Cadmium Toxicity to Microcystis aeruginosa PCC 7806 and Its Microcystin-Lacking Mutant

The adverse effects of microcystin (MC) produced by cyanobacteria have drawn considerable attention from the public. Yet it remains unclear whether MC confers any benefits to the cyanobacteria themselves. One suggested function of MC is complexation, which may influence the bioaccumulation and toxicity of trace metals. To test this hypothesis, we examined Cd toxicity to wild-type Microcystis aeruginosa PCC 7806 (WT) and its MC-lacking mutant (MT) under nutrient-enriched (+NP), phosphorus-limited (-P), and nitrogen-limited (-N) conditions. The accumulation of Cd and the biochemical parameters associated with its detoxification [total phosphorus (TP), inorganic polyphosphate (Poly-P), and glutathione (GSH) in the cells as well as intra- and extra-cellular carbohydrates] were quantified. Although the –P cyanobacteria accumulated less Cd than their +NP and –N counterparts, the different nutrient-conditioned cyanobacteria were similarly inhibited by similar free ion concentration of Cd in the medium ([Cd²⁺]_F). Such good toxicity predictability of [Cd²⁺]_F was ascribed to the synchronous decrease in the intracellular concentrations of Cd and TP. Nevertheless, Cd toxicity was still determined by the intracellular Cd to phosphorus ratio (Cd/P), in accordance with what has been reported in the literature. On the other hand, the concentrations of TP, Poly-P, and carbohydrates went up, but GSH concentration dropped down with the enhancement of [Cd²⁺]_F, indicating their association with Cd detoxification. Although the inactivation of MC peptide synthetase gene had some nutrient and Cd concentration dependent effects on the parameters above, both cyanobacterial strains showed the same Cd accumulation ability and displayed similar Cd sensitivity. These results suggest that MC cannot affect metal toxicity either by regulating metal accumulation or by altering the detoxification ability of the cyanobacteria. Other possible functions of MC need to be further investigated.     

Correlation between metallothionein and energy metabolism in sea bass,Dicentrarchus labrax,exposed to cadmium

Specimens of sea bass (Dicentrarchus labrax) were exposed to two different cadmium concentrations (0.5 and 5 μg Cd²⁺/ml seawater) for a period of 7 days. Cadmium accumulated in the tissues of D. labrax in the following order: kidney > liver > gills at both concentrations. Accumulation patterns in fish exposed to 0.5 μg Cd²⁺/ml seawater were different with respect to 5.0 μg Cd²⁺/ml seawater. At both Cd concentrations a similar stress situation occurred during the first 4 hr as noted by the depletion of glycogen stores and the increase in free glucose in the muscle; metallothionein was induced in the liver, but failed to bind all the cytosolic Cd, which was in part bound to high-molecular-weight ligands. Fish recovered from this initial stress situation within 24 hr as indicated by the increase in glycogen and the decrease of glucose. Long-term effects were clearly dependent upon metal concentration: at lower Cd exposure, metallothionein induction increased linearly with time and counteracted the toxic effect of the metal; on the other hand, when fish were exposed to 5.0 μg Cd²⁺/ml seawater a clear stress occurred at the end of the exposure, as indicated by the notable decrease of glycogen stores, the increase of free glucose, the decrease of AEC in the muscle and the increase of Cd bound to high-molecular-weight ligands in the liver.     

Rational evolution of Cd2+-specific DNAzymes with phosphorothioate modified cleavage junction and Cd2+ sensing

In vitro selection of RNA-cleaving DNAzymes is a powerful method for isolating metal-specific DNA. A few successful examples are known, but it is still difficult to target some thiophilic metals such as Cd²⁺ due to limited functional groups in DNA. While using modified bases expands the chemical functionality of DNA, a single phosphorothioate modification might boost its affinity for thiophilic metals without complicating the selection process or using bases that are not commercially available. In this work, the first such in vitro selection for Cd²⁺ is reported. After using a blocking DNA and negative selections to rationally direct the library outcome, a highly specific DNAzyme with only 12 nucleotides in the catalytic loop is isolated. This DNAzyme has a cleavage rate of 0.12 min⁻¹ with 10 μM Cd²⁺ at pH 6.0. The R_p form of the substrate is cleaved ∼100-fold faster than the S_p form. The DNAzyme is most active with Cd²⁺ and its selectivity against Zn²⁺ is over 100 000-fold. Its application in detecting Cd²⁺ is also demonstrated. The idea of introducing single modifications in the fixed region expands the scope of DNA/metal interactions with minimal perturbation of DNA structure and property.     

