Catálogo de publicaciones - libros

We designed a new toxic bioassay using the green paramecia as testing organism. is a unicellular organism that occurs widely in rivers and ponds. Since uses metabolites of endosymbiotic green algae in the cytoplasm as a nutritive source, culturing is much easier than culturing mammalian cells. The use of will thus make quicker and more convenient evaluation of toxicity of various polluting chemicals. Here, we selected thirty-two pollutants such as pesticides, toxic metals and polycyclic aromatic hydrocarbons. Those substances were added at various concentrations to the culture medium of paramecia. Then the IC values, defined as the concentrations of chemicals inhibiting the growth of organisms by 50%, obtained for both paramecia and mammalian cell cultures were compared. We found that paramecia were much highly sensitive to some chemicals such as methylmercury chloride and mercuric chloride, compared to cultured mammalian cells. We conclude that is one of the best organism for assessing the effect of chemical pollutants in the aqueous environment.

A short-term batch respirometric test coupled with ultraviolet (UV) photometry was developed to detect the presence in waste water of toxic substances such as heavy metals, cleaning and sanitising agents, and textile dyes. Tests have been performed on a waste water plant containing various toxic substances in order to assess the usefulness of this global sensor for detecting toxic events. Short-term respirometry gives an estimation of the immediate biological activity, which is influenced by the consumption of rapidly biodegradable pollutants and by potential inhibition. To separate both effects, we combined short-term respirometry with a rapid estimation of pollution by UV-spectrophotometry.

Environmental biosensors to assess the toxicity of environmental media such as water, soil, and atmosphere have been developed using various recombinant bioluminescent bacteria. Those bacteria were constructed based on specific stress-responsive promoters in bacterial cells. They are thus activated by different groups of toxicity. For continuous monitoring of water toxicity, a multichannel system having different stress-responsive strains in each channel, and composed of two-stage mini-bioreactors, was successfully developed. Soil toxicity was assessed using a soil biosensor based upon immobilization of recombinant bioluminescent bacteria that worked with the addition of rhamnolipids biosurfactant. An example of phenanthrene toxicity is shown. For the assessment of gas toxicity, an immobilization technique has been set up to allow the biosensor to come in direct contact with the toxic gas in the sensing chamber. An example of benzene toxicity is shown. This mini review will show how the recombinant bioluminescent bacteria can be utilized as environmental biosensors. With further findings and developments of new non-specific stress promoters, the potency and extensiveness of the information that can be obtained using these environmental biosensors is immense.

We investigated the toxicity of field waters using a multi-channel continuous monitoring system in Berlin, Germany. This system uses genetically engineered bioluminescent bacteria for the assessment of the toxicity of soluble chemicals. It showed easy and long-term monitoring without any system shut down due to pollution overloading. We used the bioluminescent bacterial strains DPD2794, DPD2540, TV1061 and GC2, which respond respectively to DNA-, cell membrane-, protein- and general cellular-damaging agents. The bioluminescent levels either increase for DPD2794, DPD2540 and TV1061 strains, or decrease for the GC2 strain after being mixed with toxic samples. We monitored the toxicity over a period of two to three weeks at three different sites: the Ruhleben wastewater treatment plant discharge flow, and river flows at the Teltowkanal and Fischereiamt in Berlin. At all sites the DPD2540 and TV1061 strains showed a significant increase of bioluminescence while bioluminescence decreased for the GC2 strain. This result demonstrates the occurrence of chemicals that affect the integrity of the cellular membrane; leading to either protein denaturation and inhibiting cellular metabolism or to cell death. Therefore, our findings suggest that the bioluminescent bacteria array may serve as a novel water toxicity monitoring system in outdoor fields.

We developped a microbial biosensor for detection of surfactants. The biosensor receptor is based on bacterial strains that are harboring plasmids for biodegradation of surfactants. We studied the sensitivity, stability and selectivity of the biosensor with respect to anionic, cationic and non-ionic surfactants, carbohydrates, alcohols, humic acids and toxic compounds such as aromatic xenobiotics. Under laboratory conditions the microbial biosensor was used to assess the purity level of water that was decontaminated from surfactants. On the basis of the obtained sensor characteristics, a portable microprocessor analyzer was set up. The biosensor device registers the transducer’s signals and makes it possible to process calibration dependencies and determination of the concentration of target compound in a sample.

