Fossil Geyserite and Testate Amoebae in Geothermal Spring Vent Pools: Paleoecology and Variable Preservation Quality in Jurassic Sinter of Patagonia (Deseado Massif, Argentina).

Geyserite is a type of terrestrial siliceous hot spring deposit (sinter) formed subaerially in proximal vent areas, with near-neutral pH, alkali chloride discharge fluids characterized by initial high temperatures (~73°C to up to 100°C) that fluctuate rapidly in relation to dynamic hydrology, seasonality, wind, and other environmental parameters. We analyzed sinters at the Claudia paleogeothermal field from the Late Jurassic (~150 Ma) Deseado Massif geological province, Argentinean Patagonia. The geyserite samples-with spicular to columnar to nodular morphologies-contain abundant microfossils in monotypic assemblages that occur in three diagenetic states of preservation. The best-preserved microfossils consist of vesicle-like structures with radial heteropolar symmetry (~35 µm average diameter), circular apertures, smooth walls lacking ornamentation, and disk- or beret-like shapes. Comparisons with extant, morphologically similar organisms suggest an affinity with the testate amoebae of the Arcella hemisphaerica-Arcella rotundata complex and Centropyxis aculeata strain discoides. These species occur in active geothermal pools between 22°C and 45°C, inconsistent with the temperature of formation of modern geyserites. We propose that the testate amoebae may have colonized the geyserite during cooler phases in between spring-vent eruptive cycles to prey on biofilms. Silica precipitation through intermittent bathing and splashing of fluctuating thermal fluid discharge could have led to their entrapment and fossilization. Petrographic analysis supports cyclicity in paleovent water eruptions and later diagenesis that transformed the opal into quartz. Spatially patchy degradation and modification of the silicified microorganisms resulted in variable preservation quality of the microfossils. This contribution illustrates the importance of microscale analysis to locate early silicification and identify high-quality preservation of fossil remains in siliceous hot spring deposits, which are important in early life studies on Earth and potentially Mars.

Metagenomics insight into Puga geothermal geyser located in Himalayan Geothermal Belt (Trans-Himalayan Plateau) Ladakh, India.

Puga geothermal geyser and surrounding area, located in the Himalayan Geothermal Belt of the Trans-Himalayan Plateau in Ladakh, India, are very geographically isolated and considered pristine and free of anthropogenic activities. In this study, we have conducted the first metagenomic investigation of the microbes in and around the geyser. The whole genome sequencing analysis showed the presence of a total of 44.8%, 39.7% and 41.4% bacterial phyla in the PugW, PugS, and PugSo samples respectively, 8.6% of archaeal phyla (in all the samples), unclassified (derived from other sequences, PugW: 27.6%, PugS: 27.6%, and PugSo: 15.5%) and unclassified (derived from bacteria, PugW: 12%, PugS: 13.8%, and PugSo: 13.8%). The majority of archaeal sequences were linked to Euryarchaeota (2.84%) while the majority of the bacterial communities that predominated in most geothermal locations were linked to Pseudomonadota (67.14%) and Bacteroidota (12.52%). The abundant bacterial strains at the species level included Dechloromonas aromatica, Acinetobacter baumannii, and Arcobacter butzleri, in all the samples while the most abundant archaeal species were Methanosaeta thermophile, Methanoregula boonei, and Methanosarcina berkeri. Further, this geothermal geyser metagenome has a large number of unique sequences linked to unidentified and unclassified lineages, suggesting a potential source for novel species of microbes and their products. The present study which only examined one of the many geothermal geysers and springs in the Puga geothermal area, should be regarded as a preliminary investigation of the microbiota that live in the geothermal springs on these remote areas. These findings suggest that further investigations should be undertaken to characterize the ecosystems of the Puga geothermal area, which serve as a repository for unidentified microbial lineages.

Deciphering the microbial communities of alkaline hot spring in Panamik, Ladakh, India using a high-throughput sequencing approach.