Effect of calcium chloride on heavy metal induced alteration in growth and nitrate assimilation of Sesamum indicum seedlings

In the early growth phase of Sesamum indicum cv. PB-1, the decrease in fresh and dry mass was higher with 1.0 mM Cd²⁺ than with the same level of Pb²⁺ and Cu²⁺. Recovery from the metal stress was considerable in the root fresh weight and almost completely in the root dry weight when 10.0 mM (1.9 EC), calcium chloride was supplied to the growing seedlings along with the metal salts in various combinations. Accumulation of Pb²⁺, Cd²⁺ and Cu²⁺ was differential to the metals and the plant parts when supplied without or with 10.0 mM calcium chloride. The order of endogenous metal accumulation was Cu²⁺Cd²⁺Pb²⁺ and roots accumulated more metal than the leaves in the absence, as well as in the presence, of calcium chloride. Calcium chloride could recover loss of in vivo NRA in roots caused by either of the metal combinations, whereas the salt could recover the loss in leaf NRA caused only by Pb²⁺Cd²⁺ (1.0 mM each). Response of root and leaf NRA was on the other hand, different when the enzyme was assayed directly using an in vitro assay method, and the salt accelerated the loss in enzyme activity drastically. The organic-N content of root and leaf was, however, increased significantly (p < 0.001) with calcium chloride alone and with the metals supplied in various combinations. Our data indicate that instead of a high endogenous accumulation of Cu²⁺, Cd²⁺ and Pb²⁺ in roots and leaves the metal toxicity is recovered to a great extent in the presence of 10.0 mM calcium chloride in the root environment regarding growth and nitrate reduction of the roots and leaves of young sesame seedlings.     

Bioremoval capacity of three heavy metals by some microalgae species (Egyptian Isolates)

Three fresh water microalgal isolates [Phormidium ambiguum (Cyanobacterium), Pseudochlorococcum typicum and Scenedesmus quadricauda var quadrispina (Chlorophyta)] were tested for tolerance and removal of mercury (Hg²⁺), lead (Pb²⁺) and cadmium (Cd²⁺) in aqueous solutions as a single metal species at conc. 5–100 mg / L under controled laboratory conditions. The obtained results showed that Hg²⁺ was the most toxic of the three metal ions to the test algae even at low concentration (< 20 mg/L). While lower concentration of Pb²⁺ and Cd²⁺ (5–20 mg / L) enhanced the algal growth (chlorophyll a and protein), elevated concentrations (40–100 mg / L) were inhibitory to the growth. The results also revealed that Ph. ambiguum was the most sensitive alga to the three metal ions even at lower concentrations (5 and 10 mg / L) while P. typicum and S. quadricauda were more tolerant to high metal concentrations up to 100 mg / L. The bioremoval of heavy metal ions (Hg²⁺, Pb²⁺ and Cd²⁺) by P. typicum from aqueous solution showed that the highest percentage of metal bioremoval occurred in the first 30 min of contact recording 97% (Hg²⁺), 86% (Cd²⁺) and 70% (Pb²⁺). Transmission electron microscopy (TEM) was used to study the interaction between heavy metal ions and P. typicum cells. At ultrastructural level, an electron dense layers were detected on the algal cell surfaces when exposed to Cd, Hg and Pb. At the same time, dark spherical electron dense bodies were accumulated in the vacuoles of the algal cells exposed to Pb. Excessive accumulation of starch around the pyrenoids were recorded as well as deteriorations of the algal cell organelles exposed to the three metal ions.     

