Pollution-induced Community Tolerance Research Articles

An ecological perspective in aquatic ecotoxicology: Approaches and challenges

Abstract This paper aims to highlight the considerable potential of a better integration of ecological theory in aquatic ecotoxicology. It outlines how community ecology, studies on trophic interaction and disturbance ecology could provide an enhanced theoretical basis for aquatic ecotoxicology and increase ecological relevance in environmental risk assessment of chemicals. Based on the literature and own research, approaches from aquatic ecotoxicology are presented, which are based on ecological considerations and address a higher level of biological complexity for risk assessment strategies of chemicals. The concepts of species-sensitivity distribution (SSD), pollution-induced community tolerance (PICT), the use of model ecosystems and the sediment quality triad (SQT) in ecological risk assesment as well as inputs from ecotoxicology into landscape ecology are illustrated. These examples aim to evidence aquatic ecotoxicology as a rewarding field of ecological research.

The use of leucine incorporation to determine the toxicity of phenols to bacterial communities extracted from soil

The toxicity of different phenols to the soil bacterial community was studied in the laboratory using the leucine incorporation technique. The effects of environmental factors such as pH, temperature and binding strength to soil particles were also assessed in order to deduce confounding effects due to the chemical and physical conditions in the soil from which the bacterial community was extracted. Bacterial growth varied with temperature and pH, the optima being at around 33 °C and pH 7 in a soil with a natural pH of 7. The toxicity of different phenols varied less than bacterial growth as a function of temperature and pH, but the toxicity of 2,3,6-trichlorophenol increased at lower temperatures and pH. The pH affected the toxicity in the same way in soils with naturally different pH values and when the pH was changed using buffers, showing that this was a direct effect of pH and was not due to different communities in different soils. The degree to which the bacteria were bound to soil particles had no effect on the toxicity of phenols. Neither did freezing nor thawing the soil have any effect on toxicity, although the bacterial growth was lower in frozen soil than in non-frozen soil. Increasing numbers of substitutes on the phenols increased their toxicity to soil bacteria, and chlorine substitutes were more toxic than the corresponding methyl substitutes. The toxicity of the phenols studied to the whole soil bacterial community was correlated with literature data on the toxicity to the single species Pseudomonas putida. Applying the leucine incorporation technique to the bacterial community extracted from soil was shown to be a rapid and sensitive method of estimating toxicity. The methodology also allowed differentiation between the effects of environmental factors on toxicity to the soil bacterial community and changes in the tolerance of the community. Thus, the development of pollution-induced community tolerance in phenol- and 2-chlorophenol-polluted soils could be shown.

On the limits of toxicant‐induced tolerance testing: Cotolerance and response variation of antibiotic effects

Pollution-induced community tolerance (PICT) as an ecotoxicological test system has been claimed to detect pollutant effects highly specifically and sensitively. However, the specificity might be limited by the occurrence of cotolerance. Another limitation of the application of any ecotoxicological test system lies in variation of the measured responses. We tested the variation and the occurrence of cotolerance experimentally, using antibiotics as toxicants, soil microcosms as microbial communities, and tolerance determination in Biolog plates as PICT detection test. Bacteria have been discussed as being prone to multiple tolerances due to the possible accumulation of multiple resistance genes on mobile genetic elements. However, in our experiments, cotolerance occurred only between antibiotics of the same group (oxytetracycline and tetracycline), as expected from their identical mode of action. Cotolerance between oxytetracycline and tylosin in soil microcosms exposed to oxytetracycline was low, as was cotolerance to oxytetracycline in tylosin-exposed microcosms. We conclude that tolerance development to antibiotics in soils reflects the actual selection pressure rather than a general pattern of multiple resistances. Concerning variation, the PICT effect of tetracycline was well reproducible in two consecutive years. The response variation linked to PICT experiments in controlled microcosms was comparable to that of ecotoxicological test systems of equivalent complexity. In conclusion, our results support an application of the PICT methodology as an effective means to study the soil ecotoxicology of antibiotics.