Due to their distinctive physicochemical characteristics, hot springs are extremely important. The whole genome metagenomic sequencing technology can be utilized to analyze the diverse microbial community that thrives in this habitat due to the particular selection pressure that prevails there. The current investigation emphasizes on culture-independent metagenomic study of the Panamik hot spring and its nearby areas from Ladakh, India. Based on different diversity indices, sequence analysis of the soil reservoir showed higher species richness and diversity in comparison to water and sediment samples. The mineral content and various physicochemical pameters like temperature, pH had an impact on the composition of the microbial community of the geothermal springs. The phyla Proteobacteria, Cyanobacteria, Bacteroidetes, Actinobacter, Firmicutes, and Verrucomicrobia in bacterial domain dominate the thermos-alkaline spring at Panamik in different concentrations. Economically significant microbes from the genera Actinobacter, Thermosynechoccus, Candidatus Solibacter, Chthoniobacter, Synechoccus, Pseudomonas and Sphingomonas, were prevalent in hot spring. In the archaeal domain, the most dominant phylum and genera were Euryarchaeota and Thermococcus in all the samples. Further, the most abundant species were Methanosarcina barkeri, Nitrospumilus maritimus and Methanosarcina acetivorans. The present study which only examined one of the several thermal springs present in the Himalayan geothermal area, should be regarded as a preliminary investigation of the microbiota that live in the hot springs on these remote areas. These findings suggest that further investigations should be undertaken to characterize the ecosystems of the Panamik hot spring, which serve as a repository for unidentified microbial lineages.

Microbial diversity analysis of Chumathang geothermal spring, Ladakh, India.

In light of their unique and challenging environment, the high-altitude Chumathang geothermal springs in Ladakh, India, are undeniably intriguing for microbiological study. The purpose of this study was to employ a culture-independent sequencing approach to give a comprehensive characterization of the unknown bacterial and archaeal community structure, composition and networks in water and soil from the Chumathang geothermal spring. A total of 50%, and 42.86% bacterial phyla were found in the water, and soil samples respectively and this analysis also showed a total of 9.62% and 7.94% of archaeal phyla in both the samples, respectively. Further, the presence of unclassified (derived from other sequences, water: 17.31%, and soil: 19.05%) and unclassified (derived from bacteria, water: 13.46%, and soil: 12.70%) were also observed in the current metagenomics investigation. Firmicutes and Proteobacteria were the most abundant bacterial phyla in water, whereas Proteobacteria and Bacteroidetes were the most abundant bacterial phyla in geothermal soil. Crenarchaeota and Euryarchaeota dominated archeal communities in soil and water, respectively. This metagenomic study gave a detailed insight into the microbial diversity found in Chumathang geothermal spring and surrounding area, located in Ladakh, India. Surprisingly, this finding indicated the existence of geographically distinct microbial communities that were suited to various geothermal water habitats along the Himalayan Geothermal Belt. Future studies must take into account the metabolic pathways of these microbial communities that exist in these extreme environments. This will allow us to obtain a better knowledge of the microbial metabolisms that are common at these geothermal locations, which have a lot of potential for biotechnological applications. They will also enable us to establish links between the microbial community composition and the physicochemical environment of geothermal water and area.

Hyperthermophile diversity microbes in the Calientes geothermal field, Tacna, Peru.

Hyperthermophile microorganisms have been discovered worldwide, and several studies regarding biodiversity and the potential biotechnological applications have been reported. In this work, we describe for the first time the diversity of hyperthermophile communities in the Calientes Geothermal Field (CGF) located 4400 m above sea level in Tacna Region, Perú. Three hot springs were monitored and showed a temperature around 84 to 88 °C, for the microbiome analyzed was taken by sampling of sediment and water (pH 7.3-7.6). The hyperthermophile diversity was determined by PCR, DGGE, and DNA sequencing. The sediments analyzed showed a greater diversity than water samples. Sediments showed a more abundant population of bacteria than archaea, with the presence of at least 9 and 5 phylotypes, respectively. Most interestingly, in some taxa of bacteria (Bacillus) and archaea (Haloarcula and Halalkalicoccus), any of operational taxonomic units (OTUs) have not been observed before in hyperthermophile environments. Our results provide insight in the hyperthermophile diversity and reveal the possibility to develop new biotechnological applications based on the kind of environments.