Resistance acquisition of Thiobacillus thiooxidans upon cadmium and zinc ion addition and formation of cadmium ion-binding and zinc ion-binding proteins exhibiting metallothionein-like properties

Through subcultivations of Thiobacillus thiooxidans WU-79A in autotrophic media in which the concentrations of Cd²⁺ and Zn²⁺ were increased successively, Cd²⁺-resistant (CDR) and Zn²⁺-resistant strains (ZNR) were obtained. The growth of WU-79A was inhibited by the addition of 25 mM Cd²⁺ as well as Zn²⁺. However, CDR and ZNR could grow without any lag phase in media containing 200 mM Cd²⁺ and 250 mM Zn²⁺, respectively. CDR and ZNR were able to grow even in media containing up to 400 mM Cd²⁺ and 600 mM Zn²⁺, respectively, although they exhibited lag phases. CDR could grow in medium containing up to 250 mM Zn²⁺, as could ZNR in medium containing up to 200 mM Cd²⁺. Cd²⁺-binding and Zn²⁺-binding proteins were isolated from CDR and ZNR, respectively, by gel filtration and ion exchange chromatography. The molecular weights of both proteins were estimated to be approximately 13,000 by gel filtration. The fact that there was no strong absorption at 280 nm of the proteins suggested that they had few aromatic amino acids. Broad absorption bands which are typical of mercaptide (metal thiolate) complexes were detected. The properties of the proteins were spectrophotometrically similar to those of metallothionein.     

Development, Characterization, and Application of a Cadmium-Selective Microelectrode for the Measurement of Cadmium Fluxes in Roots of Thlaspi Species and Wheat

A Cd²⁺-selective vibrating microelectrode was constructed using a neutral carrier-based Cd ionophore to investigate ion-transport processes along the roots of wheat (Triticum aestivum L.) and two species of Thlaspi, one a Zn/Cd hyperaccumulator and the other a related nonaccumulator. In simple Cd(NO₃)₂ solutions, the electrode exhibited a Nernstian response in solutions with Cd²⁺ activities as low as 50 nm. Addition of Ca²⁺ to the calibration solutions did not influence the slope of the calibration curve but reduced the detection limit to a solution activity of 1 μm Cd²⁺. Addition of high concentrations of K⁺ and Mg²⁺ to the calibration solution to mimic the ionic composition of the cytoplasm affected neither the slope nor the sensitivity of the electrode, demonstrating the pH-insensitive electrode's potential for intracellular investigations. The electrode was assayed for selectivity and was shown to be at least 1000 times more selective for Cd²⁺ than for any of those potentially interfering ions tested. Flux measurements along the roots of the two Thlaspi species showed no differences in the pattern or the magnitude of Cd²⁺ uptake within the time frame considered. The Cd²⁺-selective microelectrode will permit detailed investigations of heavy-metal ion transport in plant roots, especially in the area of phytoremediation.     

Effect of Cl on Cd uptake by Swiss chard in nutrient solutions

Swiss chard (Beta vulgaris L., cv. Fordhook Giant) was grown in nutrient solution with Cl concentrations varying between 0.01 mM and 120 mM. Solution Na concentration and ionic strength were maintained in all treatments by compensating with NaNO₃. All solutions contained Cd (50 nM, spiked with ¹⁰⁹Cd). Three different Cd²⁺ buffering systems were used. In one experiment, Cd²⁺ activity was unbuffered; its activity decreased with increased Cl concentration as a result of the formation of CdCl_n ^2–n species. In the other experiments, Cd²⁺ activity was buffered by the chelator nitrilotriacetate (NTA, 50 M) and ethylene-bis-(oxyethylenenitrilo)-tetraacetate (EGTA, 50 M) at about 10^–9 M and 10^–11 M, respectively. Plant growth was generally unaffected by increasing Cl concentrations in the three experiments. In unbuffered solutions, Cd concentrations in plant tissue decreased significantly (p<0.01) (approximately 2.4-fold) as solution Cl concentration increased from 0.01 mM to 120 mM. However, this decrease was smaller in magnitude than the 4.7-fold decrease in Cd²⁺ activity as calculated by the GEOCHEM-PC program for the same range of Cl concentrations. In solutions where Cd²⁺ activity was buffered by NTA, Cd concentrations in plant tissue increased approximately 1.4-fold with increasing Cl concentration in solution, while the Cd²⁺ activity was calculated to decrease 1.3-fold. In solutions where Cd²⁺ activity was buffered by EGTA, Cd concentrations in the roots increased 1.3-fold with increasing Cl concentration in solution but there was no effect of Cl on shoot Cd concentrations. The data suggest that either CdCl_n ^2–nspecies can be taken up by plant roots or that Cl enhances uptake of Cd²⁺ through enhanced diffusion of the uncomplexed metal to uptake sites.Abbreviations DAS days after sowing - EGTA ethylene-bis-(oxyethylenenitrilo)-tetraacetate - HBED N,N-bis(2-hydroxybenzyl)-ethylenediamine-N,N-diacetate - NTA nitrilotriacetate     

