Enhancing the Antibiotic Antibacterial Effect by Sub Lethal Tellurite Concentrations: Tellurite and Cefotaxime Act Synergistically in Escherichia coli

The emergence of antibiotic-resistant pathogenic bacteria during the last decades has become a public health concern worldwide. Aiming to explore new alternatives to treat antibiotic-resistant bacteria and given that the tellurium oxyanion tellurite is highly toxic for most microorganisms, we evaluated the ability of sub lethal tellurite concentrations to strengthen the effect of several antibiotics. Tellurite, at nM or µM concentrations, increased importantly the toxicity of defined antibacterials. This was observed with both Gram negative and Gram positive bacteria, irrespective of the antibiotic or tellurite tolerance of the particular microorganism. The tellurite-mediated antibiotic-potentiating effect occurs in laboratory and clinical, uropathogenic Escherichia coli, especially with antibiotics disturbing the cell wall (ampicillin, cefotaxime) or protein synthesis (tetracycline, chloramphenicol, gentamicin). In particular, the effect of tellurite on the activity of the clinically-relevant, third-generation cephalosporin (cefotaxime), was evaluated. Cell viability assays showed that tellurite and cefotaxime act synergistically against E. coli. In conclusion, using tellurite like an adjuvant could be of great help to cope with several multi-resistant pathogens

Catalases Are NAD(P)H-Dependent Tellurite Reductases

Reactive oxygen species damage intracellular targets and are implicated in cancer, genetic disease, mutagenesis, and aging. Catalases are among the key enzymatic defenses against one of the most physiologically abundant reactive oxygen species, hydrogen peroxide. The well-studied, heme-dependent catalases accelerate the rate of the dismutation of peroxide to molecular oxygen and water with near kinetic perfection. Many catalases also bind the cofactors NADPH and NADH tenaciously, but, surprisingly, NAD(P)H is not required for their dismutase activity. Although NAD(P)H protects bovine catalase against oxidative damage by its peroxide substrate, the catalytic role of the nicotinamide cofactor in the function of this enzyme has remained a biochemical mystery to date. Anions formed by heavy metal oxides are among the most highly reactive, natural oxidizing agents. Here, we show that a natural isolate of Staphylococcus epidermidis resistant to tellurite detoxifies this anion thanks to a novel activity of its catalase, and that a subset of both bacterial and mammalian catalases carry out the NAD(P)H-dependent reduction of soluble tellurite ion (TeO(3) (2−)) to the less toxic, insoluble metal, tellurium (Te°), in vitro. An Escherichia coli mutant defective in the KatG catalase/peroxidase is sensitive to tellurite, and expression of the S. epidermidis catalase gene in a heterologous E. coli host confers increased resistance to tellurite as well as to hydrogen peroxide in vivo, arguing that S. epidermidis catalase provides a physiological line of defense against both of these strong oxidizing agents. Kinetic studies reveal that bovine catalase reduces tellurite with a low Michaelis-Menten constant, a result suggesting that tellurite is among the natural substrates of this enzyme. The reduction of tellurite by bovine catalase occurs at the expense of producing the highly reactive superoxide radical

Bacterial Toxicity of Potassium Tellurite: Unveiling an Ancient Enigma

Biochemical, genetic, enzymatic and molecular approaches were used to demonstrate, for the first time, that tellurite (TeO(3) (2−)) toxicity in E. coli involves superoxide formation. This radical is derived, at least in part, from enzymatic TeO(3) (2−) reduction. This conclusion is supported by the following observations made in K(2)TeO(3)-treated E. coli BW25113: i) induction of the ibpA gene encoding for the small heat shock protein IbpA, which has been associated with resistance to superoxide, ii) increase of cytoplasmic reactive oxygen species (ROS) as determined with ROS-specific probe 2′7′-dichlorodihydrofluorescein diacetate (H(2)DCFDA), iii) increase of carbonyl content in cellular proteins, iv) increase in the generation of thiobarbituric acid-reactive substances (TBARs), v) inactivation of oxidative stress-sensitive [Fe-S] enzymes such as aconitase, vi) increase of superoxide dismutase (SOD) activity, vii) increase of sodA, sodB and soxS mRNA transcription, and viii) generation of superoxide radical during in vitro enzymatic reduction of potassium tellurite

