Sulphide as an inhibitor and electron donor for the cytochrome c oxidase system

Anomalies both kinetic and equilibrium in nature are described for the inhibition of cytochrome c oxidase activity by sulphide in the isolated enzyme and in submitochondrial particles. These anomalies are related to the involvement of more than 1 mol of sulphide in the blockage of one cytochrome aa3 centre. Sulphide reduces resting cytochrome a3, a reaction that results in oxygen uptake and the loss of a sulphide molecule. Sulphide can also reduce cytochromes c and a; in the former case, a part of the one-equivalent oxidation product, presumed to be the SH radical, reacts with oxygen. Such oxygen uptake is also seen under aerobic conditions when ferricyanide reacts with sulphide. Three phases are identified in the inhibitory interaction of sulphide with the cytochrome c oxidase enzyme itself: an initial rapid reaction involving sulphide oxidation, oxygen uptake, and conversion of cytochrome aa3 into the low-spin "oxyferri" form; a subsequent step in which sulphide reduces cytochrome a; and the final inhibitory step in which a third molecule of sulphide binds the a3 iron centre in the cytochrome a2+ a3 3+ (oxy) species to give cytochrome a2+ a3 3+ H2S. the initial events parallel some of the events in the interaction of the cytochrome c-cytochrome aa3 system with monothiols; the final inhibitory event resembles that with cyanide.     

Semblant Land Plants from the Middle Ordovician of the Prague Basin Reinterpreted as Animals

Two plant-like fossil are described from the middle Ordovician of the Prague Basin, Czech Republic. Both fossils bear a strong superficial resemblance to early land plants, but anatomical data indicate an affinity with animals. New evidence on internal structure demonstrates that the putative plant Boiophyton pragense Obrhel is a dendroid graptolite. The relationships of other heavily coalified, branched axes found in associated strata are more obscure. Scanning electron microscopy of surface features suggests that these plant-like remains are fossils of the collagenous stipes of gorgonacean octocorals.     

Prevention of sulphide accumulation and phosphate mobilization by the addition of iron(II) chloride to a reduced sediment: an enclosure experiment

1. In an enclosure experiment carried out in a ditch receiving sulphate-enriched seepage water, iron(II) chloride was added to the sediment. In the sediment pore water of the iron-treated enclosures sulphide levels decreased to very low values (<1 μmol 1^?1) immediately after the iron addition while in the control enclosures sulphide reached values up to 500 μmoll^?1. 2. The sulphide levels in the sediment pore water were also strongly correlated with temperature. In summer, phosphate mobilization was observed in the non-treated enclosures while in the iron-treated enclosures phosphate levels remained low. 3. Total phosphate levels increased greatly in the water layer of the non-treated enclosures, coincident with an algal bloom and increased turbidity. It is suggested that phosphate mobilization in summer is caused by the reduction of iron(III) phosphate complexes and in this high sulphate water body probably also by the reduction of iron(III) by sulphide and the consequential precipitation of iron(II) sulphide. 4. Iron addition appeared to prevent sulphide toxicity in Potamogeton acutifolius Link which was planted in the enclosures immediately after iron(II) addition. In the non-iron-treated enclosures P. acutifolius plants decayed within a few weeks probably as a result of sulphide toxicity.     

Biogeochemical interactions between iron and sulphate in freshwater wetlands and their implications for interspecific competition between aquatic macrophytes