Contribution of Cd-EDTA complexes to cadmium uptake by maize: a modelling approach

Background and aims

Chelant-enhanced phytoextraction has given variable and often unexplained experimental results. This work was carried out to better understand the mechanisms of Cd plant uptake in the presence of EDTA and to evaluate the contributions of Cd-EDTA complexes to the uptake.

Method

A 1-D mechanistic model was implemented, which described the free Cd²⁺ root absorption, the dissociation and the direct absorption of the Cd-EDTA complexes. It was used to explain Cd uptake by maize in hydroponics and in soil.

Results

In hydroponics, the addition of EDTA caused a decrease in Cd uptake by maize, particularly when the ratio of total EDTA ([EDTA] _T ) to total Cd ([Cd] _T ) was greater than 1. At [Cd] _T = 1 μM, when [EDTA] _T /[Cd] _T < 1, the model indicated that Cd uptake was predominantly due to the absorption of free Cd²⁺, whose pool was replenished by the dissociation of Cd-EDTA. When [EDTA] _T /[Cd] _T > 1, the low Cd uptake was mostly due to Cd-EDTA absorption. In soil spiked with 5 mg Cd kg^?1, Cd uptake was not affected by the various EDTA additions, because of the buffering capacity of the soil solid phase.

Conclusions

Addition of EDTA to soil increases Cd solubility but dissociation of Cd-EDTA limits the availability of the free Cd²⁺ at the root surface, which finally reduces the plant uptake of the metal.     

Solid phase speciation of Zn and Cd in zinc smelter effluent-irrigated soils

Solubility of metal in contaminated soils is a key factor which controls the phytoavailability and toxic effects of metals on soil environment. The chemical equilibria of metal ions between soil solution and solid phases govern the solubility of metals in soil. Hence, an attempt was made to identify the probable solid phases (minerals), which govern the solubility of Zn²⁺ and Cd²⁺ in zinc smelter effluent-irrigated soils. Estimation of free ion activities of Zn²⁺ (pZn²⁺) and Cd²⁺ (pCd²⁺) by Baker soil test indicated that metal ion activities were higher in smelter effluent-irrigated soils as compared to that in tubewell water-irrigated soils. Identification of solid phases further reveals that free ion activity of Zn²⁺ and Cd²⁺ in soil highly contaminated with Zn and Cd due to long-term irrigation with zinc smelter effluent is limited by the solubility of willemite (Zn₂SiO₄) in equilibrium with quartz and octavite (CdCO₃), respectively. However, in case of tubewell water-irrigated soil, franklinite (ZnFe₂O₄) in equilibrium with soil-Fe and exchangeable Cd are likely to govern the activity of Zn²⁺ and Cd²⁺ in soil solution, respectively. Formation of highly soluble minerals namely, willemite and octavite indicates the potential ecological risk of Zn and Cd, respectively in smelter effluent irrigated soil.     

Uptake and binding of cadmium and mercury to metallothionein in rat hepatocyte primary cultures

