[Leaching of copper ore of the Udokanskoe deposit at low temperatures by an association of acidophilic chemolithotrophic microorganisms].

Pure cultures of indigenous microorganisms Acidithiobacillus ferrooxidans strain TFUd, Leptospirillum ferrooxidans strain LUd, and Sulfobacillus thermotolerans strain SUd have been isolated from the oxidation zone of sulfide copper ore of the Udokanskoe deposit. Regimes of bacterial-chemical leaching of ore have been studied over a temperature range from -10 to +20 degrees C. Effects of pH, temperature, and the presence of microorganisms on the extraction of copper have been shown. Bacterial leaching has been detected only at positive values of temperature, and has been much more active at +20 than at +4 degrees C. The process of leaching was more active when the ore contained more hydrophilic and oxidized minerals. The possibility of copper ore leaching of the Udokanskoe deposit using sulfuric acid with pH 0.4 at negative values of temperature and applying acidophilic chemolithotrophic microorganisms at positive values of temperature and low pH values was shown.

[Isolation of an aboriginal bacterial community capable of utilizing cyanide, thiocyanate, and ammonia from metallurgical plant wastewater].

An aboriginal bacterial community capable of degrading cyanide (10 mg/l) and thiocyanate (2 g/l) and eliminating ammonia (120 mg/l) had been isolated from recycled water samples after blast-furnace gas purification of a metallurgical plant wastewater. It was shown that the optimal conditions for this bacterial community were as follows: temperature, 34 degrees C; pH, 8.8-9.0; available organic matter concentration (glucose equivalent), 5 g/l; and dissolved O2 concentration, 8-10 mg/l. This aboriginal community was formed by the bacteria belonging to the genus Pseudomonas.

[Restriction analysis of Thiobacillus ferrooxidans DNA using pulsed field electrophoresis]. / Restriktsionnyi analiz DNK Thiobacillus ferrooxidans s ispol'zovaniem élektroforeza v pul'siruiushchikh raznonapravlennykh élektricheskikh poliakh.

The DNA of Thiobacillus ferrooxidans digested by any of the five restriction endonucleases (DraI, EcoRI, Eco321, HindIII, XbaI) was studied by electrophoresis in the pulsating differently directed electric fields (PF). The influence of PF conditions on the sized row of the divided DNA fragments was studied. Only the XbaI restriction endonuclease was shown to cleave the Thiobacillus ferrooxidans DNA into a number of fragments permitting one to define the sizes of fragments and genome (no more than 2300 bp). The prospects of using the restriction analysis of Thiobacillus ferrooxidans wild type culture for improving its properties in obtaining heavy metals are discussed.

[Cloning and expression of thermostable secretory metalloproteinase genes from two Bacillus brevis strains in Bacillus subtilis cells]. / Klonirovanie i ékspressia genov termostabil'nykh sekretornykh metalloproteinaz dvukh shtammov Bacillus brevis v kletkakh Bacillus Subtilis.

Two closely related genes of thermostable Bac. brevis metalloproteases were cloned and expressed in Bac. subtilis cells. Their restriction maps and directions of transcription were determined. Thermostability and thermal optimum of proteolytic activity of cloned gene products are significantly lower than those of native enzymes. The authors believe that alteration of the enzymes' characteristics may be due to uncorrected folding of thermostable protein in case of its expression in mesophilic bacterial strains.

[The treatment industrial sewage of nonferrous heavy metals using biosorbents]. / Ochistka promyshlennykh stochnykh vod ot tiazhelykh tsvetnykh metallov s pomoshch'iu biosorbentov.

Producers of the antibiotics neomycin and lincomycin were most potent in sorbing nonferrous metals; they removed 99% Zn and Cd and 95% Ni. The degree of metal sorption increased with an increase in solution pH and calcium content in the biomass. Dynamic studies of biosorbents prepared from microbial biomass showed that neomycin industrial waste products are highly efficient in treating galvanic washing water. The total dynamic exchange capacities by nickel and cadmium were 6.85 and 7.16 mg/ml, respectively.

The primary structure and characteristics of the ISAfe600, an insertion sequence from Acidithiobacillus ferrooxidans strains.