Morphogenesis of digitate structures in hot spring silica sinters of the El Tatio geothermal field, Chile.

In silica-rich hot spring environments, internally laminated, digitate sinter deposits are often interpreted as bio-mediated structures. The organic components of microbial communities (cell surfaces, sheaths and extracellular polymeric substances) can act as templates for silica precipitation, therefore influencing digitate sinter morphogenesis. In addition to biologic surface-templating effects, various microenvironmental factors (hydrodynamics, local pH and fluctuating wind patterns) can also influence silica precipitation, and therefore the morphology of resulting digitate sinters. Digitate sinter morphology thus depends on the dynamic interplay between microenvironmentally driven silica precipitation and microbial growth, but the relative contributions of both factors are a topic of continuing research. Here we present a detailed study of digitate silica sinters in distal, low-temperature regimes of the El Tatio geothermal field, Chile. This high-altitude geothermal field is extremely arid and windy, and has one of the highest silica precipitation rates found in the world. We find that digitate silica sinters at El Tatio always accrete into the prevailing eastward wind direction and exhibit laminar growth patterns coinciding with day-night cycles of wind- and thermally driven evaporation and rewetting. Subaerial parts of digitate sinters lack preserved organics and sinter textures that would indicate past microbial colonization, while filamentous cyanobacteria with resistant, silicified sheaths only inhabit subaqueous cavities that crosscut the primary laminations. We conclude that, although fragile biofilms of extremophile micro-organisms may have initially been present and templated silica precipitation at the tips of these digitate sinters, the saltation of sand grains and precipitation of silica by recurrent wind- and thermally driven environmental forcing at El Tatio are important, if not dominant factors shaping the morphology of these digitate structures. Our study sheds light on the relative contributions of biogenic and abiogenic factors in sinter formation in geothermal systems, with geobiological implications for the cautious interpretation of stromatolite-like features in ancient silica deposits on Earth and Mars.

Novel hyperthermophilic crenarchaeon Infirmifilum lucidum gen. nov. sp. nov., reclassification of Thermofilum uzonense as Infirmifilum uzonense comb. nov. and assignment of the family Thermofilaceae to the order Thermofilales ord. nov.

A novel hyperthermophilic crenarchaeon, strain 3507LTT, was isolated from a terrestrial hot spring near Tinguiririca volcano, Chile. Cells were non-motile thin, slightly curved filamentous rods. It grew at 73-93 °C and pH range of 5 to 7.5 with an optimum at 85 °C and pH 6.0-6.7. The presence of culture broth filtrate of another hyperthemophilic archaeon as well as yeast extract was obligatory for growth of the novel isolate. Strain 3507LTT is an anaerobic chemoorganoheterotroph, fermenting monosaccharides, disaccharides and polysaccharides (lichenan, starch, xanthan gum, xyloglucan, alpha-cellulose and amorphous cellulose). No growth stimulation was detected when nitrate, thiosulfate, selenate or elemental sulfur were added as the electron acceptors. The complete genome of strain 3507LTT consisted of a single circular chromosome with size of 1.63 Mbp. The DNA G+C content was 53.9%. According to the 16S rRNA gene sequence as well as conserved protein sequences phylogenetic analyses, strain 3507LTT together with Thermofilum uzonense formed a separate cluster within a Thermofilaceae family (Thermoproteales/Thermoprotei/Crenarchaeota). Based on phenotypic characteristics, phylogeny as well as AAI comparisons, a novel genus and species Infirmifilum lucidum strain 3507LTT (=VKM B-3376T = KCTC 15797T) gen. nov. sp. nov. was proposed. Its closest relative, Thermofilum uzonense strain 1807-2T should be reclassified as Infirmifilum uzonense strain 1807-2T comb. nov. Finally, based on phylogenomic and comparative genome analyses of representatives of Thermofilaceae family and other representatives of Thermoproteales order, a proposal of transfer of the family Thermofilaceae into a separate order Thermofilales ord. nov. was made.

