Selected neonicotinoids and associated risk for aquatic organisms.

Neonicotinoids are one of the newest groups of systemic pesticides, effective on a wide range of invertebrate pests. The success of neonicotinoids can be assessed according to the amount used, for example, in the Czech Republic, which now accounts for 1/3 of the insecticide market. The European Union (EU) has a relatively interesting attitude towards neonicotinoids. Three neonicotinoid substances (imidacloprid, clothianidin and thiamethoxam) were severely restricted in 2013. In 2019, imidacloprid and clothianidin were banned, while thiamethoxam and thiacloprid were banned in 2020. In 2022, another substance, sulfoxaflor, was banned. Therefore, only two neonicotinoid substances (acetamiprid and flupyradifurone) are approved for outdoor use in the EU. Neonicotinoids enter aquatic ecosystems in many ways. In European rivers, neonicotinoids usually occur in nanograms per litre. Due to the low toxicity of neonicotinoids to standard test species, they were not expected to significantly impact the aquatic ecosystem until later studies showed that aquatic invertebrates, especially insects, are much more sensitive to neonicotinoids. In addition to the lethal effects, many studies point to sublethal impacts - reduced reproductive capacity, initiation of downstream drift of organisms, reduced ability to eat, or a change in feeding strategies. Neonicotinoids can affect individuals, populations, and entire ecosystems.

Safety of the anthelminthic drugs levamisole, fenbendazole, and ivermectin administered in therapeutic baths for the common carp Cyprinus carpio.

Due to the safe "off label" use of selected antiparasitic drugs in fish, we assessed the effects of a therapeutic bath with levamisole (50 mg/l), fenbendazole (25 mg/l), and ivermectin (0.031 mg/l) on the haematological and biochemical blood indices, oxidative and antioxidant variables, and gill histology of the common carp (Cyprinus carpio). Levamisole did not affect the haematological profile, but significantly increased (P < 0.01) the plasma glucose, lactate, and ammonia concentrations, alkaline phosphatase, and alanine aminotransferase activities, TBARS (muscle, liver), total superoxide dismutase activity (muscle), and catalase activity (liver) and significantly decreased (P < 0.01) plasma aspartate aminotransferase and glutathione reductase activity (gill, liver, muscle). Ivermectin led to a significantly (P < 0.01) greater muscle total superoxide dismutase activity compared to the controls, whereas the haematological and biochemical indices remained unchanged. On the other hand, fenbendazole did not affect the haematological or biochemical indices, and the oxidative stress parameters and antioxidant indices did not differ from the controls. The bath in FBZ can be recommended for safe antiparasitic treatment in carp.

Nitrogen factors for rainbow trout (Oncorhynchus mykiss) and brook trout (Salvelinus fontinalis) fillets.

Measures for consumer protection against food adulteration and misleading labelling are integrated into EU legislation, including methods for detecting misleading practices. Verification of the meat content is available for marine products, but not for salmonid fish due to the lack of standard nitrogen factors. This study aimed to establish nitrogen factors for rainbow trout (Oncorhynchus mykiss) and brook trout (Salvelinus fontinalis). The study analysed 340 fish from Czech fisheries obtained in the summer of 2018-2020. According to the established ISO methods, fillet samples with and without skin were analysed for their nitrogen content (protein), dry matter, ash, and fat. The recommended nitrogen factor for rainbow trout fillets with and without the skin is 3.07 ± 0.12 and 3.06 ± 0.14, respectively, and the nitrogen factor for fat-free rainbow trout fillets with and without the skin is 3.33 ± 0.15 and 3.29 ± 0.15, respectively. The recommended nitrogen factor for brook trout fillets with and without the skin is 3.16 ± 0.10 and 3.12 ± 0.09, respectively, and the nitrogen factor for fat-free brook trout fillets with and without the skin is 3.42 ± 0.13 and 3.36 ± 0.12, respectively. The established nitrogen factors will enable the analysis of the meat content to ensure that consumers purchase correctly described and labelled fish products.