The administration of inorganic Cd and Hg in vivo has been shown to result in markedly different metal concentrations in rat liver. Primary cultures of rat hepatocytes were utilized to gain insight into the dispositional differences between these chemically similar metals. Hepatocyte monolayer cultures were exposed to several concentrations of Cd or Hg (3, 10 and 30μm) in serum-containing medium for 30min. The cells were then washed and incubated in fresh medium for the remainder of the experiment. Hepatocytes exposed to Cd accumulated significantly more metal than hepatocytes exposed to equimolar concentrations of Hg. In cells exposed to 3μm-Cd there was an initial loss of Cd from the hepatocytes when placed in fresh medium, followed by a gradual re-uptake of metal, concomitant with increased binding to metallothionein. In hepatocytes exposed to 3 and 10μm-Cd, 87 and 77% of the intracellular Cd was bound to metallothionein within 24h. Loss of Hg from hepatocytes pulsed with 30μm-Hg was also observed upon the addition of fresh medium and continued for the duration of the experiment. No time-dependent increase in Hg binding to metallothionein was observed. A maximum of about 10% of the intracellular Hg was found associated with metallothionein in hepatocytes exposed to 30μm-Hg. Studies utilizing [³⁵S]cysteine incorporation indicated significant increases in the amount of metallothionein synthesized in hepatocytes exposed to 3 and 10μm-Cd (300% of control value) and 30μm-Hg (150% of control value) 24h after metal pulsing. Time-course studies revealed a 6–12h lag in metallothionein synthesis, followed by a significant elevation in [³⁵S]cysteine incorporation into metallothionein between 12 and 24h. These studies suggest that (a) isolated hepatocytes differentiate between Cd and Hg and preferentially accumulate the former, and (b) Cd strongly stimulates the induction of, and preferentially binds to, metallothionein, whereas Hg induces weakly, and does not preferentially bind to, metallothionein.     

Ferrochelatase of Rhodopseudomonas spheroides

Extracts of Rhodopseudomonas spheroides contain two ferrochelatases: one is soluble and forms metalloporphyrins from deuteroporphyrin and haematoporphyrin; the other is particulate and forms metalloporphyrins from protoporphyrin, mesoporphyrin, deuteroporphyrin and haematoporphyrin. Neither enzyme incorporates Mg²⁺ into porphyrins or Fe²⁺ into porphyrin cytochrome c. By using the particulate enzyme, plots of 1/v versus 1/s when one substrate was varied and the other kept constant showed that neither substrate affected the K_m of the other. The suggested sequential mechanism for the reaction is supported by derivative plots of slopes and intercepts. The K_m for deuteroporphyrin was 21.3μm and that for Co²⁺ was 6.13μm. The enzyme incorporated Co²⁺, Fe²⁺, Zn²⁺, Ni²⁺ and Mn²⁺; Cd²⁺ was not incorporated and was an inhibitor, competitive with respect to Co²⁺, non-competitive with respect to deuteroporphyrin. The K_i for Cd²⁺ was 0.73μm. Ferrochelatase was inhibited by protohaem, non-competitively with respect to Co²⁺ or with respect to deuteroporphyrin. Inhibition by magnesium protoporphyrin was non-competitive with respect to deuteroporphyrin, uncompetitive with respect to Co²⁺. The inhibitory concentrations of the metalloporphyrins are lower than those required for the inhibition of δ-aminolaevulate synthetase by protohaem. Fe²⁺ is not incorporated aerobically into porphyrins unless an electron donor, succinate or NADH, is supplied; the low aerobic rate of metalloporphyrin synthesis obtained is insensitive to rotenone and antimycin. The rate of Fe³⁺ incorporation increases as anaerobic conditions are achieved.     

Transformation of the Antibacterial Agent Norfloxacin by Environmental Mycobacteria

Because fluoroquinolone antimicrobial agents may be released into the environment, the potential for environmental bacteria to biotransform these drugs was investigated. Eight Mycobacterium sp. cultures in a sorbitol-yeast extract medium were dosed with 100 μg ml⁻¹ of norfloxacin and incubated for 7 days. The MICs of norfloxacin for these strains, tested by an agar dilution method, were 1.6 to 25 μg ml⁻¹. Cultures were extracted with ethyl acetate, and potential metabolites in the extracts were purified by high-performance liquid chromatography. The metabolites were identified using mass spectrometry and nuclear magnetic resonance spectroscopy. N-Acetylnorfloxacin (5 to 50% of the total absorbance at 280 nm) was produced by the eight Mycobacterium strains. N-Nitrosonorfloxacin (5 to 30% of the total absorbance) was also produced by Mycobacterium sp. strain PYR100 and Mycobacterium gilvum PYR-GCK. The MICs of N-nitrosonorfloxacin and N-acetylnorfloxacin were 2- to 38- and 4- to 1,000-fold higher, respectively, than those of norfloxacin for several different bacteria, including the two strains that produced both metabolites. Although N-nitrosonorfloxacin had less antibacterial activity, nitrosamines are potentially carcinogenic. The biotransformation of fluoroquinolones by mycobacteria may serve as a resistance mechanism.     

