Исследован процесс окисления флотоконцентрата сульфидной руды с высоким содержанием пирротина сообществами ацидофильных хемолитотрофных микроорганизмов при 35, 40 и 45°C. Физико-химические параметры жидкой фазы пульпы, результаты анализа твердых остатков после биоокисления и цианирования свидетельствовали о том, что с большей скоростью процесс вело сообщество микроорганизмов, сформировавшееся при 40°C. Степень извлечения золота при 35°C составила 89.34%, при 40°C – 94.59%, при 45°C – 83.25%. При 40°C в жидкой фазе пульпы отмечена самая высокая численность микроорганизмов – 6.01 ? 109 кл/мл. Изменение температуры почти не сказалось на видовом составе сообществ микроорганизмов, за исключением отсутствия Leptospirillum ferriphilum при 35°C, но соотношение видов в сообществах заметно различалось. При 40°C в сообществе микроорганизмов отмечена относительно большая численность Sulfobacillus thermosulfidooxidans, окисляющих с более высокой скоростью железо и элементную серу, чем представители других видов ацидофильных хемолитотрофных микроорганизмов.

Oxidation of flotation concentrate of a pyrrhotite-rich sulfide ore by acidophilic chemolithoautotrophic microbial communities at 35, 40, and 45°C was investigated. According to the physicochemical parameters of the liquid phase of the pulp, as well as the results of analysis of the solid residue after biooxidation and cyanidation, the community developed at 40°C exhibited the highest rate of oxidation. The degree of gold recovery at 35, 40, and 45°C was 89.34, 94.59, and 83.25%, respectively. At 40°C, the highest number of microbial cells (6.01 × 109 cells/mL) was observed. While temperature had very little effect on the species composition of microbial communities (except for the absence of Leptospirillum ferriphilum at 35°C), the shares of individual species in the communities varied with temperature. Relatively high numbers of Sulfobacillus thermosulfidooxidans, the organism oxidizing iron and elemental sulfur at higher rates than other acidophilic chemolithotrophic species, were observed at 40°C.

Optimal aeration conditions were determined and the effect of yeast extract on biooxidation of high-pyrrhotite sulfide ore flotation concentrate in the course of continuous cultivation of an acidophilic chemolithotrophic microbial community was studied in a line of four sequential laboratory reactors; the aeration rate was 3 L/(L min), yeast extract concentration was 0.02%. The gold recovery level was 96.45% at 2.23% elemental sulfur content in the solid residue. The dominant strains identified in the community responsible for biooxidation were Acidithiobacillus caldus OL13-1, At. caldus OL13-3 = At. caldus OL12-3, and an ‘Acidiferrobacter’ strain. Strains Sulfobacillus thermosulfidooxidans OL13-2 = S. thermosulfidooxidans OL12-1 and Ferroplasma acidiphilum OL13-4 = F. acidiphilum OL12-4 were isolated in pure culture and identified.

Селекционировано сообщество микроорганизмов с высокой скоростью окисления флотоконцентрата пирротинсодержащей сульфидной руды, в составе которого в качестве доминирующих культур были идентифицированы Acidithiobacillus caldus ОП-1 и Ferroplasma acidiphilum ОП-2. Отмечено также присутствие культур Acidithiobacillus ferrooxidans ОП-3, Leptospirillum ferriphilum ОП-4 и Sulfobacillus thermosulfidooxidans ОП-5. Результаты анализа твердых остатков процесса показали бльшую степень окисления элементной серы и извлечения золота (90.5%) при поддержании исходного значения рН в реакторе I на уровне 1.8–2.0, чем на уровне 1.6–1.8 (86.3%). льшую степень окисления элементной серы и извлечения золота (90.5%) при поддержании исходного значения рН в реакторе I на уровне 1.8–2.0, чем на уровне 1.6–1.8 (86.3%).

A community of acidochemolithotrophic microorganisms with a high oxidation rate of pyrrhotite-containing sulphide ore flotation concentrate was selected. The Acidithiobacillus caldus OP-1 and Ferroplasma acidiphilum OP-2 cultures were identified to be dominating members. The presence of the Acidithio- bacillusferrooxidans OP-3, Leptospirillumferriphilum OP-4, and Sulfobacillus thermosulfidooxidans OP-5 cultures in the community's composition was also mentioned. The analysis results of solid residues of the process showed a greater elemental sulfur oxidation level and gold recovery when the initial pH value in tank I was maintained at a level of 1.8-2.0 (90.5%) rather than 1.6-1.8 (86.3%).