The Microtox® bioassay, based upon the fading of light emitted by the luminescent bacteria when exposed to noxious substances, was used for studying the changes in speciation and the related changes in toxicity of two metals known as environmental pollutants. It was verified that modifications of pH and of ionic composition of the incubation medium did not affect the standard toxicity of phenol. By contrast, Cr(VI) toxicity was found to decrease as pH increased, underscoring that hydrogenchromate anion, the dominant species at low pH, is the most harmful. Cr(VI) toxicity was not modified when changing the medium composition, as this metal does not form chloro-complexes in the presence of sodium chloride. Conversely, Cd(II) toxicity was almost unaffected by pH within the 5.0–7.0 range. Replacing sodium chloride either by sodium nitrate or by sodium perchlorate resulted in changes of the measured cadmium toxicity, due to changes in speciation. Free Cd ion was found to be the most harmful toward the bacteria. In conclusion, both pH and ionic composition are factors that strongly influence the measured toxicity of environmental samples containing hexavalent chromium and/or cadmium when using the Microtox® bioassay.

The presented investigations were carried out in order to detect environmental pollutants using the HT29, HepG2 and A549 cultured human cells as biological sensors of toxicity. We measured the growth rate inhibition and the expression level of stress proteins after exposure to cadmium, nickel, chromium, ethanol, 1-propanol, benzene, toluene, xylene, dichlorobenzene and trichlorobenzenes. Threshold concentrations were determined for selected pollutants and significance as well as reliability of the results were discussed. New perspectives for developing improved biodetectors are reported.

On 12 December 1999, one third of the load of the tanker ‘Erika’, amounting to about 10 000 t crude oil flowed into sea waters close to the French Atlantic Coast (Finistère region). The spilled oil was fuel No. 6, a heavy or residual fuel that is a complex mixture of polycyclic aromatic compounds (PAC) of high molecular weight. Some polycyclic aromatic compounds are genotzoxic substances that induce carcinogenic lesions in laboratory animals. DNA adducts, reflecting genotoxic effects, are used as biomarker of early pollution. In this study, the genotoxic impact of the ‘Erika’ oil was assessed by studying the presence of DNA adduct in the liver of immature fishes () from four locations of the French Brittany coasts, two, six and nine months after the disaster. Two months after the spill, a high amount of DNA adducts was found in samples from all locations (92 to 290 DNA adduct/10 nucleotides). DNA adducts were more persistent in the North than in the South of the affected French Brittany coasts. In September, no significant difference was observed between the locations. When incubated in presence of an ‘Erika’ fuel extract, DNA adduct patterns similar to those obtained from the liver of liver are observed both, in cell culture (HePG2) and, in presence of fish liver microsomes.

The bioassay LUX-FLUORO test was developed for the rapid detection and quantification of environmental pollutants with genotoxic and/or cytotoxic effects. This bacterial test system uses two different reporter genes whose gene products and their reactions, respectively, can be measured easily and simultaneously by optical methods. Genotoxicity is measured by the increase of bioluminescence in genetically modified bacteria which carry a plasmid with a complete operon for the enzyme luciferase from the marine photobacterium under the control of a DNA-damage dependent so-called SOS promoter. If the desoxyribonucleic acid (DNA) in these bacteria is damaged by a genotoxic chemical, the SOS promoter is turned on and the operon is expressed. The newly synthesized luciferase reacts with its substrate thereby producing bioluminescence in a damage-proportional manner. In the second part of the system, genetically modified bacteria carry the gfp gene for the green fluorescent protein from the jellyfish downstream from a constitutively expressed promoter. These bacteria are fluorescent under normal conditions. If their cellular metabolism is disturbed by the action of cytotoxic chemicals the fluorescence decreases in a dose-proportional manner. The combined LUX-FLUORO test can be used for the biological assessment of the geno- and cytotoxicity of a wide variety of organic and inorganic chemicals including complex mixtures in different matrices.