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PubMed2001

Analysis of a 2,4,6-trichlorophenol-dehalogenating enrichment culture and isolation of the dehalogenating member Desulfitobacterium frappieri strain TCP-A.

An anaerobic, 2,4,6-trichlorophenol ortho-dehalogenating mixed culture was enriched from sediment of the river Saale (Germany). Two isolated dechlorinating colonies (MK1 and MK2) consisted of rods of different lengths and thicknesses, indicating heterogeneity. Following subcultivation with thiosulfate as alternative electron acceptor and cocultivation with Clostridium celerecrescensT, the 2,4,6-trichlorophenol-dehalogenating bacterium Desulfitobacterium frappieri strain TCP-A was isolated and characterized regarding its taxonomic properties and the spectrum of chlorophenols that it dehalogenated. Four other bacterial strains were coenriched and identified as organisms with closest phylogenetic relatedness to the Clostridium type strains C. indolis, C. glycolicum, C. hydroxybenzoicum and C. sporosphaeroides (16S rDNA sequence identities of 99.5, 99.2, 94.4, and 93.5%, respectively). Amplified ribosomal DNA restriction analysis of the original dehalogenating cultures MK1 and MK2 (when not exposed to thiosulfate) confirmed the microbial heterogeneity and revealed the presence of two additional species related to the type strains of C. celerecrescens and Clostridium propionicum. Only one copy of the 16S rRNA genes of Desulfitobacterium frappieri in each of the clone libraries of MK1 and MK2 (containing 136 and 56 clones, respectively) was found by dot-blot hybridization, suggesting a relatively low number of the dehalogenating bacterium within the enrichment culture.

Anaerobiosis↗
PubMed2002

A capillary-based amperometric flow immunoassay for 2,4,6-trichlorophenol.

This paper describes the development of two different capillary-based heterogeneous competitive flow immunoassay formats (capillary flow injection immunoassay (CFIIA) and capillary sequential injection immunoassay (CSIIA)) for the determination of 2,4,6-trichlorophenol (2,4,6-TCP). The assays are based on the competition between the analyte and an analyte derivative labelled with the enzyme beta-galactosidase, for an anti-TCP antibody, followed by the injection of the mixture at equilibrium into a flow stream, where separation between the fractions bound and unbound to the antibody is performed in a glass capillary containing immobilised protein A. The antibody-tracer fraction retained inside the protein A capillary was measured by injection of 4-aminophenyl- beta- D-galactoside (4-APG), followed by amperometric detection of the enzymatically generated 4-aminophenol (4-AP), leading to a negative correlation between the signal and the analyte concentration. The two immunoassay formats were compared in terms of sensitivity and speed, giving IC(50) values of 1.41+/-0.03 and 1.64+/-0.07 micro g L(-1), detection limits of 0.2 and 0.4 micro g L(-1), and sample throughputs of 6 and 4 h(-1) for the CFIIA and CSIIA system, respectively. The influence of different interfering chlorophenolic compounds in the assay was minor, with only one exception (i.e. 2,4-dichlorophenol). In addition, different water matrices were tested (surface, tap, and rain water), showing that the matrix influence was negligible, except for rainwater, which resulted in a 30% increase in sensitivity. As a conclusion, the assay is suitable for the fast screening of TCP present at low concentration levels in water samples.

Antibodies, Monoclonal↗
PubMed2002

Effects of 2,4-dichlorophenol on activated sludge.

The effects of 2,4-dichlorophenol (2,4-DCP) on both acclimated and unacclimated activated sludge were investigated in batch reactors. The IC(50) values on the basis of maximum specific growth rate ( micro(m)), percent chemical oxygen demand (COD) removal efficiency and sludge activity were found to be 72, 60 and 47 mg l(-1), respectively, for unacclimated culture. The percent COD removal efficiencies of unacclimated culture were affected adversely, even at low concentrations, whereas culture acclimated to 75 mg 2,4-DCP l(-1) could tolerate about 200 mg 2,4-DCP l(-1)on the basis of COD removal efficiency. Although yield coefficient values of unacclimated culture increased surprisingly to very high values with the addition of 2,4-DCP, a linear decrease with respect to 2,4-DCP concentrations was observed for acclimated culture. Although no removal was observed with unacclimated culture, almost complete removal of 2,4-DCP up to a concentration of 148.7 mg l(-1) was observed with acclimated culture. It was showed that the culture could use 2,4-DCP as sole organic carbon source, although higher removal efficiencies in the presence of a readily degradable substrate were observed. Culture acclimated to 4-chlorophenol used 2,4-DCP as sole organic carbon source better than those acclimated to 2,4-DCP.