Screening and Characterization of Bacillus Strains Producing Highly Thermostable Amylase from Various Hot Springs of Algeria

HIGHLIGHTS Screening extremophile Bacillus strains from various Hot Springs Characterization Of Bacillus Strains Producing Highly Thermostable Amylase Genetic identification of the best strains

Abstract Currently thermostable Amylase represents a broad biotechnological interest and desired by a various industries. In this study, forty-six bacterial strains belonging to the genus Bacillus were isolated from various hot springs in the North West of Algeria based on their ability to degrade starch and produce amylase in Starch Agar medium. The majority of isolates showed a positive amylolytic activity. In order to select the most thermostables amylase the effect of temperature on enzymes was estimated, therefore the study of amylase thermostability was culminated by the selection of Four Strains having an interesting optimum of activity and range of stability, reaching 75 °C for the strains HBH1-2, HBH1-3, HBH3-1and 85 °C for the strain HC-2, This indicates that the Enzyme produced by retained strains have optimum activity at high temperature. The identity of the selected strains was established on the basis of the morphological, biochemical characteristics and phylogenetic position as determined by 16S Ribosomal DNA gene sequencing. The whole strains belonged to the genus Bacillus and their phylogeny were also reported in this study.

Diversity and enzymatic potential of thermophilic bacteria associated with terrestrial hot springs in Algeria.

This study aims to determine the diversity of culturable thermophilic bacteria isolated from eight terrestrial hot springs in Northeastern of Algeria using the conventional methods, SDS-PAGE fingerprinting of whole-cell proteins and 16S rRNA gene sequencing. In addition, their hydrolytic enzyme activities were also investigated. A total of 293 strains were isolated from the hot springs' water and sediment using different culture media. Overall, five distinct bacterial groups were characterized by whole-cell protein pattern analysis. Based on the 16S rRNA gene sequencing of 100 selected strains, the isolates were assigned to the following three major phyla: Firmicutes (93%), Deinococcus-Thermus (5%), and Actinobacteria (2%), which included 27 distinct species belonging to 12 different phylotypes, Aeribacillus, Aneurinibacillus, Anoxybacillus, Bacillus, Brevibacillus, Geobacillus, Laceyella, Meiothermus, Saccharomonospora, Thermoactinomyces, Thermobifida, and Thermus. The screening for nine extracellular enzymes showed that 65.87% of the isolates presented at least five types of enzyme activities, and 6.48% of strains combined all tested enzymes (amylase, cellulase, pectinase, esculinase, protease, gelatinase, lipase, lecithinase, and nuclease). It was found that Bacillus, Anoxybacillus, Aeribacillus, and Aneurinibacillus were the genera showing the highest activities. Likewise, the study showed an abundant and diverse thermophilic community with novel taxa presenting a promising source of thermozymes with important biotechnological applications. This study showed that a combined identification method using SDS-PAGE profiles of whole-cell proteins and subsequent 16S rRNA gene sequence analysis could successfully differentiate thermophilic bacteria from Algerian hot springs.

Isolation of a novel thermophilic bacterium capable of producing high-yield bioemulsifier and its kinetic modelling aspects along with proposed metabolic pathway.

