Showing papers in "Extremophiles in 2017"

Adaptations of archaeal and bacterial membranes to variations in temperature, pH and pressure

Abstract: The cytoplasmic membrane of a prokaryotic cell consists of a lipid bilayer or a monolayer that shields the cellular content from the environment. In addition, the membrane contains proteins that are responsible for transport of proteins and metabolites as well as for signalling and energy transduction. Maintenance of the functionality of the membrane during changing environmental conditions relies on the cell’s potential to rapidly adjust the lipid composition of its membrane. Despite the fundamental chemical differences between bacterial ester lipids and archaeal ether lipids, both types are functional under a wide range of environmental conditions. We here provide an overview of archaeal and bacterial strategies of changing the lipid compositions of their membranes. Some molecular adjustments are unique for archaea or bacteria, whereas others are shared between the two domains. Strikingly, shared adjustments were predominantly observed near the growth boundaries of bacteria. Here, we demonstrate that the presence of membrane spanning ether-lipids and methyl branches shows a striking relationship with the growth boundaries of archaea and bacteria.

Microbial diversity and autotrophic activity in Kamchatka hot springs

Abstract: Microbial communities of Kamchatka Peninsula terrestrial hot springs were studied using molecular, radioisotopic and cultural approaches. Analysis of 16S rRNA gene fragments performed by means of high-throughput sequencing revealed that aerobic autotrophic sulfur-oxidizing bacteria of the genus Sulfurihydrogenibium (phylum Aquificae) dominated in a majority of streamers. Another widely distributed and abundant group was that of anaerobic bacteria of the genus Caldimicrobium (phylum Thermodesulfobacteria). Archaea of the genus Vulcanisaeta were abundant in a high-temperature, slightly acidic hot spring, where they were accompanied by numerous Nanoarchaeota, while the domination of uncultured Thermoplasmataceae A10 was characteristic for moderately thermophilic acidic habitats. The highest rates of inorganic carbon assimilation determined by the in situ incubation of samples in the presence of 14C-labeled bicarbonate were found in oxygen-dependent streamers; in two sediment samples taken from the hottest springs this process, though much weaker, was found to be not dependent on oxygen. The isolation of anaerobic lithoautotrophic prokaryotes from Kamchatka hot springs revealed a wide distribution of the ability for sulfur disproportionation, a new lithoautotrophic process capable to fuel autonomous anaerobic ecosystems.

Biodegradation of polycyclic aromatic hydrocarbons by an acidophilic Stenotrophomonas maltophilia strain AJH1 isolated from a mineral mining site in Saudi Arabia.

Abstract: The present study aims at analyzing the degradation of polycyclic aromatic hydrocarbons (PAHs) at acidic conditions (pH = 2) by acidophilic Stenotrophomonas maltophilia strain AJH1 (KU664513). The strain AJH1 was obtained from an enrichment culture obtained from soil samples of mining area in the presence of PAH as sole sources of carbon and energy. Strain AJH1was able to degrade low (anthracene, phenanthrene, naphthalene, fluorene) and high (pyrene, benzo(e)pyrene and benzo(k)fluoranthene) molecular weight PAHs in acidophilic mineral salt medium at pH 2, with removal rates of up to 95% (LMW PAH) and 80% (HMW PAH), respectively. In addition, strain AJH1 treated petroleum wastewater with 89 ± 1.1% COD removal under acidic condition (pH 2) in a continuously stirred reactor. Acidophilic S. maltophilia strain AJH1, hence holds the promise as an effective degrader for biological treatment of PAHs contaminated wastewater at acidic pH.

Degradation of azo dye methyl red by alkaliphilic, halotolerant Nesterenkonia lacusekhoensis EMLA3: application in alkaline and salt-rich dyeing effluent treatment.

Abstract: Effluents from textile industries are highly colored due to vast use of various azo dyes and color is the first visual indicator of pollution. Biological treatment of textile effluent is often hampered due to the alkaline pH and high salinity; a common characteristic of many textile industrial wastewaters. Considering this, the present study explores the potential of a newly isolated halotolerant and alkaliphilic bacterium Nesterenkonia lacusekhoensis EMLA3 for degradation of methyl red (MR) dye under alkaline condition. Strain EMLA3 showed 97% degradation of 50 mg L−1 MR after 16 h at initial pH of 11.5 in nutrient medium. Dye degradation by the isolate is supported by the formation of low-molecular weight metabolites as divulge through GC-MS & FTIR studies Optimum dye degradation was observed in the pH range of 8.0–11.5 and temperature range of 30–35 °C. Significant MR degrading activity of the strain could be achieved in the presence of very high salt level (100–120 g L−1 NaCl) and in co-presence of different heavy metals. Application of strain to alkaline pH, salt, and heavy metals laden-textile effluent resulted in overall 83% dye removal from the effluent after 120 h of treatment under static condition. Furthermore, the property of microbe to drop-down the pH of wastewater from 11.5 to 8.60 after treatment also lowers the need of additional neutralization treatment. The entire study thus comes out with novel application of N. lacusekhoensis—a less explored extremophilic bacterium—for treatment of alkaline and salt-rich azo dye-containing wastewaters.

