A hydrothermally precipitated catalytic iron sulphide membrane as a first step toward life

We propose that life emerged from growing aggregates of iron sulphide bubbles containing alkaline and highly reduced hydrothermal solution. These bubbles were inflated hydrostatically at sulphidic submarine hot springs sited some distance from oceanic spreading centers four billion years ago. The membrane enclosing the bubbles was precipitated in response to contact between the spring waters and the mildly oxidized, acidic and iron-bearing Hadean ocean water. As the gelatinous sulphide bubbles aged and were inflated beyond their strength they budded, producing contiguous daughter bubbles by the precipitation of new membrane. [Fe₂S₂]^+/0 or [Fe₄S₄]^2+/+ clusters, possibly bonded by hydrothermal thiolate ligands as proferredoxins, could have catalyzed oxidation of thiolates to disulphides, thereby modifying membrane properties.We envisage the earliest iron sulphide bubbles (pro botryoids) first growing by hydrostatic inflation with hydrothermal fluid, but evolving to grow mainly by osmosis (the protocellular stage), driven by (1) catabolism of hydrothermal abiogenic organics trapped on the inner walls of the membrane, catalyzed by the iron sulphide clusters; and (2) cleavage of hydrophobic compounds dissolved in the membrane to hydrophilic moieties which were translocated, by the proton motive force inherent in the acidic Hadean ocean, to the alkaline interior of the protocell. The organics were generated first within the hydrothermal convective system feeding the hot springs operating in the oceanic crust and later in the pyritizing mound developing on the sea floor, as a consequence of the reduction of CO, CO₂, and formaldehyde by Fe²⁺- and S^2–-bearing minerals.We imagine the physicochemical interactions in and on the membrane to have been sufficiently complex to have engendered auto- and cross-catalytic replication. The membrane may have been constructed in such a way that a successful parent could have informed the daughters of membrane characteristics functional for the then-current level of evolution.Correspondence to: M. J. RussellGlossary: Hollow pyrite botryoids: hollow hemispheres of cryptocrystalline pyrite (FeS₂) 0.1–1 mm across. Fischer-Tropsch syntheses: the highly exothermic catalytic hydrogenation of CO to hydrocarbons and aliphatic oxygenated compounds using finely divided iron. Greigite (Fe₃S₄): metastable iron sulphide precipitated from aqueous solution in a gel at 100°C and containing two-thirds of its iron as the high-spin ferric ion. Haber-Bosch process: the exothermic catalytic hydrogenation of nitrogen to yield ammonia. Probotryoid: a hydrostatically inflated colloidal iron monosulphide bubble; precursor to hollow botryoids and the progenitor to protocells. Proferredoxins: [Fe₂S₂] and [Fe₃MS₄] clusters (M = Fe, Mo, W, Ni, etc.) ligated by abiogenic thiols and thiolates. Protocell: a cell comprised mainly of abiogenic organics including thiols with subordinate iron sulphides, partly as proferredoxins; growth results from catabolism and osmotic pressure     

The thermal springs of Bakreswar,India physico-chemical conditions,Flora and Fauna.

The physico-chemical characteristics as well as floristic and faunistic life of the seven thermal springs and one cold spring of Bakreswar, West Bengal form the basis of this paper. A compendium pertaining to the origin, geology and morphometry of these springs has been incorporated in brief.The temperature of the thermal springs remained almost constant over the past two decades and the location of the fissures of the springs is unchanged. On the basis of temperature these thermal springs are grouped into acrothermal and euthermal. The water of all the springs was alkaline with a pH between 7.4 and 9.2 No carbon dioxide was detected during the period of study. Absence of nitrite and traces of nitrate are characteristic features of these springs.The variation of colour of the water may be related to the presence of algal mats at the bottom of the springs. The quantity of plankton was maximal and minimal in thermal springs having highest and lowest temperature respectively. Oscillatoria sp., Navicula sp., Cyclotella sp. and Tetraëdron sp. were noted at temperatures between 26° C and 51° C. It is a new record from India that diatoms like Navicula sp. and Cyclotella sp. can survive in the temperature range of 43–51° C. The thermophilic nematode, Tobrilus sp., Cyclops sp., Nauplius larvae, and Lecane sp. were recorded from Swetganga with a maximum temperature of 44° C. The mechanism of thermophily of the organisms is discussed in brief.     