Bioemulsifiers form stable emulsions and lower surface tension between two phases with potent anti-microbial activities. Some applications of bioemulsifier are performed at high temperatures and hence production of bioemulsifiers that are stable at high temperature is required. This study aimed at the production of bioemulsifier by an unexplored bacterial strain isolated from a local hot spring. The parameters tested for bioemulsifier production (emulsification ability, surface tension measurement and product formation) showed that 24 h is the optimal time for the production of bioemulsifier by strain S3 with yield of 1.4 g/l. The logistic growth curve of bacterial strain was analysed and kinetic constants for substrate utilisation and product formation were determined by Luedeking-Piret kinetic models. The bacterial strain S3 was Gram-positive and was classified as a strain of Brevibacillus borstelensis. The specific growth rate of the organism was 0.0096 h-1 with the kinetic rate constants as 11.246 (γ) and 10.626 (δ) for Luedeking-Piret substrate and 3.8423 (α) and - 1.9075 (ß) for Luedeking-Piret product. Knowledge of these values will help in estimating the substrate utilisation or bioemulsifier formed at any time point. These studies will also help in understanding internal metabolic fluxes hence rigorous analysis of metabolic pathway of bioemulsan is also performed in this study. Graphical abstract.

Enrichment and isolation of acid-tolerant sulfate-reducing microorganisms in the anoxic, acidic hot spring sediments from Copahue volcano, Argentina.

The geothermal Copahue-Caviahue (GCC) system (Argentina) is an extreme acidic environment, dominated by the activity of Copahue volcano. Environments characterised by low pH values, such as volcanic areas, are of particular interest for the search of acidophilic microorganisms with application in biotechnological processes. In this work, sulfate-reducing microorganisms were investigated in geothermal acidic, anaerobic zones from GCC system. Sediment samples from Agua del Limón (AL1), Las Máquinas (LMa2), Las Maquinitas (LMi) and Baño 9 (B9-2, B9-3) were found to be acidic (pH values 2.1-3.0) to moderate acidic (5.1-5.2), containing small total organic carbon values, and ferric iron precipitates. The organic electron donor added to the enrichment was completely oxidised to CO2. Bacteria related to 'Desulfobacillus acidavidus' strain CL4 were found to be dominant (67-83% of the total number of clones) in the enrichment cultures, and their presence was confirmed by their isolation on overlay plates. Other bacteria were also detected with lower abundance (6-20% of the total number of clones), with representatives of the genera Acidithiobacillus, Sulfobacillus, Alicyclobacillus and Athalassotoga/Mesoaciditoga. These enrichment and isolates found at low pH confirm the presence of anaerobic activities in the acidic sediments from the geothermal Copahue-Caviahue system.

Fischerella thermalis: a model organism to study thermophilic diazotrophy, photosynthesis and multicellularity in cyanobacteria.

The true-branching cyanobacterium Fischerella thermalis (also known as Mastigocladus laminosus) is widely distributed in hot springs around the world. Morphologically, it has been described as early as 1837. However, its taxonomic placement remains controversial. F. thermalis belongs to the same genus as mesophilic Fischerella species but forms a monophyletic clade of thermophilic Fischerella strains and sequences from hot springs. Their recent divergence from freshwater or soil true-branching species and the ongoing process of specialization inside the thermal gradient make them an interesting evolutionary model to study. F. thermalis is one of the most complex prokaryotes. It forms a cellular network in which the main trichome and branches exchange metabolites and regulators via septal junctions. This species can adapt to a variety of environmental conditions, with its photosynthetic apparatus remaining active in a temperature range from 15 to 58 °C. Together with its nitrogen-fixing ability, this allows it to dominate in hot spring microbial mats and contribute significantly to the de novo carbon and nitrogen input. Here, we review the current knowledge on the taxonomy and distribution of F. thermalis, its morphological complexity, and its physiological adaptations to an extreme environment.

Colombian Andean thermal springs: reservoir of thermophilic anaerobic bacteria producing hydrolytic enzymes.

