Studies on the effect of mercury and organomercurial on the growth and nitrogen fixation by mercury‐resistant Azotobacter strains
TL;DR: Five nitrogen-fixing Azotobacter strains isolated from agricultural farms in West Bengal, India, were resistant to mercuric ion and organomercurials and NADPH and GSH might have a role in suppressing the inhibition of N 2 -fixation in the presence of Hg compounds.
Abstract: Five nitrogen-fixing Azotobacter strains isolated from agricultural farms in West Bengal, India, were resistant to mercuric ion and organomercurials. Resistance of Hg-resistant bacteria to mercury compounds is mediated by the activities of mercuric reductase and organomercurial lyase in the presence of NADPH and GSH as cofactors. These bacteria showed an extended lag phase in the presence of 10-50 μmol l -1 HgCl 2 . Nitrogen-fixing ability of these isolates was slightly inhibited when the mercury-resistant bacterial cells were preincubated with 10 μmol l -1 HgCl 2 . Acetylene reduction by these bacteria was significantly inhibited (91-97%) by 50 μmol l -1 HgCl 2 . However, when GSH and NADPH were added to the acetylene reduction assay mixture containing 50 nmol l -1 HgCl 2 , only 42-50% inhibition of nitrogenase activity was observed. NADPH and GSH might have a role in suppressing the inhibition of N 2 -fixation in the presence of Hg compounds either by assisting Hg-detoxifying enzymes to lower Hg concentration in the assay mixture or by formation of adduct comprising Hg and GSH which is unable to inhibit nitrogen fixation.
Citations
More filters
TL;DR: The isolate and the enzyme detected were effective in reducing Hg (II) to Hg(0), showing the potential to develop bioremediation technologies and processes to clean-up the environment and waste contaminated with mercury.
Abstract: This study aimed to isolate mercury resistant bacteria, determine the minimum inhibitory concentration for Hg, estimate mercury removal by selected isolates, explore the mer genes, and detect and characterize the activity of the enzyme mercuric (II) reductase produced by a new strain of Pseudomonas sp. B50A. The Hg removal capacity of the isolates was determined by incubating the isolates in Luria Bertani broth and the remaining mercury quantified by atomic absorption spectrophotometry. A PCR reaction was carried out to detect the merA gene and the mercury (II) reductase activity was determined in a spectrophotometer at 340 nm. Eight Gram-negative bacterial isolates were resistant to high mercury concentrations and capable of removing mercury, and of these, five were positive for the gene merA. The isolate Pseudomonas sp. B50A removed 86% of the mercury present in the culture medium and was chosen for further analysis of its enzyme activity. Mercuric (II) reductase activity was detected in the crude extract of this strain. This enzyme showed optimal activity at pH 8 and at temperatures between 37°C and 45°C. The ions NH4+, Ba2+, Sn2+, Ni2+ and Cd2+ neither inhibited nor stimulated the enzyme activity but it decreased in the presence of the ions Ca2+, Cu+ and K+. The isolate and the enzyme detected were effective in reducing Hg(II) to Hg(0), showing the potential to develop bioremediation technologies and processes to clean-up the environment and waste contaminated with mercury.
64 citations
Cites background from "Studies on the effect of mercury an..."
...[30] found that the ability f microorganisms to remove mercury from the culture medium as affected by high concentrations of HgCl2, suggesting that it ay be due to sequestering of intracellular Hg by cell components hat bind to the metal....
[...]
01 Oct 1990
TL;DR: It was determined that the toxicities of different mercury salts were influenced by the nature of the co-ion, and complexes of Hg(II) with histidine, chloride, and cysteine were the species primarily responsible for toxicity in the M-IIY medium.