The effect of chronic exposure to chloridazon and its degradation product chloridazon-desphenyl on signal crayfish Pacifastacus leniusculus.

The effects of chloridazon (Ch) and its metabolite chloridazon-desphenyl (Ch-D) at the environmentally relevant concentrations of 0.45 µg/L and 2.7 µg/L on signal crayfish Pacifastacus leniusculus were assessed in a 30-day exposure followed by a 15-day depuration period. Locomotion, biochemical haemolymph profile, oxidative and antioxidant parameters, and histopathology were evaluated. Crayfish exposed to Ch at 0.45 µg/L and 2.7 µg/L showed significantly (p < 0.01) higher CAT activity and GSH level in hepatopancreas and gill compared to controls. The concentration of Ch at 2.7 µg/L was associated with significantly (p < 0.01) higher levels of GLU, LACT, ALT, AST in haemolymph compared to controls. Chloridazon-desphenyl exposure at both tested concentrations caused significantly higher (p < 0.01) GLU, LACT, ALT, AST, NH3, and Ca in haemolymph; lipid peroxidation (TBARS) levels in hepatopancreas; and CAT activity and GSH level in hepatopancreas and gill. Alterations of structure including focal dilatation of tubules, increased number of fibrillar cells, and haemocyte infiltration in the interstitium were observed with 2.7 µg/L Ch and with both Ch-D exposures. Locomotion patterns did not vary significantly among groups. A 15-day recovery period was insufficient to restore normal physiological parameters in exposed groups. Chloridazon and its metabolite Ch-D exerts harmful effects on crayfish.

Acute effects of neonicotinoid insecticides on Mytilus galloprovincialis: A case study with the active compound thiacloprid and the commercial formulation calypso 480 SC.

Pesticides can enter aquatic environments potentially affecting non-target organisms. Unfortunately, the effects of such substances are still poorly understood. This study investigated the effects of the active neonicotinoid substance thiacloprid (TH) and the commercial product Calypso 480 SC (CA) (active compound 40.4% TH) on Mytilus galloprovincialis after short-term exposure to sublethal concentrations. Mussels were tested for seven days to 0, 1, 5 and 10 mg L-1 TH and 0, 10, 50 and 100 mg L-1 CA. For this purpose, several parameters, such as cell viability of haemocytes and digestive cells, biochemical haemolymph features, superoxide dismutase (SOD) and catalase (CAT) enzymatic activity of gills and digestive gland, as well as histology of such tissues were analysed. The sublethal concentrations of both substances lead to abatement or completely stopping the byssal fibres creation. Biochemical analysis of haemolymph showed significant changes (P < 0.01) in electrolytes ions (Cl-, K+, Na+, Ca2+, S-phosphor), lactate dehydrogenase (LDH) enzyme activity and glucose concentration following exposure to both substances. The TH-exposed mussels showed significant imbalance (P < 0.05) in CAT activity in digestive gland and gills. CA caused significant decrease (P < 0.05) in SOD activity in gills and in CAT activity in both tissues. Results of histological analyses showed severe damage in both digestive gland and gills in a time- and concentration-dependent manner. This study provides useful information about the acute toxicity of a neonicotinoid compound and a commercial insecticide on mussels. Nevertheless, considering that neonicotinoids are still widely used and that mussels are very important species for marine environment and human consumption, further researches are needed to better comprehend the potential risk posed by such compounds to aquatic non-target species.

Effects of s-metolachlor on early life stages of marbled crayfish.

