Short-term exposure to sub-lethal doses of lindane affects developmental parameters in Chironomus riparius Meigen, but has no effect on larval glutathione-S-transferase activity

Chironomus riparius Meigen were exposed to 0, 0.01, 0.1, 0.5, 0.75 and 1.0 ppm lindane for 48 h as fourth instar larvae. Exposure had no effect on glutathione-S-transferase (GST) activity in larvae snap-frozen immediately following exposure. In contrast, exposure had longer-term consequences affecting developmental parameters. Concentrations above 0.5 ppm lindane affected larval behaviour, reduced adult body size and fecundity and delayed emergence times. The lack of significant change in GST activity when life history characters were affected by high concentrations of lindane, suggests that in C. riparius, GST is not a sensitive biomarker of pesticide exposure or effect.     

In vivo evaluation of three biomarkers in the mosquitofish (Gambusia yucatana) exposed to pesticides

In this study, the acute toxicity and the in vivo effects of commercial chlorpyrifos, carbofuran and glyphosate formulations on cholinesterase (ChE), glutathione S-transferase (GST) and lactate dehydrogenase (LDH) activities of the mosquitofish (Gambusia yucatana) were investigated. In a first phase of the study, head and muscle ChE were characterized with different substrates (acetylthiocholine iodide, s-butyrylthiocholine iodide and propionylthiocholine iodide) and the selective inhibitors eserine hemisulfate, 1,5-bis(4-allyldimethylammoniumphenyl)-pentan-3-one dibromide (BW284C51), and N,N'-diisopropylphosphorodiamic acid (iso-OMPA). The results obtained suggest that the enzyme present in both head and muscle of G. yucatana is mainly acetylcholinesterase (AChE). Acute toxicity was evaluated by exposing fish to several concentrations of single pesticides and of a mixture of chlorpyrifos/glyphosate. LC50 values were determined after 96 h of exposure, except in the case of carbofuran for which LC50 was calculated after 24 h since almost all the fish died within this period. LC50 values were 0.085 mg/l for chlorpyrifos, 17.79 mg/l for glyphosate, 0.636 mg/l for carbofuran and 0.011 mg/l for the chlorpyrifos/glyphosate mixture. A Toxic Unit approach was used to compare the toxicity of chlorpyrifos and glyphosate when occurring in a mixture with their toxicities as single compounds. Synergistic effects of chlorpyrifos and glyphosate when present in a mixture were found. At the end of each bioassay (24 h for carbofuran, 96 for the other substances/mixture), effects on biomarkers were analyzed. Muscle LDH activity was not altered by any of the three pesticides tested. Gill GST activity was significantly inhibited (40%) by carbofuran after 24 h of exposure to concentrations equal or higher than 0.06 mg/l. ChE muscle and head activity were significantly inhibited (50% and 30%, respectively) by carbofuran at concentrations equal or higher than 0.25 mg/l. Chlorpyrifos induced a significant inhibition of both muscle and head ChE (80% and 50%, respectively) after 96 h of exposure to concentrations equal or higher than 0.05 mg/l. Carbofuran did not induce significant alterations of fish ChE. The ChE EC50 determined for chlorpyrifos/glyphosate mixture (0.070 mg/l) was higher than the correspondent value calculated for chlorpyrifos alone (0.011 mg/l) suggesting an antagonistic effect of glyphosate on ChE inhibition by chlorpyrifos. ChE activity of G. yucatana seems to be a good biomarker to diagnose the exposure of wild populations of this species exposed to anticholinesterase pesticides.     

