae sp. nov.

Current -omics methods allow the collection of a large amount of information that helps in describing the microbial diversity in nature. Here, and as a result of a culturomic approach that rendered the collection of thousands of isolates from 5 different hypersaline sites (in Spain, USA and New Zealand), we obtained 21 strains that represent two new Salinibacter species. For these species we propose the names Salinibacter pepae sp. nov. and Salinibacter grassmerensis sp. nov. (showing average nucleotide identity (ANI) values < 95.09% and 87.08% with Sal. ruber M31T, respectively). Metabolomics revealed species-specific discriminative profiles. Sal. ruber strains were distinguished by a higher percentage of polyunsaturated fatty acids and specific N-functionalized fatty acids; and Sal. altiplanensis was distinguished by an increased number of glycosylated molecules. Based on sequence characteristics and inferred phenotype of metagenome-assembled genomes (MAGs), we describe two new members of the genus Salinibacter. These species dominated in different sites and always coexisted with Sal. ruber and Sal. pepae. Based on the MAGs from three Argentinian lakes in the Pampa region of Argentina and the MAG of the Romanian lake Fara Fund, we describe the species Salinibacter pampae sp. nov. and Salinibacter abyssi sp. nov. respectively (showing ANI values 90.94% and 91.48% with Sal. ruber M31T, respectively). Sal. grassmerensis sp. nov. name was formed according to the rules of the International Code for Nomenclature of Prokaryotes (ICNP), and Sal. pepae, Sal. pampae sp. nov. and Sal. abyssi sp. nov. are proposed following the rules of the newly published Code of Nomenclature of Prokaryotes Described from Sequence Data (SeqCode). This work constitutes an example on how classification under ICNP and SeqCode can coexist, and how the official naming a cultivated organism for which the deposit in public repositories is difficult finds an intermediate solution.

Diversity, distribution and organic substrates preferences of microbial communities of a low anthropic activity cave in North-Western Romania.

Introduction: Karst caves are characterized by relatively constant temperature, lack of light, high humidity, and low nutrients availability. The diversity and functionality of the microorganisms dwelling in caves micro-habitats are yet underexplored. Therefore, in-depth investigations of these ecosystems aid in enlarging our understanding of the microbial interactions and microbially driven biogeochemical cycles. Here, we aimed at evaluating the diversity, abundance, distribution, and organic substrate preferences of microbial communities from Peștera cu Apa din Valea Leșului (Leșu Cave) located in the Apuseni Mountains (North-Western Romania). Materials and Methods: To achieve this goal, we employed 16S rRNA gene amplicon sequencing and community-level physiological profiling (CLPP) paralleled by the assessment of environmental parameters of cave sediments and water. Results and Discussion: Pseudomonadota (synonym Proteobacteria) was the most prevalent phylum detected across all samples whereas the abundance detected at order level varied among sites and between water and sediment samples. Despite the general similarity at the phylum-level in Leșu Cave across the sampled area, the results obtained in this study suggest that specific sites drive bacterial community at the order-level, perhaps sustaining the enrichment of unique bacterial populations due to microenvironmental conditions. For most of the dominant orders the distribution pattern showed a positive correlation with C-sources such as putrescine, γ-amino butyric acid, and D-malic acid, while particular cases were positively correlated with polymers (Tween 40, Tween 80 and α-cyclodextrin), carbohydrates (α-D-lactose, i-erythritol, D-mannitol) and most of the carboxylic and ketonic acids. Physicochemical analysis reveals that sediments are geochemically distinct, with increased concentration of Ca, Fe, Al, Mg, Na and K, whereas water showed low nitrate concentration. Our PCA indicated the clustering of different dominant orders with Mg, As, P, Fe, and Cr. This information serves as a starting point for further studies in elucidating the links between the taxonomic and functional diversity of subterranean microbial communities.

Competition-cooperation in the chemoautotrophic ecosystem of Movile Cave: first metagenomic approach on sediments.