The Alkaline Solution to the Emergence of Life: Energy,Entropy and Early Evolution

The Earth agglomerates and heats. Convection cells within the planetary interior expedite the cooling process. Volcanoes evolve steam, carbon dioxide, sulfur dioxide and pyrophosphate. An acidulous Hadean ocean condenses from the carbon dioxide atmosphere. Dusts and stratospheric sulfurous smogs absorb a proportion of the Sun’s rays. The cooled ocean leaks into the stressed crust and also convects. High temperature acid springs, coupled to magmatic plumes and spreading centers, emit iron, manganese, zinc, cobalt and nickel ions to the ocean. Away from the spreading centers cooler alkaline spring waters emanate from the ocean floor. These bear hydrogen, formate, ammonia, hydrosulfide and minor methane thiol. The thermal potential begins to be dissipated but the chemical potential is dammed. The exhaling alkaline solutions are frustrated in their further attempt to mix thoroughly with their oceanic source by the spontaneous precipitation of biomorphic barriers of colloidal iron compounds and other minerals. It is here we surmise that organic molecules are synthesized, filtered, concentrated and adsorbed, while acetate and methane—separate products of the precursor to the reductive acetyl-coenzyme-A pathway—are exhaled as waste. Reactions in mineral compartments produce acetate, amino acids, and the components of nucleosides. Short peptides, condensed from the simple amino acids, sequester ‘ready-made’ iron sulfide clusters to form protoferredoxins, and also bind phosphates. Nucleotides are assembled from amino acids, simple phosphates carbon dioxide and ribose phosphate upon nanocrystalline mineral surfaces. The side chains of particular amino acids register to fitting nucleotide triplet clefts. Keyed in, the amino acids are polymerized, through acid–base catalysis, to alpha chains. Peptides, the tenuous outer-most filaments of the nanocrysts, continually peel away from bound RNA. The polymers are concentrated at cooler regions of the mineral compartments through thermophoresis. RNA is reproduced through a convective polymerase chain reaction operating between 40 and 100°C. The coded peptides produce true ferredoxins, the ubiquitous proteins with the longest evolutionary pedigree. They take over the role of catalyst and electron transfer agent from the iron sulfides. Other iron–nickel sulfide clusters, sequestered now by cysteine residues as CO-dehydrogenase and acetyl-coenzyme-A synthase, promote further chemosynthesis and support the hatchery—the electrochemical reactor—from which they sprang. Reactions and interactions fall into step as further pathways are negotiated. This hydrothermal circuitry offers a continuous supply of material and chemical energy, as well as electricity and proticity at a potential appropriate for the onset of life in the dark, a rapidly emerging kinetic structure born to persist, evolve and generate entropy while the sun shines. An erratum to this article can be found at     

Serpentinization as a source of energy at the origin of life

For life to have emerged from CO₂, rocks, and water on the early Earth, a sustained source of chemically transducible energy was essential. The serpentinization process is emerging as an increasingly likely source of that energy. Serpentinization of ultramafic crust would have continuously supplied hydrogen, methane, minor formate, and ammonia, as well as calcium and traces of acetate, molybdenum and tungsten, to off‐ridge alkaline hydrothermal springs that interfaced with the metal‐rich carbonic Hadean Ocean. Silica and bisulfide were also delivered to these springs where cherts and sulfides were intersected by the alkaline solutions. The proton and redox gradients so generated represent a rich source of naturally produced chemiosmotic energy, stemming from geochemistry that merely had to be tapped, rather than induced, by the earliest biochemical systems. Hydrothermal mounds accumulating at similar sites in today’s oceans offer conceptual and experimental models for the chemistry germane to the emergence of life, although the ubiquity of microbial communities at such sites in addition to our oxygenated atmosphere preclude an exact analogy.     