Bioreactors↗
PubMed2004

The generation of high biomass from chlororespiring bacteria using a continuous fed-batch bioreactor.

A continuous fed-batch reactor system was developed to rapidly obtain dense chlororespiring cultures of Anaeromyxobacter dehalogenans strain 2CP-C. A syringe pump continuously delivered concentrated 2,6-dichlorophenol (50-150 mM) to an anaerobic reactor vessel at a rate that sustained linear growth but prevented the substrate toxicity of chlorophenol. Dechlorination was not significantly inhibited by end product phenol up to 8 mM. A cell density of 76.8 mg protein l(-1) was obtained in 24 h. Specific growth rates averaged 0.033 h(-1)at 50% substrate limitation, which was in agreement with the maximum specific growth rate of 0.068 h(-1). This reactor system provides an efficient, cost-effective, and convenient method to rapidly obtain dense dechlorinating biomass and is promising to accelerate investigations of enzymes involved in chlororespiration.

Anaerobiosis↗
PubMed2006

Characterization of genes involved in the initial reactions of 4-chloronitrobenzene degradation in Pseudomonas putida ZWL73.

The genes encoding enzymes involved in the initial reactions during degradation of 4-chloronitrobenzene (4CNB) were characterized from the 4CNB utilizer Pseudomonas putida ZWL73, in which a partial reductive pathway was adopted. A DNA fragment containing genes coding for chloronitrobenzene nitroreductase (CnbA) and hydroxylaminobenzene mutase (CnbB) were PCR-amplified and subsequently sequenced. These two genes were actively expressed in Escherichia coli, and recombinant E. coli cells catalyzed the conversion of 4CNB to 2-amino-5-chlorophenol, which is the ring-cleavage substrate in the degradation of 4CNB. Phylogenetic analyses on sequences of chloronitrobenzene nitroreductase and hydroxylaminobenzene mutase revealed that these two enzymes are closely related to the functionally identified nitrobenzene nitroreductase and hydroxylaminobenzene mutase from Pseudomonas strains JS45 and HS12. The nitroreductase from strain ZWL73 showed a higher specific activity toward 4CNB than nitrobenzene (approximately at a ratio of 1.6:1 for the recombinant or 2:1 for the wild type), which is in contrast to the case where the nitroreductase from nitrobenzene utilizers Pseudomonas pseudoalcaligenes JS45 with an apparently lower specific activity against 4CNB than nitrobenzene (0.16:1) [Kadiyala et al. Appl Environ Microbiol 69:6520-6526, 2003]. This suggests that the nitroreductase from 4-chloronitrobenzene utilizer P. putida ZWL73 may have evolved to prefer chloronitrobenzene to nitrobenzene as its substrate.

Bacterial Proteins↗
PubMed2000

Volatile organic compounds associated with microbial growth in automobile air conditioning systems.

Volatile organic compounds from Penicillium viridicatum and Methylobacterium mesophilicum growing on laboratory media and on component materials of automobile air conditioners were analyzed with gas chromatography and mass spectrometry. P. viridicatum produced compounds such as 4-methyl thiazole, terpenes and alcohols, whereas M. mesophilicum produced dimethyl disulfide, dimethyl trisulfide, and chlorophenol with growth on laboratory media. In comparison with laboratory media, fewer volatiles were detected from colonized foam insulation materials. Biofilms of M. mesophilicum on aluminum evaporator components produced mainly dimethyl disulfide. These biofilms, after inoculation with P. viridicatum, produced offensive smelling alcohols and esters such as 2-methyl propanol, 3-penten-2-ol, and the ethyl ester of butanoic acid. The moisture and substrates innate to the automobile air conditioning systems provided an environment suitable for microbial biofilm development and odor production. Reduction of retained moisture in the air conditioning system coupled with use of less susceptible or antimicrobial substrates are advised for remediation of the noxious odors.

Air Conditioning↗
PubMed2000

Human toxicokinetics of inhaled monochlorobenzene: latest experimental findings regarding re-evaluation of the biological tolerance value.