BACKGROUND: Movile Cave (SE Romania) is a chemoautotrophically-based ecosystem fed by hydrogen sulfide-rich groundwater serving as a primary energy source analogous to the deep-sea hydrothermal ecosystems. Our current understanding of Movile Cave microbiology has been confined to the sulfidic water and its proximity, as most studies focused on the water-floating microbial mat and planktonic accumulations likely acting as the primary production powerhouse of this unique subterranean ecosystem. By employing comprehensive genomic-resolved metagenomics, we questioned the spatial variation, chemoautotrophic abilities, ecological interactions and trophic roles of Movile Cave's microbiome thriving beyond the sulfidic-rich water. RESULTS: A customized bioinformatics pipeline led to the recovery of 106 high-quality metagenome-assembled genomes from 7 cave sediment metagenomes. Assemblies' taxonomy spanned 19 bacterial and three archaeal phyla with Acidobacteriota, Chloroflexota, Proteobacteria, Planctomycetota, Ca. Patescibacteria, Thermoproteota, Methylomirabilota, and Ca. Zixibacteria as prevalent phyla. Functional gene analyses predicted the presence of CO2 fixation, methanotrophy, sulfur and ammonia oxidation in the explored sediments. Species Metabolic Coupling Analysis of metagenome-scale metabolic models revealed the highest competition-cooperation interactions in the sediments collected away from the water. Simulated metabolic interactions indicated autotrophs and methanotrophs as major donors of metabolites in the sediment communities. Cross-feeding dependencies were assumed only towards 'currency' molecules and inorganic compounds (O2, PO43-, H+, Fe2+, Cu2+) in the water proximity sediment, whereas hydrogen sulfide and methanol were assumedly traded exclusively among distant gallery communities. CONCLUSIONS: These findings suggest that the primary production potential of Movile Cave expands way beyond its hydrothermal waters, enhancing our understanding of the functioning and ecological interactions within chemolithoautotrophically-based subterranean ecosystems.

Asgard archaea in saline environments.

Members of candidate Asgardarchaeota superphylum appear to share numerous eukaryotic-like attributes thus being broadly explored for their relevance to eukaryogenesis. On the contrast, the ecological roles of Asgard archaea remains understudied. Asgard archaea have been frequently associated to low-oxygen aquatic sedimentary environments worldwide spanning a broad but not extreme salinity range. To date, the available information on diversity and potential biogeochemical roles of Asgardarchaeota mostly sourced from marine habitats and to a much lesser extend from true saline environments (i.e., > 3% w/v total salinity). Here, we provide an overview on diversity and ecological implications of Asgard archaea distributed across saline environments and briefly explore their metagenome-resolved potential for osmoadaptation. Loki-, Thor- and Heimdallarchaeota are the dominant Asgard clades in saline habitats where they might employ anaerobic/microaerophilic organic matter degradation and autotrophic carbon fixation. Homologs of primary solute uptake ABC transporters seemingly prevail in Thorarchaeota, whereas those putatively involved in trehalose and ectoine biosynthesis were mostly inferred in Lokiarchaeota. We speculate that Asgardarchaeota might adopt compatible solute-accumulating ('salt-out') strategy as response to salt stress. Our current understanding on the distribution, ecology and salt-adaptive strategies of Asgardarchaeota in saline environments are, however, limited by insufficient sampling and incompleteness of the available metagenome-assembled genomes. Extensive sampling combined with 'omics'- and cultivation-based approaches seem, therefore, crucial to gain deeper knowledge on this particularly intriguing archaeal lineage.

Intact ribosomal DNA arrays of Potentilla origin detected in Erythronium nucleus suggest recent eudicot-to-monocot horizontal transfer.

During our initial phylogenetic study of the monocot genus Erythronium (Liliaceae), we observed peculiar eudicot-type internal transcribed spacer (ITS) sequences in a dataset derived from genomic DNA of Erythronium dens-canis. This raised the possibility of horizontal transfer of a eudicot alien ribosomal DNA (rDNA) into the Erythronium genome. In this work we aimed to support this hypothesis by carrying out genomic, molecular, and cytogenetic analyses. Genome skimming coupled by PacBio HiFi sequencing of a bacterial artificial chromosome clone derived from flow-sorted nuclei was used to characterise the alien 45S rDNA. Integration of alien rDNA in the recipient genome was further proved by Southern blotting and fluorescence in situ hybridization using specific probes. Alien rDNA, nested among Potentilla species in phylogenetic analysis, likely entered the Erythronium lineage in the common ancestor of E. dens-canis and E. caucasicum. Transferred eudicot-type rDNA preserved its tandemly arrayed feature on a single chromosome and was found to be transcribed in the monocot host, albeit much less efficiently than the native counterpart. This study adds a new example to the rarely documented nuclear-to-nuclear jumps of DNA between eudicots and monocots while holding the scientific community continually in suspense about the mode of DNA transfer.