Методом создания и анализа библиотек клонов гена 16S рРНК изучен видовой состав двух термотолерантных сообществ ацидофильных хемолитотрофных микроорганизмов (АХМ), выделенных из пульпы лабораторных реакторов, в которых проводились процессы окисления концентратов разных золотосодержащих руд. Первое сообщество сформировалось в процессе окисления пирит-арсенопиритного концентрата руды Кючусского месторождения. Клоны бактериального компонента первого сообщества принадлежали к родам Sulfobacillus и Leptospirillum. 32 клона Sulfobacillus были разделены на три группы: Sb. thermosulfidooxidans, Sb. benefaciens и Sb. thermotolerans. 33 клона Leptospirillum были близки к виду Leptospirillum ferriphilum. Все клоны архейного компонента соответствовали виду Ferroplasma acidiphilum. Микроорганизмы данного сообщества были использованы как инокулят для процесса биоокисления другого по составу минерального концентрата – пирротинсодержащего пирит-арсенопиритного концентрата руды Олимпиадинского месторождения, и состав сообщества, сформировавшегося в данном процессе был также исследован. Клоны бактериального компонента второго сообщества также принадлежали к родам Sulfobacillus и Leptospirillum. 14 клонов Sulfobacillus были разделены на две группы: Sb. thermosulfidooxidans (13 клонов) и Sb. thermotolerans (1 клон). 48 клонов рода Leptospirillum были близки к виду L. ferriphilum. Все клоны архейного компонента соответствовали виду F. acidiphilum. В процессе адаптации сообщества к новому окисляемому минеральному субстрату произошли изменения, как в его составе, так и в численном соотношении видов микроорганизмов.

A correlation was observed between the rate of oxidation of pure sulfide minerals (pyrite, pyrrhotite, and arsenopyrite) by communities of acidophilic chemolithotrophic microorganisms (ACM) and the mineral substrate where these communities were formed. The ACM community formed during continuous oxidation of the pyrite-arsenopyrite ore concentrate (Kyuchus deposit) exhibited the highest rate of pyrite oxidation. The highest rate of pyrrhotite oxidation was observed for the ACM community developed during semicontinuous oxidation of the pyrrhotite-containing pyrite-arsenopyrite ore concentrate (Olympiadinskoe deposit), by the communities isolated from the pyrrhotite concentrate, and ore of the Shanuch deposit. In the case of arsenopyrite oxidation, the ACM community isolated during oxidation of the Olympiadinskoe ore concentrate grew without a lag phase. Other communities commenced arsenopyrite oxidation at various rates only after a two-day lag phase. The similarity of the mineralogical characteristics of pure sulfide minerals with those of the minerals in the substrates where the ACM communities developed may affect the rates of oxidation.

Construction and analysis of the 16S rDNA clone libraries was used to investigate the species composition of two thermotolerant communities of acidophilic chemolithotrophic microorganisms (ACM) isolated from the pulp of laboratory reactors used for oxidation of different gold-containing ore concentrates. The first community was formed during oxidation of the pyrite-arsenopyrite ore concentrate from the Kyuchus deposit. The clones of the bacterial component of this community belonged to the genera Sulfobacillus (32 clones) and Leptospirillum (33 clones). The Sulfobacillus clones belonged to three groups: Sb. thermosulfidooxidans, Sb. benefaciens, and Sb. thermotolerans . All Leptospirillum clones were closely related to L. ferriphilum . All clones of the archaeal component belonged to Ferroplasma acidiphilum . The microorganisms of this community were used as inoculum for biooxidation of a different mineral concentrate, the pyrrhotite-containing pyrite-arsenopyrite ore concentrate from the Olympiadinskoe deposit, and the structure of the community formed in the process was investigated. The clones of the bacterial component of the second community also belonged to the genera Sulfobacillus (14 clones) and Leptospirillum (48 clones). The Sulfobacillus clones belonged to the species Sb. thermosulfidooxidans (13 clones) and Sb. thermotolerans (1 clone). All Leptospirillum clones were closely related to L. ferriphilum . All clones of the archaeal component belonged to Ferroplasma acidiphilum . During the adaptation of the community to a new oxidized mineral substrate, both the composition and the ratio of the constituent microbial species changed.

