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PubMed2000

Functional analysis of gapped microbial genomes: amino acid metabolism of Thiobacillus ferrooxidans.

A gapped genome sequence of the biomining bacterium Thiobacillus ferrooxidans strain ATCC23270 was assembled from sheared DNA fragments (3.2-times coverage) into 1,912 contigs. A total of 2,712 potential genes (ORFs) were identified in 2.6 Mbp (megabase pairs) of Thiobacillus genomic sequence. Of these genes, 2,159 could be assigned functions by using the WIT-Pro/EMP genome analysis system, most with a high degree of certainty. Nine hundred of the genes have been assigned roles in metabolic pathways, producing an overview of cellular biosynthesis, bioenergetics, and catabolism. Sequence similarities, relative gene positions on the chromosome, and metabolic reconstruction (placement of gene products in metabolic pathways) were all used to aid gene assignments and for development of a functional overview. Amino acid biosynthesis was chosen to demonstrate the analytical capabilities of this approach. Only 10 expected enzymatic activities, of the nearly 150 involved in the biosynthesis of all 20 amino acids, are currently unassigned in the Thiobacillus genome. This result compares favorably with 10 missing genes for amino acid biosynthesis in the complete Escherichia coli genome. Gapped genome analysis can therefore give a decent picture of the central metabolism of a microorganism, equivalent to that of a complete sequence, at significantly lower cost.

Amino Acids↗

PubMed1994

Characterization of Thiobacillus caldus sp. nov., a moderately thermophilic acidophile.

Two isolates of a novel, moderately thermophilic Thiobacillus species have been studied. The isolates, KU and BC13, are Gram-negative, motile bacteria having a pH optimum for growth of 2-2.5 and an optimum growth temperature of 45 degrees C. Both isolates are capable of chemolithotrophic growth on reduced sulfur substrates. They can also use molecular hydrogen as an electron donor. These two isolates can grow mixotrophically with sulfur or tetrathionate and yeast extract or glucose. The G+C content is 63.1-63.9 mol% and the isolates exhibit no significant DNA homology to any other Thiobacillus species. Strains KU and BC13 both contain ubiquinone Q-8. 16S rRNA analysis indicates that these strains belong to a group of bacteria which includes other chemolithotrophic sulfur oxidizers such as T. ferrooxidans and T. thiooxidans. These characteristics distinguish KU and BC13 from any other species described previously and they thus represent the first acidophilic, thermophilic Thiobacillus species, named T. caldus sp. nov., to be described. The type strain, referred to as strain KU in this paper, has been deposited in the Deutsche Sammlung von Mikroorganismen, Braunschweig, FRG, with the accession number DSM 8584.

Base Composition↗

PubMed1984

Ornithine-containing lipids in Thiobacillus A2 and Achromobacter sp.

20 bacterial strains (corresponding to 16 species) were screened for ornithine lipids. Only two species (Thiobacillus A2 and Achromobacter sp.) turned out to contain ornithine lipids (2.71 mmol/100 g and 0.38 mmol/100 g bacterial dry weight, respectively). In both ornithine lipids, a 3-hydroxy fatty acid was amide-linked to the alpha-amino group of ornithine, a normal fatty acid was ester-linked to the 3-hydroxy group of the former. The predominant fatty acids were 18:1(11) and 3-hydroxy-20:1(13) in Thiobacillus A2, 16:0 and 3-hydroxy-18:1(11) in Achromobacter sp. All monounsaturated fatty acids (with one exception) belonged to the (n-7) family. 11, 12-Epoxy octadecanoic acid was identified among the ester-linked fatty acids of Thiobacillus A2. Phosphatidylcholine was the principal phospholipid in both bacterial species.

Alcaligenes↗

PubMed2000

Development and application of small-subunit rRNA probes for assessment of selected Thiobacillus species and members of the genus Acidiphilium.