Comprehensive mineralogical and physicochemical characterization of recent sapropels from Romanian saline lakes for potential use in pelotherapy.

In this study we aimed to compare the mineralogical, thermal, physicochemical, and biological characteristics of recent organic carbon-rich sediments ('sapropels') from three geographically distant Romanian lakes (Tekirghiol and Amara, SE Romania, and Ursu, Central Romania) with distinct hydrogeochemical origins, presently used for pelotherapy. The investigated lakes were classified as inland brackish Na-Cl-sulfated type (Amara), coastal moderately saline and inland hypersaline Na-Cl types (Tekirghiol and Ursu, respectively). The settled organic matter is largely composed of photosynthetic pigments derived from autochthonous phytoplankton. Kerogen was identified in the sapropel of coastal Tekirghiol Lake suggesting its incipient maturation stage. The mineral composition was fairly similar in all sapropels and mainly consisted of quartz, calcite, and aragonite. Smectite, illite, mixed layer smectite/illite appeared as major clay components. Potentially toxic elements were found in low concentrations. The physical properties (i.e., specific heat, thermal conductivity and retentivity) and cation exchange capacity are comparable to other peloids used for therapy. This study is the first comprehensive multi-approached investigation of the geochemical nature of recent sapropels in Romanian saline lakes and thus contributes to expanding our knowledge on the origin and physicochemical qualities of organic matter-rich peloids with therapeutic uses.

Mitochondrial DNA Profiles of Individuals from a 12th Century Necropolis in Feldioara (Transylvania).

The genetic signature of modern Europeans is the cumulated result of millennia of discrete small-scale exchanges between multiple distinct population groups that performed a repeated cycle of movement, settlement, and interactions with each other. In this study we aimed to highlight one such minute genetic cycle in a sea of genetic interactions by reconstructing part of the genetic story of the migration, settlement, interaction, and legacy of what is today the Transylvanian Saxon. The analysis of the mitochondrial DNA control region of 13 medieval individuals from Feldioara necropolis (Transylvania region, Romania) reveals a genetically heterogeneous group where all identified haplotypes are different. Most of the perceived maternal lineages are of Western Eurasian origin, except for the Central Asiatic haplogroup C seen in only one sample. Comparisons with historical and modern populations describe the contribution of the investigated Saxon settlers to the genetic history of this part of Europe.

Spatio-temporal insights into microbiology of the freshwater-to-hypersaline, oxic-hypoxic-euxinic waters of Ursu Lake.

Ursu Lake is located in the Middle Miocene salt deposit of Central Romania. It is stratified, and the water column has three distinct water masses: an upper freshwater-to-moderately saline stratum (0-3 m), an intermediate stratum exhibiting a steep halocline (3-3.5 m), and a lower hypersaline stratum (4 m and below) that is euxinic (i.e. anoxic and sulphidic). Recent studies have characterized the lake's microbial taxonomy and given rise to intriguing ecological questions. Here, we explore whether the communities are dynamic or stable in relation to taxonomic composition, geochemistry, biophysics, and ecophysiological functions during the annual cycle. We found: (i) seasonally fluctuating, light-dependent communities in the upper layer (≥0.987-0.990 water-activity), a stable but phylogenetically diverse population of heterotrophs in the hypersaline stratum (water activities down to 0.762) and a persistent plate of green sulphur bacteria that connects these two (0.958-0.956 water activity) at 3-3.5 to 4 m; (ii) communities that might be involved in carbon- and sulphur-cycling between and within the lake's three main water masses; (iii) uncultured lineages including Acetothermia (OP1), Cloacimonetes (WWE1), Marinimicrobia (SAR406), Omnitrophicaeota (OP3), Parcubacteria (OD1) and other Candidate Phyla Radiation bacteria, and SR1 in the hypersaline stratum (likely involved in the anaerobic steps of carbon- and sulphur-cycling); and (iv) that species richness and habitat stability are associated with high redox-potentials. Ursu Lake has a unique and complex ecology, at the same time exhibiting dynamic fluctuations and stability, and can be used as a modern analogue for ancient euxinic water bodies and comparator system for other stratified hypersaline systems.

