Characteristics of a moderately thermophilic and acidophilic iron-oxidizing Thiobacillus

Summary Thiobacillus TH1 is an acidophilic chemolithotrophic heterotroph growing at temperatures up to about 50°C on media containing ferrous iron or pyrite when supplemented with yeast extract or glutathione. Virtually no carbon dioxide fixation occurred during growth on iron with yeast extract. Its DNA contains 48 mol % guanine + cytosine. The organism effects the thermophilic leaching of metals from pyrite, chalcopyrite, CuS, and copper concentrates. Oxidation of soluble ferrous iron at pH 1.6 was competitively inhibited by ferric iron and had a K_m of 7.3 mM FeSO₄.     

Genomic organization of the acidophilic chemolithoautotrophic bacterium Thiobacillus ferrooxidans ATCC 21834.

The genomic organization of the acidophilic chemolithoautotrophic bacterium Thiobacillus ferrooxidans ATCC 21834 has been studied by pulsed-field gel electrophoresis (PFGE). Analysis of its intact DNA, as well as the restriction patterns obtained with several endonucleases, allowed the characterization of one circular chromosome of 2.9 Mb and one plasmid of 8.6 kb. The first complete and highly resolved physical map (86 restriction sites) of the chromosome of an acidophilic obligate chemolithoautotrophic bacterium has been constructed by using endonucleases PmeI, SwaI, XbaI, and SpeI. The rRNA and str operons have been located on the chromosomal physical map.     

A new solid medium for the isolation and enumeration of Thiobacillus ferrooxidans and acidophilic heterotrophic bacteria

A solid medium (FeTSB) was developed for the simultaneous isolation and enumeration of the iron-oxidising bacterium Thiobacilluls ferrooxidans and acidophilic heterotrophic bacteria. The medium consisted of ferrous sulfate, tryptone soya broth and basal salts, solidified with agarose, although bacteriological agar could be substituted for some strains. The medium has been used to isolate bacteria from natural environments and also in laboratory studies of characterised strains. The factors which influence the success of colony growth on solid media are discussed.     

Cloning and expression in Escherichia coli of a recA-like gene from the acidophilic autotroph Thiobacillus ferrooxidans

A recombinant plasmid, pRSR100, containing the functional analogue of the Escherichia coli recA gene was isolated from a genomic library of Thiobacillus ferrooxidans ATCC 33020. The plasmid complemented defects in DNA repair and homologous recombination in E. coli recA mutant strains. Antiserum raised against E. coli RecA protein reacted with the native but defective E. coli HB101 RecA protein; it did not react with protein extracts from the recA deletion mutant E. coli JK696, but it reacted with two protein bands in extracts of E. coli JK696(pRSR100). A single band with an apparent Mr equal to the higher-Mr band in E. coli JK696(pRSR100) was detected in T. ferrooxidans cell extracts with the E. coli RecA antiserum.     

Cytochrome oxidase of an acidophilic iron-oxidizing bacterium, Thiobacillus ferrooxidans, functions at pH 3.5

Cytochrome oxidase of Thiobacillus ferrooxidans was partially purified. The oxidase preparation had haems a and c, and oxidized ferrocytochrome c-552 of the bacterium. The optimal pH of the reaction was 3.5. The enzyme also oxidized the reduced form of rusticyanin, a copper protein of the bacterium. Our results indicate that the reduction of molecular oxygen by this enzyme may occur in the periplasm.     

Effect of ferric sulfate on activity of moderately thermophilic acidophilic iron-oxidizing microorganisms

The effect of high ferric sulfate concentrations on the organisms predominating in biohydrometallurgical processes (bacteria of genus Sulfobaсillus and archaea of the genus Acidiplasma) was studied. Ability of the studied strains to grow and oxidize ferrous iron in the media with 125 to 500 mM ferric sulfate was determined. High concentrations of ferric sulfate significantly inhibited the oxidative activity and growth of the studied microorganisms. Bacteria of the genus Sulfobaсillus were found to be incapable of active iron oxidation in the presence of ferric iron sulfate at concentrations exceeding 250 mM. Archaea of the genus Acidiplasma oxidized ferrous iron completely in the presence of 500 mM Fe³⁺. Microbial growth was suppressed by relatively low ferric sulfate concentrations. Almost no growth occurred at ferric sulfate concentrations exceeding 199 mM, while lysis of the cells of all studied strains was observed at higher Fe³⁺ concentrations. Archaea (genus Acidiplasma, family Ferroplasmaceae) were shown to be more tolerant to high ferric sulfate concentrations than bacteria of the genus Sulfobaсillus. The results obtained may be used for improvement of biohydrometallurgical technologies and are also important for the understanding of the patterns of formation of microbial communities carrying out the technological processes.     

Effect of external pH perturbations on in vivo protein synthesis by the acidophilic bacterium Thiobacillus ferrooxidans.