1. Wetlands are threatened by desiccation, eutrophication and changing water quality, generally leading to greatly altered biogeochemical processes. Sulphate pollution can lead to severe eutrophication and sulphide toxicity, but may also interact with the availability of iron and other metals. 2. In the present study, we examined the biogeochemical interactions between sulphate and iron availability, and their effects on aquatic macrophytes, in a field experiment with enclosures. The natural iron supply by groundwater was mimicked by adding iron to the sediment, and the effect of increased sulphate concentrations in the surface water was also studied. The enclosure experiment was performed in a mesotrophic, anaerobic ditch in a peat meadow reserve in the Netherlands. In all enclosures, three Stratiotes aloides plants were introduced to serve as indicator species. 3. Addition of sulphate led to the mobilisation of phosphate, whereas addition of iron or both iron and sulphate did not affect P mobilisation. Growth of S. aloides was decreased by both iron addition and sulphate addition (sulphide toxicity). Addition of iron under sulphidic conditions, however, led to mutual detoxification of both toxicants (iron and sulphide) and did not decrease S. aloides growth. The uptake of metals was highest in the treatment involving sulphate addition, probably as a result of increased mineralisation of the peat soil. 4. Growth of Elodea nuttallii, which grew naturally in the enclosures, was stimulated by iron or iron plus sulphate addition. It did not, however, grow in the enclosures with sulphate addition, as a result of sulphide toxicity or sulphide‐induced iron deficiency. Under iron‐rich conditions, E. nuttallii appeared to be a better competitor than S. aloides and depressed the growth of the latter species. 5. We propose that the growth of S. aloides is directly regulated by interactions between sulphide and iron and indirectly by the effects of both compounds on the competitive strength of E. nuttallii. In general, we conclude that biogeochemical interactions between sulphate and iron can have a strong influence on plant species composition in freshwater wetlands, because of direct effects or changes in the competitive strength of plant species related to differential sensitivity to either iron or sulphide.     

Sulphide release from anoxic sediments in relation to iron availability and organic matter recalcitrance and its effects on inorganic phosphorus recycling

This research aims to analyse the sediment capacity to buffer free sulphide release in three coastal lagoons which differ in terms of eutrophication level, tide influence and primary producer communities. A preliminary estimate of soluble reactive phosphorus (SRP) regeneration coupled with sulphide fluxes is also made. Sediment profiles of ferrous and ferric iron and reduced sulphur pools were determined in three stations in the Bassin d'Arcachon (South West France), in one site in the Etang du Prévost lagoon (Southern France), and in three stations in the Sacca di Goro lagoon (Northern Italy). Laboratory experiments were also conducted by incubating sediment slurries. Slurries from the French lagoons were also enriched with about 2% d.w. of organic detritus obtained from the dominant macrophytes of each site, namely Zostera noltii and Ruppia cirrhosa (Bassin d'Arcachon), and Ulva rigida (Etang du Prévost). In the Sacca di Goro, slurry experiments were conducted at two sites with different salinity range, sediment composition and hydrodynamics.Field data showed that concentrations of available iron (Fe(II)+Fe(III)) ranged from a minimum of 28.5 µmol cm^–3 (Etang du Prévost) to a maximum of 275.7 µmol cm^–3 (Sacca di Goro). Moreover, in the French lagoons, acid volatile sulphide (AVS) accumulation in the superficial sediment was related to ferrous iron concentrations. Laboratory experiments showed that in spite of strong reducing conditions, sulphide and SRP release was weaker in iron-rich sediments and in those enriched with the most refractory organic matter. The highest fluxes were detected in sediment slurries from the Etang du Prévost, which had the lowest iron content, supplied by 2% of the labile detritus from Ulva rigida. In this case, SRP release was directly related to sulphide production.Two factors seem significant to evaluate the buffer capacity against free sulphide and SRP release from anoxic sediment: organic matter biodegradability, which forces sediment toward reducing conditions, and iron availability, which can affect sulphide mobility as well as the iron hydroxide-phosphate-sulphide system.     

How soil characteristics and water quality influence the biogeochemical response to flooding in riverine wetlands

Although phosphate concentrations have been reduced, the rivers Meuse and Rhine are still polluted with sulphate, which most probably affects vegetation development in newly created riverine wetlands. The influence of flooding with river water rich in sulphate was tested on three soil types from floodplains of the river Meuse using flow-through and batch experiments. Soils were selected for contrasting concentrations of iron and organic matter and originated from a floating fen (iron-poor, organic), an alder carr (iron-rich, organic) and a clay pit (iron-rich, low in organic matter). Flooding induced mobilisation of phosphate. Sulphate only enhanced this effect in the alder carr soil, where sulphide and phosphate competed for binding to iron. Only in the floating fen soil did the addition of sulphate result in the formation of free sulphide, which reduced the growth of Glyceria maxima, serving as a phytometer. In addition, the floating soil started to sink, due to falling methane concentrations. In the different soil types methane production was hampered by the presence of more favourable electron acceptors such as sulphate in the water and Fe(III) in the soil. It was concluded that the effects of inundation with sulphate-polluted water strongly depend on the soil type: under iron-poor circumstances, free sulphide may accumulate, leading to phytotoxicity, while in soils rich in iron, sulphide toxicity is prevented, but phosphate availability may be increased. In addition, shortage of easily degradable organic matter can limit the formation of potential toxicants such as ammonium, iron and sulphide. Results are discussed in terms of their implications for nature management.     