The main representatives of acidophilic chemolithotrophs oxidizing sulfide minerals, ferrous iron, elemental sulfur, and reduced sulfur compounds and forming microbial communities in the natural and technogenic ecosystems with low pH values and high concentrations of heavy metal ions are listed. The species and strain diversity of the communities and environmental factors affecting their composition (temperature, pH value, energy substrate, mineralogical composition of sulfide ore concentrates, the presence of organic substances, and level of aeration) are analyzed. Involvement of mobile genetic elements (IS elements and plasmids) in the structural changes of the chromosomal DNA in the course of switching microbial metabolism to the oxidation of new energy substrates or under increased concentrations of metal ions is shown to be a probable mechanism responsible for the intraspecific genetic heterogeneity of the populations. Importance of determination of the dominant strains of different microbial species in the communities and of their physiological peculiarities for stabilization, optimization, and enhancement of efficiency of biotechnological processes for sulfide mineral oxidation is stressed.

The growth of a microbial community and the oxidation of iron- and sulfur-containing substrates in batch culture during the leaching/oxidation of the flotation concentrate of refractory gold-arsenic sulfide ore were optimized with respect to the following medium parameters: temperature, pH, and requirement in organic substances. It was revealed that the optimum mode is (i) to maintain the pH at 1.6–1.7 and the temperature at 34–35 and 38°C and (ii) to add Corg in the form of yeast extract (0.02%). Mutually beneficial or competitive relationships among groups of microorganisms of the community were established, depending on the cultivation conditions.

Aboriginal and experimental (constructed of pure microbial cultures) communities of acidophilic chemolithotrophs have been studied. The oxidation of elemental sulfur, sodium thiosulfate, and potassium tetrathionate as sole sources of energy has been monitored. The oxidation rate of the experimental community is higher as compared to the aboriginal community isolated from a flotation concentrate of pyrrhotine-containing pyrite-arsenopyrite gold-arsenic sulfide ore. The degree of oxidation of the mentioned S substrates amounts to 17.91, 68.30, and 93.94% for the experimental microbial community and to 10.71, 56.03, and 79.50% for the aboriginal community, respectively. The degree of oxidation of sulfur sulfide forms in the ore flotation concentrate is 59.15% by the aboriginal microbial community and 49.40% by the experimental microbial community. Despite a higher rate of oxidation of S substrates as a sole source of energy by the experimental microbial community, the aboriginal community oxidizes S substrates at a higher rate in the flotation concentrate of pyrrhotine-containing pyrite-arsenopyrite gold-arsenic sulfide ore, from which it was isolated. Bacterial-chemical oxidation of the flotation concentrate by the aboriginal microbial community allows for the extraction of an additional 32.3% of gold from sulfide minerals, which is by 5.7% larger compared to the yield obtained by the experimental microbial community.

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°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°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.

A technology for tank biooxidation of refractory gold-bearing concentrate under variable temperature conditions has been improved: the temperature of the first of two stages was changed from 30°C to 34–36°C. Gold in this concentrate is mainly associated with sulfide minerals: arsenopyrite and pyrite, which underlies a low gold recovery (16.68%) as a result of cyanidation. To resolve the problem, an association of mesophilic acidophilic chemolithotrophic microorganisms and moderately thermophilic bacteria of the Sulfobacillus genus were used for the concentrate oxidation. The composition of the used microbial association was studied; it was shown that it depends upon temperature: at 42°C, the population of the mesophilic thiobacteria decreased, whereas that of thermophilic sulfobacilli enhanced as compared to 36°C. The accepted scheme of the process ensures a high extent of gold recovery (94.6%) within a short space of time for biooxidation (96 h).

The species composition of the microbial association involved in industrial tank biooxidation of the concentrate of refractory pyrrhotite-containing pyrite-arsenopyrite gold-arsenic ore of the Olympiadinskoe deposit at 39°C was studied by cultural and molecular biological techniques. Pure microbial cultures were isolated, their physiological characteristics were investigated, and their taxonomic position was determined by 16S rRNA gene sequencing. The library of 16S rRNA gene clones obtained from the total DNA isolated from the biomass of the pulp of industrial reactors was analyzed. The diversity of microorganisms revealed by cultural techniques in the association of acidophilic chemolithotrophs (Acidithiobacillus ferrooxidans, Leptospirillum ferriphilum, Sulfobacillus thermosulfidooxidans, Ferroplasma acidiphilum, Alicyclobacillus tolerans, and Acidiphilium cryptum) was higher than the diversity of the 16S rDNA clone library (At. ferrooxidans, L. ferriphilum, and F. acidiphilum). The combination of microbiological and molecular biological techniques for the investigation of the biodiversity in natural and anthropogenic microbial communities promotes detection of new phylogenetic microbial groups in these communities.