Discriminating between effects of metals and natural variables in terrestrial bacterial communities

This study aimed to assess effects of metals on bacterial communities in grassland soil and to discriminate these effects from natural variability in soil properties. Changes in gross parameters of bacterial communities were investigated by the determination of 14C-leucine and 3H-thymidine incorporation rates, CO 2 evolution, biomass indicators and N mineralization rates. 14C-leucine incorporation rate and CO 2 evolution showed correlations with all metals tested as well as with organic matter content. 3H-thymidine incorporation rates, biomass indicators and N mineralization rates did not strongly correlate with any physical or chemical parameters. Further, community-level physiological profiles (CLPP) and polymerase chain reaction (PCR)-amplified denaturing gradient gel electrophoresis (DGGE) of 16S rDNA were performed. Monte-Carlo permutation testing was performed to analyse CLPP and DGGE data to allow for a stringent discrimination between sources of effects. CLPP changes correlated with the Pb concentration and pH in the soil. DGGE changes correlated with Pb and Cu concentrations and organic matter content. Pollution-induced community tolerance (PICT) was not found for any of the metals assessed. A causal relation between the effects observed on bacterial communities and the presence of metals was not established with PICT. The negative outcome of PICT can probably be attributed to indirect effects or to methodological problems. We concluded that the observed shifts in CLPP and DGGE patterns are a strong indication of metals effects on bacterial communities in this grassland. These effects could only be filtered from the total variation by multivariate analyses.

Microbial Biomass and Tolerance of Microbial Community on an Aged Heavy Metal Polluted Floodplain in Japan

The objective of the present study was to increase understanding of the effects of heavy metal pollution and soil properties on microorganisms in relation to the biomass and microbial functional community. Soil samples were collected from aged polluted and reference sites on a floodplain. The soil Cu, Zn and Pb total concentrations were much higher at the polluted sites (average 231.6–309.9 mg kg−1, 195.7–233.0 mg kg−1, and 72.4–86.0 mg kg−1, respectively) than at the reference site (average 33.3–44.0 mg kg−1, 76.7–98.0 mg kg−1, and 30.8–41.6 mg kg−1, respectively), while the available heavy metal concentrations in CaCl2 extraction were similar in all sites. Small seasonal variations in the size of microbial biomass were observed. Ambient soil properties (e.g. total C, N, pH, moisture content, and CEC) affected the soil microbial biomass more than the heavy metal pollution. However, the aged pollution tended to impact on the composition of the microbial community. PICT (pollution-induced community tolerance) test using BIOLOG Ecoplates showed enhanced tolerance of the microbial community to Cu stress in the polluted site. In non polluted but low nutrient, low pH and low moisture soil, the microbial biomass was lower and the microbial community was more vulnerable to Cu stress. In spite of the low heavy metal availability due to ageing, the BIOLOG technique provided sensitive detection of microbial community level changes in PICT analysis.

Effects on the function of three trophic levels in marine plankton communities under stress from the antifouling compound zinc pyrithione

This study aimed to investigate functional responses of natural marine planktonic communities to stress from the antifouling compound zinc pyrithione (ZPT). Isotope labelling techniques ( 14C) were applied to study bacterial incorporation of leucine, photosynthetic activity of phytoplankton and grazing of labelled prey by zooplankton communities for 6 days after exposures to nominal concentrations of 0, 5, 25, 50 nM ZPT in a mesocosm experiment in Isefjord, Denmark. Significant direct effects were visible on chlorophyll a concentrations, which decreased in all exposed communities, to between 48 and 36% of control concentrations on Day 3, 1 day after the last exposure. Phytoplankton activities were also significantly affected on Day 3 with activities between 9 and 26% of control levels, as was zooplankton activities in the 25 and 50 nM exposures. In the 50 nM exposure the total community zooplankton activity was reduced to 25 ± 4%, and per individual to 46 ± 11% of control levels. Bacterial communities showed positive indirect effects with high activities (up to 183 ± 40%) due to higher amounts of available substrate from algal death. Pollution induced community tolerance analyses performed on phytoplankton and bacterial communities at the end of the experiment indicated a development of increased tolerance for phytoplankton in the 50 nM exposed communities, whereas there were no changes in tolerance in the bacterial communities. Multivariate analysis of the integrated functional response by the plankton communities revealed a significant difference ( p < 0.05) between exposed communities compared to controls in the first 3 days after last exposure and in the end of the experiment. The study provides evidence of diverse effects on the functions of marine plankton communities under stress from a pollutant. Direct effects lead to cascading indirect effects throughout the community, eventually causing different developments. Continuous exposure to ZPT could lead to severe long-term effects, causing more permanent changes in structure and function than observed here. The study demonstrates that it is possible to assess the functional effects of a stressor in a complex mesocosm system, and to determine effects in a complex plankton community, which were not predictable from laboratory studies.