OBJECTIVE: The aim of this study was to obtain toxicokinetic data on the absorption and elimination of monochlorobenzene (MCB) in blood and its main metabolite 4-chlorocatechol (4-ClCat) as well as on the isomeric chlorophenols (o-ClPh, m-ClPh, and especially p-CIPh as the main ClPh metabolite) in urine for reevaluation of the biological tolerance (BAT) value of MCB. METHODS: Eight subjects performed 8-h inhalation tests daily over five successive days in an exposure chamber, at a maximum allowable concentration at the workplace (MAK) value of 10 ppm MCB. Five and two probands carried out the test series during physical activity levels of 75 and 50 W, respectively, for 10 min/h on a bicycle ergometer, and one subject was exposed continuously while at rest. MCB and its metabolites were analyzed by gas chromatography in combination with mass spectrometry. RESULTS: The mean MCB blood concentration of the five subjects exposed during physical activity of 75 W was 217 +/- 42 microg/l. The relationship of the mean blood concentration measured under the conditions of rest or 50 and 75 W activity levels was in a ratio of about 1:1.7:2.8. The half-life values in the first hour after ending the exposures were 53 min and 150 min for the ensuing period, with steady-state being reached after 45 min. The mean 4-ClCat concentration in urine at the end of the five days was 150 +/- 13 mg/g creatinine in the case of the subjects exposed at 75 W, which decreased to 25 mg/g creatinine at the beginning of the next exposure. The analogous p-ClPh concentrations were 25 +/- 2 and 9 +/- 2 mg/g creatinine. The elimination half-life values of the ClPh isomers ranged from 12.4 to 16.5 h, and the half-life of 4-ClCat was 6.4 h. There was no apparent tendency for MCB and its metabolites to accumulate in blood or urine. CONCLUSIONS: The results are in accordance with relevant field and laboratory studies. Taken into consideration with the 95th percentile, the evaluated BAT values should be set at levels of 300 microg MCB/l blood, 175 mg 4-ClCat/g creatinine or alternatively at 30 mg p-ClPh/g creatinine in urine after the end of a shift. At the beginning of the next shift, the BAT values of the metabolites should be 35 and 15 mg/g creatinine, respectively.

Adult↗
PubMed2005

A new pharmacological screening assay with Trypanosoma cruzi epimastigotes expressing beta-galactosidase.

We have developed a new pharmacological screening assay for epimastigotes of Trypanosoma cruzi (clone CL-B5) that express the Escherichia coli LacZ gene. The assay is based on determining the activity of the cytoplasmic beta-galactosidase released into the culture on membrane lysis in the presence of the substrate chlorophenol red beta-D-galactopyranoside (CPRG). The experimental conditions were adjusted to find those in which the relationship between epimastigote number and CPRG absorbance was linear over the widest possible range. Absorbance was significantly correlated with the number epimastigote from 5x10(3) to 1.2x10(6) parasites/ml (r=0.98, P<0.01). The optimal final concentration of CPRG was 200 microM and the optimal incubation period was 6 h when parasites were incubated for 3 days. Once the assay was standardized, the trypanocidal activities of nifurtimox and benznidazole were determined both by CPRG assay and microscopic counting, demonstrating the methods utility for drug-screening. The efficacy obtained was comparable to that obtained with the manual method.

Animals↗
PubMed2005

Cells of Pseudomonas putida and Enterobacter sp. adapt to toxic organic compounds by increasing their size.

The phenol-degrading solvent-tolerant bacterium Pseudomonas putida P8 changed its cell shape when grown in the presence of aromatic compounds such as phenol and 4-chlorophenol. The sizes of cells that had been growing after addition of different concentrations of the toxic compounds were measured using a coulter counter that calculates the sizes of the rod-shaped bacteria to diameters of virtual spheres. The cells showed an increase in the diameter depending on the toxic effects of the applied concentrations of both solvents. The same effect was measured for an alkanol degrading bacterium, Enterobacter sp. VKGH12, in the presence of n-butanol. The reaction of the cells to different concentrations of n-butanol was examined by scanning electron microscopy. With this technique it could be shown that the size of the bacteria increased with increasing concentrations of n-butanol. These changes in cell size were dependent on the cellular activity and occurred only after addition of non-lethal concentrations. In the presence of lethal concentrations that completely inhibited cell growth, the cell sizes were similar to those of cells without intoxication. Taking into account the mathematical formula for spherical and cylindrical diameter and surface, respectively, the cells reacted to the presence of organic solvents by decreasing the ratio between surface and volume of the cells and therefore reducing their relative surfaces. As the cell surface and especially the cytoplasmic membrane are the major targets for the toxic effects of membrane-active compounds, this reduction of the relative surface represents an adaptive response to the presence of such compounds.