On the origins of cells: a hypothesis for the evolutionary transitions from abiotic geochemistry to chemoautotrophic prokaryotes,and from prokaryotes to nucleated cells

All life is organized as cells. Physical compartmentation from the environment and self-organization of self-contained redox reactions are the most conserved attributes of living things, hence inorganic matter with such attributes would be life's most likely forebear. We propose that life evolved in structured iron monosulphide precipitates in a seepage site hydrothermal mound at a redox, pH and temperature gradient between sulphide-rich hydrothermal fluid and iron(II)-containing waters of the Hadean ocean floor. The naturally arising, three-dimensional compartmentation observed within fossilized seepage-site metal sulphide precipitates indicates that these inorganic compartments were the precursors of cell walls and membranes found in free-living prokaryotes. The known capability of FeS and NiS to catalyse the synthesis of the acetyl-methylsulphide from carbon monoxide and methylsulphide, constituents of hydrothermal fluid, indicates that pre-biotic syntheses occurred at the inner surfaces of these metal-sulphide-walled compartments, which furthermore restrained reacted products from diffusion into the ocean, providing sufficient concentrations of reactants to forge the transition from geochemistry to biochemistry. The chemistry of what is known as the RNA-world could have taken place within these naturally forming, catalyticwalled compartments to give rise to replicating systems. Sufficient concentrations of precursors to support replication would have been synthesized in situ geochemically and biogeochemically, with FeS (and NiS) centres playing the central catalytic role. The universal ancestor we infer was not a free-living cell, but rather was confined to the naturally chemiosmotic, FeS compartments within which the synthesis of its constituents occurred. The first free-living cells are suggested to have been eubacterial and archaebacterial chemoautotrophs that emerged more than 3.8 Gyr ago from their inorganic confines. We propose that the emergence of these prokaryotic lineages from inorganic confines occurred independently, facilitated by the independent origins of membrane-lipid biosynthesis: isoprenoid ether membranes in the archaebacterial and fatty acid ester membranes in the eubacterial lineage. The eukaryotes, all of which are ancestrally heterotrophs and possess eubacterial lipids, are suggested to have arisen ca. 2 Gyr ago through symbiosis involving an autotrophic archaebacterial host and a heterotrophic eubacterial symbiont, the common ancestor of mitochondria and hydrogenosomes. The attributes shared by all prokaryotes are viewed as inheritances from their confined universal ancestor. The attributes that distinguish eubacteria and archaebacteria, yet are uniform within the groups, are viewed as relics of their phase of differentiation after divergence from the non-free-living universal ancestor and before the origin of the free-living chemoautotrophic lifestyle. The attributes shared by eukaryotes with eubacteria and archaebacteria, respectively, are viewed as inheritances via symbiosis. The attributes unique to eukaryotes are viewed as inventions specific to their lineage. The origin of the eukaryotic endomembrane system and nuclear membrane are suggested to be the fortuitous result of the expression of genes for eubacterial membrane lipid synthesis by an archaebacterial genetic apparatus in a compartment that was not fully prepared to accommodate such compounds, resulting in vesicles of eubacterial lipids that accumulated in the cytosol around their site of synthesis. Under these premises, the most ancient divide in the living world is that between eubacteria and archaebacteria, yet the steepest evolutionary grade is that between prokaryotes and eukaryotes.     

Isolation of Halotolerant Thermus spp. from Submarine Hot Springs in Iceland

Thermophilic, aerobic bacteria of the genus Thermus were isolated from submarine alkaline hot springs in Iceland. Five submarine hot springs were sampled, and all had viable counts of Thermus spp. of about 10³ CFU/ml. All submarine strains grew in the presence of NaCl at 3% or higher, but no strains from terrestrial hot springs would grow at concentrations higher than 1% NaCl. The growth rate of submarine Thermus strains was not stimulated by NaCl and was reduced at NaCl concentrations higher than 1%. The pattern of growth of these isolates on single carbon sources was similar to that of terrestrial isolates.     