Combined effects of copper and food on the midge Chironomus riparius in whole-sediment bioassays

Effects observed in whole-sediment bioassays must be seen as the joint effect of all sediment characteristics. In whole-sediment bioassays, however, adverse effects on test organisms are usually attributed to the presence of contaminants and effects of food are often ignored. The aim of this study was to analyze the response of the midge Chironomus riparius to sediment spiked with different combinations of food and copper. The responses of C. riparius to these spiked sediments were assessed in 10-day whole-sediment bioassays. Decreases in survival, dry weight, and length of C. riparius were observed with increasing copper concentrations. However, an increase in the amount of food resulted in an increase of larval dry weight and length until copper concentrations reached a critical threshold of 200 mg/kg. In addition, an increase in the amount of food resulted in a decrease of accumulated copper in the larvae. The present study demonstrated that the combination of copper and food in the sediment determines the performance of C. riparius in whole-sediment bioassays. The dependency of C. riparius on high feeding levels, which mask toxic effects, questions its suitability as a test organism for whole-sediment bioassays. Because benthic communities in polluted ecosystems are often exposed to varying levels of both food and toxicants it is concluded that the trophic state of the ecosystem may alter the ecological risk of sediment-bound toxicants to opportunistic benthic invertebrates such as C. riparius.     

Inhibition, recovery and oxime-induced reactivation of muscle esterases following chlorpyrifos exposure in the earthworm Lumbricus terrestris

Assessment of wildlife exposure to organophosphorus (OP) pesticides generally involves the measurement of cholinesterase (ChE) inhibition, and complementary biomarkers (or related endpoints) are rarely included. Herein, we investigated the time course inhibition and recovery of ChE and carboxylesterase (CE) activities in the earthworm Lumbricus terrestris exposed to chlorpyrifos, and the ability of oximes to reactivate the phosphorylated ChE activity. Results indicated that these esterase activities are a suitable multibiomarker scheme for monitoring OP exposure due to their high sensitivity to OP inhibition and slow recovery to full activity levels following pesticide exposure. Moreover, oximes reactivated the inhibited ChE activity of the earthworms exposed to 12 and 48 mg kg⁻¹ chlorpyrifos during the first week following pesticide exposure. This methodology is useful for providing evidence for OP-mediated ChE inhibition in individuals with a short history of OP exposure (≤1 week); resulting a valuable approach for assessing multiple OP exposure episodes in the field.     

Enzymatic biomarkers in the crab Carcinus maenas from the Minho River estuary (NW Portugal) exposed to zinc and mercury

Zinc and mercury are common contaminants of estuaries and coastal ecosystems where they may induce adverse effects on the biota. Carcinus maenas is a key-species in several European estuaries, living in close association with the sediment where a considerable number of environmental contaminants, including zinc and mercury may accumulate. In the present study, the acute effects of zinc and mercury on C. maenas from the Minho River Estuary (NW Portugal) were investigated by using the activity of the enzymes cholinesterase (ChE), lactate dehydrogenase (LDH) and glutathione S-transferases (GST) as effects' criteria. Crabs were exposed for 96h to several concentrations of Zn(2+) (0, 1.84, 3.71, 7.39 and 14.79mg/l) or Hg(2+) (0, 0.09, 0.19, 0.37 and 0.74mg/l). The choice of Zn(2+) test concentrations was based on the LC50 value (14.86mg/l) determined in a first part of the study, while the choice of Hg(2+) concentrations was based on values from the literature. At the end of the bioassays, eye, muscle and hepatopancreas tissues were isolated and used for ChE, LDH and GST determinations, respectively. Zinc significantly inhibited ChE activity (p<0.05, EC50=14.68mg/l), caused significant alterations of GST activity (p<0.05) and induced LDH activity (p<0.05) at the exposure of 14.79mg/l. Mercury significantly inhibited ChE activity (p<0.001, LOEC=0.09mg/l, EC50=0.235mg/l) and increased both GST (p<0.05, LOEC=0.774mg/l) and LDH activities (p<0.05, LOEC=0.119mg/l). These results suggest that both metals interfere with cholinergic neurotransmission in C. maenas by inhibiting ChE activity. In addition, they also interfere with GST activity and this may reduce the capacity of detoxification of some chemicals and/or to increase the probability of oxidative stress to occur. Furthermore, both metals increase LDH activity, suggesting an interference with energy production pathways. Therefore, the presence of zinc and mercury in estuaries at concentrations in the mg/l or mug/l range, respectively, may have a negative impact on C. maenas.     