Anaerobic cultivable microbial communities in thermal springs producing hydrolytic enzymes were studied. Thermal water samples from seven thermal springs located in the Andean volcanic belt, in the eastern and central mountain ranges of the Colombian Andes were used as inocula for the growth and isolation of thermophilic microorganisms using substrates such as starch, gelatin, xylan, cellulose, Tween 80, olive oil, peptone and casamino acids. These springs differed in temperature (50-70 °C) and pH (6.5-7.5). The predominant ion in eastern mountain range thermal springs was sulphate, whereas that in central mountain range springs was bicarbonate. A total of 40 anaerobic thermophilic bacterial strains that belonged to the genera Thermoanaerobacter, Caloramator, Anoxybacillus, Caloranaerobacter, Desulfomicrobium, Geotoga, Hydrogenophilus, Desulfacinum and Thermoanaerobacterium were isolated. To investigate the metabolic potential of these isolates, selected strains were analysed for enzymatic activities to identify strains than can produce hydrolytic enzymes. We demonstrated that these thermal springs contained diverse microbial populations of anaerobic thermophilic comprising different metabolic groups of bacteria including strains belonging to the genera Thermoanaerobacter, Caloramator, Anoxybacillus, Caloranaerobacter, Desulfomicrobium, Geotoga, Hydrogenophilus, Desulfacinum and Thermoanaerobacterium with amylases, proteases, lipases, esterases, xylanases and pectinases; therefore, the strains represent a promising source of enzymes with biotechnological potential.

Plastic Silica Conglomerate with an Extremophile Microbial Matrix in a Hot-Water Stream Paleoenvironment.

A new and unusual type of fossil, siliceous hot-spring deposit (sinter)-comprising monomictic, quartzose conglomerate encrusted with silicified microbial laminates-has been recognized in distal portions of Jurassic and Miocene paleo-geothermal fields of South and North America, respectively. The siliceous clasts are inferred to have originated as conduit-delivered hydrothermal silica gel, owing to their general plastic morphologies, which were then locally reworked and redistributed in geothermally influenced stream paleoenvironments. Today, hot-spring-fed streams and creeks, in places with silica-armored pavements, host microbial mats coating streambeds and/or growing over, and silicifying at, stream air-water interfaces, for example, in Yellowstone National Park (USA) and Waimangu Volcanic Valley (New Zealand). However, the modern deposits do not contain the plastically deformed silica cobbles evident in Mesozoic and Cenozoic examples described herein. Moreover, the fossil microbial laminates of this study are relatively dense and strongly coat the silica cobbles, suggesting the mats stabilized the clasts under fully submerged and hot, high-energy conditions. Thus, this new sinter facies, typically found a few kilometers from main spring-vent areas, is a perhaps unexpected extreme environment in which life took hold in hydrothermal-fluvial settings of the past, and may serve as an additional target in the search for fossil biosignatures of early Earth and possibly Mars.

Arsenic-tolerant microbial consortia from sediments of Copahue geothermal system with potential applications in bioremediation.

Although arsenic (As) is recognized as a toxic element for living species, some microorganisms have the ability to tolerate and transform it; recent studies have proposed to take advantage of such capacity to develop sustainable bioremediation strategies. In this study, we evaluated the adaptation to increasing concentrations of As(III) and As(V) of three metabolically different microbial cultures (heterotrophic, autotrophic-acidophilic, and anaerobic) obtained from a sample with low-soluble As content from the Copahue geothermal system. At the end of the adaptation process, the heterotrophic culture was able to grow at 20 mM and 450 mM of As(III) and As(V), respectively; the autotrophic-acidophilic culture showed tolerance to 15 mM of As(III) and 150 mM of As(V), whereas the anaerobic culture only developed in As(V) at concentrations up to 50 mM. The most tolerant consortia were characterized by their growth performance, complexity, and the presence of genes related to As metabolism and resistance. Regarding the consortia complexity, the predominant genera identified were: Paenibacillus in both heterotrophic consortia, Acidithiobacillus in the autotrophic-acidophilic consortium tolerant to As(III), Acidiphilium in the autotrophic-acidophilic consortium tolerant to As(V), and Thiomonas and Clostridium in the anaerobic consortium. This study is the first report of As tolerance microorganisms obtained from Copahue and reasserts the versatility and flexibility of the community of this natural extreme environment; also, it opens the door to the study of possible uses of these consortia in the design of biotechnological processes where the As concentration may fluctuate.