The inevitable journey to being

Life is evolutionarily the most complex of the emergent symmetry-breaking, macroscopically organized dynamic structures in the Universe. Members of this cascading series of disequilibria-converting systems, or engines in Cottrell''s terminology, become ever more complicated—more chemical and less physical—as each engine extracts, exploits and generates ever lower grades of energy and resources in the service of entropy generation. Each one of these engines emerges spontaneously from order created by a particular mother engine or engines, as the disequilibrated potential daughter is driven beyond a critical point. Exothermic serpentinization of ocean crust is life''s mother engine. It drives alkaline hydrothermal convection and thereby the spontaneous production of precipitated submarine hydrothermal mounds. Here, the two chemical disequilibria directly causative in the emergence of life spontaneously arose across the mineral precipitate membranes separating the acidulous, nitrate-bearing CO₂-rich, Hadean sea from the alkaline and CH₄/H₂-rich serpentinization-generated effluents. Essential redox gradients—involving hydrothermal CH₄ and H₂ as electron donors, CO₂ and nitrate, nitrite, and ferric iron from the ambient ocean as acceptors—were imposed which functioned as the original ‘carbon-fixing engine’. At the same time, a post-critical-point (milli)voltage pH potential (proton concentration gradient) drove the condensation of orthophosphate to produce a high energy currency: ‘the pyrophosphatase engine’.     

Prebiotic Synthesis of Glycine from Ethanolamine in Simulated Archean Alkaline Hydrothermal Vents

Submarine hydrothermal vents are generally considered as the likely habitats for the origin and evolution of early life on Earth. In recent years, a novel hydrothermal system in Archean subseafloor has been proposed. In this model, highly alkaline and high temperature hydrothermal fluids were generated in basalt-hosted hydrothermal vents, where H₂ and CO₂ could be abundantly provided. These extreme conditions could have played an irreplaceable role in the early evolution of life. Nevertheless, sufficient information has not yet been obtained for the abiotic synthesis of amino acids, which are indispensable components of life, at high temperature and alkaline condition. This study aims to propose a new method for the synthesis of glycine in simulated Archean submarine alkaline vent systems. We investigated the formation of glycine from ethanolamine under conditions of high temperature (80–160 °C) and highly alkaline solutions (pH = 9.70). Experiments were performed in an anaerobic environment under mild pressure (0.1–8.0 MPa) at the same time. The results suggested that the formation of glycine from ethanolamine occurred rapidly and efficiently in the presence of metal powders, and was favored by high temperatures and high pressures. The experiment provides a new pathway for prebiotic glycine formation and points out the phenomenal influence of high-temperature alkaline hydrothermal vents in origin of life in the early ocean.     

Serpentinite and the dawn of life

Submarine hydrothermal vents above serpentinite produce chemical potential gradients of aqueous and ionic hydrogen, thus providing a very attractive venue for the origin of life. This environment was most favourable before Earth's massive CO(2) atmosphere was subducted into the mantle, which occurred tens to approximately 100 Myr after the moon-forming impact; thermophile to clement conditions persisted for several million years while atmospheric pCO(2) dropped from approximately 25 bar to below 1 bar. The ocean was weakly acid (pH ～ 6), and a large pH gradient existed for nascent life with pH 9-11 fluids venting from serpentinite on the seafloor. Total CO(2) in water was significant so the vent environment was not carbon limited. Biologically important phosphate and Fe(II) were somewhat soluble during this period, which occurred well before the earliest record of preserved surface rocks approximately 3.8 billion years ago (Ga) when photosynthetic life teemed on the Earth and the oceanic pH was the modern value of approximately 8. Serpentinite existed by 3.9 Ga, but older rocks that might retain evidence of its presence have not been found. Earth's sequesters extensive evidence of Archaean and younger subducted biological material, but has yet to be exploited for the Hadean record.     

Seasonal Changes in Reproductive Activity in the Potter's angelfish (Centropyge potteri) in Kaneohe Bay, Hawaii

Potter's angelfish, Centropyge potteri, is a protogynous hermaphrodite, with the alpha female of a harem becoming male under the proper social conditions. Gonads and plasma samples were collected from females every 6–9 days for 1 year, and then for every 4–6 weeks for another year. The gonadosomatic index (GSI) of females remained below 1% from July until December. Starting from the first week of December, large spikes occurred in the GSI, fluctuating from 1.4% to 4.1% and falling below 1% in June. Plasma concentrations of estradiol-17 were relatively low (1–2ngml^–1) between July and November. Beginning in the third week of November, large spikes of plasma estradiol-17 were observed, fluctuating from 0.5 to 5ngml^–1. This pattern continued until the third week of May, and then estradiol-17 levels remained low for the rest of the year. Estradiol-17 levels showed a highly significant correlation with GSI. Estradiol-17 levels during late vitellogenesis and final maturation were significantly greater than those of the other stages. These results suggest that the spawning season of the Potter's angelfish is from December to June. One of the causes of the large fluctuations in GSI and plasma estradiol-17 within the spawning season appears to be due to the fact that Potter's angelfish is an asynchronous daily spawner.     