In situ assay with the midge Kiefferulus calligaster for contamination evaluation in aquatic agro-systems in central Thailand

The aims of this study were to verify the suitability of in situ tests using the tropical midge Kiefferulus calligaster and to evaluate the most sensitive endpoint for the assessment of aquatic pesticide contamination. In situ tests were carried out in freshwater drainage channels (farm channels) that supply vegetable crops and receive considerable pesticide spray drift, and at channels outside farms (main channels). Moreover a pesticide-free farm was used as reference site. The endpoints analysed were: survival of the larvae, body length increment, capsule width increment, cholinesterase activity and glutathione S-transferase activity. Seasonal change was investigated as rainy season and dry season. Deleterious effects were observed at some farms especially during the rainy season when farmers apply heavier doses of pesticides. However, high mortality rates observed in main channels suggest that these water bodies are also affected by other impacts besides pesticide use. This work shows the potential of the in situ assay with K. calligaster as a tool for the environmental quality assessment of tropical aquatic ecosystems.     

Biochemical biomarkers in environmental studies—lessons learnt from enzymes catalase,glutathione <Emphasis Type="Italic">S</Emphasis>-transferase and cholinesterase in two crustacean species

Background, aim and scope  

For reliable environmental risk assessment of pollutants, knowledge on the effects at different levels of biological organisation is needed. During the early days of biomarker research in environmental studies approximately two decades ago, biochemical biomarkers were considered as the most promising tool for such purposes. Among these, three enzymes have often been studied: catalase (CAT), glutathione S-transferase (GST) and cholinesterase (ChE). However, despite their intensive research, their measurements in invertebrates have not been commonly applied in environmental risk assessment (ERA) or for regulatory purposes.     

Cholinesterase characterization and effects of the environmental contaminants chlorpyrifos and carbofuran on two species of marine crabs,Carcinus maenas and Pachygrapsus marmoratus

Among the most frequent targets for toxic effects of modern pesticides, namely organophosphates and carbamates, one may find cholinesterases (ChEs). ChEs exist in a wide variety of animals and have been used actively to discriminate among the environmental effects of different pollutant groups, including the aforementioned pesticides. This study had three purposes, namely (i) identifying the ChE forms present in tissues (eyes and walking legs muscle) of two crab species, Carcinus maenas and Pachygrapsus marmoratus; to (ii) determine the in vitro toxicological effects, and (iii) compare the sensitivity of such enzymatic forms towards commonly used anti-ChE pesticides, namely the organophosphate chlorpyrifos and the carbamate carbofuran. Our results showed that there was not a clear preference for any of the tested substrates in any of the tissues from both species. Furthermore, the ChE activity was almost completely suppressed following incubation with eserine and with the specific inhibitor BW284C51 in all tissues from both species. In vitro exposure to chlorpyrifos promoted a significant decrease in ChE activity in both species. Furthermore, the ChE activity was completely suppressed following incubation with carbofuran and chlorpyrifos. These results suggest that the major ChE forms present in tissues of both crab species show intermediate structural properties and activity patterns, halfway between classic acetylcholinesterase and pseudocholinesterases. However, the sensitivity of the found forms towards ChE inhibitors was established, and the responsiveness of such forms towards common anti-ChE chemicals was established. Both tested species seem to be promising test organisms to be used in marine and coastal scenarios of putative contaminations by anti-ChE chemicals, considering the here reported patterns of response.

     

Cholinesterase activities as potential biomarkers: characterization in two freshwater snails, Potamopyrgus antipodarum (Mollusca, Hydrobiidae, Smith 1889) and Valvata piscinalis (Mollusca, Valvatidae, Müller 1774)