Induction of atrazine tolerance in a natural soil assemblage of microalgae reared in the laboratory

We investigated the occurrence of tolerance to atrazine (2-chloro-4-ethylamino-6-isopropylamino- s-triazine) in soil microalgae by means of the pollution-induced community tolerance methodology. To this end, a natural soil assemblage of microalgae, reared under laboratory conditions, was used as experimental model. Experimental cultures were exposed to 0.46, 0.93, 1.85, 2.70, and 5.40 mg atrazine/L medium for 40 days. After this chronic exposure, both untreated and atrazine-pretreated cultures were subjected to a short-term dose–effect study with atrazine using average growth rate as endpoint. Results showed that chronic exposure to atrazine induced significant changes at the community level, increasing its atrazine tolerance. In addition, atrazine-pretreated assemblages seemed to display, on average, a reduced growth rate compared with untreated assemblages in the absence of atrazine. Response to the presence of atrazine could thus be achieved with some fitness cost. This suggests that the soil microalgal productivity could be diminished after exposure to atrazine.

Pollution-induced community tolerance of microorganisms from forest soil organic layers polluted with Zn or Cu

The long-term accumulation of heavy metals in forest soil organic layers may adversely affect soil microorganisms. The objectives of the study were to assess the effects of high contents of Cu and Zn on the soil microbial communities and their tolerance to metal pollution. Several microbial indices such as microbial biomass (C mic), the basal soil respiration (BAS), the community level physiological profiles (CLPP) and the pollution-induced community tolerance (PICT) approach, based on Biolog ® EcoPlate assay, were used. The soil organic layer samples were taken in the Cu-polluted (CuP), Zn-polluted (ZnP) and unpolluted (UP) Scots pine forests. Despite very high total heavy metal concentrations in soils C mic, BAS and CLPPs at the polluted sites did not differ from those at the UP sites. A damaging effect of heavy metals was detected only by the tolerance measurements with Biolog ® plates. PICT to Cu at the CuP sites was found for five substrates. At the ZnP sites significant PICT to Zn was found for 10 out of 31 substrates. Furthermore, the CuP samples exhibited lower resistance to Zn and the ZnP samples to Cu compared with the UP samples, indicating that at the polluted sites microbial resistance to additional stress caused by another metal was decreased. Since the tolerance measurements using Biolog ® assay appeared to be more sensitive than other methods used, we concluded that this approach could be useful in environmental risk assessment.

Influence of soil zinc concentrations on zinc sensitivity and functional diversity of microbial communities

Pollution induced community tolerance (PICT) is based on the phenomenon that toxic effects reduce survival of the most sensitive organisms, thus increasing community tolerance. Community tolerance for a contaminant is thus a strong indicator for the presence of that contaminant at the level of adverse concentrations. Here we assessed PICT in 11 soils contaminated with zinc runoff from galvanised electricity pylons and 11 reference soils sampled at 10 m distance from these pylons. Using PICT, the influence of background concentration and bioavailability of zinc on zinc sensitivity and functional diversity of microbial communities was assessed. Zinc sensitivity of microbial communities decreased significantly with increasing zinc concentrations in pore water and calcium chloride extracted fraction while no significant relationship was found with total zinc concentration in the soil. It was also found that functional diversity of microbial communities decreased with increasing zinc concentrations, indicating that increased tolerance is indeed an undesirable phenomenon when environmental quality is considered. The hypothesis that zinc sensitivity of microbial communities is related to background zinc concentration in pore water could not be confirmed.