1-Butanol↗
PubMed2005

Ability of 13 chemical agents used in dental practice to induce sister-chromatid exchanges in Syrian hamster embryo cells.

To evaluate the genotoxic potential of 13 chemical agents used in dental practice, the abilities of these agents to induce sister-chromatid exchanges (SCEs) were examined using Syrian hamster embryo (SHE) cells. Statistically significant increases in the frequencies of SCEs were observed in SHE cells treated with all seven of the chemical agents used as endodontic medicaments: p-chlorophenol, m-cresol, formaldehyde, guaiacol, hydrogen peroxide, p-phenolsulfonic acid, and sodium hypochlorite (P < 0.01; Student t test). Assessment of two chemical agents that are applied to the oral mucosa as antiseptics showed that SCEs were induced by iodine (P < 0.01), but not by chlorhexidine. Of three chemical agents that are used as dyes for disclosing dental plaque, erythrosine B had no effect on SCE induction, while acid fuchsin and basic fuchsin increased the SCE frequencies in SHE cells (P < 0.01). Glutaraldehyde, which is used as a disinfectant for dental instruments and impressions, also induced SCEs (P < 0.01). Because SCE assays are used as a sensitive indicator for evaluating genetic toxicity of chemicals, the chemical agents that had a positive response in the present study are potentially genotoxic to mammalian cells.

Animals↗
PubMed2005

Assessment of pesticide residues in two arable soils from the semi-arid and subtropical regions of China.

The residues of 31 chlorinated hydrocarbons (CHCs), 25 chlorophenols (CPs), 30 organophosphorus (OP) and pyrethroid (PRT) in two arable soils from the semi-arid and subtropical regions of China were assessed. Data obtained indicate that the main compounds of CHC pesticide residues in the semi-arid soil were 4,4'-DDE (25.3 ng/g) and beta-HCH (14.1 ng/g), which totally accounted to about 90% of total CHC residues detected. The total content of CHC residues detected in the subtropical soil was only 3.1 ng/g, of which approximately 50% was beta-HCH. However, the total content of CP residues in both of the soils was about 11 ng/g. In the semi-arid soil, only parathion-methyl amongst the 30 compounds of OP and PRT residues was detected (32.5 ng/g), whilst malathion and parathion-methyl (8.7 and 17.7 ng/g, respectively) detected in the subtropical soil. Based on these results, it was suggested the environmental risk of pesticide residues ranked in an order as CHCs (mainly as 4,4'-DDE, beta-HCH) > OP (parathion-methyl) > CPs for the semi-arid soil, and as OPs (parathion-methyl and malathion) > CPs > CHCs (beta-HCH) for the subtropical soil.

China↗
PubMed1978

Effects of halogenated antibacterials on the erythrocyte membrane.

The influence of halogenated antibacterials on membrane structure and function was investigated using the human erythrocyte membrane as a model. Measurements of hemolysis in isotonic solution, altered membrane permeability, and stabilization against hypotonic hemolysis resulting from exposure of erythrocytes to halogenated antibacterials served as criteria of membrane-related effects. The hemolytic potency of the compounds studied differed widely, decreasing in the order hexachlorophene (HCP) greater than 2,2'-methylenebis(3,5-dichlorophenol) (3,5-TCP) greater than 2,2'-methylenebis(3,4-dichlorophenol) (3,4-TCP) approximately equal to 2,2'-methylenebis(4,6-dichlorophenol) (4,6-TCP) greater than 2,2'-methylenebis(4-chlorophenol) (DCP) greater than 3,4'-tribromosalicylanilide (TBS) approximately equal to 3,3',4',5-tetrachlorosalicylanilide (TCSA). Each of the antibacterials tested stabilized the erythrocyte against hypotonic hemolysis, although there were marked differences in the concentrations required to afford maximum stabilization as well as in the extent of protection. The observed order of protective effectiveness was HCP greater than 3,4-TCP greater than 4,6-TCP greater than DCP approximately equal to TCS greater than TBS. As shown by measurements of the first-order rate constant for K+ efflux, the permeability of the erythrocyte membrane to K+ was increased upon exposure to the antibacterials, with the effect of HCP greater than 3,4-TCP greater than 4,6-TCP approximately equal to 3,4-TCP greater than DCP approximately equal to TCS greater than TBS. These results indicate that halogenated antibacterials are capable of perturbing mammalian membranes, a feature which may account in part for their mammalian toxicity.