Anti-cholinesterase insecticides constitute a major portion of modern synthetic pesticides and the assessment of cholinesterase (ChE) inhibition is widely used as a specific biomarker for evaluating the exposure of non-target organisms to these pollutants. However, most studies on this biomarker were developed on vertebrates and among invertebrates, gastropod mollusks are rarely used. Gastropods are important members of aquatic habitats and therefore present a high ecological relevance for freshwater ecosystems. In this context, ChE activities were characterized in two freshwater gastropod mollusks, Potamopyrgus antipodarum and Valvata piscinalis, in order to ascertain their value as sentinel species. Firstly, characterization of ChE activities was performed using different substrates (acetylcholine iodide, butyrylcholine iodide and propionylcholine iodide) and specific inhibitors (eserine, iso-OMPA and BW284c51). Secondly, in vivo effect of a widely used organophosphate insecticide, chlorpyrifos, was tested on ChE activity in both species. Results suggested that P. antipodarum possesses two isoforms of cholinesterases, one isoform which properties are intermediate between an acetyl and a propionyl ChE, and one minor isoform which correspond to a butyryl ChE, while V. piscinalis seems to possess only one isoform which displays typical properties of an acetyl ChE. Chlorpyrifos induced no effect on V. piscinalis ChE. In contrast, P. antipodarum activity was significantly decreased by environmental realistic chlorpyrifos concentrations (2.86 and 14.2 nM) after seven days of contact. The present study suggests that P. antipodarum may be employed as a biological indicator for assessing pesticide contamination.     

Binary mixtures of azinphos-methyl oxon and chlorpyrifos oxon produce in vitro synergistic cholinesterase inhibition in Planorbarius corneus

In this study, the cholinesterase (ChE) and carboxylesterase (CES) activities present in whole organism homogenates from Planorbarius corneus and their in vitro sensitivity to organophosphorous (OP) pesticides were studied. Firstly, a characterization of ChE and CES activities using different substrates and selective inhibitors was performed. Secondly, the effects of azinphos-methyl oxon (AZM-oxon) and chlorpyrifos oxon (CPF-oxon), the active oxygen analogs of the OP insecticides AZM and CPF, on ChE and CES activities were evaluated. Finally, it was analyzed whether binary mixtures of the pesticide oxons cause additive, antagonistic or synergistic ChE inhibition in P. corneus homogenates. The results showed that the extracts of P. corneus preferentially hydrolyzed acetylthiocholine (AcSCh) over propionylthiocholine (PrSCh) and butyrylthiocholine (BuSCh). Besides, AcSCh hydrolyzing activity was inhibited by low concentrations of BW284c51, a selective inhibitor of AChE activity, and also by high concentrations of substrate. These facts suggest the presence of a typical AChE activity in this species. However, the different dose-response curves observed with BW284c51 when using PrSCh or BuSCh instead of AcSCh suggest the presence of at least another ChE activity. This would probably correspond to an atypical BuChE. Regarding CES activity, the highest specific activity was obtained when using 2-naphthyl acetate (2-NA), followed by 1-naphthyl acetate (1-NA); p-nitrophenyl acetate (p-NPA), and p-nitrophenyl butyrate (p-NPB). The comparison of the IC(50) values revealed that, regardless of the substrate used, CES activity was approximately one order of magnitude more sensitive to AZM-oxon than ChE activity. Although ChE activity was very sensitive to CPF-oxon, CES activity measured with 1-NA, 2-NA, and p-NPA was poorly inhibited by this pesticide. In contrast, CES activity measured with p-NPB was equally sensitive to CPF-oxon than ChE activity. Several specific binary combinations of AZM-oxon and CPF-oxon caused a synergistic effect on the ChE inhibition in P. corneus homogenates. The degree of synergism tended to increase as the ratio of AZM-oxon to CPF-oxon decreased. These results suggest that synergism is likely to occur in P. corneus snails exposed in vivo to binary mixtures of the OPs AZM and CPF.     