Effects of antibiotics on soil microorganisms: time and nutrients influence pollution-induced community tolerance

A method for the detection of the effects of antibiotics on soil microbial communities was optimised in the present study. We investigated the influence of measurement time and nutrient status on the pollution-induced community tolerance (PICT), using the sulfonamide sulfachloropyridazine (SCP) as model compound. The tolerance development in soil microcosms that were exposed to SCP under different conditions was compared with the background tolerance in SCP-unexposed microcosms. The tolerance of bacterial extracts from the soil microcosms was determined in Biolog ® multiwell plates as the SCP sensitivity of a range of physiological processes. The background tolerance was not affected by soil nutrient amendments, but an influence of the inoculum density in the microtiter plates was observed. Still, the variance of the background tolerance was low, which is in favour of the usage of community tolerance measurements for a selective detection of a toxicant impact. In line with the assumptions of the PICT concept, soil amendment with SCP led to an increase in community tolerance. This tolerance development was enhanced upon additional soil amendment with fresh pig slurry, and less by alfalfa meal addition. Tolerance increases were observed after a soil exposure to SCP of only 7 days, possibly because nutrient input facilitates the fast adaptation of the soil microbial community. However, a further increase in exposure time led to variable changes in the observed tolerance. Prolonged tolerance detection in the microtiter plates (11 days) enabled a clearer differentiation between different soil treatments, as it better resolved the EC 50 values of processes with a high tolerance to bacteriostatic antimicrobial compounds. For the detection of antibiotic effects on soil microbial communities, it is therefore recommended to use nutrient amendments (possibly fresh pig slurry), to standardize the soil exposure time, and to extend the period of Biolog plate measurement beyond 7 days.

Predicting and observing responses of algal communities to photosystem ii‐herbicide exposure using pollution‐induced community tolerance and species‐sensitivity distributions

Various test strategies are in use in ecotoxicology to assess the potential risks of toxicants on aquatic communities. The species-sensitivity distribution concept (SSD) works by arranging single-species laboratory test data in a cumulative frequency distribution. The pollution-induced community tolerance concept (PICT) uses observable community responses by measuring increases in community tolerance caused by the replacement of sensitive species after exposure. The aim of this study was to compare these two concepts in assessing the effects of three herbicides. Atrazine, prometryn, and isoproturon were found to increase community tolerance by a factor up to six. Atrazine increased community tolerance only at higher test concentrations (0.125 mg L(-1)). Species-sensitivity distributions correspond well to community responses: The median effective concentrations (EC50s) of untreated periphyton communities tested covered 55 to 65% of affected species represented in the SSD. The sensitivities of tolerant algal communities shifted to the right end of the SSDs. In the microcosm experiments, higher test concentrations affected biomass, species numbers, and community structure. Community tolerance could not be induced any further, suggesting that these concentrations represent a maximum of functional redundancy of a functional group. At higher concentrations, even the least-sensitive species are affected. These results can be interpreted as a confirmation of the SSD concept by observed algal community responses, when applied to photosystem II (PSII)-inhibiting herbicides.

Effects of copper and temperature on aquatic bacterial communities

The present study aimed to characterise effects of copper and temperature on bacterial communities in photosynthetic biofilms using a suit of supplementary methods: pollution-induced community tolerance (PICT), DNA profiles with denaturing gradient gel electrophoresis (DGGE) and physiological profiles with community-level physiological profiling (CLPP). Biofilms of algae and bacteria were grown in a ditch of a Dutch polder and exposed in the laboratory to copper (3 μM and a reference) at three different temperatures (10, 14 and 20 °C). Bacterial communities sampled from the field showed heterogeneity in their physiological profiles, however the heterogeneity decreased during laboratory incubation. After 3 days laboratory incubation, the copper treated biofilms were different from the reference biofilms, as revealed by DGGE and CLPP analyses. Effects of temperature were not observed in the CLPPs, or in the DGGE profiles. PICT was observed for the bacterial communities at all temperatures. The copper-tolerance at 10 and 14 °C increased about 3 times, whereas copper-tolerance at 20 °C increased about 6 times. Temperature had an effect on the community tolerance, but not on the structure or on the physiological profile, suggesting that temperature was not a major factor causing successional changes under these laboratory conditions. In contrast, temperature had an effect on tolerance development indicating that the exposure to copper was enhanced at higher temperature.