Thermoplasmatales and sulfur-oxidizing bacteria dominate the microbial community at the surface water of a CO2-rich hydrothermal spring located in Tenorio Volcano National Park, Costa Rica.

Here we report the chemical and microbial characterization of the surface water of a CO2-rich hydrothermal vent known in Costa Rica as Borbollones, located at Tenorio Volcano National Park. The Borbollones showed a temperature surrounding 60 °C, a pH of 2.4 and the gas released has a composition of ~ 97% CO2, ~ 0.07% H2S, ~ 2.3% N2 and ~ 0.12% CH4. Other chemical species such as sulfate and iron were found at high levels with respect to typical fresh water bodies. Analysis by 16S rRNA gene metabarcoding revealed that in Borbollones predominates an archaeon from the order Thermoplasmatales and one bacterium from the genus Sulfurimonas. Other sulfur- (genera Thiomonas, Acidithiobacillus, Sulfuriferula, and Sulfuricurvum) and iron-oxidizing bacteria (genera Sideroxydans, Gallionella, and Ferrovum) were identified. Our results show that CO2-influenced surface water of Borbollones contains microorganisms that are usually found in acid rock drainage environments or sulfur-rich hydrothermal vents. To our knowledge, this is the first microbiological characterization of a CO2-dominated hydrothermal spring from Central America and expands our understanding of those extreme ecosystems.

Temperature modulates Fischerella thermalis ecotypes in Porcelana Hot Spring.

In the Porcelana Hot Spring (Northern Patagonia), true-branching cyanobacteria are the dominant primary producers in microbial mats, and they are mainly responsible for carbon and nitrogen fixation. However, little is known about their metabolic and genomic adaptations at high temperatures. Therefore, in this study, a total of 81 Fischerella thermalis strains (also known as Mastigocladus laminosus) were isolated from mat samples in a thermal gradient between 61-46°C. The complementary use of proteomic comparisons from these strains, and comparative genomics of F. thermalis pangenomes, suggested that at least two different ecotypes were present within these populations. MALDI-TOF MS analysis separated the strains into three clusters; two with strains obtained from mats within the upper temperature range (61 and 54°C), and a third obtained from mats within the lower temperature range (51 and 46°C). Both groups possessed different but synonymous nifH alleles. The main proteomic differences were associated with the abundance of photosynthesis-related proteins. Three F. thermalis metagenome assembled genomes (MAGs) were described from 66, 58 and 48°C metagenomes. These pangenomes indicated a divergence of orthologous genes and a high abundance of exclusive genes at 66°C. These results improved the current understanding of thermal adaptation of F. thermalis and the evolution of these thermophilic cyanobacterial species.

Description of Alicyclobacillus montanus sp. nov., a mixotrophic bacterium isolated from acidic hot springs.