Anti-Infective Agents↗
PubMed1990

Interaction of chlorinated phenols with thyroxine binding sites of human transthyretin, albumin and thyroid binding globulin.

Previous results (Brouwer and van den Berg, Toxicol. Appl. Pharmacol., 85 (1986) 301) indicated preferential binding of a hydroxylated metabolite of tetrachlorobiphenyl to transthyretin (TTR) a carrier of thyroxine (T4). In the present study it was investigated whether the T4 binding site of TTR could be occupied specifically by hydroxylated chlorinated aromatic compounds using chlorinated phenol congeners as model compounds in a competition assay with [125I]T4. Chlorinated aromatics such as 2,3-dichlorobenzene and 3,4,3',4'-tetrachlorobiphenyl, and phenols such as 4-hydroxybiphenyl and phenol were inefficient competitors. All chlorinated phenols tested were competitors for the T4 binding site of TTR. The ranking in competition was pentachlorophenol (PCP) greater than trichlorophenols greater than dichlorophenols greater than monochlorophenols. Structures with chlorine in both ortho positions to the hydroxyl group were more efficient competitors. The relative affinity of binding of pentachlorophenol (PCP) to TTR was about twice that of T4. Scatchard analysis showed that PCP mainly decreased the affinity constant K11 while the binding capacity R1 was not altered, indicating a competitive type of inhibition. PCP was also able to compete with T4 sites on albumin with a relative affinity of 0.25. T4 binding to thyroid binding globulin (TBG) was much less affected by interference of PCP (relative affinity 0.001). The results indicate a specific interaction of chlorophenols with the T4 binding site of TTR.

Binding Sites↗
PubMed1995

Analysis of toxicological risks from local contamination by PCDDs and PCDFs: importance of isomer distributions and toxic equivalents.

The concentrations of 2,3,7,8-chlorinated dibenzo-p-dioxins and furans (PCDD/PCDFs) at Finnish contaminated sites associated with chlorophenolic (CP) wood preservatives were measured by isomer-specific GC/HRMS methods, and the implications of the spatial and structural distributions of the isomers and of toxic equivalent (TEF) systems for toxicological risks were investigated. High concentrations (maximum value of over 90 ppb and arithmetic mean value of 19 ppb I-TEQs in dry weight) of PCDD/Fs were found in soil samples from sites where wood preservative consisting of 2346-TeCP, PeCP and 246-TCP have been used. The PCDD/Fs were distributed heterogeneously between soil layers, the concentrations in topsoil samples being generally smaller (mean 9.1 ppb I-TEQ d.w.) than those in subsoil (mean 28 ppb I-TEQ d.w.). Concentrations of PCDD/Fs were unrelated to the CP contents in the soil samples. Hexa-, hepta- and octachlorinated dibenzofurans were the dominant congeners by concentration in these and in most other samples, also in the wood preservative. The isomer distribution differed considerably between study sites, and even between samples of soil and waste at the same site. The relative importance of the samples and isomers was affected by the selection of TEFs; the importance of some of the CP-related samples was increased when applying TEFs based on lethality to juvenile fish, due to the abundance of 123478-HxCDF.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Photochemical conversion of chlorinated phenolic substances in aquatic media as studied by AOX and Microtox tests.

Six chlorophenolic substances were photochemically converted by illumination with xenon light. The AOX (adsorbable organic halogens) contents were determined after illumination for various periods. Pure chlorinated phenolic substances were readily detected by AOX. Both adsorbable and non-adsorbable chlorinated products were formed. Chloride analysis indicated that most (if not all) of the non-adsorbable products consisted of chloride ions. Microtox, a bacterial bioluminescence test, was used as a screening test for toxic properties of the photolysis products. Products from 2,4,6-trichlorophenol were more toxic than the original substance. Other compounds tested had less toxic products as measured by their combined effect on the Microtox system. Products of 2,4-dichlorophenol, 2,3,4-trichlorophenol, 4,5,6-trichloroguaiacol and 2,3,4,5-tetrachlorophenol did not seem to contribute to the combined toxic properties after partial photolysis of the parent compound. Pentachlorophenol was converted into products which had substantial effects on the Microtox system.