Brain lysosomal hydrolases: I. Solubilization and electrophoretic behavior of acid hydrolases in nerve-ending and mitochondrial-lysosomal fractions from rat brain. Effects of autolysis,neuraminidase, and storage

In solubility studies of 7 acid hydrolases, the extent of solubilization by sonic disruption varied with the enzyme species and increased with increasing pH and Triton X-100 concentration of the suspension medium. Hydrolases in the nerve-ending (NE) fraction were more resistant to solubilization than those in the mitochondrial-lysosomal (M-L) fraction, but nearly quantitative solubilization was attained by sonication in an alkaline buffer containing 0,5% Triton X-100. Polyacrylamide gel electrophoresis of extracts revealed multiple components of acid phosphatase, acid esterase, arylsulfatase,-glucuronidase, and-N-acetyl-hexosaminidase. The enzyme patterns varied with the subcellular fraction and the composition of the medium. In general, the acidic (anodic) forms of these hydrolases were more readily solubilized by sonication in acidic buffer, alkaline pH and Triton X-100 being required to solubilize the basic (cationic) components. The acidic forms of these enzymes were converted to less anodic or cathodic forms, or both, during autolysis at pH 6 at 0 and 37°C, and during storage at –20°C.     

Polyphasic Characterization of a Thermotolerant Siderophilic Filamentous Cyanobacterium That Produces Intracellular Iron Deposits

Despite the high potential for oxidative stress stimulated by reduced iron, contemporary iron-depositing hot springs with circum-neutral pH are intensively populated with cyanobacteria. Therefore, studies of the physiology, diversity, and phylogeny of cyanobacteria inhabiting iron-depositing hot springs may provide insights into the contribution of cyanobacteria to iron redox cycling in these environments and new mechanisms of oxidative stress mitigation. In this study the morphology, ultrastructure, physiology, and phylogeny of a novel cyanobacterial taxon, JSC-1, isolated from an iron-depositing hot spring, were determined. The JSC-1 strain has been deposited in ATCC under the name Marsacia ferruginose, accession number BAA-2121. Strain JSC-1 represents a new operational taxonomical unit (OTU) within Leptolyngbya sensu lato. Strain JSC-1 exhibited an unusually high ratio between photosystem (PS) I and PS II, was capable of complementary chromatic adaptation, and is apparently capable of nitrogen fixation. Furthermore, it synthesized a unique set of carotenoids, but only chlorophyll a. Strain JSC-1 not only required high levels of Fe for growth (≥40 μM), but it also accumulated large amounts of extracellular iron in the form of ferrihydrite and intracellular iron in the form of ferric phosphates. Collectively, these observations provide insights into the physiological strategies that might have allowed cyanobacteria to develop and proliferate in Fe-rich, circum-neutral environments.Cyanobacteria inhabiting ferrous iron-rich hot springs with circum-neutral pH represent unique models for examining the mechanisms by which early organisms evolved to cope with such habitats common on early Earth. Such organisms have previously been shown to be resistant to Fe²⁺ (37) or Fe³⁺ (6, 7) at concentrations in the micromolar to millimolar range. Moreover, high Fe concentrations (apparent optimum of ∼0.5 mM) stimulated the growth of these cyanobacteria, which were described as siderophilic (having an affinity for iron) cyanobacteria (7).The cyanobacteria inhabiting the Chocolate Pots hot springs in Yellowstone National Park, Wyoming, were shown to have played at least a passive role in contributing to iron deposition by serving as nucleation sites for the accumulation of iron minerals and associated silica deposits (36, 38). The precipitation of external iron that encrusts the cyanobacterial cells inhabiting this hot spring appears to be dependent on the species composition and chemistry of the mat (36, 38); however, multiple anoxygenic phototrophs found in the Chocolate Pots hot springs (8) could also contribute to the formation of Fe oxides (21, 49). Therefore, only iron mineralization experiments with model cyanobacterial strains can demonstrate the role of siderophilic cyanobacteria in the formation of specific, iron-bearing minerals.An additional common feature of circum-neutral iron-depositing hot springs is elevated concentrations of hydrogen peroxide (50). Shcolnick and coauthors (41) showed that a wild type of Synechococcus sp. PCC 6803 was resistant to 8 mM H₂O₂ if grown with 0.3 μM Fe³⁺, while the same concentration of hydrogen peroxide completely inhibited the growth of this cyanobacterium if it was grown with 10 μM Fe³⁺. If a similar correlation between iron concentration and the magnitude of an externally applied oxidative stress were the case for siderophilic cyanobacteria, iron-depositing hot springs should be free of cyanobacteria. However, such springs are very rich with cyanobacteria (38, 7, 36), which suggests that siderophilic cyanobacteria may possess unusual mechanisms of iron homeostasis maintenance and oxidative stress mitigation. Additionally, understanding iron tolerance and phenomena associated with siderophily in oxygenic prokaryotes is also important because such siderophilic organisms might help us find applications for bioremediation of waters polluted with iron.The current work describes the morphology, ultrastructure, physiology, and phylogeny of a previously undescribed, siderophilic cyanobacterium. The results of this polyphasic characterization led to the conclusion that strain JSC-1 represents a new operational taxonomic unit (OTU). (The epithet for JSC-1, Marsacia ferruginose, was chosen in honor of Nicole Tandeau de Marsac.) Additionally, biomineralization of intracellular iron by a cyanobacterium is demonstrated for the first time.     