Visualization of Lokiarchaeia and Heimdallarchaeia (Asgardarchaeota) by Fluorescence In Situ Hybridization and Catalyzed Reporter Deposition (CARD-FISH).

Metagenome-assembled genomes (MAGs) of Asgardarchaeota have been recovered from a variety of habitats, broadening their environmental distribution and providing access to the genetic makeup of this archaeal lineage. The recent success in cultivating the first representative of Lokiarchaeia was a breakthrough in science at large and gave rise to new hypotheses about the evolution of eukaryotes. Despite their singular phylogenetic position at the base of the eukaryotic tree of life, the morphology of these bewildering organisms remains a mystery, except for the report of an unusual morphology with long, branching protrusions of the cultivated Lokiarchaeion strain "Candidatus Prometheoarchaeum syntrophicum" MK-D1. In order to visualize this elusive group, we applied a combination of fluorescence in situ hybridization and catalyzed reporter deposition (CARD-FISH) and epifluorescence microscopy on coastal hypersaline sediment samples, using specifically designed CARD-FISH probes for Heimdallarchaeia and Lokiarchaeia lineages, and provide the first visual evidence for Heimdallarchaeia and new images of a lineage of Lokiarchaeia that is different from the cultured representative. Here, we show that while Heimdallarchaeia are characterized by a uniform cellular morphology typified by a centralized DNA localization, Lokiarchaeia display a plethora of shapes and sizes that likely reflect their broad phylogenetic diversity and ecological distribution.IMPORTANCE Asgardarchaeota are considered to be the closest relatives to modern eukaryotes. These enigmatic microbes have been mainly studied using metagenome-assembled genomes (MAGs). Only very recently, a first member of Lokiarchaeia was isolated and characterized in detail; it featured a striking morphology with long, branching protrusions. In order to visualize additional members of the phylum Asgardarchaeota, we applied a fluorescence in situ hybridization technique and epifluorescence microscopy on coastal hypersaline sediment samples, using specifically designed probes for Heimdallarchaeia and Lokiarchaeia lineages. We provide the first visual evidence for Heimdallarchaeia that are characterized by a uniform cellular morphology typified by an apparently centralized DNA localization. Further, we provide new images of a lineage of Lokiarchaeia that is different from the cultured representative and with multiple morphologies, ranging from small ovoid cells to long filaments. This diversity in observed cell shapes is likely owing to the large phylogenetic diversity within Asgardarchaeota, the vast majority of which remain uncultured.

Schizorhodopsins: A family of rhodopsins from Asgard archaea that function as light-driven inward H+ pumps.

Schizorhodopsins (SzRs), a rhodopsin family first identified in Asgard archaea, the archaeal group closest to eukaryotes, are present at a phylogenetically intermediate position between typical microbial rhodopsins and heliorhodopsins. However, the biological function and molecular properties of SzRs have not been reported. Here, SzRs from Asgardarchaeota and from a yet unknown microorganism are expressed in Escherichia coli and mammalian cells, and ion transport assays and patch clamp analyses are used to demonstrate SzR as a novel type of light-driven inward H+ pump. The mutation of a cytoplasmic glutamate inhibited inward H+ transport, suggesting that it functions as a cytoplasmic H+ acceptor. The function, trimeric structure, and H+ transport mechanism of SzR are similar to that of xenorhodopsin (XeR), a light-driven inward H+ pumping microbial rhodopsins, implying that they evolved convergently. The inward H+ pump function of SzR provides new insight into the photobiological life cycle of the Asgardarchaeota.

Physiological response to silver toxicity in the extremely halophilic archaeon Halomicrobium mukohataei.

