Growth inhibition by tungsten in the sulfur-oxidizing bacterium Acidithiobacillus thiooxidans.

Growth of five strains of sulfur-oxidizing bacteria Acidithiobacillus thiooxidans, including strain NB1-3, was inhibited completely by 50 microM of sodium tungstate (Na(2)WO(4)). When the cells of NB1-3 were incubated in 0.1 M beta-alanine-SO(4)(2-) buffer (pH 3.0) with 100 microM Na(2)WO(4) for 1 h, the amount of tungsten bound to the cells was 33 microg/mg protein. Approximately 10 times more tungsten was bound to the cells at pH 3.0 than at pH 7.0. The tungsten binding to NB1-3 cells was inhibited by oxyanions such as sodium molybdenum and ammonium vanadate. The activities of enzymes involved in elemental sulfur oxidation of NB1-3 cells such as sulfur oxidase, sulfur dioxygenase, and sulfite oxidase were strongly inhibited by Na(2)WO(4). These results indicate that tungsten binds to NB1-3 cells and inhibits the sulfur oxidation enzyme system of the cells, and as a result, inhibits cell growth. When portland cement bars supplemented with 0.075% metal nickel and with 0.075% metal nickel and 0.075% calcium tungstate were exposed to the atmosphere of a sewage treatment plant containing 28 ppm of H(2)S for 2 years, the weight loss of the portland cement bar with metal nickel and calcium tungstate was much lower than the cement bar containing 0.075% metal nickel.

Existence of a tungsten-binding protein in Acidithiobacillus ferrooxidans AP19-3.

A tungsten-binding protein was purified from a plasma membrane preparation of the iron-oxidizing bacterium, Acidithiobacillus ferrooxidans AP19-3 in an electrophoretically homogenous state. The protein was composed of two subunits with apparent molecular masses of 12 and 20.7 kDa. The molecular mass of the native protein was estimated to be 26.4 kDa in the presence of 1.5% 1-o-octyl-D -glucopyranoside (OGL), indicating that the native tungsten-binding protein is a heterodimeric protein. The amounts of tungsten bound to 1 mg of plasma membranes of A. ferrooxidans AP19-3 and the purified tungsten-binding protein at pH 3.0 were 191 and 1506 mug, respectively. In contrast, the amounts of tungsten bound to 1 mg of albumin, aldolase, catalase, chymotrypsinogen A, ferritin, and ferredoxin at pH 3.0 were 13.1, 18.6, 12.8, 16.6, 11.4, and 6.1 mug, respectively. Incubation of the tungsten-binding protein for 1 h with 10 mM Na(2)WO(4) plus 10 mM metal ion, such as NaVO(3), Na(2)MoO(4), CuSO(4), NiSO(4), MnSO(4), CoSO(4), or CdCl(2), did not markedly affect the amount of tungsten bound to the tungsten-binding protein, suggesting that the protein specifically binds tungsten.

Volatilization of mercury by an iron oxidation enzyme system in a highly mercury-resistant Acidithiobacillus ferrooxidans strain MON-1.

A highly mercury-resistant strain Acidithiobacillus ferrooxidans MON-1, was isolated from a culture of a moderately mercury-resistant strain, A. ferrooxidans SUG 2-2 (previously described as Thiobacillus ferrooxidans SUG 2-2), by successive cultivation and isolation of the latter strain in a Fe2+ medium with increased amounts of Hg2+ from 6 microM to 20 microM. The original stain SUG 2-2 grew in a Fe2+ medium containing 6 microM Hg2+ with a lag time of 22 days, but could not grow in a Fe2+ medium containing 10 microM Hg2+. In contrast, strain MON-1 could grow in a Fe2+ medium containing 20 microM Hg2+ with a lag time of 2 days and the ability of strain MON-1 to grow rapidly in a Fe2+ medium containing 20 microM Hg2+ was maintained stably after the strain was cultured many times in a Fe2+ medium without Hg2+. A similar level of NADPH-dependent mercury reductase activity was observed in cell extracts from strains SUG 2-2 and MON-1. By contrast, the amounts of mercury volatilized for 3 h from the reaction mixture containing 7 microM Hg2+ using a Fe(2+)-dependent mercury volatilization enzyme system were 5.6 nmol for SUG 2-2 and 67.5 nmol for MON-1, respectively, indicating that a marked increase of Fe(2+)-dependent mercury volatilization activity conferred on strain MON-1 the ability to grow rapidly in a Fe2+ medium containing 20 microM Hg2+. Iron oxidizing activities, 2,3,5,6-tetramethyl-p-phenylenediamine (TMPD) oxidizing activities and cytochrome c oxidase activities of strains SUG 2-2 and MON-1 were 26.3 and 41.9 microl O2 uptake/mg/min, 15.6 and 25.0 microl O2 uptake/mg/min, and 2.1 and 6.1 mU/mg, respectively. These results indicate that among components of the iron oxidation enzyme system, especially cytochrome c oxidase activity, increased by the acquisition of further mercury resistance in strain MON-1. Mercury volatilized by the Fe(2+)-dependent mercury volatilization enzyme system of strain MON-1 was strongly inhibited by 1.0 mM sodium cyanide, but was not by 50 nM rotenone, 5 microM 2-n-heptyl-4-hydroxy-quinoline-N-oxide (HQNO), 0.5 microM antimycin A, or 0.5 microM myxothiazol, indicating that cytochrome c oxidase plays a crucial role in mercury volatilization of strain MON-1 in the presence of Fe2+.

Volatilization and recovery of mercury from mercury wastewater produced in the course of laboratory work using Acidithiobacillus ferrooxidans SUG 2-2 cells.

The iron-oxidizing bacterium Acidithiobacillus ferrooxidans SUG 2-2 is markedly resistant to mercuric chloride and can volatilize mercury (Hg0) from mercuric ion (Hg2+) under acidic conditions. To develop a microbial technique to volatilize and recover mercury from acidic and organic compound-containing mercury wastewater, which is usually produced in the course of everyday laboratory work in Okayama University, the effects of organic and inorganic chemicals on the mercury volatilization activity of A. ferrooxidans cells were studied. Among 55 chemicals tested, the mercury volatilization from a reaction mixture (pH 2.5) containing resting cells of SUG 2-2 (1 mg of protein) and mercury chloride (14 nmol) was strongly inhibited by AgNO3 (0.05 mM), K2CrO7 (1.0 mM), cysteine (1.0 mM), trichloroethylene (1 microM), and commercially produced detergents (0.05%). However, the strong inhibition by trichloroethylene and detergents was not observed when these organic compounds were chemically decomposed using Fenton's method before the treatment of the wastewater with SUG 2-2 cells. When 20 ml of water acidified with sulfuric acid (pH 2.5) containing ferrous sulfate (3%), diluted mercury wastewater (17.5 nmol of Hg2+) and SUG 2-2 cells (0.05 mg of protein) were incubated for 10 d at 30 degrees C, 47% of the total mercury in the wastewater was volatilized and recovered into a trapping reagent for metal mercury. However, when the organic compounds in the mercury wastewater were decomposed using Fenton's method and then treated with A. ferrooxidans cells, approximately 100% of the total mercury in the wastewater was volatilized and recovered.