The influence of metal cations and pH on the heat sensitivity of photosynthetic oxygen evolution and chlorophyll fluorescence in spinach chloroplasts

The heat-sensitivity of photosynthetic oxygen evolution of thylakoids isolated from spinach increases by increasing the pH above neutral value. The temperature for inactivation (transition temperature) is lowered from about 45° C (pH 6.0–7.4) to 33°C (pH 8.5). Similar results are obtained with intact chloroplasts. At pH 7.0 the transition temperature of washed thylakoids decreases by lowering the salt concentration below 20 mM with monovalent cations (Li⁺, Na⁺, K⁺) and below 3–4 mM with divalent cations (Mg²⁺, Ca²⁺, Sr²⁺). Illumination decreases the heat-sensitivity of oxygen evolution in intact chloroplasts, but even increases the heat-sensitivity in uncoupled chloroplasts. In intact chloroplasts the transition temperature of the heat-induced rise in chlorophyll fluorescence yield (F_o; see Schreiber and Armond 1978) decreases from 44° C to 38° C when the pH of the suspending medium is increased from 6.5 to 8.5. At 20° C, F_o is almost insensitive to pH (6.0–8.5). At 40° C, however, F_o is constant between 6.0 and 7.0, but strongly increases by increasing the pH above neutral value. The results are discussed in terms of a close relation between electrostatic forces at the thylakoid membrane and thermal sensitivity of photosynthetic apparatus. It is suggested that the heat-sensitivity of the photosystem II complex partially depends on the ionization state of fixed groups having alkaline pK. The packed volume of thylakoids suspended in a low salt medium increases when the temperature is increased above 30° C (pH 7.0) and above 20° C (pH 8.0), respectively. This result suggests a heat-induced increase in surface charge density of the thylakoid membrane.Abbreviations HEPES N-2-hydroxyethylpiperazine-N-2-ethane sulfonic acid - MES morpholinoethane sulfonic acid - MOPS 2-N-morpholinopropane sulfonic acid - TRICIN N-[tris(hydroxymethyl)-methyl] glycine     

Model experiments on nitrite and nitrate in simulated primeval conditions.

In experiments on the prebiotic formation of nitric oxides, anoxic mixtures of N2 and water vapour were sparked in contact with phosphate buffer solutions at various pH values. Nitrite was found in the aqueous phase, and nitrate grew from it, presumably by reaction with H2O2. In acid solutions, these anions were reduced and destroyed by Fe2+, and the same was true of nitrite in solutions kept at a pH value similar to that of the contemporary ocean (8.2) with HEPES buffer. Nitrate was not destroyed in short-term experiments, but as in sparking nitrate is formed only vianitrite, neither anion could accumulate. In further sparking experiments with alkaline sulphide, both nitrite and nitrate were reduced entirely. It is concluded that it is unlikely that the primeval ocean contained appreciable concentrations of nitrite or nitrate either at the reducing or at the redox-neutral stage.     