Toxic effects of isoproturon on periphyton communities – a microcosm study

Coastal and estuarine ecosystems are increasingly exposed to herbicide contamination. To study the potential risks of these pollutants, the establishment of periphyton communities under exposure of a concentration series of isoproturon in a range of 0.0024–0.312 mg L −1 was investigated in a microcosm study. After two weeks of growth, chronic effects on biomass development, measured as Chl a fluorescence, taxonomic composition and photosynthetic capacity of PS II of attached microalgae were analysed using a pulse-amplitude modulated (PAM) fluorescence-based method. Algal classes were differentiated according to their pigment systems using four excitation wavelengths. Biomass remained constant up to pre-exposure concentrations of 0.02 mg L −1 isoproturon but decreased within one order of magnitude at the highest test concentration. Algal classes shifted from diatoms to chlorophytes at pre-exposure concentrations of 0.02–0.39 mg L −1. Community structure of diatoms representing the dominant group of attached algae at lower test concentrations was studied by microscopic analysis revealing that species numbers were highest at lowest test concentrations as compared to controls. At higher test concentrations Navicula halophila dominated with 89% of the diatom biomass but was abnormally shaped. Pollution-induced community tolerance (PICT) of periphyton to isoproturon was determined by short-term inhibition tests of photosynthesis. Pre-exposed communities increased their effect concentrations (EC 50) up to threefold within the 0.039 mg L −1 pre-treatment. Our results suggest that attached microalgal communities have a high potential of tolerance development to isoproturon at lower levels of contamination due to the replacement of sensitive species by more tolerant algae, although these species seemed to be affected too. We conclude that herbicide concentrations in river basins, estuaries or coastal zones may change community structure and primary production of microphytobenthos even when the contamination does not exceed levels of acute toxicity.

Pollution‐induced community tolerance of soil microbes in response to a zinc gradient

The long-term accumulation of Zn in soils provides the microbial community time to adapt to this heavy metal. To assess the effects of long-term exposure to Zn on the metabolic diversity and tolerance to Zn of soil microbial community, the pollution-induced community tolerance (PICT) method, based on the Biolog microtitre plate system, was used. It especially is useful to study gradients of pollutants for detecting PICT. Such a Zn gradient was found by selecting soils at increasing distances from galvanized electricity pylons at two sites where Zn accumulation had occurred over a period of decades. Soil metabolic profiles showed a humpbacked response to increasing soil Zn concentrations, indicating that accumulation of Zn up to 2,000 mg/kg did not decrease the metabolic biodiversity in the culturable fraction of the microbial community. This fraction of the microbial community showed significant PICT, because the effective concentration that reduces the metabolic biodiversity by 50% (EC50), values for Zn added to the Biolog wells increased by up to two orders of magnitude with increasing soil-Zn concentration along the transects. Significant PICT was detectable at soil Zn concentrations above approximately 300 mg/kg. The EC50 values correlated more closely with soil total Zn than with soil pore water Zn. The results suggest that, although long-term exposure to Zn imposes stress on soil microbes, resulting in an increased tolerance, detectable PICT does not necessarily lead to a decrease in metabolic diversity.