Accumulation of cadmium by hairy-root cultures of Solanum nigrum

Summary Cadmium uptake by cultures of transformed hairy-roots of Solanum nigrum was studied. The effect of pH, buffer type, temperature, exposure time and Cd²⁺ content ranging from 0.2 to 2000 ppm was measured. Cd²⁺ uptake was dependent on increasing metal concentration and it was time dependent. From the variety of buffers tested, MES buffer and borate ions were found to be beneficial for the Cd²⁺ uptake. The high effectivness of Cd²⁺ accumulation in the roots decreased significantly after increasing the Cd²⁺ content in the buffer over 2 ppm.     

The Role of Iron-Deficiency Stress Responses in Stimulating Heavy-Metal Transport in Plants

Plant accumulation of Fe and other metals can be enhanced under Fe deficiency. We investigated the influence of Fe status on heavy-metal and divalent-cation uptake in roots of pea (Pisum sativum L. cv Sparkle) seedlings using Cd²⁺ uptake as a model system. Radiotracer techniques were used to quantify unidirectional ¹⁰⁹Cd influx into roots of Fe-deficient and Fe-sufficient pea seedlings. The concentration-dependent kinetics for ¹⁰⁹Cd influx were graphically complex and nonsaturating but could be resolved into a linear component and a saturable component exhibiting Michaelis-Menten kinetics. We demonstrated that the linear component was apoplastically bound Cd²⁺ remaining in the root cell wall after desorption, whereas the saturable component was transporter-mediated Cd²⁺ influx across the root-cell plasma membrane. The Cd²⁺ transport system in roots of both Fe-deficient and Fe-sufficient seedlings exhibited similar Michaelis constant values, 1.5 and 0.6 μm, respectively, for saturable Cd²⁺ influx, whereas the maximum initial velocity for Cd²⁺ uptake in Fe-deficient seedlings was nearly 7-fold higher than that in Fe-grown seedlings. Investigations into the mechanistic basis for this response demonstrated that Fe-deficiency-induced stimulation of the plasma membrane H⁺-ATPase did not play a role in the enhanced Cd²⁺ uptake. Expression studies with the Fe²⁺ transporter cloned from Arabidopsis, IRT1, indicated that Fe deficiency induced the expression of this transporter, which might facilitate the transport of heavy-metal divalent cations such as Cd²⁺ and Zn²⁺, in addition to Fe²⁺.     

Temporal variability of solution Cd2+ concentration in metal-contaminated soils as affected by soil temperature: consequences on lettuce (Lactuca sativa L.) exposure

The aim of this study was to assess how the solubility and the speciation of Cd in soil solution were affected over time by the soil temperature for three metal-contaminated soils. The changes of solution Cd concentration (either total or free ionic) and other physico-chemical parameters (e.g. pH, ionic strength, the concentrations of ${\text{NO}}_3^ - $ , ${\text{SO}}_4^{2 - } $ , Ca, Mg and dissolved organic carbon) were monitored over a 28-day culture of lettuce (Lactuca sativa L.) in soils incubated at 10°C, 20°C or 30°C. The major result of this study was that Cd²⁺ concentration greatly varied over time in soil solution. The Cd²⁺ concentration declined over time in soil solution as did the concentration of cations that may compete for adsorption (Ca²⁺, Mg²⁺). The rise in soil temperature primarily impacted on the concentration of Cd²⁺ via promoting the microbial C-degradation and, thus, the complexation of Cd in soil solution. The integration of the temporal variations in Cd²⁺ concentration through the calculation of the root exposure to solution Cd (E _Cd) provided a fairly close and robust prediction of Cd concentration in lettuce roots. The present work thus provided new insights on the fate of Cd in contaminated soils that may be relevant for predicting the root uptake of Cd.     