The composition was studied of the microbial association involved in tank biooxidation of the concentrate of a refractory pyrrhotite-containing pyrite-arsenopyrite gold-arsenic ore from the Olympiadinskoe deposit at 50°C. The two Sulfobacillus thermosulfidooxidans strains predominant in the association were phylogenetically different from the strains used as inocula. The isolates were found to differ significantly both from each other and from the strains that dominated in the processes of biooxidation of a similar concentrate by traditional tank technology at 39°C or at 39°C with treatment of the concentrate with ferric iron prior to biooxidation. These results indicate the strain and species diversity of sulfobacilli in microbial associations involved in biooxidation of the concentrates under different technological modes.

Laboratory experiments in percolators have demonstrated effective bioleaching of copper and zinc, as well as recovery of precious metals from flotation tailings of the Svyatogor concentrator. The defined culture constructed from strains of acidophilic chemolithotrophic microorganisms exhibited higher rates of bioleaching than the indigenous culture obtained from the acid mine drainage. Treatment of the substrate with sulfuric acid (pH 1.24) resulted in recovery of 48.6% of zinc, 28.2% of copper, and 2.8% the total iron. Acidic percolation leaching and biooxidation for 134days with the indigenous and defined cultures resulted in solubilization of 59.5 and 87.0% zinc, 46.0 and 54.5% copper, and recovery of 56.4 and 57.8% gold and 50.5 and 50.9% silver, respectively.

Изучен состав ассоциации микроорганизмов, участвующих в процессе чанового биоокисления при 50°С концентрата упорной пирротинсодержащей пиритно-арсенопиритной золотомышьяковой руды Олимпиадинского месторождения. В составе ассоциации доминировали два штамма Sulfobacillus thermosulfidooxidans, отличающиеся по филогенетическому положению от штаммов, использованных в качестве инокулята. При изучении их свойств были показаны значительные отличия изолятов как друг от друга, так и от штаммов, преобладающих в процессах биоокисления аналогичного по составу концентрата в других условиях: при 39°С с использованием традиционной чановой технологии или при 39°С с введением перед биоокислением дополнительной стадии химической обработки окисляемого концентрата раствором трехвалентного железа. Полученные результаты свидетельствуют о штаммовом и видовом разнообразии сульфобацилл в ассоциациях микроорганизмов, участвующих в процессах биоокисления концентратов в разных технологических режимах.

Quantitative abundance of microbial species within an association was found to depend on the energy substrate and the oxidation temperature of sulfide minerals. The number of microbial cells varied depending on the position of reactor in the chain, i.e., the stage of the energy substrate oxidation. Microbial associations oxidized the energy substrate more efficiently than any of their individual components. The increase in pulp density up to the solid : liquid ratio of 1 : 2.5 had an unfavorable effect on microorganisms comprising microbial associations.

In the process of biooxidation at 39°C in a continuous mode of the gold-arsenic concentrate from the Olympiadinskoe deposit, which was pretreated by chemical leaching with ferric ions, by a microbial association from the BIO department reactors of the Polyus gold mining company, a bacterial culture designated as strain HT-4 was isolated. The bacterium was a spore-forming rod 0.5–0.6 × 1.4–2.0 μm with a flagellum. The optimal temperature for growth and Fe 2+ oxidation was 55°C. The strain grew in the pH range from 1.21 to 2.10 with the optimum at pH 1.6. The organism was incapable of lithotrophic and organotrophic growth. It grew mixotrophically by Fe 2+ oxidation in the presence of 0.02% yeast extract. The DNA G+C base content was 48.6 mol %. Based on comparative phylogenetic analysis of 1472-bp nucleotide sequences of 16S rRNA genes, strain HT-4 was classified as Sulfobacillus thermosulfidooxidans . Analysis by pulse-field gel electrophoresis revealed a unique profile of the Not I fragments of the chromosomal DNA. These results demonstrate the strain and species diversity of sulfobacilli in microbial associations involved in biooxidation of concentrates in different technological conditions. The strain “ S. olympiadicus S-5” dominated in the process of biooxidation of original concentrate not treated with ferric iron, while S. thermosulfidooxidans HT-4 was predominant in biooxidation of the chemically leached concentrate.