The response of the obligate acidophilic bacterium Thiobacillus ferrooxidans to external pH changes is reported. When T. ferrooxidans cells grown at pH 1.5 were shifted to pH 3.5, there were several changes in the general protein synthesis pattern, including a large stimulation of the synthesis of a 36-kDa protein (p36). The apparent low isoelectric point of p36, its location in the membrane fraction, and its cross-reaction with anti-OmpC from Salmonella typhi suggested that it may be a porin whose expression is regulated by extracellular pH.     

Selective Adhesion of Thiobacillus ferrooxidans to Pyrite

Bacterial adhesion to mineral surfaces plays an important role not only in bacterial survival in natural ecosystems, but also in mining industry applications. Selective adhesion was investigated with Thiobacillus ferrooxidans by using four minerals, pyrite, quartz, chalcopyrite, and galena. Escherichia coli was used as a control bacterium. Contact angles were used as indicators of hydrophobicity, which was an important factor in the interaction between minerals and bacteria. The contact angle of E. coli in a 0.5% sodium chloride solution was 31°, and the contact angle of T. ferrooxidans in a pH 2.0 sulfuric acid solution was 23°. E. coli tended to adhere to more hydrophobic minerals by hydrophobic interaction, while T. ferrooxidans selectively adhered to iron-containing minerals, such as pyrite and chalcopyrite. Ferrous ion inhibited the selective adhesion of T. ferrooxidans to pyrite competitively, while ferric ion scarcely inhibited such adhesion. When selective adhesion was quenched by ferrous ion completely, adhesion of T. ferrooxidans was controlled by hydrophilic interactions. Adhesion of E. coli to pyrite exhibited a liner relationship on langmuir isotherm plots, but adhesion of T. ferrooxidans did not. T. ferrooxidans recognized the reduced iron in minerals and selectively adhered to pyrite and chalcopyrite by a strong interaction other than the physical interaction.     

Response of Thiobacillus ferrooxidans to phosphate limitation

Abstract: Phosphate ion is an essential nutrient for all cells. Consequently, starvation for this component may constitute a stressing condition which affects the bioleaching capacity of the biomining microorganisms. Therefore, we have studied the manner in which the chemolithotroph Thiobacillus ferrooxidans responds to phosphate limitation. Under these circumstances the bacteria reduced its growth rate, capacity to oxidize ferrous iron and to fix CO,. Concomitant with these changes, the cells showed an increased synthesis of several proteins, some of which were exclusively synthesized during phosphate starvation. When intact cells grown in the absence of phosphate were labelled with ¹²⁵I, several proteins were iodinated in addition of those observed in control cells, suggesting that the lack of phosphate induces some proteins located in the membranes or the periplasmic space of the bacteria. It is expected that by measuring the levels of expression of some of the proteins induced by the shortage of phosphorus, it might be possible to estimate in situ the relative physiological condition of the bacteria in a given bioleaching operation.     

Plasmid and transposon transfer to Thiobacillus ferrooxidans.

The broad-host-range IncP plasmids RP4, R68.45, RP1::Tn501, and pUB307 were transferred to acidophilic, obligately chemolithotrophic Thiobacillus ferrooxidans from Escherichia coli by conjugation. A genetic marker of kanamycin resistance was expressed in T. ferrooxidans. Plasmid RP4 was transferred back to E. coli from T. ferrooxidans. The broad-host-range IncQ vector pJRD215 was mobilized to T. ferrooxidans with the aid of plasmid RP4 integrated in the chromosome of E. coli SM10. pJRD215 was stable, and all genetic markers (kanamycin/neomycin and streptomycin resistance) were expressed in T. ferrooxidans. By the use of suicide vector pSUP1011, transposon Tn5 was introduced into T. ferrooxidans. The influence of some factors on plasmid transfer from E. coli to T. ferrooxidans was investigated. Results showed that the physiological state of donor cells might be important to the mobilization of plasmids. The transfer of plasmids from E. coli to T. ferrooxidans occurred in the absence of energy sources for both donor and recipient.     

氧化亚铁硫杆菌的形态及对Fe2+的氧化研究

在纯培养的条件下,对江西德兴铜矿酸性矿坑水中分离出的一株氧化亚铁硫杆菌(Thiobacillus ferrooxidans)的细胞形态、生长条件以及对Fe2 的氧化进行了初步研究。透射电子显微镜检查的结果表明,其成熟菌体大小均一,有较好的运动性;采用光学显微镜对微生物进行菌群观测和利用血小板计数器法对细菌计数的结果表明,在摇床转速为160r/min的条件下,T.f.菌在9K液体培养基中最适生长条件为温度30℃左右,最佳初始pH 2.0;用重铬酸钾滴定法测定铁的结果表明,在摇床转速为160r/min的条件下,pH值1.7,温度30℃时T.f.菌对Fe2 的氧化速率最大,约为0.58g/L·h。