Carotenoids from the extreme halophilic archaeon Haloterrigena turkmenica: identification and antioxidant activity.

Abstract: Haloterrigena turkmenica was able to synthesize carotenoids when grown in halobacteria medium. These molecules have antioxidant properties and find application in food, cosmetic, and pharmaceutical fields. The carotenoids were extracted with methanol, separated by RP-HPLC, and identified by mass spectrometry and UV/Vis spectra analyses. The C50 carotenoids were the main pigments, and C30, C40, and C51 carotenoids were also detected. Seven geometric isomers were distinguished for bacterioruberin, monoanhydrobacterioruberin, and bisanhydrobacterioruberin. The assignment to a specific isomer was tentatively attempted through the analysis of the corresponding UV/Vis spectrum, the intensity of the cis peak, and its spectral fine structure. Lycopene, phytoene, and lycopersene were among the minor carotenoids further identified. The extract displayed antioxidant power higher than alpha-tocopherol, butylhydroxytoluene, and ascorbic acid used as reference compounds. Our studies identified for the first time seven geometric isomers of bacterioruberin derivatives and 30 carotenoids in a haloarchaeon.

Searching for novel photolyases in UVC-resistant Antarctic bacteria.

Abstract: Ultraviolet (UV) light irradiation has serious consequences for cell survival, including DNA damage by formation of cyclobutane pyrimidine dimers (CPD) and pyrimidine (6,4) pyrimidone photoproducts. In general, the Nucleotide Excision Repair pathway repairs these lesions; however, all living forms, except placental mammals and some marsupials, produce a flavoprotein known as photolyase that directly reverses these lesions. The aim of this work was the isolation and identification of Antarctic UVC-resistant bacteria, and the search for novel photolyases. Two Antarctic water samples were UVC-irradiated (254 nm; 50–200 J m− 2) and 12 UVC-resistant bacteria were isolated and identified by 16S rDNA amplification/analysis as members of the genera Pseudomonas, Janthinobacterium, Flavobacterium, Hymenobacter and Sphingomonas. The UVC 50% lethal dose and the photo-repair ability of isolates were analyzed. The occurrence of photolyase coding sequences in Pseudomonas, Hymenobacter and Sphingomonas isolates were searched by PCR or by searching in the draft DNA genome. Results suggest that Pseudomonas and Hymenobacter isolates produce CDP-photolyases, and Sphingomonas produces two CPD-photolyases and a 6,4-photolyase. Results suggest that the Antarctic environment is an important source of genetic material for the identification of novel photolyase genes with potential biotechnological applications.

Soda pans of the Pannonian steppe harbor unique bacterial communities adapted to multiple extreme conditions

Abstract: Soda pans of the Pannonian steppe are unique environments regarding their physical and chemical characteristics: shallowness, high turbidity, intermittent character, alkaline pH, polyhumic organic carbon concentration, hypertrophic condition, moderately high salinity, sodium and carbonate ion dominance. The pans are highly productive environments with picophytoplankton predominance. Little is known about the planktonic bacterial communities inhabiting these aquatic habitats; therefore, amplicon sequencing and shotgun metagenomics were applied to reveal their composition and functional properties. Results showed a taxonomically complex bacterial community which was distinct from other soda lakes regarding its composition, e.g. the dominance of class Alphaproteobacteria was observed within phylum Proteobacteria. The shotgun metagenomic analysis revealed several functional gene components related to the harsh and at the same time hypertrophic environmental conditions, e.g. proteins involved in stress response, transport and hydrolase systems targeting phytoplankton-derived organic matter. This is the first detailed report on the indigenous planktonic bacterial communities coping with the multiple extreme conditions present in the unique soda pans of the Pannonian steppe.

Antarctomyces pellizariae sp. nov., a new, endemic, blue, snow resident psychrophilic ascomycete fungus from Antarctica.

Abstract: In the present study, we have identified and characterised a new snow resident ascomycete blue stain fungus from Antarctica named Antarctomyces pellizariae sp. nov. Menezes, Godinho, Porto, Goncalves and Rosa, using polyphasic taxonomy techniques. This fungal species was recovered from the seasonal snow of the Antarctic Peninsula. Antarctomyces pellizariae displayed different macro- and micromorphology when compared with A. psychrotrophicus Stchigel and Guarro, the only other Antarctomyces species reported until date. Antarctomyces pellizariae showed psychrophilic behavior and very low growth rate at 22–25 °C, quite different from A. psychrotrophicus that has a higher growth rate at mesophilic temperatures. In addition, micromorphological characteristics and the analysis of the nuclear rDNA internal transcribed spacer, β-tubulin, and RNA polymerase II regions revealed that A. pellizariae is a new species that is related to A. psychrotrophicus and Thelebolus species. Since the Antarctic Peninsula is reported to be one of the main regions of the earth experiencing the effects of global change in climate, species, such as A. pellizariae, might provide information about these effects on the endemic Antarctic biota. In addition, A. pellizariae displayed psychrophilic behavior and might be a source of interesting anti-freeze compounds that might prove useful in biotechnological processes.