The effects of s-metolachlor chronic exposure at concentrations of 1.1 µg/L (maximal real environmental concentration in the Czech Republic), 11 µg/L (environmental relevant concentration) and 110 µg/L on early life stages of marbled crayfish (Procambarus virginalis) was evaluated under laboratory conditions. All s-metolachlor exposures resulted in higher mortality, delay ontogenetic development with accompanied slower growth and excited behaviour (increase of total distance moved and walking speed). Significantly lower superoxide dismutase, catalase, glutathione S-transferase activity and reduced glutathione level was observed at two higher tested concentrations (11 and 110 µg/L) of s-metolachlor compared with the control. S-metolachlor in concentrations 110 µg/L showed alteration of the tubular system of hepatopancreas including focal disintegration of tubular epithelium and notable reduction in epithelial cells number, especially B-cells. In conclusion, potential risk associated with using of s-metolachlor in agriculture, due to effects on non-target aquatic organisms as documented on early life stages of marbled crayfish in this study, should be taken into account.

Chronic dietary toxicity of zinc oxide nanoparticles in common carp (Cyprinus carpio L.): Tissue accumulation and physiological responses.

Concerns regarding the potential toxic effects of zinc oxide nanoparticles (ZnO NPs) on aquatic organisms are growing due to the fact that NPs may be released into aquatic ecosystems. This study aimed to investigate the effects of dietary exposure to ZnO NPs on juvenile common carp (Cyprinus carpio). Fish were fed a spiked diets at doses 50 and 500mg of ZnO NPs per kg of feed for 6 weeks followed by a 2-week recovery period. Fish were sampled every 2 weeks for haematology trends, blood biochemistry measures, histology analyses, and determination of the accumulation of zinc in tissues. At the end of the exposure and post-exposure periods, fish were sampled for an assessment of lipid peroxidation levels. Dietborne ZnO NPs had no effects on haematology, blood biochemistry, and lipid peroxidation levels during the exposure period. After the recovery period, aspartate aminotransferase activity significantly (p < 0.05) increased and alanine transferase activity significantly (p < 0.05) decreased in the higher exposure group. The level of lipid peroxidation significantly (p < 0.05) decreased in liver of treated fish after 2 weeks post-exposure period. A histological examination revealed mild histopathological changes in kidneys during exposure. Our results did not show a significant increase of zinc content at the end of experiment in any of tested organs. However, chronic dietary exposure to ZnO NPs might affect kidney and liver function.

The chronic effects of terbuthylazine-2-hydroxy on early life stages of marbled crayfish (Procambarus fallax f. virginalis).

This study assessed the chronic effects of terbuthylazine-2-hydroxy (T2H), one of the main terbuthylazine degradation products, on early life stages of marbled crayfish (Procambarus fallax f. virginalis) by means of mortality, growth rate, early ontogeny, oxidative stress, antioxidant defence and histopathology. The crayfish were exposed to four concentrations of the tested substance as follows: 0.75µg/l (environmental concentration), 75, 375 and 750µg/l for 62days. Concentrations over 75µg/l caused lower weight compared to the control group. T2H at 750µg/l caused delay in ontogenetic development. Levels of thiobarbituric acid reactive substances and total superoxide dismutase activity were significantly (p<0.01) lower in groups exposed to 375 and 750µg/l T2H. Crayfish in these treatments also showed alteration of tubular system including disintegration of tubular epithelium with complete loss of structure in some places of hepatopancreas and wall thinning up to disintegration of branchial filaments with focal infiltrations of hemocytes. In conclusion, chronic terbuthylazine-2-hydroxy exposure in concentrations up 75µg/l (100 times higher than environmental concentration) affected growth, ontogenetic development, antioxidant system, caused oxidative stress and pathological changes in hepatopancreas of early life stages of marbled crayfish.

Histological changes and antioxidant enzyme activity in signal crayfish (Pacifastacus leniusculus) associated with sub-acute peracetic acid exposure.