Isolation of thermophilic obligately autotrophic hydrogen-oxidizing bacteria,similar to Hydrogenobacter thermophilus,from Icelandic hot springs

Thermophilic obligately autotrophic hydrogen-oxidizing bacteria were isolated from several alkaline hot springs in Iceland. The bacteria were Gram negative rods, 0.4–0.5 m in diameter and 3–4 m long but 6–7 m long cells without septa were often seen. Long and short laments are formed. Spores, flagella or lipid granules were not observed. Strains H1 and H12 grew optimally at 70° C and pH 6.5 under mixture of air plus 0.6 atm H₂ and 0.1 atm CO₂. The cells contained cytochromes and carotenoid-like pigments. They would not grow on agar or silicia gel plates. The cells would not grow heterotrophically on organic substrates and were inhibited by most of these same organic compounds and agar in low concentrations. They were very sensitive to common antibiotics. The role of these bacteria in the hot spring ecosystem is discussed.     

Xylanases of marine fungi of potential use for biobleaching of paper pulp

Microbial xylanases that are thermostable, active at alkaline pH and cellulase-free are generally preferred for biobleaching of paper pulp. We screened obligate and facultative marine fungi for xylanase activity with these desirable traits. Several fungal isolates obtained from marine habitats showed alkaline xylanase activity. The crude enzyme from NIOCC isolate 3 (Aspergillus niger), with high xylanase activity, cellulase-free and unique properties containing 580 U l^–1 xylanase, could bring about bleaching of sugarcane bagasse pulp by a 60 min treatment at 55°C, resulting in a decrease of ten kappa numbers and a 30% reduction in consumption of chlorine during bleaching. The culture filtrate showed peaks of xylanase activity at pH 3.5 and pH 8.5. When assayed at pH 3.5, optimum activity was detected at 50°C, with a second peak of activity at 90°C. When assayed at pH 8.5, optimum activity was seen at 80°C. The crude enzyme was thermostable at 55°C for at least 4 h and retained about 60% activity. Gel filtration of the 50–80% ammonium sulphate-precipitated fraction of the crude culture filtrate separated into two peaks of xylanase with specific activities of 393 and 2,457 U (mg protein)^–1. The two peaks showing xylanase activity had molecular masses of 13 and 18 kDa. Zymogram analysis of xylanase of crude culture filtrate as well as the 50–80% ammonium sulphate-precipitated fraction showed two distinct xylanase activity bands on native PAGE. The crude culture filtrate also showed moderate activities of -xylosidase and -l-arabinofuranosidase, which could act synergistically with xylanase in attacking xylan. This is the first report showing the potential application of crude culture filtrate of a marine fungal isolate possessing thermostable, cellulase-free alkaline xylanase activity in biobleaching of paper pulp.     

Xylanases of thermophilic bacteria from Icelandic hot springs

Thermophilic, aerobic bacteria isolated from Icelandic hot springs were screened for xylanase activity. Of 97 strains tested, 14 were found to be xylanase positive. Xylanase activities up to 12 nkat/ml were produced by these strains in shake flasks on xylan medium. The xylanases of the two strains producing the highest activities (ITI 36 and ITI 283) were similar with respect to temperature and pH optima (80°C and pH 8.0). Xylanase production of strain ITI 36 was found to be induced by xylan and xylose. Xylanase activity of 24 nkat/ml was obtained with this strain in a laboratory-scale-fermentor cultivation on xylose medium. -Xylosidase activity was also detected in the culture filtrate. The thermal half-life of ITI 36 xylanase was 24 h at 70°C. The highest production of sugars from hydrolysis of beech xylan was obtained at 70°C, although xylan depolymerization was detected even up to 90°C. Correspondence to: M. Rättö     

Chemolithotrophic metabolism of the newly-isolated moderately thermophilic,obligately autotrophic Thiobacillus tepidarius