Phenol degradation and heavy metal tolerance of Antarctic yeasts.

Abstract: In cold environments, biodegradation of organic pollutants and heavy metal bio-conversion requires the activity of cold-adapted or cold-tolerant microorganisms. In this work, the ability to utilize phenol, methanol and n-hexadecane as C source, the tolerance to different heavy metals and growth from 5 to 30 °C were evaluated in cold-adapted yeasts isolated from Antarctica. Fifty-nine percent of the yeasts were classified as psychrotolerant as they could grow in all the range of temperature tested, while the other 41% were classified as psychrophilic as they only grew below 25 °C. In the assimilation tests, 32, 78, and 13% of the yeasts could utilize phenol, n-hexadecane, and methanol as C source, respectively, but only 6% could assimilate the three C sources evaluated. In relation to heavy metals ions, 55, 68, and 80% were tolerant to 1 mM of Cr(VI), Cd(II), and Cu(II), respectively. Approximately a half of the isolates tolerated all of them. Most of the selected yeasts belong to genera previously reported as common for Antarctic soils, but several other genera were also isolated, which contribute to the knowledge of this cold environment mycodiversity. The tolerance to heavy metals of the phenol-degrading cold-adapted yeasts illustrated that the strains could be valuable as inoculant for cold wastewater treatment in extremely cold environments.

Avoidance of protein oxidation correlates with the desiccation and radiation resistance of hot and cold desert strains of the cyanobacterium Chroococcidiopsis

Abstract: To investigate the relationship between desiccation and the extent of protein oxidation in desert strains of Chroococcidiopsis a selection of 10 isolates from hot and cold deserts and the terrestrial cyanobacterium Chroococcidiopsis thermalis sp. PCC 7203 were exposed to desiccation (air-drying) and analyzed for survival. Strain CCMEE 029 from the Negev desert and the aquatic cyanobacterium Synechocystis sp. PCC 6803 were further investigated for protein oxidation after desiccation (drying over silica gel), treatment with H2O2 up to 1 M and exposure to γ-rays up to 25 kGy. Then a selection of desert strains of Chroococcidiopsis with different survival rates after prolonged desiccation, as well as Synechocystis sp. PCC 6803 and Chroococcidiopsis thermalis sp. PCC 7203, were analyzed for protein oxidation after treatment with 10 and 100 mM of H2O2. Results suggest that in the investigated strains a tight correlation occurs between desiccation and radiation tolerance and avoidance of protein oxidation.

Diversity of bacteria and archaea from two shallow marine hydrothermal vents from Vulcano Island

Abstract: To obtain new insights into community compositions of hyperthermophilic microorganisms, defined as having optimal growth temperatures of 80 °C and above, sediment and water samples were taken from two shallow marine hydrothermal vents (I and II) with temperatures of 100 °C at Vulcano Island, Italy. A combinatorial approach of denaturant gradient gel electrophoresis (DGGE) and metagenomic sequencing was used for microbial community analyses of the samples. In addition, enrichment cultures, growing anaerobically on selected polysaccharides such as starch and cellulose, were also analyzed by the combinatorial approach. Our results showed a high abundance of hyperthermophilic archaea, especially in sample II, and a comparable diverse archaeal community composition in both samples. In particular, the strains of the hyperthermophilic anaerobic genera Staphylothermus and Thermococcus, and strains of the aerobic hyperthermophilic genus Aeropyrum, were abundant. Regarding the bacterial community, e-Proteobacteria, especially the genera Sulfurimonas and Sulfurovum, were highly abundant. The microbial diversity of the enrichment cultures changed significantly by showing a high dominance of archaea, particularly the genera Thermococcus and Palaeococcus, depending on the carbon source and the selected temperature.