Glutathione, glutathione S-transferase, and glutathione conjugates, complementary markers of oxidative stress in aquatic biota

Contaminants are ubiquitous in the environment and their impacts are of increasing concern due to human population expansion and the generation of deleterious effects in aquatic species. Oxidative stress can result from the presence of persistent organic pollutants, metals, pesticides, toxins, pharmaceuticals, and nanomaterials, as well as changes in temperature or oxygen in water, the examined species, with differences in age, sex, or reproductive cycle of an individual. The antioxidant role of glutathione (GSH), accompanied by the formation of its disulfide dimer, GSSG, and metabolites in response to chemical stress, are highlighted in this review along with, to some extent, that of glutathione S-transferase (GST). The available literature concerning the use and analysis of these markers will be discussed, focusing on studies of aquatic organisms. The inclusion of GST within the suite of biomarkers used to assess the effects of xenobiotics is recommended to complement that of lipid peroxidation and mixed function oxygenation. Combining the analysis of GSH, GSSG, and conjugates would be beneficial in pinpointing the role of contaminants within the plethora of causes that could lead to the toxic effects of reactive oxygen species.     

Characterisation of cholinesterases and evaluation of the inhibitory potential of chlorpyrifos and dichlorvos to Artemia salina and Artemia parthenogenetica

In this study, the acute toxicity of the organophosphorous pesticides dichlorvos and chlorpyrifos to two different species of Artemia (A. salina and A. parthenogenetica) was evaluated. In addition, the in vivo effect of these two pesticides on cholinesterase (ChE) activity of both A. salina and A. parthenogenetica was also determined. The characterisation of the ChE, using different substrates and specific inhibitors, and the normal range of activity in non-exposed individuals were previously investigated for both species. The results obtained indicate that the ChE of A. salina is different from that of A. parthenogenetica and that both enzymes cannot be classified neither as acetylcholinesterase nor as butyrylcholinesterase since they show intermediary characteristics between the two vertebrate forms. The range of normal ChE activity was 2.65+/-0.15 U/mg protein for A. salina, and 3.69+/-0.17 U/mg protein for A. parthenogenetica. Significant in vivo effects of both pesticides on Artemia ChE activity were found, at concentrations between 5.38 and 9.30 mg/l for dichlorvos and between 1.85 and 3.19 mg/l for chlorpyrifos. Both Artemia species are resistant to these pesticides and they are able to survive with more than 80% ChE inhibition. However, A. parthenogenetica is more resistant than A. salina, with about a 95% reduction in its ChE activity respect to the control for nauplii exposed to the median lethal concentrations (LC50), without lethal effects after 24 h of exposure.     

In situ bioassay chambers and procedures for assessment of sediment toxicity with Chironomus riparius

The purpose of this study was to develop an in situ sediment bioassay chamber and respective procedures, suitable for performing toxicity bioassays with benthic invertebrates, using the midge Chironomus riparius. It was also our objective to compare the responses obtained under controlled conditions (laboratory 10-day larval growth and survival test) with those obtained in situ. Clean sand and a formulated sediment were incorporated in the in situ bioassay, along with local sediments, as a way of minimizing natural variability due to physicochemical differences among sediments or due to interactions with indigenous organisms. Recovery of organisms was good (80-100% in the control and reference site), indicating that the developed chamber and protocol were suitable for exposing and retrieving C. riparius in situ. Results also showed differences between responses obtained with formulated and natural sediment in situ, as well as between laboratory and in situ.     

Evidence of differences in the biotransformation of organic contaminants in three species of freshwater invertebrates

Acute static bioassays were performed using three freshwater invertebrate species (the oligochaete Lumbriculus variegatus, the fingernail clam Sphaerium corneum and the larvae Chironomus riparius) exposed separately to a variety of 14C radiolabelled contaminants. The aim of this work was to investigate if the chemicals remained as parent compounds after the treatments. Chemicals used were 2,4-dichlorophenol; 2,4,5-trichlorophenol; pentachlorophenol; pyrene; Fenpropidin, and Trifluralin. Homogenates of the whole body tissue of each organism were prepared and total radioactivity was measured. Contaminants were then extracted into organic solvents and analysed by high-pressure liquid chromatography techniques. Chromatograms showed that most of the substances extracted were present as parent compounds in S. corneum and in L. variegatus. In contrast, for C. riparius a low proportion of the chemicals was recovered as parent compounds. These results suggest that different metabolic processes could take place in the different species.     