In-Feed Use of Heavy Metal Micronutrients in U.S. Swine Production Systems and Its Role in Persistence of Multidrug-Resistant Salmonellae

The study aimed to characterize the role of heavy metal micronutrients in swine feed in emergence of heavy-metal-tolerant and multidrug-resistant Salmonella organisms. We conducted a longitudinal study in 36 swine barns over a 2-year period. The feed and fecal levels of Cu²⁺ and Zn²⁺ were measured. Salmonella was isolated at early and late finishing. MICs of copper sulfate and zinc chloride were measured using agar dilution. Antimicrobial susceptibility was tested using the Kirby-Bauer method, and 283 isolates were serotyped. We amplified pcoA and czcD genes that encode Cu²⁺ and Zn²⁺ tolerance, respectively. Of the 283 isolates, 113 (48%) showed Cu²⁺ tolerance at 24 mM and 164 (58%) showed Zn²⁺ tolerance at 8 mM. In multivariate analysis, serotype and source of isolates were significantly associated with Cu²⁺ tolerance (P < 0.001). Fecal isolates were more likely to be Cu²⁺ tolerant than those of feed origin (odds ratio [OR], 27.0; 95% confidence interval [CI], 2.8 to 250; P = 0.0042) or environmental origin (OR, 5.8), implying the significance of gastrointestinal selective pressure. Salmonella enterica serotypes Typhimurium and Heidelberg, highly significant for public health, had higher odds of having >20 mM MICs of Cu²⁺ than did “other” serotypes. More than 60% of Salmonella isolates with resistance type (R-type) AmStTeKm (32 of 53) carried pcoA; only 5% with R-type AmClStSuTe carried this gene. czcD gene carriage was significantly associated with a higher Zn²⁺ MIC (P < 0.05). The odds of having a high Zn²⁺ MIC (≥8 mM) were 14.66 times higher in isolates with R-type AmClStSuTe than in those with R-type AmStTeKm (P < 0.05). The findings demonstrate strong association between heavy metal tolerance and antimicrobial resistance, particularly among Salmonella serotypes important in public health.     

Effect of Cadmium on gamma-Glutamylcysteine Synthesis in Maize Seedlings

Cysteine, γ-glutamylcysteine, and glutathione and the extractable activity of the enzymes of glutathione biosynthesis, γ-glutamylcysteine synthetase (EC 6.3.2.2) and glutathione synthetase (EC 6.3.2.3), were measured in roots and leaves of maize seedlings (Zea mays L. cv LG 9) exposed to CdCl₂ concentrations up to 200 micromolar. At 50 micromolar Cd²⁺, γ-glutamylcysteine contents increased continuously during 4 days up to 21-fold and eightfold of the control in roots and leaves, respectively. Even at 0.5 micromolar Cd²⁺, the concentration of γ-glutamylcysteine in the roots was significantly higher than in the control. At 5 micromolar and higher Cd²⁺ concentrations, a significant increase in γ-glutamylcysteine synthetase activity was measured in the roots, whereas in the leaves this enzyme activity was enhanced only at 200 micromolar Cd²⁺. Labeling of isolated roots with [³⁵S]sulfate showed that both sulfate assimilation and glutathione synthesis were increased by Cd. The accumulation of γ-glutamylcysteine in the roots did not affect the root exudation rate of this compound. Our results indicate that maize roots are at least in part autonomous in providing the additional thiols required for phytochelatin synthesis induced by Cd.     

Multiple antibiotic resistances in metal tolerant E. coli from hospital waste water

Study of antibiotic resistance was done among the metal tolerant E. coli isolates from hospital wastewater at Lucknow city. Metal tolerance was determined in terms of visible growth on metal amended plates at their varying concentrations. MICs were also determined among all metal tolerant E. coli isolates. All the isolates showed their MIC in between 100-2000 µg/ml while maximum isolates demonstrated their MICs at 400, 800 and 1600 µg/ml against all the metal tested. 23.07% of the isolates showed their MIC at 2000 µg/ml against Ni³⁺. Multiple antibiotic resistances were recorded among all the metal resistant E.coli isolates. A high level of resistance was observed against Methicillin (86.53%) followed by penicillin (73.07%), Cephradin (57.69%), Rifampicin (34.61%), Erythromycin (26.92%), Nalidixic acids (25%), Chloramphenicol (3.84%) and least to Gentamycine (1.92%). Streptomycin was recorded most effective against E.coli isolates among the entire antibiotic tested. Antimicrobial resistance observed among the bacteria from the aquatic system contaminated with hospital wastes may be threatful for the environment and public health both.