Three morphologically similar thermo-acidophilic strains, USBA-GBX-501, USBA-GBX-502 and USBA-GBX-503T, were isolated from acidic thermal springs at the National Natural Park Los Nevados (Colombia). All isolates were spore-forming, Gram-stain-positive and motile, growing aerobically at 25-55 °C (optimum ~45 °C) and at pH 1.5-4.5 (optimum pH ~3.0). Phylogenetic analysis of the 16S rRNA gene sequences of these isolates showed an almost identical sequence (99.0 % similarity) and they formed a robust cluster with the closest relative Alicyclobacillus tolerans DSM 16297T with a sequence similarity of 99.0 %. Average similarity to other species of the genus Alicyclobacillus was 93.0 % and average similarity to species of the genus Effusibacillus was 90 %. In addition, the level of DNA-DNA hybridization between strain USBA-GBX-503T and Alicyclobacillus tolerans DSM 16297T was 31.7 %. The genomic DNA G+C content of strain USBA-GBX-503T was 44.6 mol%. The only menaquinone was MK-7 (100.0 %). No ω-alicyclic fatty acids were detected in strain USBA-GBX-503T, and the major cellular fatty acids were C18 : 1ω7c, anteiso-C17 : 0 and iso-C17 : 0. Based on phenotypic and chemotaxonomic characteristics, phylogenetic analysis and DNA-DNA relatedness values, along with low levels of identity at the whole genome level (ANIb and ANIm values of <67.0 and <91.0 %, respectively), it can be concluded that strain USBA-GBX-503T represents a novel species of the genus Alicyclobacillus, for which the name Alicyclobacillus montanus sp. nov. is proposed. The type strain is USBA-GBX-503T (=CMPUJ UGB U503T=CBMAI1927T).

Physiological and gene expression responses to nitrogen regimes and temperatures in Mastigocladus sp. strain CHP1, a predominant thermotolerant cyanobacterium of hot springs.

Cyanobacteria are widely distributed primary producers with significant implications for the global biogeochemical cycles of carbon and nitrogen. Diazotrophic cyanobacteria of subsection V (Order Stigonematales) are particularly ubiquitous in photoautotrophic microbial mats of hot springs. The Stigonematal cyanobacterium strain CHP1 isolated from the Porcelana hot spring (Chile) was one of the major contributors of the new nitrogen through nitrogen fixation. Further morphological and genetic characterization verified that the strain CHP1 belongs to Stigonematales, and it formed a separate clade together with other thermophiles of the genera Fischerella and Mastigocladus. Strain CHP1 fixed maximum N2 in the light, independent of the temperature range. At 50°C nifH gene transcripts showed high expression during the light period, whereas the nifH gene expression at 45°C was arrhythmic. The strain displayed a high affinity for nitrate and a low tolerance for high ammonium concentrations, whereas the narB and glnA genes showed higher expression in light and at the beginning of the dark phase. It is proposed that Mastigocladus sp. strain CHP1 would represent a good model for the study of subsection V thermophilic cyanobacteria, and for understanding the adaptations of these photoautotrophic organisms inhabiting microbial mats in hot springs globally.

Description of a new anaerobic thermophilic bacterium, Thermoanaerobacterium butyriciformans sp. nov.

Strain USBA-019T, an anaerobic and thermophilic strain, was identified as a new member of the genus Thermoanaerobacterium. USBA-019T cells are gram-positive, strictly anaerobic, thermophilic, chemoorganotrophic, moderately acidophilic, non-motile, endospore-forming, slightly curved, and rod-shaped. Cells measure 0.4×3.0-7.0µm. Optimal growth occurs at 50-55°C (35-65°C). Optimum pH is 5.0-5.5 (4.0-8.5). Thiosulfate, elemental sulfur and nitrate were utilized as electron acceptors. Fermentation of glucose, lactose, cellobiose, galactose, arabinose, xylose, starch and xylan primarily produced acetate and butyrate. Xylan, starch and cellobiose produced ethanol and starch, cellobiose, galactose, arabinose and mannose produced lactic acid. Phylogenetic analyses based on 16S rRNA gene sequence comparison and genomic relatedness indices show the close relation of USBA-019T to Thermoanaerobacterium thermostercoris and Thermoanaerobacterium aotearoense (similarity value: 99%). Hybridization of USBA-019T, Th. thermostercoris DSM22141T and Th. aotearoense DMS10170T found DNA-DNA relatedness of 33.2% and 18.2%, respectively. Based on phenotypic, chemotaxonomic and phylogenetic evidence, along with low identity at whole genome level, USBA-019T is a novel species of the genus Thermoanaerobacterium which we propose to name Thermoanaerobacterium butyriciformans sp. nov. The type strain is USBA-019T (=CMPUJ U-019T=DSM 101588T).