Peracetic acid (PAA) is a powerful disinfectant recently adopted as a therapeutic agent in aquaculture. A concentration of 10 mg L(-1) PAA effectively suppresses zoospores of Aphanomyces astaci, the agent of crayfish plague. To aid in establishing safe therapeutic guideline, the effects of PAA on treated crayfish were investigated through assessment of histological changes and oxidative damage. Adult female signal crayfish Pacifastacus leniusculus (n = 135) were exposed to 2 mg L(-1) and 10 mg L(-1) of PAA for 7 days followed by a 7 day recovery period in clean water. Superoxide dismutase activity was significantly lower in gill and hepatopancreas after three days exposure to 10 mg L(1) PAA than in the group treated with 2 mg L(-1) PAA and a control in only clean water. Catalase activity in gill and hepatopancreas remained unaffected by both exposures. Glutathione reductase was significantly decreased in gill of 10 mg L(-1) PAA treated crayfish and increased in group exposed to 2 mg L(-1) compared to control after 7 days exposure. Antioxidant enzyme activity in exposed groups returned to control values after recovery period. Gill, hepatopancreas, and antennal gland showed slight damage in crayfish treated with 2 mg L(-1) of PAA compared to the control group. The extent and frequency of histological alterations were more pronounced in animals exposed to 10 mg L(-1). The gill was the most affected organ, infiltrated by granular hemocytes and displaying malformations of lamella tips and disorganization of epithelial cells. After a 7 day recovery period, the infiltrating cells in affected tissues of the exposed crayfish began to return to normal levels. Results suggested that the given concentrations could be applied to signal crayfish against crayfish plague agent in aquaculture; however, further studies are required for safe use.

Acute toxicity effect of cypermethrin on common carp (Cyprinus carpio).

OBJECTIVES: The present study aimed to assess the acute effect of cypermethrin on common carp (Cyprinus carpio). Evaluation of toxicity effects of cypermethrin on carp was performed on based assessment of hematological profile, antioxidant and oxidative biomarkers and histopathology. METHODS: For testing of toxicity was used the commercial product Cyperkill 25 EC - CY, which contains 250 g.L-1 of cypermethrin. Carp were exposed to two concentrations (1CY - 4.57 µg.L-1 and 2CY - 45.7 µg.L-1) of CY and a control for 96 hours. RESULTS: Significant reduction (p<0.05) in leukocyte count were in fish exposed to concentration 45.7 µg.L-1 (2CY). The both exposure of CY lead to significant differences (p<0.01) in antioxidants biomarkers (superoxide dismutase, catalase, glutathione reductase, reduced glutathione and glutathione S-transferase) and in lipid peroxidation in carp tissues. Many of these changes were observed in liver, gills, muscle, intestine, brain and kidney. Additionally, exposure to CY caused many histological changes in gills, liver and caudal kidney. CONCLUSION: Exposure to CY caused hematological, biochemical and histopathological changes in carp. This study provides and complements other important results for evaluating the toxicity effect of pyrethroids on fish.

The effects of cypermethrin on oxidative stress and antioxidant biomarkers in marbled crayfish (Procambarus fallax f. virginalis).

OBJECTIVES: Pyrethroid insecticides are known to be highly toxic to non-target aquatic organisms. Toxic effects of different types of pyrethroids to fish has been well described but there is a dearth of data on their effects to aquatic crustaceans. The aim of this study was to assess the effects of a commercial product Cyperkill 25 EC (25% of cypermethrin as an active substance) on oxidative stress and antioxidants activity in marbled crayfish. DESIGN: The juvenile crayfish were exposed to two concentrations of Cyperkill 25 EC: 0.02 µg.L-1 (10% 96hLC50) and 0.05 µg.L-1 (30% 96hLC50) for 72 hours. Activity of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione-S-transpherase (GST) and reduced glutathione (GSH), as well as levels of thiobarbituric acid reactive substances (TBARS) were assessed in the whole-body homogenate. RESULTS: Cyperkill 25 EC in both tested concentrations caused significant (p<0.01) decreased in levels of TBARS. Changes in the antioxidants activity were observed in every examined parameters except for the catalase. Cyperkill 25 EC in concentration 0.05 µg.L-1 caused significant increase (p<0.05) of SOD activity and significant decrease of GST (p<0.01) and GSH (p<0.05) activity. The activity of GR was significantly (p<0.01) decreased in both treated groups. CONCLUSIONS: This study suggests that Cyperkill 25 EC causes oxidative stress and also disruption of antioxidant systems in crayfish. Expanding to our previous knowledge, obtained results describe effects of cypermethrin also to non-target aquatic invertebrates and suggest a need of deeper understanding of mechanisms involved in this process.