Adaptive strategies responsible for heavy metal tolerance were explored in the extremely halophilic archaeon Halomicrobium mukohataei DSM 12286. The tested strain was seemingly able to overcome silver-induced oxidative stress (assessed by malondialdehyde quantification, catalase assay and total antioxidant capacity measurement) mainly through non-enzymatic antioxidants. Energy dispersive spectrometry analysis illustrated the presence of colloidal silver in Hmc. mukohataei cultures exposed to AgNO3. Bright-field and transmission electron microscopy images, as well as dynamic light scattering analysis, demonstrated the presence of intracellular nanoparticles, mostly spherical, within a size range of 20-100 nm. As determined by the zeta potential measurement, the biosynthesized nanoparticles were highly stable, with a negative surface charge. Our research is a first attempt in the systematic study of the oxidative stress and intracellular silver nanoparticle accumulation, generated by exposure to silver ions, in members of Halobacteria class, thus broadening our knowledge on mechanisms supporting heavy metal tolerance of microbial cells living under saline conditions.

Casting light on Asgardarchaeota metabolism in a sunlit microoxic niche.

Recent advances in phylogenomic analyses and increased genomic sampling of uncultured prokaryotic lineages have brought compelling evidence in support of the emergence of eukaryotes from within the archaeal domain of life (eocyte hypothesis)1,2. The discovery of Asgardarchaeota and its supposed position at the base of the eukaryotic tree of life3,4 provided cues about the long-awaited identity of the eocytic lineage from which the nucleated cells (Eukaryota) emerged. While it is apparent that Asgardarchaeota encode a plethora of eukaryotic-specific proteins (the highest number identified yet in prokaryotes)5, the lack of genomic information and metabolic characterization has precluded inferences about their lifestyles and the metabolic landscape that favoured the emergence of the protoeukaryote ancestor. Here, we use advanced phylogenetic analyses for inferring the deep ancestry of eukaryotes, and genome-scale metabolic reconstructions for shedding light on the metabolic milieu of Asgardarchaeota. In doing so, we: (1) show that Heimdallarchaeia (the closest eocytic lineage to eukaryotes to date) are likely to have a microoxic niche, based on their genomic potential, with aerobic metabolic pathways that are unique among Archaea (that is, the kynurenine pathway); (2) provide evidence of mixotrophy within Asgardarchaeota; and (3) describe a previously unknown family of rhodopsins encoded within the recovered genomes.

Ecology and physiology of halophilic microorganisms - Thematic issue based on papers presented at Halophiles 2019 - 12th International Conference on Halophilic Microorganisms, Cluj-Napoca, Romania, 24-28 June, 2019.

In June 2019, the 12th International Conference on Halophilic Microorganisms - Halophiles 2019, was held in Cluj-Napoca, Romania. This thematic issue of FEMS Microbiology Letters contains papers based on lectures and posters presented at the conference. We here provide a short overview of past research on hypersaline environments in Romania and the microorganisms inhabiting them, and briefly present the papers published in this thematic issue.

NaCl-saturated brines are thermodynamically moderate, rather than extreme, microbial habitats.

NaCl-saturated brines such as saltern crystalliser ponds, inland salt lakes, deep-sea brines and liquids-of-deliquescence on halite are commonly regarded as a paradigm for the limit of life on Earth. There are, however, other habitats that are thermodynamically more extreme. Typically, NaCl-saturated environments contain all domains of life and perform complete biogeochemical cycling. Despite their reduced water activity, ∼0.755 at 5 M NaCl, some halophiles belonging to the Archaea and Bacteria exhibit optimum growth/metabolism in these brines. Furthermore, the recognised water-activity limit for microbial function, ∼0.585 for some strains of fungi, lies far below 0.755. Other biophysical constraints on the microbial biosphere (temperatures of >121°C; pH > 12; and high chaotropicity; e.g. ethanol at >18.9% w/v (24% v/v) and MgCl2 at >3.03 M) can prevent any cellular metabolism or ecosystem function. By contrast, NaCl-saturated environments contain biomass-dense, metabolically diverse, highly active and complex microbial ecosystems; and this underscores their moderate character. Here, we survey the evidence that NaCl-saturated brines are biologically permissive, fertile habitats that are thermodynamically mid-range rather than extreme. Indeed, were NaCl sufficiently soluble, some halophiles might grow at concentrations of up to 8 M. It may be that the finite solubility of NaCl has stabilised the genetic composition of halophile populations and limited the action of natural selection in driving halophile evolution towards greater xerophilicity. Further implications are considered for the origin(s) of life and other aspects of astrobiology.

Hypersaline sapropels act as hotspots for microbial dark matter.