Culture-dependent studies have implicated sulfur-oxidizing bacteria as the causative agents of acid mine drainage and concrete corrosion in sewers. Thiobacillus species are considered the major representatives of the acid-producing bacteria in these environments. Small-subunit rRNA genes from all of the Thiobacillus and Acidiphilium species catalogued by the Ribosomal Database Project were identified and used to design oligonucleotide DNA probes. Two oligonucleotide probes were synthesized to complement variable regions of 16S rRNA in the following acidophilic bacteria: Thiobacillus ferrooxidans and T. thiooxidans (probe Thio820) and members of the genus Acidiphilium (probe Acdp821). Using (32)P radiolabels, probe specificity was characterized by hybridization dissociation temperature (T(d)) with membrane-immobilized RNA extracted from a suite of 21 strains representing three groups of bacteria. Fluorochrome-conjugated probes were evaluated for use with fluorescent in situ hybridization (FISH) at the experimentally determined T(d)s. FISH was used to identify and enumerate bacteria in laboratory reactors and environmental samples. Probing of laboratory reactors inoculated with a mixed culture of acidophilic bacteria validated the ability of the oligonucleotide probes to track specific cell numbers with time. Additionally, probing of sediments from an active acid mine drainage site in Colorado demonstrated the ability to identify numbers of active bacteria in natural environments that contain high concentrations of metals, associated precipitates, and other mineral debris.

Acetobacteraceae↗

PubMed1980

Presence of two subunit types in ribulose 1,5-bisphosphate carboxylase from Thiobacillus intermedius.

Ribulose bisphosphate carboxylase (EC 4.1.1.39) has been purified to homogeneity from glutamate-CO2-thiosulfate-grown Thiobacillus intermedius by pelleting the protein from the 93,000 X g supernatant fluid followed by ammonium sulfate fractionation and sedimentation into a discontinuous sucrose density gradient. The molecular weight of the native protein approximated that of the higher plant enzyme (550,000) based on its relative electrophoretic mobility in polyacrylamide disc gels compared with that of standards of known molecular weight, including crystalline tobacco ribulose bisphosphate carboxylase. Sodium dodecyl sulfate electrophoresis in 12% polyacrylamide disc gels and Sephadex G-100 chromatography in the presence of sodium dodecyl sulfate indicated that the purified Thiobacillus protein, like the tobacco enzyme, consisted of two types of nonidentical subunits. The molecular weights of the large and small subunits were estimated to be about 55,000 and 13,000, respectively, by means of sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The carboxylase activity of the protein purified from spinach leaves and T. intermedius responded similarly to the effectors reduced nicotinamide adenine dinucleotide phosphate and 6-phosphogluconate. Contrary to a previous report (K. Purohit, B. A. McFadden, and A. L. Cohen, J. Bacteriol. 127:505-515, 1976), these results indicate that ribulose bisphosphate carboxylase purified from Thiobacillus intermedius closely resembles the higher plant enzyme with respect to quaternary structure, molecular weight, and regulatory properties.

Carboxy-Lyases↗

PubMed1975

Thiobacillus acidophilus sp. nov.; isolation and some physiological characteristics.

After a brief exposition to glucose, Thiobacillus acidophilus was isolated from a culture of iron-grown T. ferrooxidans. Physicochemical analysis of its DNA showed a G+C content of 62.9-63.2%. The new isolate grows best at 25-30 degrees C and at pH 3.0. Growth is possible between pH 1.5 and 6.0. Thiobacillus acidophilus is apparently strictly aerobic. Ammonium salts are the only suitable source of nitrogen. The bacterium is a facultative autotroph. In addition to elemental sulfur, it obtains energy from organic compounds such as D-glucose, D-galactose, D-fructose, D-mannitol, D-xylose, D-ribose, D-arabinose, L-arabinose, sucrose, sodium citrate, malic acid,dl-aspartic acid, and dl-glutamic acid. Thiobacillus acidophilus possesses the key enzymes of the tricarboxylic acid (TCA) cycle including NAD-and NADP-linked isocitric dehydrogenase and alpha-ketoglutarate dehydrogenase, and the key enzymes of the hexose monophosphate pathway (glucose-6-phosphate and 6-phosphogluconate dehydrogenase, and fructose 1,6-diphosphate aldolase). NADH oxidase has been found in particulate fraction of extracts. Rhodanese and thiosulfate oxidase have also been detected.