Thiobacillus tepidarius, isolated from the hot springs at Bath, Avon, UK, grew optimally at 43–45°C and pH 6.0–7.5 on thiosulphate or tetrathionate. In batch culture, thiosulphate was oxidized stoichiometrically to tetrathionate, with a rise in pH. The tetrathionate was then oxidized to sulphate, supporting growth and producing a fall in pH to a minimum of ph 4.8. The organism contained high levels of thiosulphate-oxidizing enzyme, rhodanese and ribulose bisphosphate carboxylase. It was obligately chemolithotrophic and autotrophic. In chemostat culture, T. tepidarius grew autotrophically with the following sole energy-substrates: sulphide, thiosulphate, trithionate, tetrathionate, hexathionate or heptathionate. Thiocyanate, dithionate and sulphite were not used as sole substrates, although sulphite enhanced growth yields in the presence of thiosulphate. Maximum specific growth rate on tetrathionate was 0.44 h^-1. True growth yields (Y _max) and maintenance coefficients (m) were calculated for sulphide, thiosulphate, trithionate and tetrathionate and observed yields at a single fixed dilution rate compared with those on hexathionate and heptathionate. Mean values for Y _max, determined from measurements of absorbance, dry wt, total organic carbon and cell protein, were similar for sulphide, thiosulphate and trithionate (10.9 g dry wt/mol substrate) as expected from their equivalent oxygen consumption for oxidation. Y _max for tetrathionate (20.5) and the relative Y _o values (as g dry wt/g atom oxygen consumed) for thiosulphate and all four polythionates indicated that substrate level phosphorylation did not contribute significantly to energy conservation. These Y _max values were 40–70% higher than any of those previously reported for obligately aerobic thiobacilli. Mean values for m were 6.7 mmol substrate oxidized/g dry wt·h for sulphide, thiosulphate and trithionate, and 2.6 for tetrathionate.Abbreviation PIPES Piperazine-N,N-bis(ethane sulphonic acid)     

Inadequate iron supply and high bicarbonate impair the symbiosis of peanuts (Arachis hypogaea L.) with different Bradyrhizobium strains

In the present study, we examined the effects of iron deficiency in an acid solution and in an alkaline solution containing bicarbonate on the growth and nodulation of peanuts inoculated with different bradyrhizobial strains or supplied with fertilizer nitrogen.Inadequate iron supply in acid solution decreased the number of nodule initials, nodule number and nodule mass. Alleviating the iron deficiency increased acetylene reduction but not bacteroid numbers in nodules. Nitrogen concentrations in shoots of inoculated plants increased as iron concentrations in solution increased when determined at day 30 but not at day 50. Higher iron concentrations in solution were required for maximum growth of plants reliant on symbiotic nitrogen fixation than for those receiving fertilizer nitrogen.Adding bicarbonate to the solution with 7.5 M Fe markedly depressed nodule formation. This effect was much more severe than that of inadequate iron supply alone. Bicarbonate also decreased nitrogenase activity but did not decrease bacteroid concentrations in nodules.Both NC92 and TAL1000 nodulated peanuts poorly when bicarbonate was present. However, an interaction between iron concentrations in acid solutions and Bradyrhizobium strains on nodulation of peanuts was observed. Alleviating iron deficiency increased the number of nodule initials and nodules to a much greater extent for plants inoculated with TAL1000 than for plants inoculated with NC92.     

The Cold Origin of Life: A. Implications Based On The Hydrolytic Stabilities Of Hydrogen Cyanide And Formamide

It has been suggested that hydrogen cyanide(HCN) would not have been present in sufficient concentrationto polymerize in the primitive ocean to produce nucleic acidbases and amino acids. We have measured the hydrolysis ratesof HCN and formamide over the range of 30–150 °C and pH 0–14,and estimated the steady state concentrations in theprimitive ocean. At 100 °C and pH 8, the steady stateconcentration of HCN and formamide were calculated to be7 × 10^-13 M and 1 × 10^-15 M, respectively. Thus, itseems unlikely that HCN could have polymerized in a warmprimitive ocean. It is suggested that eutectic freezing mighthave been required to have concentrated HCN sufficiantly forit to polymerize. If the HCN polymerization was important forthe origin of life, some regions of the primitive earth mighthave been frozen.