PALAEOSTACHYA DIRCEI N. SP., AN AUTHIGENICALLY CEMENTED EQUISETALEAN STROBILUS FROM THE MIDDLE PENNSYLVANIAN OF SOUTHERN ILLINOIS

Equisetalean strobili normally are encountered as disarticulated organs. This condition has necessitated the creation of 12 form genera to accommodate the various morphological architectures and anatomical configurations. Taxonomically useful characteristics, which are discernable in permineralized specimens, are rarely distinguishable in coalified compressions due to their destruction during diagenesis and coalification. Therefore, genera established on the position of sporangiophore-trace origin, such as Schimperia Remy and Remy, are untenable when dealing with coalified compressions. Although the two largest genera of strobili, Calamostachys Schimper and Palaeostachya Weiss, may be synonymous, it is proposed that these form genera be retained when dealing with coalified compression specimens. Each genus provides a particular architectural concept for specimens which may not be assignable to the species level. Calamostachys and Palaeostachya are highly overspeciated genera. It is suggested that characteristics necessary to delimit new species include bract height, degree of bract fusion, disposition of bracts, bract: sporangiophore ratio, number of sporangia per sporangiophore, and sexual status.     

Corroding iron as a hydrogen source for sulphate reduction in growing cultures of sulphate-reducing bacteria

Summary In anaerobic corrosion experiments, hydrogenase-positiveDesulfovibrio strains, grown with limiting lactate concentrations in the presence of steel wool, formed more sulphide than expected or observed with lactate alone. The additional sulphide obviously originated from sulphate reduction with cathodically formed hydrogen from the steel surface. The hydrogenasenegativeD. sapovorans did not produce additional sulphide. The observations agree with the theory of von Wolzogen Kühr and van der Vlugt (1934) that explains anaerobic corrosion as a cathodic depolarization of iron surfaces by hydrogen-consuming sulphate-reducing bacteria. The influence of the iron surface area, the salt concentration and the pH-value on the utilization of cathodically formed hydrogen was investigated. The significance of an additional organic electron donor for the corrosion of iron in aqueous environments is discussed.     

Role of Thiobacillus ferrooxidans and sulphur (sulphide)-dependent ferric-ion-reducing activity in the oxidation of sulphide minerals

 Thiobacillus ferrooxidans oxidized the sulphide minerals e.g., pyrite, pyrrhotite and copper concentrate under anaerobic conditions in the presence of ferric ion as sole electron acceptor. Copper and iron were solubilized from sulphide ores by the sulphur (sulphide)-dependent ferric-ion oxidoreductase activity. Treatment of resting cells of T. ferrooxidans with 0.5% phenol for 30 min completely destroyed the iron- and copper-solubilizing activity. The above treatment destroyed the sulphur(sulphide)-dependent ferric-ion-reducing activity completely but did not affect the iron-oxidizing activity. The results suggest that sulphur(sulphide)-dependent ferric-ion-reducing activity actively participates in the oxidation of sulphide minerals under anaerobic conditions. The activity of sulphur(sulphide)-dependent ferric ion reduction in the solubilization of iron and copper from the sulphide ores were also observed under aerobic conditions in presence of sodium azide (0.1 μmol), which completely inhibits the iron-oxidizing activity. Received: 23 May 1995/Received revision: 10 October 1995/Accepted: 16 October 1995     

Indirect bioleaching of covellite by Thiobacillus thiooxidans with an oxidant agent

Summary High copper extraction by the action of Thiobacillus thiooxidans (T.t.) on covellite in presence of iron(III) is explained by indirect mechanism, in which T.t. only oxidizes the layer of sulphur that covers the sulphide surface allowing sulphide oxidation by iron(III). In cultures on elemental sulphur with iron(III) T.t. is not able to use iron(III) as an acceptor of electrons in sulphur oxidation; iron(III) only oxidizes those intermediates which were generated in the aerobic oxidation of sulphur.     