The thermophilic biomass-degrading fungus Thielavia terrestris Co3Bag1 produces a hyperthermophilic and thermostable β-1,4-xylanase with exo- and endo-activity

Abstract: A hyperthermophilic and thermostable xylanase of 82 kDa (TtXynA) was purified from the culture supernatant of T. terrestris Co3Bag1, grown on carboxymethyl cellulose (CMC), and characterized biochemically. TtXynA showed optimal xylanolytic activity at pH 5.5 and at 85 °C, and retained more than 90% of its activity at a broad pH range (4.5-10). The enzyme is highly thermostable with a half-life of 23.1 days at 65 °C, and active in the presence of several metal ions. Circular dichroism spectra strongly suggest the enzyme gains secondary structures when temperature increases. TtXynA displayed higher substrate affinity and higher catalytic efficiency towards beechwood xylan than towards birchwood xylan, oat-spelt xylan, and CMC. According to its final hydrolysis products, TtXynA displays endo-/exo-activity, yielded xylobiose, an unknown oligosaccharide containing about five residues of xylose and a small amount of xylose on beechwood xylan. Finally, this report represents the description of the first fungal hyperthermophilic xylanase which is produced by T. terrestris Co3Bag1. Since TtXynA displays relevant biochemical properties, it may be a suitable candidate for biotechnological applications carried out at high temperatures, like the enzymatic pretreatment of plant biomass for the production of bioethanol.

Effect of environmental parameters on biodiversity of the fungal component in lithic Antarctic communities.

Abstract: A wide sampling of rocks, colonized by microbial epi–endolithic communities, was performed along an altitudinal gradient from sea level to 3600 m asl and sea distance from the coast to 100 km inland along the Victoria Land Coast, Antarctica. Seventy-two rock samples of different typology, representative of the entire survey, were selected and studied using denaturing gradient gel electrophoresis to compare variation in fungal diversity according to environmental conditions along this altitudinal and sea distance transect. Lichenized fungi were largely predominant in all the samples studied and the biodiversity was heavily influenced even by minimal local variations. The n-MDS analysis showed that altitude and sea distance affect fungal biodiversity, while sandstone allows the communities to maintain high biodiversity indices. The Pareto-Lorenz curves indicate that all the communities analyzed are highly adapted to extreme conditions but scarcely resilient, so any external perturbation may have irreversible effects on these fragile ecosystems.

Microbial community analysis of pH 4 thermal springs in Yellowstone National Park.

Abstract: The pH of the majority of thermal springs in Yellowstone National Park (YNP) is from 1 to 3 and 6 to 10; relatively few springs (~5%) have a pH range of 4–5. We used 16S rRNA gene pyrosequencing to investigate microbial communities sampled from four pH 4 thermal springs collected from four regions of YNP that differed in their fluid temperature and geochemistry. Our results revealed that the composition of bacterial communities varied among the sites, despite sharing similar pH values. The taxonomic composition and metabolic functional potential of the site with the lowest temperature (55 °C), a thermal spring from the Seven Mile Hole (SMH) area, were further investigated using shotgun metagenome sequencing. The taxonomic classification, based on 372 Mbp of unassembled metagenomic reads, indicated that this community included a high proportion of Chloroflexi, Bacteroidetes, Proteobacteria, and Firmicutes. Functional comparison with other YNP thermal spring metagenomes indicated that the SMH metagenome was enriched in genes related to energy production and conversion, transcription, and carbohydrate transport. Analysis of genes involved in nitrogen metabolism revealed assimilatory and dissimilatory nitrate reduction pathways, whereas the majority of genes involved in sulfur metabolism were related to the reduction of sulfate to adenylylsulfate, sulfite, and H2S. Given that pH 4 thermal springs are relatively less common in YNP and thermal areas worldwide, they may harbor novel microbiota and the communities that inhabit them deserve further investigation.

Taxonomy, phylogeny and ecology of cultivable fungi present in seawater gradients across the Northern Antarctica Peninsula.

Abstract: Thirty-six seawater samples collected at different depths of the Gerlache and Bransfield Straits in the Northern Antarctic Peninsula were analyzed, and the average of the total fungal counts ranged from 0.3 to >300 colony forming units per liter (CFU/L) in density. The fungal were purified and identified as 15 taxa belonged to the genera Acremonium, Aspergillus, Cladosporium, Cystobasidium, Exophiala, Glaciozyma, Graphium, Lecanicillium, Metschnikowia, Penicillium, Purpureocillium and Simplicillium. Penicillium chrysogenum, Cladosporium sphaerospermum, and Graphium rubrum were found at high densities in at least two different sites and depths. Our results show at the first time that in the seawater of Antarctic Ocean occur diverse fungal assemblages despite extreme conditions, which suggests the presence of a complex aquatic fungi food web, including species reported as barophiles, symbionts, weak and strong saprobes, parasites and pathogens, as well as those found in the polluted environments of the world. Additionally, some taxa were found in different sites, suggesting that the underwater current might contribute to fungal (and microbial) dispersal across the Antarctic Ocean, and nearby areas such as South America and Australia.