Aerobiosis↗

PubMed2001

[Tn5037-a Tn21-like mercury transposon, detected in Thiobacillus ferrooxidans].

The 6645-bp mercury resistance transposon of the chemolithotrophic bacterium Thiobacillus ferrooxidans was cloned and sequenced. This transposon, named Tn5037, belongs to the Tn21 branch of the Tn21 subgroup, many members of which have been isolated from clinical sources. Having the minimum set of the genes (merRTPA), the mercury resistance operon of Tn5037 is organized similarly to most of the Gram-negative bacteria mer operons and is closest to that of Thiobacillus 3.2. The operator-promoter region of the mer operon of Tn5037 also has the common (Tn21/Tn501-like) structure. However, its inverted, presumably MerR protein binding repeats in the operator/promoter element are two base pairs shorter than in Tn21/Tn501. In the merA region, this transposon shares 77.4, 79.1, 83.2 and 87.8% identical bases with Tn21, Tn501, T. ferrooxidance E-15, and Thiobacillus 3.2, respectively. No inducibility of the Tn5037 mer operon was detected in the in vivo experiments. The transposition system (terminal repeats plus gene tnpA) of Tn5037 was inactive in Escherichia coli K12, in contrast to its resolution system (res site plus gene tnpR). However, transposition of Tn5037 in this host was provided by the tnpA gene of Tn5036, a member of the Tn21 subgroup. Sequence analysis of the Tn5037 res site suggested its recombinant nature.

Base Sequence↗

PubMed2003

[Isolation of Thiobacillus ferrooxidans and its application on heavy metal bioleaching from sewage sludge].

An iron-oxidizing strain, Thiobacillus ferrooxidans, was isolated from sewage sludge through incubation in the 9K-selected liquor medium followed by inoculation in Leathen solid plate. The strain can survive and propagate rapidly in sewage sludge with the addition of ferrous sulfate or sulfur powder as its energy substance. The pH of the sewage sludge inoculated with Thiobacillus ferrooxidans can be reduced to 1.80 from initial 7.86 within 6 days for bioleaching. Compared to the control without the addition of the strain, the treatment with Thiobacillus ferrooxidans exhibited a high ability to oxide Fe2+ and sulfur in the first 6 days during bioleaching. A batch experiment further demonstrated that as high as 100%, 100% and 80% of the removal efficiency of Cu, Zn and Cr in sewage sludge could be obtained, respectively, in the period of 4-10 days bioleaching by using the isolated strain.

Bioreactors↗

PubMedDate not supplied

[Restriction analysis of Thiobacillus ferrooxidans DNA using pulsed field electrophoresis].

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.

DNA, Bacterial↗

PubMed1992

Thiobacillus ferrooxidans detection using immunoelectron microscopy.

A specific, fast and very sensitive immunoelectron microscopy method was developed to morphologically and serologically distinguish different cultures of iron oxidizers. Bacteria isolated from the acidic waters of "Matahambre" and "Mina Delita" mines (Cuba) were characterized. An antiserum specific to Thiobacillus ferrooxidans did not react with other bacteria also present in the acidic waters of mine drainage. Our results suggest the occurrence of some strains of Thiobacillus ferrooxidans, Thiobacillus thiooxidans and Leptospirillum ferrooxidans in these waters.

Antibodies, Bacterial↗

PubMed2003

[Study on the rejuvenating by isolation and the immobilization of Thiobacillus ferrooxidans].