A late Wenlock flora from Co. Tipperary, Ireland

An assemblage of macroplants preserved as highly coalified compressions which lack anatomy is described from a Wenlock locality in County Tipperary, Ireland. Most of the fertile specimens are assigned to Cooksonia Lang. The taxonomic status of this genus is discussed. Some poorly preserved palynomorphs, including miospores, acritarchs, chitinozoans and a variety of tubes, have been isolated from associated sediments, but the age of the flora is based on graptolites. Sedimentological and palaeontological studies of the region are summarized. They provide little direct evidence for the habitats of the plants which are considered to have been terrestrial. The relevance of this flora to the current debate on the colonization of the land is evaluated and it is concluded that these plants provide the earliest record of erect fertile land plants of possible pteridophyte affinity.     

Novel conducting tissues in Lower Devonian plants

Elongate cells presumed to comprise water-conducting tissues are described from the central regions of short lengths of two naked, stomatiferous, coalified, axial fossils from Lochkovian (Lower Devonian) fluvial rocks in the Welsh Borderland. In one, a discrete central strand is predominantly composed of uniformly thickened cells that are compared with central tissues in coeval plants, e.g. Aglaophyton , and the hydroids of extant mosses. The other has at least two types of cells with pits of plasmodesmata dimensions that perforate only the inner layer of a bilayered wall. These are compared with liverwort and Takakia hydroids and the coeval S-type tracheids that characterize the Rhyniopsida. The affinities of the two axes remain equivocal. The relevance of plasmodesmata-derived pits to the evolution of diversity in water-conducting elements in early cmbryophytes is discussed.     

A case study for late Archean and Proterozoic biogeochemical iron‐ and sulphur cycling in a modern habitat—the Arvadi Spring

As a consequence of Earth's surface oxygenation, ocean geochemistry changed from ferruginous (iron(II)‐rich) into more complex ferro‐euxinic (iron(II)‐sulphide‐rich) conditions during the Paleoproterozoic. This transition must have had profound implications for the Proterozoic microbial community that existed within the ocean water and bottom sediment; in particular, iron‐oxidizing bacteria likely had to compete with emerging sulphur‐metabolizers. However, the nature of their coexistence and interaction remains speculative. Here, we present geochemical and microbiological data from the Arvadi Spring in the eastern Swiss Alps, a modern model habitat for ferro‐euxinic transition zones in late Archean and Proterozoic oceans during high‐oxygen intervals, which enables us to reconstruct the microbial community structure in respective settings for this geological era. The spring water is oxygen‐saturated but still contains relatively elevated concentrations of dissolved iron(II) (17.2 ± 2.8 μM) and sulphide (2.5 ± 0.2 μM) with simultaneously high concentrations of sulphate (8.3 ± 0.04 mM). Solids consisting of quartz, calcite, dolomite and iron(III) oxyhydroxide minerals as well as sulphur‐containing particles, presumably elemental S⁰, cover the spring sediment. Cultivation‐based most probable number counts revealed microaerophilic iron(II)‐oxidizers and sulphide‐oxidizers to represent the largest fraction of iron‐ and sulphur‐metabolizers in the spring, coexisting with less abundant iron(III)‐reducers, sulphate‐reducers and phototrophic and nitrate‐reducing iron(II)‐oxidizers. 16S rRNA gene 454 pyrosequencing showed sulphide‐oxidizing Thiothrix species to be the dominating genus, supporting the results from our cultivation‐based assessment. Collectively, our results suggest that anaerobic and microaerophilic iron‐ and sulphur‐metabolizers could have coexisted in oxygenated ferro‐sulphidic transition zones of late Archean and Proterozoic oceans, where they would have sustained continuous cycling of iron and sulphur compounds.     