Microbial diversity in degraded and non-degraded petroleum samples and comparison across oil reservoirs at local and global scales

Abstract: Microorganisms have shown their ability to colonize extreme environments including deep subsurface petroleum reservoirs. Physicochemical parameters may vary greatly among petroleum reservoirs worldwide and so do the microbial communities inhabiting these different environments. The present work aimed at the characterization of the microbiota in biodegraded and non-degraded petroleum samples from three Brazilian reservoirs and the comparison of microbial community diversity across oil reservoirs at local and global scales using 16S rRNA clone libraries. The analysis of 620 16S rRNA bacterial and archaeal sequences obtained from Brazilian oil samples revealed 42 bacterial OTUs and 21 archaeal OTUs. The bacterial community from the degraded oil was more diverse than the non-degraded samples. Non-degraded oil samples were overwhelmingly dominated by gammaproteobacterial sequences with a predominance of the genera Marinobacter and Marinobacterium. Comparisons of microbial diversity among oil reservoirs worldwide suggested an apparent correlation of prokaryotic communities with reservoir temperature and depth and no influence of geographic distance among reservoirs. The detailed analysis of the phylogenetic diversity across reservoirs allowed us to define a core microbiome encompassing three bacterial classes (Gammaproteobacteria, Clostridia, and Bacteroidia) and one archaeal class (Methanomicrobia) ubiquitous in petroleum reservoirs and presumably owning the abilities to sustain life in these environments.

Namib Desert edaphic bacterial, fungal and archaeal communities assemble through deterministic processes but are influenced by different abiotic parameters.

Abstract: The central Namib Desert is hyperarid, where limited plant growth ensures that biogeochemical processes are largely driven by microbial populations. Recent research has shown that niche partitioning is critically involved in the assembly of Namib Desert edaphic communities. However, these studies have mainly focussed on the Domain Bacteria. Using microbial community fingerprinting, we compared the assembly of the bacterial, fungal and archaeal populations of microbial communities across nine soil niches from four Namib Desert soil habitats (riverbed, dune, gravel plain and salt pan). Permutational multivariate analysis of variance indicated that the nine soil niches presented significantly different physicochemistries (R 2 = 0.8306, P ≤ 0.0001) and that bacterial, fungal and archaeal populations were soil niche specific (R 2 ≥ 0.64, P ≤ 0.001). However, the abiotic drivers of community structure were Domain-specific (P < 0.05), with P, clay and sand fraction, and NH4 influencing bacterial, fungal and archaeal communities, respectively. Soil physicochemistry and soil niche explained over 50% of the variation in community structure, and communities displayed strong non-random patterns of co-occurrence. Taken together, these results demonstrate that in central Namib Desert soil microbial communities, assembly is principally driven by deterministic processes.

In-depth characterization of bacterial and archaeal communities present in the abandoned Kettara pyrrhotite mine tailings (Morocco)

Abstract: In Morocco, pollution caused by closed mines continues to be a serious threat to the environment, like the generation of acid mine drainage. Mine drainage is produced by environmental and microbial oxidation of sulfur minerals originating from mine wastes. The fundamental role of microbial communities is well known, like implication of Fe-oxidizing and to a lesser extent S-oxidizing microorganism in bioleaching. However, the structure of the microbial communities varies a lot from one site to another, like diversity depends on many factors such as mineralogy, concentration of metals and metalloids or pH, etc. In this study, prokaryotic communities in the pyrrhotite-rich tailings of Kettara mine were characterized using the Illumina sequencing. In-depth phylogenetic analysis revealed a total of 12 phyla of bacteria and 1 phyla of Archaea. The majority of sequences belonged to the phylum of Proteobacteria and Firmicutes with a predominance of Bacillus, Pseudomonas or Corynebacterium genera. Many microbial populations are implicated in the iron, sulfur and arsenic cycles, like Acidiferrobacter, Leptospirillum, or Alicyclobacillus in Fe; Acidiferrobacter and Sulfobacillus in S; and Bacillus or Pseudomonas in As. This is one of the first description of prokaryotic communities in pyrrhotite-rich mine tailings using high-throughput sequencing.

Bacterial diversity in fumarole environments of the Paricutin volcano, Michoacan (Mexico)

Abstract: Active volcanoes are among the most extreme environments on Earth. The extreme temperatures, presence of toxic heavy metals and low nutrient bioavailability favor the development of extremophiles. We characterized the physical-chemical parameters of and bacterial communities (T-RFLP and 16S rRNA gene libraries) inhabiting fumarole niches of the Paricutin volcano located in Michoacan (Mexico). This volcano, which surged in 1943, is one of the youngest volcanoes on Earth and the microbial diversity in this area is yet to be characterized. The sampling stations were characterized in a pH range from 5.34 to 7.89 and showed different temperatures (soil, 27-87 °C; air, 13.6-56 °C) with high concentrations of metals such as iron and arsenic. The most abundant bacterial populations, confirmed by T-RFLP and 16S rRNA gene libraries, were related to members of Firmicutes and Proteobacteria phyla including sequences associated with thermophiles and sulfate reducing bacteria. Overall, the Paricutin volcano showed low bacterial diversity and its prokaryotic diversity was characterized by the impossibility of amplifying Archaea-related sequences.