Location‐specific ecotoxicological risk assessment of metal‐polluted soils

When chemical analysis indicates metal pollution, a second-tier method is needed to evaluate whether toxic effects occur at the polluted sites. A method based on pollution-induced community tolerance (PICT) was developed using samples taken from locations polluted with sewage more than 20 years ago. Microorganisms extracted from soil samples were exposed to a concentration range of zinc, nickel, copper, chromium (III), or chromium (VI) salts in a buffer suspension. The remaining activity of the intoxicated microorganisms was determined by color formation with 31 different organic substrates in microtiter plates. Microorganisms from moderately Zn-polluted sites (>45 mg/kg) showed an increased tolerance for zinc. Nickel tolerance was observed at 51 mg Ni/kg soil, chromium (VI) tolerance at 923 mg Cr/kg. In most cases, tolerance also was observed at higher concentrations. High concentrations of 1,494 mg Cu/kg or 3,935 mg Cr/kg did not show PICT, indicating a limited bioavailability of Cu and Cr at these sites. The benefits of our method are its greater sensitivity compared to other tests used at these sites, and its specificity for those metals that exceed allowable levels.

Microbial community-level toxicity testing of linear alkylbenzene sulfonates in aquatic microcosms

Complex microbial communities may serve as ideal and ecologically relevant toxicity indicators. We here report an assessment of frequently used methods in microbial ecology for their feasibility to detect toxic effects of the environmentally important surfactant linear alkylbenzene sulfonate (LAS) on microbial communities in lake water and treated waste water. The two microbial communities were evaluated for changes in community structure and function over a period of 7 weeks in replicated aquatic microcosms amended with various levels of LAS (0, 0.1, 1, 10 or 100 mg l −1) and inorganic nutrients. In general, the two communities behaved similarly when challenged with LAS. Following lag periods of 1–3 weeks, LAS was degraded to non-toxic substances. Denaturing gradient gel electrophoresis of 16S rRNA gene fragments and [ 3H]leucine incorporation were the most sensitive assays with effect levels of 0–1 and 1–10 mg LAS l −1, respectively. Community-level physiological profiles and pollution-induced community tolerance determinations using Biolog microplates demonstrated less sensitivity with effect levels of 10–100 mg LAS l −1. Total cell counts and net uptake of inorganic N and P were unaffected even at 100 mg LAS l −1. Interestingly, different microbial communities developed in some replicate microcosms, indicating the importance of stochastic events for community succession. We conclude that microbial community-level toxicity testing holds great promise and suggest a polyphasic approach involving a range of independent methods targeting both the structure and function of the tested microbial communities.

Pollution-induced community tolerance of soil microbial communities caused by the antibiotic sulfachloropyridazine.

Little is known about the environmental hazards linked to the treatment of farm animals with antibiotics and subsequent spreading of manure, especially regarding soil microbial communities. In this investigation, pollution-induced community tolerance (PICT) of bacteria from soils artificially spiked with the sulfonamide sulfachloropyridazine (SCP) was investigated. Tolerance of the bacterial communities after 3 weeks' exposure to SCP was determined by analyzing the sensitivity of 31 microbial metabolic processes in microtiter plates. Bacterial suspensions extracted from soils containing higher concentrations of SCP showed an increased tolerance of their metabolic activities to this antibiotic. An increase in tolerance by 10% was found at 7.3 mg/kg dw SCP. The PICT effect could be demonstrated by both a shift in the tolerance of the average of all metabolic activities and a shift of the physiological process sensitivity distributions made up from the single metabolic processes. The PICT effect was accompanied by smaller changes in the community-level physiological profile (CLPP). To conclude, PICT has been shown to be a versatile and illustrative method for the detection of the effects of antibacterial agents on soil microorganisms.

Pollution induced community tolerance (PICT) and analysis of 16S rRNA genes to evaluate the long‐term effects of herbicides on methanotrophic communities in soil

SummaryThere is an increasing interest in agricultural systems in which the use of herbicides is forbidden. Therefore, soils treated with herbicides atrazine and metolachlor for the last 20 years were compared with soil samples from the same field that had never been treated (control soil). We determined the pollution induced community tolerance (PICT) by evaluating the methane oxidation capacity of soil samples after adding increasing amounts of a methane oxidation inhibitor, 2,4‐dichlorophenoxyacetic acid (2,4‐D). Denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes assessed whether the soil methanotrophic community differed between the two treatments. Addition of 60 µg 2,4‐D per g soil clearly inhibited methane oxidation in both soils but increased the time needed to oxidize 5% methane in the headspace by 250% for the control soil compared with 175% for the herbicide‐treated soil. This indicates that the soil with a long‐term herbicide history had a greater tolerance to the methane oxidation inhibitor than did the control soil. The DGGE of 16S rRNA genes amplified directly from soil community DNA could also distinguish the two treatments. The banding patterns of the Type I methanotrophs contained fewer bands in the herbicide‐treated soil. It seems that both the PICT approach and DGGE analysis are effective assays to distinguish a long‐term herbicide‐treated soil from an untreated soil.