Corrosion-enhancing potential of Shewanella putrefaciens isolated from industrial cooling waters

Microbially influenced corrosion (MIC) is catalysed by a series of metabolic activities of selected micro-organisms, notably by oxidation of cathodic hydrogen by hydrogenase, by hydrogen sulphide and by reduction of ferric iron. The sulphate-reducing bacteria are considered to be the most common catalyst of MIC, whereas the role of other bacteria has been neglected. This study examined the corrosive potential of the facultative sulphide producer, Shewanella putrefaciens , isolated from an industrial cooling water system. Shewanella putrefaciens was shown to reduce ferric iron and sulphite under anaerobic conditions and with ferric iron being the preferred electron acceptor. The isolate could utilize cathodic hydrogen as an energy source, especially when using sulphite as a terminal electron acceptor. In pure culture corrosion experiments, the highest mass loss of mild steel was observed in the presence of sulphite as sole electron acceptor, although mass loss was also detected where ferric iron was the sole electron acceptor. Our data indicate that S. putefaciens plays a role in MIC as it was able to catalyse a variety of corrosion-promoting reactions and to corrode mild steel under pure culture conditions.     

The purification and properties of the respiratory-chain reduced nicotinamide–adenine dinucleotide dehydrogenase of Torulopsis utilis

1. An NADH-ferricyanide reductase activity has been isolated from the respiratory chain of Torulopsis utilis by using detergents. The isolated enzyme contains non-haem iron, acid-labile sulphide and FMN in the molar proportions 27.5:28.4:1. The preparation is free of FAD and largely free of cytochrome. 2. The enzyme catalyses ferricyanide reduction by NADPH at about 1% of the rate with NADH, and reacts poorly with acceptors other than ferricyanide. The rates of reduction of some acceptors are, as percentages of the rate with ferricyanide: menadione, 0.35%; lipoate, 0.01%; cytochrome c, 0.065%; dichlorophenolindophenol, 0.35%; ubiquinone-1, 0.08%. 3. Several properties of submitochondrial particles of T. utilis (non-haem iron, acid-labile sulphide, FMN and an NADH-reducible electron-paramagnetic-resonance signal) were found to co-purify with the NADH-ferricyanide reductase activity. Thus about 70% of the FMN and, within the limits of accuracy of the experiments, 100% of the non-haem iron and acid-labile sulphide of submitochondrial particles derived from T. utilis cells grown under conditions of glycerol limitation (but relatively low iron availability) can be attributed to the NADH-ferricyanide reductase. 4. It was also shown that the component of submitochondrial particles specifically bleached at 460nm by NADH [species 1 of Ragan & Garland (1971)] co-purifies with the NADH-ferricyanide reductase. 5. This successful purification of an NADH dehydrogenase from T. utilis forms a starting point for investigating the molecular properties of phenotypically modified mitochondrial NADH oxidation pathways that lack energy conservation between NADH and the cytochromes.     

On-line measurement of dissolved and gaseous hydrogen sulphide in anaerobic biogas reactors

The paper presented demonstrates the calibration of a silicone membrane probe for measurement of hydrogen sulphide in liquid and gas phase. The probe is connected to a quadrupole mass spectrometer as detector. The calibration of the probe results in linear calibration functions for different liquids and the gas phase. An example of the application of the measuring device for on-line measurement is reported for an experiment where sulphide is precipitated as iron sulphide by the addition of ferrous chloride. As a consequence of the addition of ferrous chloride, the concentration of H₂S in the biogas rapidly decreases from 4.2% to 1.0% (by volume). The inhibition of the anaerobic treatment process is calculated on the basis of the reduction of dissolved total organic carbon before and during the experiment. The reduction of dissolved total organic carbon before the experiment starts is constant at 60%, rising to a maximum of 70% during the addition of FeCl₂. The difference in the conversion rate corresponds to an inhibition of about 14%. The gas production increases from 7.51 l^–1 day^–1 to 8.51 l^–1 day^–1. This inhibition observed before the addition of FeCl₂ is caused by 65 mg/l undissociated hydrogen sulphide in the liquid phase as calculated from the data obtained after precipitation of sulphide as zinc sulphide. The data show clearly that the conversion of acetic acid to methane is inhibited by dissolved H₂S. The concentration of acetic acid drops sharply from about 25 mM to 15 mM after the FeCl₂ dosage has been started. The concentration of propionic acid decreases slightly from 12 mM to 9 mM. Most of the iron introduced during the experiment is immediately precipitated. The maximum concentration of dissolved iron measured in the effluent is 93 mg/l.     