Effects of prometryne on early life stages of common carp (Cyprinus carpio L.).

Toxicity of prometryne to early life stages of common carp was assessed. On the basis of accumulated mortality in the experimental groups lowest observed-effect concentration (LOEC) was estimated as 1100 µg/l; and no observed-effect concentration (NOEC) was 850 µg/l. Fulton's condition factor was significantly lower than in controls in fish exposed to 4000 µg/l after 7, 14, and 21 days. By day 14, fish exposed to 4000 µg/l prometryne showed significantly lower mass and total length compared to controls. Fish exposed the 1200 and 4000 µg/l showed delay in development, severe hyperaemia in gill, liver, and caudal and cranial kidney. Subchronic prometryne exposure of early-life stages of common carp at concentrations of 1200 and 4000 µg/l affected their survival, growth rate, early ontogeny, and histology.

Effects of acute exposure to deltamethrin and recovery time on common carp (Cyprinus carpio L.).

OBJECTIVES: The aim of this study is to evaluate effects of the insecticide Decis Mega (DM; active substance deltamethrin 50 g.L(-1)) on common carp on the basis of haematological profile, oxidative stress, antioxidant enzymes and histopathology. DESIGN: Fish were exposed two concentrations of DM 6.56 µg.L(-1) (1DM) and 65.6 µg.L(-1) (2DM) for 96 h. Then the remaining fish were transferred into DM-free water for depuration for another period of 96 h. RESULTS: Exposure to 1DM and 2DM proved effect on enzymatic activity of superoxide dismutase, catalase, glutathione reductase and on oxidative damage of cells in gills, liver and kidney (p<0.05, p<0.01). Exposure to 1DM showed differences (p<0.05, p<0.01) in hematocrit and hemoglobin in blood. Histopathological changes were observed after acute exposure to DM as well as to DM-free water in gills, liver and kidney. CONCLUSION: This study concludes that deltamethrin has influence on the haematological parameters, activity of antioxidant enzymes and caused oxidative damage, and histopathological changes in the fish. However, antioxidant balance in the body was restored after placing the fish in clean water for 4 days, however, this time was not sufficient complete regeneration.

Effects of terbuthylazine on early life stages of common carp.

OBJECTIVES: The aim of this study was to assess the toxicity of terbuthylazine in different developmental stages of common carp (Cyprinus carpio) on the basis of mortality, early ontogeny, occurrence of morphological anomalies, growth rate, and Fulton's condition factor during and at the conclusion of the test. DESIGN: The toxicity tests were performed on carp according to OECD 210 methodology. The developmental stages of carp were exposed to terbuthylazine at four concentrations, 2.9 (reported environmental concentration in Czech rivers); 70; 1,400; and 3,500 µg.L(-1) for 35 days and compared to carps in a non-treated control group. RESULTS: Terbuthylazine in concentration 1,400 and 3,000 µg.L(-1) caused significant (p<0.01) decrease of mass, total length and delayed in development of carp. Fish exposed to terbuthylazine showed alteration of tubular system of caudal kidney. On the basis of histopathological changes the values of LOEC=2.9 µg.L(-1) terbuthylazine were estimated. CONCLUSIONS: Chronic terbuthylazine exposure of early-life stages of common carp affected their growth rate, early ontogeny and histology. Some of the changes were observed only at higher exposures, but change founded in caudal kidney was affected in fish exposed to the real environmental concentration tested (i.e., 2.9 µg.L(-1)).