Present-day terrestrial analogue sites are crucial ground truth proxies for studying life in geochemical conditions close to those assumed to be present on early Earth or inferred to exist on other celestial bodies (e.g. Mars, Europa). Although hypersaline sapropels are border-of-life habitats with moderate occurrence, their microbiological and physicochemical characterization lags behind. Here, we study the diversity of life under low water activity by describing the prokaryotic communities from two disparate hypersaline sapropels (Transylvanian Basin, Romania) in relation to geochemical milieu and pore water chemistry, while inferring their role in carbon cycling by matching taxa to known taxon-specific biogeochemical functions. The polyphasic approach combined deep coverage SSU rRNA gene amplicon sequencing and bioinformatics with RT-qPCR and physicochemical investigations. We found that sapropels developed an analogous elemental milieu and harbored prokaryotes affiliated with fifty-nine phyla, among which the most abundant were Proteobacteria, Bacteroidetes and Chloroflexi. Containing thirty-two candidate divisions and possibly undocumented prokaryotic lineages, the hypersaline sapropels were found to accommodate one of the most diverse and novel ecosystems reported to date and may contribute to completing the phylogenetic branching of the tree of life.

Diversity and Biomineralization Potential of the Epilithic Bacterial Communities Inhabiting the Oldest Public Stone Monument of Cluj-Napoca (Transylvania, Romania).

In this study, we investigated the biomineralization potential and diversity of the epilithic bacterial communities dwelling on the limestone statue of Saint Donatus, the oldest public monument of Cluj-Napoca city (Transylvania region, NW Romania). Their spatial distribution together with phylogenetic and metabolic diversity, as well as their capacity to precipitate calcium carbonate was evaluated by combining molecular and phenotypic fingerprinting methods with X-ray diffraction, Fourier transform infrared spectroscopy, and scanning electron-microscopy analyses. The results of real-time quantitative PCR, molecular fingerprinting and community-level physiological profiling showed that diverse and abundant bacterial assemblages that differ in relation to their collection site colonized the statue. The cultivation and molecular identification procedures allowed the characterization of 79 bacterial isolates belonging to Proteobacteria (73.4%), Firmicutes (19%), and Actinobacteria (7.6%). Amongst them, the 22 strains identified as being capable of calcium carbonate precipitation were found to belong mostly to Bacillus and Pseudomonas genera. We found that bacteria acted as nucleation sites, inducing the formation of nanoscale aggregates that were shown to be principally composed of vaterite. Furthermore, we expanded the current knowledge on culturable diversity of carbonatogenic bacteria by providing evidence for biogenic vaterite/calcite formation mediated by: Pseudomonas synxantha, P. graminis, Brevibacterium iodinum, Streptomyces albidoflavus, and Stenotrophomonas chelatiphaga. Overall, this study highlights the need to evaluate the carbonatogenetic potential of all the bacterial communities present on stone artwork prior to designing an efficient conservation treatment based on biomineralization.

Mismatch repair deficiency increases the transfer of antibiosis and antixenosis properties against Colorado potato beetle in somatic hybrids of Solanum tuberosum + S. chacoense.

BACKGROUND: Colorado potato beetle (CPB) has become the biggest enemy of cultivated potato worldwide. One of the most effective sources of resistance to CPB is Solanum chacoense, an accession with a high leptine glycoalkaloid content. The aim of our study was to assay the repellence and toxicity of S. chacoense, its somatic hybrids (SHs) and their backcross progenies (BC1 ) with potato for CPB adults and larvae. Transgenic S. chacoense, deficient in DNA mismatch repair (MMR), was also used to produce SHs, in order to increase homeologous recombination and hence introgression of wild-species DNA into the potato gene pool. RESULTS: Wild-type SH was highly resistant to CPB. Resistance to CPB of BC1 progenies showed a 1:3 inheritance pattern. MMR-deficient SHs performed better in the resistance analysis. Most MMR-deficient SHs had a similar toxicity as S. chacoense and an intensely repellent effect on CPB adults. Resistance of SHs and BC1 clones may be attributed to leptine biosynthesis, which was confirmed using a RAPD marker. CONCLUSION: This is the first report of SHs and their progenies exhibiting both antibiosis and antixenosis against CPB. Resistant SHs are an important step forward in combating this voracious pest of potato. © 2016 Society of Chemical Industry.