A strain of Thiobacillus ferrooxidation T1 with stronger oxidation ability and more exuberant vitality was isolated from the liquid with regressive Thiobacillus ferrooxidans by method of dilution butteron on plate. A fixed-bed bioreactor was constructed by immobilized cells of Thiobacillus ferrooxidans with H-2 carriers. In the state of bioreactor operation, a maximum Fe2+ oxidation rate of 7.67g[Fe2+] x h(-1) x L(-1) was attained. We studied the precipitates formed on the surface of the carriers during the operation of the bioreactor. By X-ray diffraction analyses we proved the precipitates were jarosite.

Biocatalysis↗

PubMed1990

[Isolation of transcription promoters from Thiobacillus ferrooxidans and T. acidophilus and its introduction by conjugation to T. intermedius].

The chemolithotrophic acidophilic bacteria, Thiobacillus ferrooxidans is considered as the most important microorganism in relation to copper and uranium bioleaching ability. Since T. ferrooxidans is a strict chemolithotrophic microorganism, its genetic manipulation is a very hard task. Until now, all efforts have been unsuccessful. Therefore, we decided to approach the problem in steps, trying initially to manipulate some T. ferrooxidans related strains. We chose Thiobacillus acidophilus that shares its habitat with T. ferrooxidans and Thiobacillus intermedius, for its heterotrophic nature that makes them easier to grow and suitable for heterologous conjugation. The main objective of this work was directed towards the isolation and characterization of transcription promoters from T. acidophilus and T. ferrooxidans genomic DNA. Four different promoters from T. acidophilus and four from T. ferrooxidans were isolated and sequenced. In order to test their functional capacity in bacterial systems different from E. coli, they were subcloned and transferred to other bacteria. One of these recombinant plasmids was successfully transferred to T. intermedius and from it to Ps. putida. The subcloned promoter was able to confer streptomycin resistance to Ps. putida.

Base Sequence↗

PubMed1984

R68.45 plasmid mediated conjugation in Thiobacillus A2.

Plasmid R68.45 classified into the IncP-1 group which is characterized by a broad host range, was transferred from Pseudomonas aeruginosa PA025 or Escherichia coli into Thiobacillus A2 by conjugation. The R plasmid was stably maintained in Thiobacillus A2 but only neomycin/kanamycin resistance was fully expressed in the new host. Conjugational transfer of R68.45 between different Thiobacillus A2 strains was observed. R plasmid mediated transfer of chromosomal markers has been demonstrated.

Carbenicillin↗

PubMed1963

Attachment of Thiobacillus thiooxidans to sulfur crystals.

Schaeffer, W. I. (Rutgers, The State University, New Brunswick, N.J.), P. E. Holbert, and W. W. Umbreit. Attachment of Thiobacillus thiooxidans to sulfur crystals. J. Bacteriol. 85:137-140. 1963.-Electron micrographs of replicas of sulfur crystals before and after attack by Thiobacillus thiooxidans show that the microorganisms erode the crystal in the area immediately adjacent to the cell. When there are many cells, the entire crystal surface appears eroded.

Acidithiobacillus thiooxidans↗

PubMed1963

ISOLATION AND PROPERTIES OF AN IRON-OXIDIZING THIOBACILLUS.

Razzell, W. E. (British Columbia Research Council, Vancouver, Canada) and P. C. Trussell. Isolation and properties of an iron-oxidizing Thiobacillus. J. Bacteriol. 85:595-603. 1963. - An organism isolated from acidic copper-leaching waters has been shown to oxidize ferrous ions, sulfur, and metallic sulfides but exhibit peculiar responses to thiosulfate. The name Thiobacillus ferrooxidans has been used to describe it. A pH of 2.5 is optimal for growth on iron, sulfur, and metallic sulfides, but cells free from iron can be obtained from growth at pH 1.6, and sulfur cultures adjusted to pH 5.5 readily attain a pH of 1.8. A stationary cultivation procedure appears superior to percolation techniques for studying the oxidation of finely divided metallic sulfides. Concentrations of soluble copper in excess of 1 g per liter were obtained from chalcopyrite in less than 4 weeks. Chalcocite oxidation proceeded in the absence of iron. Sodium chloride inhibits iron oxidation without preventing oxidation of metallic sulfides by the organism.