Comparison of the effect of four anaesthetics on haematological profiles, oxidative stress and antioxidant enzymes in barbel (Barbus barbus).

OBJECTIVES: Currently, many questions regarding the effect of anaesthetics to fish remain unresolved. Fish species may differ widely in their response to an anaesthetic, the screening of dosages is often necessary. The aim of this study was to compare the effect of tricaine methane sulphonate (MS 222), clove oil, 2-phenoxyethanol and Propiscin on haematological profiles, oxidative stress biomarkers and antioxidant enzymes in barbel (Barbus barbus). DESIGN: The haematological profiles, oxidative stress biomarkers and antioxidant enzymes of barbel were evaluated immediately after a 10 min anaesthesia (MS 222--100 mg.L(-1), clove oil--33 mg.L(-1), 2-phenoxyethanol--0.4 mg.L(-1), Propiscin--1.0 mg.L(-1)), and 24 h after anaesthesia. RESULTS: The 10 min exposure in the recommended concentrations of tested anaesthetics have no significant effect on haematological profiles, levels of thiobarbituric acid reactive substances, and activity of glutathione reductase of barbel. The activity of superoxide dismutase (SOD) was significantly decreased (p<0.01) in the muscle in all experimental groups. The activity of SOD showed a significant decrease (p<0.01) in the liver 24 h after all anaesthetics; however in the gill the activity of SOD was significantly increased (p<0.01) in Propiscin (10 min). The activity of catalase (CAT) was significantly increased (p<0.05) in the muscle 24 h after all anaesthetics. CONCLUSIONS: The observed effects on barbel antioxidant systems may be a defence against oxidative damage. The results of this study suggest that the antioxidant systems of barbel are altered by Propiscin anaesthesia, but are slightly affected by MS 222, clove oil, and 2-phenoxyethanol anaesthesia.

Evaluation of the toxic effect of peracetic acid on grass carp (Ctenopharyngodon idella) juveniles.

OBJECTIVES: The aim of present study was to evaluate the effect of peracetic acid (PAA) on haematotological and biochemical indices, antioxidant status, micronucleus induction and histopathological alterations of liver and gill in grass carp. METHODS: Grass carp (Ctenopharyngodon idella) juveniles were exposed to therapeutic concentrations (1, and 3 mg x l(-1)) of PAA for a period of 10 days. Selected haematotological indices--the erythrocyte count (RBC), haematocrit (PCV), haemoglobin (Hb), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC) and leukocyte count (WBC), and biochemical indices--glucose (Glu), total protein (TP), aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatine kinase (CK), and lactate dehydrogenase (LDH) were evaluated in plasma. Activity of superoxide dismutase (SOD), catalase (CAT) and glutathione reductase (GR), as well as levels of thiobarbituric acid reactive substances (TBARS) were assessed in gill and liver. Micronucleus frequency in peripheral erythrocytes was counted in control and experimental fish. Histological examinations of gill and liver were performed. RESULTS: No significant differences were found in haematological parameters measured. Statistically significant (p < 0.05) alterations in the activities of AST, CK and LDH were found in treated fish compared to control groups. Fish exposed to 1 mg x l(-1) of PAA showed significantly lower (p < 0.05) SOD activity in liver and gill while catalase activity indicated a significant decrease (p < 0.05) only in gill tissue. Other significant changes were observed in GR activity in gill in both PAA exposed groups, while GR activity in liver remained unchanged. There was no significant difference in the count of micronuclei between control and exposed fish. Haemorrhage, fusion of primary lamellae, degeneration of secondary lamellae, some clubbing on primary and secondary lamellae tips, and lifting of epithelial cells were found in gill tissues in both control and treated fish. CONCLUSION: The results show that PAA could induce alterations in biochemical parameters in blood plasma, antioxidant enzymes response and histopathological changes in gill; however, it seems that these changes are reversible. Subsequently, lower concentration (1 mg x l(-1)) is useable as a treatment concentration for grass carp.