A new IS-like element (604 bp) was revealed in the genome of several Acidithiobacillus ferrooxidans strains isolated from diverse biotopes. It includes 26-bp imperfectly matched terminal inverted repeats (TIRs), similar in structure to the TIRs of ISAfel insertion element. The 60-bp DNA fragment adjacent to the right TIR (TIRR) exhibits pronounced homology with the similarly located DNA fragments in ISAfel and IST445, as well as with the internal fragment of ISAfel encoding the transposase gene (nucleotides from 254 to 311 bp). The central section of ISAfe600 is unique and exhibits no homology with any prokaryotic DNA. A duplication of 8 bp of the target DNA was found in the ISAfe600 insertion site. One to four copies of ISAfe600 were revealed by Southern hybridization in the genome of A. ferrooxidans strains studied. The number of ISAfe600 copies varies depending on the growth conditions (energy substrate). Since the open reading frames big enough to encode transposase are not presert in the structure of ISAfe600, it may be a deficient IS element; its translocation is possibly achieved under control of the ISAfel transposase.

[Genotypic and phenotypic polymorphism of environmental strains of the moderately thermophilic bacterium Sulfobacillus sibiricus].

Five cultures of moderately thermophilic spore-forming acidophilic chemolithotrophic bacteria were isolated from the zones of spontaneous heating of pyrrhotine-containing pyrite-arsenopyrite gold-arsenic sulfide ores in an operating open pit (strains B1, B2, B3, OFO, and SSO). Analysis of the chromosomal DNA structure revealed differences between these cultures at the strain level (apart from B3 and SSO, which had identical restriction profiles). All the strains had a similar G + C DNA molar content (47.4-48.3%). The level of DNA reassociation was 85 to 95%. The similarity between the DNA of the type strain Sulfobacillus sibiricus N1 isolated from arsenopyrite ore concentrate and that of these strains (83-93%) indicates that they belong to the same species. The strains had similar values of pH and temperature optimal for growth on ferrous iron (1.6-2.0 and 45-55 degrees C, respectively). They were mixotrophs; Fe(II), S0, and sulfide minerals along with organic compounds were used as energy sources and electron donors. However, the kinetic parameters of growth and substrate oxidation varied from strain to strain. Genetic variety of the strains from diverse ecosystems and environments is possibly the result of the different rates of microevolution processes.

[The dependence of intracellular ATP level on the nutrition mode of the acidophilic bacteria Sulfobacillus thermotolerans and Alicyclobacillus tolerans].

The dynamics of the ATP pool in the aerobic spore-forming acidothermophilic mixotrophic bacteria Sulfobacillus thermotolerans Kr1T and Alicyclobacillus tolerans K1T were studied in the course of their chemolithoheterotrophic, chemoorganoheterotrophic, and chemolithoautotrophic growth. It was established that, during mixotrophic growth, the maximum ATP concentrations in the cells of S. thermotolerans Kr1 and A. tolerans K1 were 3.8 and 0.6 nmol/mg protein, respectively. The ATP concentrations in sulfobacilli and alicyclobacilli during organotrophic growth were 2.2 and 3.1 nmol/mg protein, respectively. In the cells of the obligately heterotrophic bacterium Alicyclobacillus cycloheptanicus 4006T, the maximum ATP concentration was several times higher and reached 12.3 nmol/mg protein. During lithotrophic growth, the maximum values of the ATP concentration in the cells of S. thermotolerans Kr1 and A. tolerans K1 were 0.3 and <0.1 nmol/mg protein, respectively; in the cells of the autotrophic bacterium Acidithiobacillus ferrooxidans TFBk, the ATP content was about 60-300 times higher (17.0 nmol/mg protein). It is concluded that low ATP content is among the possible causes of growth cessation of S. thermotolerans Kr1 and A. tolerans K1 under auto- and heterotrophic conditions after several culture transfers.

[Lithotrophic microorganisms of the oxidative cycles of sulfur and iron].

The review deals with sulfur bacteria (the first chemolithotrophs ever studied) and with the acidophilic bacteria of sulfur and iron cycles which were investigated as a result of Winogradsky's discovery. The diversity of these organisms and the factors and mechanism of its origin are emphasized; their metabolic functions and nutritional regulation are discussed.

[Mechanism of cyanide and thiocyanate decomposition by an association of Pseudomonas putida and Pseudomonas stutzeri strains].