Multifactorial level of extremostability of proteins: can they be exploited for protein engineering?

Abstract: Research on extremostable proteins has seen immense growth in the past decade owing to their industrial importance. Basic research of attributes related to extreme-stability requires further exploration. Modern mechanistic approaches to engineer such proteins in vitro will have more impact in industrial biotechnology economy. Developing a priori knowledge about the mechanism behind extreme-stability will nurture better understanding of pathways leading to protein molecular evolution and folding. This review is a vivid compilation about all classes of extremostable proteins and the attributes that lead to myriad of adaptations divulged after an extensive study of 6495 articles belonging to extremostable proteins. Along with detailing on the rationale behind extreme-stability of proteins, emphasis has been put on modern approaches that have been utilized to render proteins extremostable by protein engineering. It was understood that each protein shows different approaches to extreme-stability governed by minute differences in their biophysical properties and the milieu in which they exist. Any general rule has not yet been drawn regarding adaptive mechanisms in extreme environments. This review was further instrumental to understand the drawback of the available 14 stabilizing mutation prediction algorithms. Thus, this review lays the foundation to further explore the biophysical pleiotropy of extreme-stable proteins to deduce a global prediction model for predicting the effect of mutations on protein stability.

100 kGy gamma-affected microbial communities within the ancient Arctic permafrost under simulated Martian conditions.

Abstract: This research aimed to investigate the viability and biodiversity of microbial communities within ancient Arctic permafrost after exposure to a gamma-radiation dose of 100 kGy at low temperature (- 50 °C), low pressure (1 Torr) and dehydration conditions. The main objective was to assess the possibility for long-term survival of Earth-bound microorganisms in the subsurface of Martian regolith or inside small space bodies at constant absorption and accumulation of the gamma radiation dose. Investigated microbial communities had shown high resistance to a simulated Martian environment. After irradiation the total count of prokaryotic cells and number of metabolically active bacterial cells remained at the control level, while the number of bacterial CFUs decreased by 2 orders of magnitude, and the number of metabolically active cells of archaea decreased threefold. Besides, the abundance of culturable bacteria after irradiation was kept at a high level: not less than 3.7 × 105 cells/g. Potential metabolic activity of irradiated microbial communities in general were higher than in the control sample. A fairly high biodiversity of bacteria was detected in the exposed sample of permafrost, although the microbial community structure underwent significant changes after irradiation. In particular, actinobacteria populations of the genus Arthrobacter, which was not revealed in the control samples, became predominant in bacterial communities following the exposure. The results of the study testify that long-term preservation of microbial life inside Martian permafrost is possible. The data obtained can also be evaluated from the perspective of the potential for discovering viable Earth-bound microorganisms on other objects in the Solar system and inside of small bodies in outer space.

Characterization of two novel heat-active α-galactosidases from thermophilic bacteria.

Abstract: Two genes (agal1 and agal2) encoding α-galactosidases were identified by sequence-based screening approaches. The gene agal1 was identified from a data set of a sequenced hot spring metagenome, and the deduced amino-acid sequence exhibited 99% identity to an α-galactosidase from the thermophilic bacterium Dictyoglomus thermophilum. The gene agal2 was identified from the whole genome sequence of the thermophile Meiothermus ruber. The amino-acid sequences exhibited structural motifs typical for glycoside hydrolase (GH) family 36 members and were also differentiated into different subgroups of this family. Recombinant production of the heat-active GH36b enzyme Agal1 (87 kDa) and GH36bt enzyme Agal2 (57 kDa) was carried out in E. coli. Agal1 exhibited a specific activity of 1502.3 U/mg at 80 °C, pH 6.5, and Agal2 225.4 U/mg at 60–70 °C, pH 6.5. Half-lives of 14 h (Agal1) and 39 h (Agal2) were obtained at 50 °C, and Agal1 showed half-lives of 4 and 2 h at 70 and 80 °C, respectively. In addition to the natural substrates melibiose, raffinose, and stachyose, 4NP α-d-galactopyranoside was hydrolyzed. Galactose was also liberated from locust bean gum. Both heat-active enzymes are attractive candidates for application in food and feed industry for high-temperature processes for the degradation of raffinose family oligosaccharides.

Pectinolytic yeasts from cold environments: novel findings of Guehomyces pullulans , Cystofilobasidium infirmominiatum and Cryptococcus adeliensis producing pectinases

Abstract: Fil: Cavello, Ivana Alejandra. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - La Plata. Centro de Investigacion y Desarrollo en Fermentaciones Industriales. Universidad Nacional de la Plata. Facultad de Ciencias Exactas. Centro de Investigacion y Desarrollo en Fermentaciones Industriales; Argentina

Aspergillus atacamensis and A. salisburgensis: two new halophilic species from hypersaline/arid habitats with a phialosimplex-like morphology.