The effect of subchronic metribuzin exposure to signal crayfish (Pacifastacus leniusculus Dana 1852).

OBJECTIVES: The aim of the study was to investigate effects of the triazine herbicide metribuzin on signal crayfish Pacifastacus leniusculus Dana by determining oxidative stress (thiobarbituric acid reactive substances) and antioxidant indices (total superoxide dismutase, catalase, glutathione reductase) in hepatopancreas, muscle, and gill as well as assessing their histopathology. DESIGN: Crayfish were exposed to metribuzin concentrations of 0.52 µg x l(-1) (realistic environmental concentration) and 3.06 mg x l(-1) (10% 96hLC50) for 10 and 30 days followed by a 30-day depuration period without exposure to metribuzin. RESULTS: In the thiobarbituric acid reactive substances, superoxide dismutase, and catalase were observed differences in all examined tissues compared to the control group. Differences from control were observed in glutathione reductase activity in hepatopancreas after 10 days for both exposure concentrations and after 30 days at 3.06 mg x l(-1). Histological examination revealed extensive focal autolytic disintegration of tubular epithelium in hepatopancreas of crayfish exposed to metribuzin for 30 days. CONCLUSIONS: Chronic exposure of metribuzin resulted in oxidative damage to cell lipids, in changes of antioxidant activity in crayfish tissue, and pathological changes in hepatopancreas. The results suggest that selected oxidative stress biomarkers, antioxidant enzymes, and pathologies of hepatopancreas may have potential as biomarkers for monitoring residual triazine herbicides in the aquatic environment.

Resistance of common carp (Cyprinus carpio L.) to oxidative stress after chloramine-T treatment is increased by microalgae carotenoid-rich diet.

OBJECTIVES: In fish aquaculture, disinfectants are used against bacterial and protozoal infections. These compounds cause oxidative stress that may stimulate the generation of reactive oxygen species, and subsequently the alteration in antioxidant systems of exposed organisms. Antioxidants like carotenoids present in microalgae increase carp resistance to oxidative stress after chemical treatment. DESIGN: The aim of these experiments was to prove increased resistance of common carp (Cyprinus carpio L.) juveniles fed on experimental diets with microalgae biomass supplement (Algadiets) to oxidative stress caused by a disinfectant chloramine-T. In indoor experiments fish were fed on laboratory-prepared extruded diets containing supplement of Chlorella spp. (cf. C. vulgaris Beijerinck) biomass which contains antioxidants (carotenoids) like lutein. The young-of-the-year-old fish were acclimatized and fed on basal diet (control group) and the on diets containing 1, 2, 5 and 10% (w/w) of spray-dried Chlorella biomass (Algadiet 1, 2, 5 and 10) for 14 days followed by 6 weeks. Consequently, fish were treated daily with chloramine-T (Chl-T) at concentration of 10 mg x l(-1) for 1 h in three consecutive days. After this treatment, the indices of oxidative stress and antioxidant enzyme activity were assayed in fish gill, muscle and hepatopancreas. RESULTS: The fish fed on different Algadiets had increased antioxidant enzyme activities of glutathione peroxidase, glutathione reductase and catalase in flesh after the exposure to Chl-T. Higher activities of superoxide dismutase, glutathione peroxidase and glutathione reductase were also observed in the hepatopancreas in all tested concentrations compared to the control group fed on the basal diet. The increased production and activity of antioxidant enzymes confirmed improved protection ability of fish tissues against oxidative damage when microalgae biomass was supplemented to the fish diet which was more pronounced by higher microalgae supplement in Algadiet 5 and 10 where the content of carotenoids was 105 mg and 214 mg per kilogram of fed, respectively. CONCLUSION: The results show the positive effect of carotenoids from microalgae biomass to maintain the antioxidant capacity which increases resistance of fish to oxidative stress.