The intermediate and terminal products of cyanide and thiocyanate decomposition by individual strains of the genus Pseudomonas, P. putida strain 21 and P. stutzeri strain 18, and by their association were analyzed. The activity of the enzymes of nitrogen and sulfur metabolism in these strains was compared with that of the collection strains P. putida VKM B-2187T and P. stutzeri VKM B-975T. Upon the introduction of CN- and SCN- into cell suspensions of strains 18 and 21 in phosphate buffer (pH 8.8), the production of NH4+ was observed. Due to the high rate of their utilization, NH3, NH4+, and CNO- were absent from the culture liquids of P. putida strain 21 and P. stutzeri strain 18 grown with CN- or SCN-. Both Pseudomonas strains decomposed SCN- via cyanate production. The cyanase activity was 0.75 micromol/(min mg protein) for P. putida strain 21 and 1.26 micromol/(min mg protein) for P. stutzeri strain 18. The cyanase activity was present in the cells grown with SCN- but absent in cells grown with NH4+. Strain 21 of P. putida was a more active CN- decomposer than strain 18 of P. stutzeri. Ammonium and CO2 were the terminal nitrogen and carbon products of CN- and SCN- decomposition. The terminal sulfur products of SCN- decomposition by P. stutzeri strain 18 and P. putida strain 21 were thiosulfate and tetrathionate, respectively. The strains utilized the toxic compounds in the anabolism only, as sources of nitrogen (CN- and SCN-) and sulfur (SCN-). The pathway of thiocyanate decomposition by the association of bacteria of the genus Pseudomonas is proposed based on the results obtained.

[Dependence of the genotypic characteristics of Acidithiobacillus ferrooxidans on the physical, chemical, and electrophysical properties of pyrites].

This study focused on the effect of physical, chemical, and electrophysical properties of two pyrites, pyrite 1, which had hole-type (p-type) conductivity, and pyrite 2, with electron-type (n-type) conductivity, on the genotypic characteristics of Acidithiobacillus ferrooxidans strains TFV-1 and TFBk, which were isolated from different substrates. After the adaptation of the strains to the pyrites at a pulp density of 1%, pulsed-field electrophoresis revealed changes in the chromosomal DNA of strain TFV-1 adapted to pyrite 1 and strain TFBk adapted to either of the pyrite types. In pyrite-adapted strain TFBk, the plasmid composition was the same as after growth on a medium containing ferrous iron, whereas, in strain TFV-1, changes in plasmid sizes or both in plasmid sizes and plasmid number occurred. After an increase in the density of the pyrite 2 pulp from 1 to 10%, the plasmid number increased from three to four, and, after an increase in the density of the pyrite 1 pulp from 1 to 7%, the plasmid number increased from two to six.

[Dependence of the genotypic characteristics of Acidithiobacillus ferrooxidans on the physical, chemical, and electrophysical properties of pyrites].

Comparison of Acidithiobacillus ferrooxidans strains TFV-1 and TFBk with respect to their capacity to oxidize pyrite 1, with hole-type (p-type) conductivity, or pyrite 2, with an electron-type (n-type) conductivity, showed that, at a pulp density of 1%, both before and after its adaptation to the pyrites, strain TFBk, isolated from a substrate with a more complex mineral composition, grew faster and oxidized the pyrites of both conductivity types more efficiently than strain TFV-1, which was isolated from a mineralogically simple ore. At a pulp density of 3-5%, the oxidation of pyrite 1 by strain TFV-1 and both of the pyrites by strain TFBk began only after an artificial increase in Eh to 600 mV. If the pulp density was increased gradually, strain TFBk could oxidize the pyrites at its higher values than strain TFV-1, with the rate of pyrite 2 oxidation being higher than that of pyrite 1. During chemical oxidation of both of the pyrites, an increase was observed in the absolute values of the coefficients of thermoelectromotive force (KTEMF); during bacterial-chemical oxidation, the KTEMF of pyrite 1 changed insignificantly, whereas the KTEMF of pyrite 2 decreased.

[Effect of pH and temperature on the kinetics of Fe2+ oxidation by Thiobacillus ferrooxidans]. / Vliianie pH i temperatury na kinetiku okisleniia Fe2+ Thiobacillus ferrooxidans.

A correlation has been established between the action of pH and temperature on the activity of Fe2+ oxidation by Thiobacillus ferrooxidans. The rate of Fe2+ oxidation by the bacterium is inhibited at 12 degrees C mainly due to an increase in the pH. Autooxidation of the medium upon hydrolysis of Fe3+ is inhibited at low temperatures. Apparently, the enzymes of Thiobacillus ferrooxidans involved in Fe2+ oxidation are only slightly susceptible to a decrease of temperature to 10--12 degrees C.