Acids↗

PubMed1964

GROWTH OF THIOBACILLUS THIOOXIDANS IN SHAKEN CULTURE.

Cook, Thomas M. (Rutgers, The State University, New Brunswick, N.J.). Growth of Thiobacillus thiooxidans in shaken culture. J. Bacteriol. 88:620-623. 1964.-When flasks of sulfur-containing medium were inoculated with Thiobacillus thiooxidans and placed on a shaking machine immediately after inoculation, growth was markedly inhibited. If various wetting agents were provided, or if the ratio of inoculum to sulfur was increased, growth under shaking conditions was possible. Evidence cited shows that the natural wetting agent (or other externally supplied surfactants) is necessary to effect a bacteria-sulfur contact which is stable to shaking conditions.

Acidithiobacillus thiooxidans↗

PubMed1964

EVIDENCE FOR THE PRESENCE OF CERTAIN TRICARBOXYLIC ACID CYCLE ENZYMES IN THIOBACILLUS THIOPARUS.

Cooper, Robert C. (Michigan State University, East Lansing). Evidence for the presence of certain tricarboxylic acid cycle enzymes in Thiobacillus thioparus. J. Bacteriol. 88:624-629. 1964.-Various tricarboxylic acid cycle enzymes appear to be present in Thiobacillus thioparus. Cell-free extracts of T. thioparus were active for a number of tricarboxylic acid cycle enzymes, including aconitase, isocitric dehydrogenase, and malic dehydrogenase. Tests for the presence of fumarase and the condensing enzyme, citrogenase, were inconclusive. Citrate was shown to be active in the metabolism of T. thioparus, but the actual mechanism involved in its formation was not clear. The enzyme, isocitratase, appeared to be absent. Evidence for the presence of succinic dehydrogenase was found in experiments with whole cells. From these results, it would appear that T. thioparus has a terminal respiration pathway similar to that found in many heterotrophic microorganisms.

Carbonic Anhydrases↗

PubMed1964

EFFECT OF PHOSPHATE ION AND 2,4-DINITROPEHENOL ON THE ACTIVITY OF INTACT CELLS OF THIOBACILLUS FERROOXIDANS.

Beck, Jay V. (Brigham Young University, Provo, Utah), and Fred M. Shafia. Effect of phosphate ion and 2,4-dinitrophenol on the activity of cell suspensions of Thiobacillus ferrooxidans. J. Bacteriol. 88:850-857. 1964.-The rate of oxidation of ferrous salts or elemental sulfur by aged cell suspensions, phosphate-depleted cells, or 2,4-dinitrophenol (DNP)-treated cells of Thiobacillus ferrooxidans was increased by addition of orthophosphate salts. The effect was found to be transitory, with the rate gradually approaching that observed prior to phosphate ion addition. The total increased oxygen uptake was observed to be roughly proportional to the amount of phosphate salt added. The efficiency of CO(2) fixation accompanying oxidation of ferrous salts was found to be about 1.7 mumoles of CO(2) fixed per 100 mumoles of O(2) absorbed, in contrast to a value of about 8.0 mumoles of CO(2) fixed per 100 mumoles of O(2) uptake during sulfur oxidation. The rate of oxidation did not affect the CO(2) fixation efficiency. Whereas addition of phosphate salts to aged or phosphate-depleted cells increased slightly the already high efficiency of CO(2) fixation, it did not affect the complete inhibition of CO(2) fixation observed in the presence of 10(-5)m DNP. The results indicate that the phosphate ion is essential for oxidation of the ferrous ion, and that dinitrophenol and other so-called upcoupling agents interfere with phosphate metabolism. The latter may be a result of action at the site of assimilation of the ferrous ion or it may be an effect on the electron-transport system. In any event, it seems obvious that the phosphate ion is converted into a nonactive form in the presence of dinitrophenol-treated cells, because additional quantities of orthophosphate salts cause an immediate, marked restoration of oxidative activity.

2,4-Dinitrophenol↗