The importance of sediment phosphorus release in the restoration of very shallow lakes (The Norfolk Broads,England) and implications for biomanipulation

Phosphorus release from the sediments of very shallow lakes, the Norfolk Broads, can be as high as 278 mgP m^-2 d^-1. These high rates are associated with high total sediment Fe:P ratios and occur when sulphide from sulphate reduction removes Fe(II) from the pore water. There is also evidence that bioturbation from benthic chironomids can enhance phosphorus release rates, particularly in sediments low in total iron. The release of phosphorus from the sediments of these lakes is delaying restoration following the control of phosphorus from sewage discharges. Biomanipulation is being used in these lakes to create clear water and re-establish aquatic macrophytes. This removal of fish has allowed larger populations of benthic chironomid larvae to develop which may result in an increase in the rate of phosphorus release and changes to the pore profiles of dissolved phosphorus, soluble iron and free sulphide.     

The effect of pressure and temperature on sulphate-reducing bacteria and the action of biocides in oilfield water injection systems

The effects of elevated pressures and temperatures on the growth, morphology and metabolic activity of sulphate-reducing bacteria, isolated from the North Sea, are described. Pressure/temperature profiles, growth curves and sulphate reduction rates are presented for several isolates. The maximum pressure and temperature that supported growth were 65 000 KPa and 45°C respectively. The results are discussed in connection with water injection into oil-bearing reservoirs where there is a concern that generation of hydrogen sulphide by sulphate-reducing bacteria may lead to increased hydrogen sulphide levels (souring) in oil and gas, and to corrosion problems in production facilities. The bacteriostatic effects of a number of commercial biocides were enhanced at elevated hydrostatic pressures and temperatures.     

Bioleaching of metallic sulphides withThiobacillus ferrooxidans in the absence of iron (II)

Bioleaching of metallic sulphides withThiobacillus ferrooxidans in the absence of iron (II) was studied using pure sulphides and mixtures. The direct mode of bacterial action was analysed with respect to sulphide solubility, exposed solid surfaces and bacterial attachment to the solids. Bioleaching of mixed sulphides showed enhancement of metal extraction in comparison with pure sulphides which suggests metal extractions would be better from polymetallic sulphide ores than from similar matrices with only one sulphide.     

Isolation and biochemical characterization of maleic-acid hydratase, an iron-requiring hydro-lyase

A procedure for the isolation of maleic acid hydratase (D-malate hydro-lyase, EC 4.2.1.31) of about 95% purity from rabbit kidneys is described. The enzyme consists of a single polypeptide chain of 582 amino-acid residues with an approximate molecular mass of 68 kDa. The enzyme is very unstable and has an absolute requirement for chloride ions. Addition of sodium sulphide during the purification process was essential to maintain the enzyme in an activatable state. The pure preparation has low activity but responds to activation with Fe2+ ions, Na2S and a thiol. The sequence of adding the activating reagents is critical to achieve optimal activity. Ni2+ and to a lesser extent Co2+ can replace iron in the activation process. The enzyme incorporates 4-5 mol iron/mol and 4.5-6 mol sulphide/mol during activation. In this process an [Fe-S] cluster appears to be built up, as indicated by optical and electron paramagnetic resonance (EPR) spectroscopy. In activated samples exposed to air the [Fe-S] cluster is EPR-detectable through an axial signal with g = 2.01 and g = 2.029 whose temperature and power saturation characteristics were similar to those of other [3Fe-xS] clusters. The activated enzyme, however, is readily inactivated even upon minor manipulation with destruction of the iron-sulfur core.