Abstract: Integration of physicochemical procedures for studying mercury(II) speciation with microbiological procedures for studying the effects of mercury on bacterial growth allows evaluation of ionic factors (e.g., pH and ligand species and concentration) which affect biotoxicity. A Pseudomonas fluorescens strain capable of methylating inorganic Hg(II) was isolated from sediment samples collected at Buffalo Pound Lake in Saskatchewan, Canada. The effect of pH and ligand species on the toxic response (i.e., 50% inhibitory concentration (IC{sub 50})) of the P. fluorescens isolate to mercury were determined and related to the aqueous speciation of Hg(II). It was determined that the toxicities of different mercury salts were influenced by the nature of the co-ion. At a given pH level, mercuric acetate and mercuric nitrate yielded essentially the same IC{sub 50} s; mercuric chloride, on the other hand, always produced lower IC{sub 50}s. For each Hg salt, toxicity was greatest at pH 6.0 and decreased significantly at pH 7.0. Increasing the pH to 8.0 had no effect on the toxicity of mercuric acetate or mercuric nitrate but significantly reduced the toxicity of mercuric chloride. The aqueous speciation of Hg(II) in the synthetic growth medium M-IIY (a minimal salts medium amended to contain 0.1% yeast extract andmore » 0.1% glycerol) was calculated by using the computer program GEOCHEM-PC with a modified data base.« less
46 citations
TL;DR: Titration with 5,5′‐dithiobis (2‐nitrobenzoate) demonstrated that two enzyme–SH groups become kinetically accessible on reduction with NADPH, and showed a single band on polyacrylamide gel electrophoresis of the freshly prepared enzyme.
Abstract: Mercury resistance determinants in bacteria are often plasmid-borne or transposon-mediated. Mercuric reductase, one of the proteins encoded by the mercury resistance operon, catalyses a unique reaction in which mercuric ions, Hg (II), are reduced to mercury metal Hg(O) using NADPH as a source of reducing power. Mercuric reductase was purified from Azotobacter chroococcum SS2 using Red A dye matrix affinity chromatography. Freshly purified preparations of the enzyme showed a single band on polyacrylamide gel electrophoresis under non-denaturing conditions. After SDS-polyacrylamide gel electrophoresis of the freshly prepared enzyme, two protein bands, a major and a minor one, were observed with molecular weight 69 000 and 54 000, respectively. The molecular weight of the native enzyme as determined by gel filtration in Sephacryl S-300 was 142 000. The Km of Hg2+-reductase for HgCl2 was 11·11 μmol l−1. Titration with 5,5′-dithiobis (2-nitrobenzoate) demonstrated that two enzyme–SH groups become kinetically accessible on reduction with NADPH.
15 citations
Cites methods from "Studies on the effect of mercury an..."
...Mercuric reductase activity was determined by measuring Hg2¦-induced NADPH oxidation spectrophotometrically at 340 nm following the standard method (Izaki et al. 1974; Ghosh et al. 1996a, b, c)....
[...]
Journal Article•
TL;DR: In the current study, mercury-resistant nitrogen fixing bacterial strains were isolated by growing them on selective medium (NFM) to be used as biofertilizers and for the bioremediation of mercury from polluted soils and waters.
Abstract: In the current study, mercury-resistant nitrogen fixing bacterial (NFB) strains were isolated by growing them on selective medium (NFM) to be used as biofertilizers and for the bioremediation of mercury from polluted soils and waters Bacterial strains isolated from the soil around root nodules were checked for resistance to mercury by growing on yeast extract mannitol (YEM) medium supplemented with different concentrations of HgCl2 Mercury resistant bacterial strains were primarily screened by well plate method Mercury resistant NFB strains were further checked for their H2S production by growing on lead acetate (LA) medium Selected nitrogen fixing and mercury resistant bacterial strains were characterized using different biochemical tests and found to belong to genera Pseudomonas, Cronobacter and Bacillus Quantification of detoxified mercury by selected bacterial strains was done by using dithizone method Cronobacter species were found to be significantly the most efficient in the detoxification of mercury that reached up to 95% (p<005)
12 citations
Cites background from "Studies on the effect of mercury an..."
...Limited amount of information is available on mercury-resistance in N2-fixing bacteria (Ghosh et al., 1996)....
[...]
TL;DR: In this article, the feasibility of using residual glycerol as a carbon source for Hg removal by bacteria prospected from contaminated environments was shown for the first time, using eight bacterial isolates.
Abstract: Bacteria that harbor the mer operon in their genome are able to enzymatically reduce mercury (II) to the volatile form of mercury Hg (0). Detoxification of contaminated waste by using these bacteria may be an alternative to conventional methods for mercury removal. Residual glycerol from the biodiesel industry can be used as a carbon source to accelerate the process. This work shows for the first time the feasibility of using residual glycerol as a carbon source for Hg removal by bacteria prospected from contaminated environments. Eight bacterial isolates were able to remove mercury and degrade glycerol in mineral medium and residual glycerol. Mercury removal was monitored by atomic absorption spectroscopy and glycerol degradation by high performance liquid chromatography. The best results of mercury removal and glycerol degradation were obtained using isolates of Serratia marcescens M25C (85 and 100 %), Klebsiella pneumoniae PLB (90 and 100 %), Klebsiella oxytoca (90 and 100 %), and Arthrobacter sp. U3 (80 and 75 %), with addition of 0.5 g L−1 yeast extract. The Arthrobacter sp. U3 isolate is common in soils and has proven to be a promising candidate for environment applications due to its low pathogenicity and higher Hg removal and glycerol degradation rates.