Cholinesterase and glutathione S-transferase activities of three mollusc species from the NW Portuguese coast in relation to the ‘Prestige’ oil spill

In November 2002, the tanker ‘Prestige’ released about 19,000 tonnes of a heavy fuel oil (no. 6) before sinking with about 58,000 tonnes of its cargo, 135 miles from Cabo Finisterra (Spain). A considerable part of the released fuel oil reached the Galician coast, causing a heavy black tide and an ecological disaster. Although the black tide did not reach the NW coast of Portugal, it is possible that some of the fuel oil or its components also arrived to this area directly through the sea water and/or indirectly through the food chain. Therefore, the aim of this study was to investigate possible changes in two widely used biomarkers, the activity of the enzymes cholinesterases (ChE) and glutathione S-transferases (GST), of three molluscs (Mytilus galloprovincialis, Nucella lapillus and Monodonta lineata) from wild populations of the NW Portuguese coast in relation to the ‘Prestige’ oil spill. Molluscs were collected seasonally before (autumn 2002) and after (winter 2002/2003), spring and summer 2003) the oil spill at several sites along the Portuguese NW coast. Enzymatic activities determined before the accident were compared with those determined at different times after the oil spill taking into consideration abiotic factors. Information from different parameters was integrated by Redundancy Analysis and Principal Response Curves (PRC). Results show that GST and ChE activities were influenced by abiotic factors. Despite this influence, the results of PRC analysis also suggest that some of the fuel oil reached the NW Portuguese coast changing the patterns of ChE and GST activities of local populations of rocky shore species. Furthermore, the present study highlights the need of long-term monitoring with wild populations to assess both historical and punctual effects of pollution in the marine environment.     

Neurotoxic effect and metabolic responses induced by a mixture of six pesticides on the earthworm Aporrectodea caliginosa nocturna

The effects of a mixture of insecticides and/or fungicides at different environmental concentrations were investigated on a Aporrectodea caliginosa nocturna population. This laboratory experiment was carried out in order to reproduce Gaillac (France) vineyard conditions. Neurotoxicity (cholinesterase), metabolisation (glutathione-S-transferase) and oxidative stress (catalase) enzymes were studied as biomarkers in earthworms after short-term exposure in terraria. The aim was to observe the global effects of pesticide exposure, as in a vineyard, rather than focus on each isolated biomarker variation, or on each compound's impact. ChE activity was inhibited after a few days of insecticide and/or fungicide exposure, indicative of a neurotoxic effect in earthworms. The significant increase in GST and CAT activities revealed the metabolisation of these products resulting in the production of reactive oxygen species. After a long period of exposure or high concentrations, earthworms were physiologically damaged: they could not cope with the high toxicity (cellular dysfunction, protein catabolism...). Chemical analysis showed that pesticide bioaccumulation in earthworm tissues, even in those exposed to the highest concentrations and for the longest periods, was very low (under LOD) or absent. However, the study of pesticide residues in terraria after 34 days in a climate chamber suggested that earthworms participate in soil pesticide breakdown.     

The measurement of cholinesterase activities as a biomarker in chub (Leuciscus cephalus): the fish length should not be ignored

Biomarkers are early warning systems of the exposure of aquatic organisms to pollutants. Among them, the measurement of the cholinesterase (ChE) activities in fish muscle is a biomarker of the exposure to organophosphosphates and carbamates pesticides. As such it has been used in numerous field studies both in marine and continental waters. Cyprinids (chub, Leuciscus cephalus) were sampled in river sites (France) in relatively clean and polluted areas. We performed the statistical analysis of the ChE activities and we generally observed a statistical relationship between ChE activities and fish length, the larger fish having the lower ChE activities. We concluded that the great majority of the significant differences in ChE activities between sites could be due in fact to differences in fish length between field samples. We stress the importance of taking into account the fish length whenever differences in ChE levels between field sites must be interpreted.     