[Sulfobacillus, a new genus of thermophilic sporulating bacteria]. / Sulfobacillus--novyi rod termofil'nykh sporoobrazuiushchikh bakterii.

An aerobic facultative thermophilic bacterium was isolated from the ore of the Nikolaev copper-zinc-pyrite deposit in the Eastern Kazakhstan. The bacterium is similar to Thiobacillus ferooxidans in the ability to use various energy subsrates (Fe2+, sulphide minerals and So) and in the acidophilic properties, but differs from it in the thermophilic nature, the structure of a cell wall, the ability to produce spores, and a lower GC content in DNA. Consequently, the organism has been identified as a new species of a new genus, Sulfobacillus thermosulfidooxidans gen. nov., sp. nov. The organism is presumed to play an active role in oxidation of sulphide ores and their warming in deposits.

[Function of surface membrane structures in Thiobacillus thiooxidans]. / O funktsii poverkhnostnykh membrannykh struktur Thiobacillus thiooxidans

The function of the surface membrane structures was studied with cytochemical techniques on ultrathin sections of Thiobacillus thiooxidans. The transport of elementary sulphur inside the cell involves the surface membrane structures, while oxidation of the sulphur to sulphuric acid takes place on the outer surface of the cytoplasmic membrane. The surface membrane structures are supposed also to participate in the primary dissolution of elementary sulphur at the site of contact of the cells with the mineral.

[Nature of the sulfur-containing component and its function in Thiobacillus ferrooxidans]. / O prirode serosoderzhashchego komponenta i ego funktsii v kletkakh Thiobacillus ferrooxidans.

As was shown using various reagents (Ag+, Cd2+) and solvents (ethanol, methanol), Thiobacillus ferrooxidans cells accumulate colloidal sulfur when they grow in the medium 9K containing elemental sulfur. Colloidal sulfur is accumulated in the periplasmic space, in large, bipolarly arranged spherical structures and in simple invaginates of the cytoplasmic membrane. T. ferrooxidans cells accumulate the sulfur at a highest rate during the stationary phase of growth and can use it as a source of energy under the conditions of starvation. The factors causing sulfur accumulation in T. ferrooxidans cells are discussed.

[Thiobacillus ferrooxidans pili]. / Pili Thiobacillus ferrooxidans.

The surface structures of Thiobacillus ferrooxidans were studied. When growing on a medium containing elemental sulphur, the cells possess peritrichously located filaments (piles) whose diameter varies from 4.5 to 7.0 nm and length, from 0.7 to 3.0 mcm. The cells of T. ferrooxidans do not have piles on a medium with ferrous iron. The physiological role of these structures for thiobacilli is discussed.

[Spheroplast study of Thiobacillus ferrooxidans]. / Izuchenie sferoplastov Thiobacillus ferrooxidans.

Spheroplasts were prepared, under the action of lysozyme (200 and 400 mcg/ml) from the cells of Thiobacillus ferrooxidans grown on the Waksman medium with elementary sulfur after their treatment with 0.5M NaCl solutions and exact exposure to sucrose (0.5M). Intracellular structures were revealed on the spheroplasts which could not be detected on intact cells.

[Effect of Fe3+ ions on Thiobacillus ferrooxidans oxidation of ferrous oxide at various temperatures]. / Vliianie ionov Fe3+ na okislenie Thiobacillus ferrooxidans zakisnogo zheleza pri razlichnoi temperature.

The inhibition of the rate of ferrous iron oxidation by Thiobacillus ferrooxidans with ferric ions was shown to depend on their concentration, the temperature of the medium, and the phase of the cultural growth. Ferrous iron oxidation was inhibited with ferric ions both at low (below 1.0 g/l) and high (ca. 10 g/l) concentrations, the process being temperature dependent: the rate of inhibition decreased with a fall of the temperature. The bacterium was most sensitive to unfavourable Fe3+ concentrations at 8-26 degrees C during the lag phase; Fe2+ oxidation was inhibited during the exponential phase of growth only at 26 degrees C. The Ki for Fe3+ at 27, 10 and 5 degrees C were 3.09-3.39, 11.4-22.8 and 46.0 g/l Fe3+, respectively. Therefore, the affinity of the enzyme for the inhibitor Fe3+ decreases with a fall of the temperature.