Chlorides↗
PubMed1985

Joint toxicity of mixtures of groups of organic aquatic pollutants to the guppy (Poecilia reticulata).

In this study acute lethal concentrations (LC50) to the guppy (Poecilia reticulata) were determined for mixtures of 4 groups of aquatic pollutants. The groups were composed of 11 nonreactive, nonionized organic chemicals, 11 chloroanilines, 11 chlorophenols, and 9 reactive organic halides. Earlier studies indicated that the joint toxicity within each of these groups was concentration additive, probably because of a similar mode of action. The joint toxicity of combinations of one representative from each group showed a high variance, but generally tended to be partially additive to concentration additive. This high variance is probably caused by the low number of compounds in these mixtures. Experiments with mixtures of whole groups gave more accurate results. The toxicity of a mixture of the first three groups, containing 33 well-known aquatic pollutants, was almost completely concentration additive. Concentrations of 0.04 of the individual LC50 values contributed to the toxicity of this mixture.

Aniline Compounds↗
PubMed1988

Subchronic metabolic effects and toxicity of a simulated pulp mill effluent on juvenile lake trout, Salmo trutta m. lacustris.

Juvenile lake trout (Salmo trutta m. lacustris) were exposed for 7 weeks to 0.05X and 0.2X 96-hr LC50 concentrations of simulated bleached kraft pulp mill effluent (KME - Sa + CP). A sulfate soap preparation, composed mainly of resin and fatty acids, with added chlorophenols (CP, tri-, tetra-, and penta-CP) was used as the toxicant mixture. Concentrations of free CP in plasma and free and conjugated CP in bile were proportional to their concentrations in the water. The greatest total gradient between bile and water CP was 5.2 X 10(4) for pentachlorophenol. The activity of a liver polysubstrate monooxygenase (PSMO) system, assayed with three model substrates, increased 40 to 67% due to KME - Sa + CP. However, the increase was not directly dependent on the exposure concentration. In contrast to PSMO, activities of conjugating enzymes (p-nitrophenol UDP-glucuronosyl and glutathione transferases) were decreased in the liver. Increased concentration of glutathione was noted in the liver and kidney. In addition, a small (9%) but significant decrease in blood hemoglobin concentration was observed at the higher exposure concentration. Although growth rate of lake trout was markedly decreased due to KME - Sa + CP, hydromineral balance and carbohydrate metabolism in fish were unaffected, indicating possible physiological compensation. On the other hand, lethality tests with lake trout preexposed to KME - Sa + CP at 0.2 X LC50 revealed decreased tolerance, whereas at the lower exposure concentration it was unchanged. We therefore conclude that various physiological adjustments in trout during subchronic exposures were not adaptive in terms of short-term tolerance.

7-Alkoxycoumarin O-Dealkylase↗
PubMed1984

Ecoepidemiology--a casuistic discipline describing ecological disturbances and damages in relation to their specific causes: exemplified by chlorinated phenols and chlorophenoxy acids.

Ecoepidemiology is a new concept created in analogy to human epidemiology, and aims at the study of ecotoxicological effects at the levels of ecosystems, biological communities, and populations in relation to causative environmental exposures, mostly by chemicals. By way of example, ecoepidemiology is described by presenting an example of unintentional dissipation and possible discharges of chlorophenols and phenoxy acids into the terrestrial environment and--as a more specific case--a marine, aquatic area, viz., the Køge Bay immediately to the south of Copenhagen, Denmark. The examples are illustrative of the complex situation which characterizes most ecoepidemiological cases. Difficulties with which the ecoepidemiologist are confronted are not only the identification of possible causative and confounding chemicals, and the description of ecoepidemiological effects per se, but also the assessment of critical pathways of multimedium pollutants. Biomonitoring, computer-based handling of data from natural localities, and determination of a variety of anthropogenic impact factors are necessary elements of ecoepidemiological studies.

Animals↗