Variability in zinc tolerance, measured as incorporation of radio-labeled carbon dioxide and thymidine, in periphyton communities sampled from 15 European river stretches.

Fifteen European rivers and streams belonging to watersheds in Sweden, the Netherlands, and Spain respectively, were sampled by allowing periphyton to colonize submerged glass substrata. Their zinc tolerances were quantified in short-term laboratory tests, where inhibition of photosynthesis in microalgae and thymidine incorporation in bacterial DNA was measured, and expressed as EC50 values. The variability in zinc tolerances was high reaching 1.5-2.5 orders of magnitude, ranging from 25-8145 microM for photosynthesis and 15-467 microM for thymidine assays. Based on the observed variability, uncertainty factors were estimated for the extrapolation of zinc toxicity data from river to river, both regionally and interregionally. Under the assumption to protect 95% of the observed communities the regional uncertainty factors were 1.7-4.3 and the interregional 2.4-8.6. The sampling sites were characterized in terms of biotope physiography, water chemistry, periphyton biomass, trace element content, and species composition. Multivariate analysis of the data using PLS (Projection to Latent Structure), was used to generate hypotheses about the relation between periphyton zinc tolerance and the 123 so-called predictor variables. Zinc contamination, phosphate, nitrogen nutrients, pH, calcium, bicarbonate, dissolved organic carbon, and various diatom species are important predictors for zinc tolerance in the entire data set representing all 15 river stretches. Regional models suggested that very different factors determined the zinc tolerance in the Swedish and Dutch periphyton. The results are interpreted in terms of Pollution-Induced Community Tolerance (PICT) and the bioavailability of zinc.

A Critical Review of Procedures and Approaches Used for Assessing Pollution-Induced Community Tolerance (PICT) in Biotic Communities

Pollution-induced community tolerance (PICT) is used for the detection of minor effects of toxicants in biotic communities. Organisms survive in toxic environments only if they are tolerant to the chemicals present in their habitat. In the selection phase, toxicants hinder the success of sensitive individuals and species and replace them by more tolerant ones. The resulting increase in community tolerance is quantified in the detection phase by short-term toxicity tests. In this way PICT can establish causal linkages between contaminants and effects. An increase in community tolerance compared to the baseline tolerance at reference sites suggests that the community has been adversely affected by toxicants. PICT has been used in aquatic and terrestrial environments with communities of periphyton, phytoplankton, bacteria, nematodes and insects. A variety of methods have been used for quantification of community tolerance including photosynthesis, sulfolipid synthesis, respiration, thymidine and leucine incorporation, survival, and substrate utilisation patterns. PICT has been observed for copper, zinc, nickel, mercury, cadmium, arsenate, monomethylarsonic acid, diuron, tributyl tin, 4,5,6-trichloroguaiacol, irgarol 1051, seanine 211, atrazine, and trinitrotoluene. It is necessary to validate PICT, at least by showing that it is related to the preexposure concentration of the toxicants and that it is coupled to a toxicant-induced succession (TIS) in the community. Care must also be taken to ascertain that PICT interpretation is not confounded by co-tolerance or bioavailability differences. Co-tolerance patterns, which are indicative of the specificity of PICT, have only been investigated for arsenate, diuron and a few metals. For the further improvement of PICT methodology special attention should be given to co-tolerance patterns and development of new integrating short-term tests for quantification of tolerance. It is also important to broaden the scope of organisms and toxicants used. Properly validated, PICT is a powerful tool for detection of community effects and its use in monitoring and site-specific risk assessment should be encouraged.