Abstract: Halophilic fungal strains isolated from historical wooden staircase in a salt mine in Austria, and from wall biofilm and soil of a cave in the Coastal Range of the hyperarid Atacama Desert in Chile were characterised and described newly as Aspergillus salisburgensis and Aspergillus atacamensis. Morphological characters including solitary phialides producing solitary conidia and conidia in chains and/or heads suggested affinity to Aspergillus subgenus Polypaecilum. Strains required salt for growth, grew optimally on media with 10–25% NaCl and at 15–28 °C. These values are similar to those observed for Aspergillus salinarus comb. nov. (Phialosimplex salinarum), while the ex-type strains of Aspergillus sclerotialis, Aspergillus chlamydosporus and Aspergillus caninus (all belonging to Aspergillus subgen. Polypaecilum) grew optimally at 0–5% NaCl and showed fastest growth at 28–37 °C. Phylogenetic analyses on the basis of rDNA sequences, RAPD-PCR fingerprint patterns, and cellobiohydrolase gene (cbh-I) polymorphism clustered the strains into three groups and supported their taxonomic recognition as A. salinarus, A. atacamensis and A. salisburgensis. On the basis of phylogenetic inferences, also Sagenomella keratitidis is newly combined as Aspergillus keratitidis and inferred as a species of Aspergillus subgenus Polypaecilum.

Membrane lipids and soluble sugars dynamics of the alkaliphilic fungus Sodiomyces tronii in response to ambient pH

Abstract: Alkaliphily, the ability of an organism to thrive optimally at high ambient pH, has been well-documented in several lineages: archaea, bacteria and fungi. The molecular mechanics of such adaptation has been extensively addressed in alkaliphilic bacteria and alkalitolerant fungi. In this study, we consider an additional property that may have enabled fungi to prosper at alkaline pH: altered contents of membrane lipids and cytoprotectant molecules. In the alkaliphilic Sodiomyces tronii, we showed that at its optimal growth pH 9.2, the fungus accumulates abundant cytosolic trehalose (4–10% dry weight) and phosphatidic acids in the membrane lipids, properties not normally observed in neutrophilic species. At a very high pH 10.2, the major carbohydrate, glucose, was rapidly substituted by mannitol and arabitol. Conversely, lowering the pH to 5.4–7.0 had major implications both on the content of carbohydrates and membrane lipids. It was shown that trehalose dominated at pH 5.4. Fractions of sphingolipids and sterols of plasma membranes rapidly elevated possibly indicating the formation of membrane structures called rafts. Overall, our results reveals complex dynamics of the contents of membrane lipids and cytoplasmic sugars in alkaliphilic S. tronii, suggesting their adaptive functionality against pH stress.

Thermal adaptation strategies of the extremophile bacterium Thermus filiformis based on multi-omics analysis

Abstract: Thermus filiformis is an aerobic thermophilic bacterium isolated from a hot spring in New Zealand. The experimental study of the mechanisms of thermal adaptation is important to unveil response strategies of the microorganism to stress. In this study, the main pathways involved on T. filiformis thermoadaptation, as well as, thermozymes with potential biotechnological applications were revealed based on omics approaches. The strategy adopted in this study disclosed that pathways related to the carbohydrate metabolism were affected in response to thermoadaptation. High temperatures triggered oxidative stress, leading to repression of genes involved in glycolysis and the tricarboxylic acid cycle. During heat stress, the glucose metabolism occurred predominantly via the pentose phosphate pathway instead of the glycolysis pathway. Other processes, such as protein degradation, stringent response, and duplication of aminoacyl-tRNA synthetases, were also related to T. filiformis thermoadaptation. The heat-shock response influenced the carotenoid profile of T. filiformis, favoring the synthesis of thermozeaxanthins and thermobiszeaxanthins, which are related to membrane stabilization at high temperatures. Furthermore, antioxidant enzymes correlated with free radical scavenging, including superoxide dismutase, catalase and peroxidase, and metabolites, such as oxaloacetate and α-ketoglutarate, were accumulated at 77 °C.