Oxidative stress and detoxification biomarker responses in aquatic freshwater vertebrates exposed to microcystins and cyanobacterial biomass

Cyanobacterial blooms represent a serious threat to the aquatic environment. Among other effects, biochemical markers have been studied in aquatic vertebrates after exposures to toxic cyanobacteria. Some parameters such as protein phosphatases may serve as selective markers of exposure to microcystins, but under natural conditions, fish are exposed to complex mixtures, which affect the overall biomarker response. This review aims to provide a critical summary of biomarker responses in aquatic vertebrates (mostly fish) to toxic cyanobacteria with a special focus on detoxification and oxidative stress. Detoxification biomarkers such as glutathione (GSH) and glutathione-S-transferase (GST) showed very high variability with poor general trends. Often, stimulations and/or inhibitions and/or no effects at GSH or GST have been reported, even within a single study, depending on many variables, including time, dose, tissue, species, etc. Most of the oxidative stress biomarkers (e.g., superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase) provided more consistent responses, but only lipid peroxidation (LPO) seemed to fulfill the criteria needed for biomarkers, i.e., a sufficiently long half-life and systematic response. Indeed, reviewed papers demonstrated that toxic cyanobacteria systematically elevate levels of LPO, which indicates the important role of oxidative damage in cyanobacterial toxicity. In summary, the measurement of biochemical changes under laboratory conditions may provide information on the mode of toxic action. However, comparison of different studies is very difficult, and the practical use of detoxification or oxidative stress biomarkers as diagnostic tools or early warnings of cyanobacterial toxicity is questionable.     

Short-term effects of dimethoate on metabolic responses in Chrysolina pardalina (Chrysomelidae) feeding on Berkheya coddii (Asteraceae), a hyper-accumulator of nickel

Berkheya coddii Roessler (Asteraceae) is a hyper-accumulator of nickel, which can be used in phytomining and phytoremediation. Chrysolina pardalina Fabricius (Chrysomelidae) is a phytophagous leaf beetle, which may be useful in controlling population levels of B. coddii after it has been introduced into a new habitat. The aim of this study was to investigate the response of C. pardalina to topical application of dimethoate. Data recorded included the activity of acetylcholinesterase (AChE), the concentration of glutathione (GSH), and the activity of selected enzymes connected with GSH metabolism. Assays were carried out several times during the first 24h after exposure to dimethoate. At the dosages used in this study, dimethoate was not as toxic as expected. AChE activity was significantly decreased 14 and 24h after application. GST activity was significantly decreased 24h after application. GSTPx activity was significantly decreased 2, 14 and 24h after application. GR activity was significantly increased 4h after application. GSH concentration was significantly increased 24h after application. Long-term exposure to high levels of nickel may have caused adaptive changes in the enzymes that enable C. pardalina to deal with other stressors, including organophosphate pesticides.     

Effects of organophosphate compounds on a soil protist, Colpoda inflata (Ciliophora, Colpodidae)

Many investigations on protists indicate that they play an important role in agricultural soils. We have tested the effects of three organophosphate (OP) pesticides, basudin, cidial, and fenix, on the soil ciliate Colpoda inflata, and examined its viability, fission rate, ability to excyst and extrude macronuclear chromatin into cytoplasm. Exposure to these OPs caused a dose-dependent effect on cell viability, and significantly reduced the mean fission rate at a concentration of 1/10(5) v/v. After exposure of resting cysts to 1/10(5) v/v or 1/10(6) v/v concentrations of basudin or cidial, the number of excysted cells was significantly lower than that of the controls. Conversely, exposure to a 1/10(5) v/v fenix concentration did not affect excystment and exposure to 1/10(6) v/v was found to promote excystment. Moreover, exposure to these OPs (1/10(4) v/v or 1/10(5) v/v) interferred with the ability to extrude macronuclear chromatin. The median lethal concentration in 60 min for each OPs tested was at least a hundred times lower than the doses recommended by the manufacturer. Finally, as the inhibition of cholinesterase (ChE) activity is the first target of OPs, the presence of ChE activity was checked in C. inflata. Three ChE activities were found, hydrolyzing the substrates acetyl-beta-methyl thiocholine iodide, propionyl thiocholine iodide and butyryl thiocholine iodide, that appeared to be very low and not inhibited by OP-exposure.