10 citations
Cites result from "Studies on the effect of mercury an..."
...The authors found that the Hg removal ability of the microorganisms was related to their MIC values, corroborating the findings of Ghosh et al. (1996)....
[...]
References
More filters
Journal Article•
TL;DR: Procedures are described for measuring protein in solution or after precipitation with acids or other agents, and for the determination of as little as 0.2 gamma of protein.
Abstract: Since 1922 when Wu proposed the use of the Folin phenol reagent for the measurement of proteins, a number of modified analytical procedures utilizing this reagent have been reported for the determination of proteins in serum, in antigen-antibody precipitates, and in insulin. Although the reagent would seem to be recommended by its great sensitivity and the simplicity of procedure possible with its use, it has not found great favor for general biochemical purposes. In the belief that this reagent, nevertheless, has considerable merit for certain application, but that its peculiarities and limitations need to be understood for its fullest exploitation, it has been studied with regard to effects of variations in pH, time of reaction, and concentration of reactants, permissible levels of reagents commonly used in handling proteins, and interfering substances. Procedures are described for measuring protein in solution or after precipitation with acids or other agents, and for the determination of as little as 0.2 gamma of protein.
289,852 citations
TL;DR: A review of the chemical and biochemical effects of mercury, acadmium and lead is presented and the means available to identify their biochemical sites of action are discussed.
Abstract: A review is presented of the chemical and biochemical effects of mercury, acadmium and lead. Similarities and diversities are emphasized and the means available to identify their biochemical sites of action are discussed. Toxic effects and alterations in enzyme activity are described. 551 references.
1,429 citations
TL;DR: In this paper, an extremely sensitive and accurate method for the detn. of Hg down to 1.0 ppb has been described and applied to Ni and Co metal as well as rock samples and soil samples contg. materials.
Abstract: The procedure outlined describes an extremely sensitive and accurate method for the detn. of Hg down to 1.0 ppb. in soln. This procedure has been applied to Ni and Co metal as well as rock samples and soil samples contg. org. materials. The sample is taken into soln. by an oxidizing acid attack. Hg is then reduced to the elemental state and aerated from soln. in a closed system. The Hg vapor passes through a quartz absorption cell of an at. absorption spectrophotometer where its concn. is measured. The proceure is free from interferences due to org. matter or other volatile constituents of the sam ple. Large amts. of easily reducible elements must be absent from -- AATA
1,122 citations
TL;DR: The biochemical basis of the assay is described along with relevant characteristics including Km, C2H2/N2 conversion factor, and specific N2[C2H 2]-fixing activities obtained with various systems, and methods of measurement of N2 fixation are compared.
Abstract: A comprehensive report of the acetylene reduction assay for measurement of N2 fixation is presented. The objective is to facilitate the effective use and identify some potential limitations of the method. The report is based on more than 200 accounts of the use of this technique in 15 countries during the last 5 years. Methods of measurement of N2 fixation are compared. Nomenclature, e.g., N2[C2H2] fixed, is introduced to identify values of N2 fixation determined by C2H2-C2H2 assay. The biochemical basis of the assay is described along with relevant characteristics including Km, C2H2/N2 conversion factor, and specific N2[C2H2]-fixing activities obtained with various systems. Effects of combined nitrogen, temperature, light, pO2, N2, pC2H2 and water on activity are summarized. Available methods for sample preparation, assay chamber, gas phase, assay condition, termination of reaction, C2H4 analysis and expression of results are compared. The many uses of the C2H2-C2H4 assay for investigations of the biochemistry of nitrogenase and physiology of N2-fixing organisms, definition of N2-fixing organisms and measurement of field N2 fixation by legume, non-legume, soil, marine, rhizosphere, phylloplane and mammalian samples are tabulated.
1,021 citations
TL;DR: The mercury cycle in the biosphere and biological methylation of mercury and microbial resistance to mercury and organomercurials are studied.
Abstract: BIOTRANSFORMA nONS OF TOXIC MET AL CAnONS . Mercury . The mercury cycle in the biosphere .. Biological methylation of mercury . Microbial resistance to mercury and organomercurials .
413 citations