The Geobacillus stearothermophilus V iscS Gene, Encoding Cysteine Desulfurase, Confers Resistance to Potassium Tellurite in Escherichia coli K-12

Many eubacteria are resistant to the toxic oxidizing agent potassium tellurite, and tellurite resistance involves diverse biochemical mechanisms. Expression of the iscS gene from Geobacillus stearothermophilus V, which is naturally resistant to tellurite, confers tellurite resistance in Escherichia coli K-12, which is naturally sensitive to tellurite. The G. stearothermophilus iscS gene encodes a cysteine desulfurase. A site-directed mutation in iscS that prevents binding of its pyridoxal phosphate cofactor abolishes both enzyme activity and its ability to confer tellurite resistance in E. coli. Expression of the G. stearothermophilus iscS gene confers tellurite resistance in tellurite-hypersensitive E. coli iscS and sodA sodB mutants (deficient in superoxide dismutase) and complements the auxotrophic requirement of an E. coli iscS mutant for thiamine but not for nicotinic acid. These and other results support the hypothesis that the reduction of tellurite generates superoxide anions and that the primary targets of superoxide damage in E. coli are enzymes with iron-sulfur clusters

Tellurite-mediated antibiotic-potentiating effect in clinical isolates.

<p>Antibiotic susceptibility, in the absence or presence of the indicated tellurite concentrations, was assessed by growth inhibition zones (cm²) as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035452#s4" target="_blank">Methods</a>. Parentheses indicate per cent of susceptibility increase regarding the respective control.</p

Minimal tellurite concentration causing a cefotaxime-potentiating effect in <i>E. coli</i>.

<p>Inhibition growth zones were determined as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035452#s4" target="_blank">Methods</a> using LB plates amended with the indicated sub lethal tellurite concentrations (nM).</p

Cefotaxime and potassium tellurite acts synergistically in <i>E. coli</i>.

<p>Growth curves (left panels) and cell viability (right panels) were determined at the indicated time intervals for <i>E. coli</i> exposed to 0.065 (A, sublethal), 0.13 (B, MIC) and 0.5 µg/ml (C, lethal) CTX in the absence or presence of 200 nM tellurite. Controls contained no tellurite or cefotaxime. Data represent the mean of at least 3 independent trials. Refer to inset in panel A for symbol meaning.</p

Tellurite-mediated antibiotic-potentiating effect in different bacteria.

<p>Antibiotic-mediated inhibition growth zones were determined for <i>E. coli</i> (A), <i>P. aeruginosa</i> (B) and <i>S. aureus</i> (C) grown in the absence (white bars) or presence of the indicated tellurite (T) concentrations as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035452#s4" target="_blank">Methods</a>. Values represent the average of at least 4 independent trials and significance was determined using t-test analysis (p<0.05). Significance values are (*) p<0.05, (**) p<0.01 and (***) p<0.001.</p

The product of tellurite reduction by catalase is elemental tellurium.

<div><p>Absorption spectra of products formed upon the reduction of K₂TeO₃<i>in vitro</i> by <b>(A)</b> 2-mercaptoethanol, <b>(B)</b> a crude extract prepared from <i>S. epidermidis</i> CH cells prior to chromatographic enrichment, and <b>(C)</b> purified bovine liver catalase, were resolved by Induced Coupled Plasma-Optical Emission spectroscopy <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0000070#pone.0000070-Grotti1" target="_blank">[30]</a>.</p> <p>All products show an absorption maximum at 214.281 nm, the peak wavelength of the Te° standard (triangle in panel A).</p> <p>We note that the scale used to measure absorbance in the crude extract differs by a factor of ten from those used to measure absorbance in chemically prepared tellurium (A) and in the product of tellurite reduction by bovine liver catalase (C).</p> <p>This is because the crude extract includes a plethora of components with absorption maxima at or near this wavelength.</p></div