Effect of prometryne on early life stages of marbled crayfish (Procambarus fallax f. virginalis).

OBJECTIVES: The aim of this study is to assess the toxicity of prometryne in early life stages of marbled crayfish (Procambarus fallax f. virginalis) on the basis of mortality, early ontogeny, growth rate, and histopathology during and at the end of the test. DESIGN: The early life stages of marbled crayfish were exposed to prometryne at four concentrations, 0.51, (reported concentration in Czech rivers), 144, 1440, and 4320 µg x l(-1) for 53 days and compared to crayfish in a non-treated control group. RESU LTS: Prometryne in concentration 144, 1444 and 4320 µg x l(-1) caused decrease of weight and specific growth rates of crayfish. Crayfish exposed the highest concentration 4320 µg x l(-1) showed delay in ontogeny development. All crayfish groups exposed to prometryne showed histopathological changes in gill. On the basis of histopathological changes the values of LOEC = 0.51 µg x l(-1) and NOEC = for 0.10 µg x l(-1) of prometryne for marbled crayfish juveniles was estimated. CONCLUSIONS: Chronic exposure of prometryne on early life stages of crayfish has affected their mortality, growth rate, and histology. Some of the changes were observed only at higher exposures (144, 1444 and 4320 µg x l(-1)), but histopathological changes in gills were observed also in crayfish exposed to the real environmental concentration in Czech rivers (i.e. 0.51 µg x l(-1)), which is about 9 times lower than maximal concentration (4.40 µg x l(-1)) reported in surface waters of Greece. Concentrations of prometryne in World rivers have been reported to generally vary in the range of 0.1-4.40 µg x l(-1).

Efficacy of different anaesthetics for pikeperch (Sander lucioperca L.) in relation to water temperature.

OBJECTIVES: The objectives of the study were to compare the different doses of clove oil, Propiscin, and tricaine methane sulphonate (MS 222) in relation to water temperature in pikeperch aquaculture. DESIGN: For assessment of this experiment 168 fish (10.77 ± 0.59 cm total body length and 7.88 ± 1.74 g body weight) were used. Three different anaesthetic treatments (Propiscin, clove oil and MS 222) were used. Three doses of each anaesthetic treatment (Propiscin: 0.5; 1; 1.5 ml x L(-1), clove oil: 15; 30; 60 mg x L(-1), MS 222: 50; 100; 150 mg x L(-1)) were compared at three different temperatures 9.5; 15.5 and 23 degrees C. RESULTS: In comparison of these doses of anaesthetic in different temperature, the significantly shortest time to attain phase A7 (total complete anaesthesia) was observed for Propiscin (1.5 ml L(-1)) 0:31 ± 0:04 min (23 degrees C) to 0:33 ± 0:25 min (9.5 degrees C) compared to MS 222 (150 mg x L(-1)) 1:04 ± 0:21 min (23 degrees C) to 1:54 ± 0:32 min (9.5 degrees C) and clove oil (60 mg x L(-1)) 1:05 ± 0:17 min (23 degrees C) to 3:05 ± 0:31 min (9.5 degrees C). On the other hand, the longest time of anaesthesia recovery was attained using Propiscin (1.5 ml x L(-1)) 10:35 ± 1:40 min (23 degrees C) to 32:30 ± 1:10 min (9.5 degrees C) compared to clove oil (60 mg x L(-1)) 2:39 ± 0:50 min (23 degrees C) to 9:36 ± 2:34 min (60 mg x L(-1), 9.5 degrees C) and MS 222 (150 mg x L(-1)) 2:26 ± 1:27 min (23 degrees C) to 4:59 ± 0:39 min (9.5 degrees C). CONCLUSIONS: The results from this study showed that the optimal and sufficient doses in all tested temperatures for pikeperch are 30 mg x L(-1) of clove oil, 100 mg x L(-1) of MS 222 and 0.5 ml x L(-1) of Propiscin.