Identification of a novel copper-activated and halide-tolerant laccase in Geobacillus thermopakistaniensis

Abstract: Genome search of Geobacillus thermopakistaniensis, formerly Geobacillus sp. SBS-4S, revealed the presence of an open reading frame (ESU71923) annotated as laccase. However, the gene product did not display any laccase-like activity against the substrates examined. The laccase activity was, therefore, purified from G. thermopakistaniensis cells and N-terminal amino acid residues of the enzyme were determined. These residues matched the N-terminal sequence of an open reading frame annotated as a copper oxidase (ESU72270). In order to characterize the enzyme, recombinant ESU72270 was prepared in Escherichia coli. The recombinant protein was found to exhibit a negligible amount of laccase activity when produced in the absence of copper in the growth medium. However, the recombinant protein exhibited significantly high laccase activity when produced in the presence of copper. The recombinant enzyme showed highest activity at 60 °C and a pH of 7–7.5. The purified enzyme was highly tolerant to various halides and organic solvents, thus having a potential for various industrial applications. To the best of our knowledge, this is the first characterization of a laccase from genus Geobacillus which identifies a gene responsible for functional laccase in this genus.

Antarctic rocks from continental Antarctica as source of potential human opportunistic fungi

Abstract: We assessed the diversity of culturable fungi associated with rocks of continental Antarctica to evaluate their physiological opportunistic virulence potential in vitro. The seventy fungal isolates obtained were identified as nine species of Acremonium, Byssochlamys, Cladosporium, Debaryomyces, Penicillium, and Rhodotorula. Acremonium sp., D. hansenii, P. chrysogenum, P. citrinum, P. tardochrysogenum, and R. mucilaginosa were able to grow at 37 °C; in addition, B. spectabilis displayed a high level of growth at 37 and 45 °C. Thirty-one isolates of P. chrysogenum, P. citrinum, and P. tardochrysogenum were able to produce partial haemolysis on blood agar at 37 °C. Acremonium sp., P. citrinum, and P. tardochrysogenum showed spore sizes ranging from 2.81 to 5.13 µm diameters at 37 °C. Of these, P. chrysogenum and P. tardochrysogenum displayed macro- and micro morphological polymorphism. Our results suggest that rocks of the ultra-extreme cold and dry environment of Antarctica harbour cryptic fungi phylogenetically close to opportunistic pathogenic and mycotoxigenic taxa with physiologic virulence characteristics in vitro.

Reverse gyrase is essential for microbial growth at 95 °C

Abstract: Reverse gyrase is an enzyme that induces positive supercoiling in closed circular DNA in vitro. It is unique to thermophilic organisms and found without exception in all microorganisms defined as hyperthermophiles, that is, those having optimal growth temperatures of 80 °C and above. Although its in vivo role has not been clearly defined, it has been implicated in stabilizing DNA at high temperatures. Whether or not it is absolutely required for growth at these high temperatures has yet to be fully determined. In a previous study with an organism that has an optimal growth temperature of 85 °C, it was shown that the enzyme is not a prerequisite for life at extreme temperatures as disruption of its gene did not result in a lethal phenotype at the supraoptimal growth temperature of 90 °C. Herein we show that the enzyme is absolutely required for microbial growth at 95 °C, which in this case is a suboptimal growth temperature. Deletion of the gene encoding the reverse gyrase of the model hyperthermophilic archaeon Pyrococcus furiosus, which has an optimal growth temperature of 100 °C, revealed that the gene is required for growth at 95 °C, as well as at 100 °C. The results suggest that a temperature threshold above 90 °C exists, wherein the activity of reverse gyrase is absolutely necessary to maintain a correct DNA twist for any organism growing at such temperature extremes.

Isolation and complete genome sequence of Halorientalis hydrocarbonoclasticus sp. nov., a hydrocarbon-degrading haloarchaeon

Abstract: Bioremediation in hypersaline environments is particularly challenging since the microbes that tolerate such harsh environments and degrade pollutants are quite scarce. Haloarchaea, however, due to their inherent ability to grow at high salt concentrations, hold great promise for remediating the contaminated hypersaline sites. This study aimed to isolate and characterize novel haloarchaeal strains with potentials in hydrocarbon degradation. A haloarchaeal strain IM1011 was isolated from Changlu Tanggu saltern near Da Gang Oilfield in Tianjin (China) by enrichment culture in hypersaline medium containing hexadecane. It could degrade 57 ± 5.2% hexadecane (5 g/L) in the presence of 3.6 M NaCl at 37 °C within 24 days. To get further insights into the mechanisms of petroleum hydrocarbon degradation in haloarchaea, complete genome (3,778,989 bp) of IM1011 was sequenced. Phylogenetic analysis of 16S rRNA gene, RNA polymerase beta-subunit (rpoB’) gene and of the complete genome suggested IM1011 to be a new species in Halorientalis genus, and the name Halorientalis hydrocarbonoclasticus sp. nov., is proposed. Notably, with insights from the IM1011 genome sequence, the involvement of diverse alkane hydroxylase enzymes and an intact β-oxidation pathway in hexadecane biodegradation was predicted. This is the first hexadecane-degrading strain from Halorientalis genus, of which the genome sequence information would be helpful for further dissecting the hydrocarbon degradation by haloarchaea and for their application in bioremediation of oil-polluted hypersaline environments.