Foodborne fluoxetine impacts the immune response in rainbow trout (Oncorhynchus mykkis).

The aim of this study was to reveal the effects of foodborne fluoxetine on morphological and condition profile, hematological profile, biochemical and oxidative stress indices on juvenile rainbow trout. The study was performed according to OECD Guideline No. 215. Fluoxetine was incorporated into Biomar 921 pellets at a dose of 0.047 mg/kg (environmental concentration), 0.577 mg/kg and 6.7 mg/kg. There was statistically significant change in hematological profile, including an increasing trend in neutrophil/lymphocyte ratio and a decreasing trend in the number of lymphocytes. Measurements of oxidative stress indicated decreased activity of the detoxifying enzyme glutathione-S-transferase in the liver and kidney. However, the measurement of GR, GPx, CAT, SOD activity, and TBARS showed no changes. Histopathological examination revealed damage to the proximal tubules of caudal kidney in exposed groups. This study confirms that fluoxetine has a significant effect on immune response.

The effect of foodborne sertraline on rainbow trout (Oncorhynchus mykiss).

The worldwide consumption of antidepressants is raising as well as their concentrations in the aquatic environment. This increases the risk of food chain contamination and bioaccumulation in aquatic biota. The aim of this study was to describe a potential risk of sertraline as a pollutant from water environment, wherein rainbow trout (Oncorhynchus mykiss) has been chosen as the test organism, because predatory fish are on the top of the food chain in the aquatic environment. The effects of foodborne sertraline were tested on rainbow trout during a 28-day toxicity test according to OECD 215 method. Sertraline was incorporated in commercial feed at a dose of 4.4 µg/kg (environmental concentration), 42 µg/kg and 400 µg/kg. The results confirmed that sertraline has a significant effect on fish behaviour, resulting in suppression of the escape reflex and increased resistance to stress. Moreover, increased Fultons condition factor was found in fish fed with the highest concentration of sertraline. Haematological analysis revealed a statistically significant increase in the number of neutrophilic bands and neutrophil/lymphocyte ratio, and decreased number of lymphocytes. The results of biochemical examination showed a statistically significant decrease in ammonia and lactate concentrations and histological examination revealed changes in gills and caudal kidney. Although sertraline reduces stress in fish, the decline in nonspecific immunity is a risk to fish population stability.

The effect of the antidepressant venlafaxine on gene expression of biotransformation enzymes in zebrafish (Danio rerio) embryos.

The effect of venlafaxine, a pharmaceutical commonly found in aquatic environment, was analyzed on non-target organism, Danio rerio (Hamilton, 1822). D. rerio embryos were treated by two different concentrations of venlafaxine: either concentration relevant in aquatic environment (0.3 µg/L) or concentration that was two orders of magnitude higher (30 µg/L) for the evaluation of dose-dependent effect. Time-dependent effect was rated at 24, 96, and 144 h post-fertilization (hpf). For gene expression, genes representing one of the phases of xenobiotic biotransformation (0 to III) were selected. The results of this study showed that the effect of venlafaxine on the zebrafish embryos is the most evident at hatching (96 hpf). At this time, the results showed a downregulation of gene expression in each phase of biotransformation and in both tested concentrations. In contrast, an upregulation of most of the genes was observed 144 hpf for both tested venlafaxine concentrations. The study shows that venlafaxine can affect the gene expression of biotransformation enzymes in D. rerio embryos even in the environmentally relevant concentration and thus disrupt the process of biotransformation. Moreover, the pxr regulation of genes seems to be disrupted after venlafaxine exposure in dose- and time-dependent manner.

Effects of antidepressants with different modes of action on early life stages of fish and amphibians.

Drugs are excreted from the human body as both original substances and as metabolites and enter aquatic environment through waste water. The aim of this study was to widen the current knowledge considering the effects of waterborne antidepressants with different modes of action-amitriptyline, venlafaxine, sertraline-on embryos of non-target aquatic biota-fish (represented by Danio rerio) and amphibians (represented by Xenopus tropicalis). The tested concentrations were 0.3; 3; 30; 300 and 3000 µg/L in case of amitriptyline and venlafaxine and 0.1; 1; 10; 100 and 1000 µg/L for sertraline. Test on zebrafish embryos was carried out until 144 h post fertilization, while test on Xenopus embryos was terminated after 48 h. Lethal and sublethal effects as well as swimming alterations were observed at higher tested concentrations that are not present in the environment. In contrast, mRNA expression of genes related to heart, eye, brain and bone development (nkx2.5, otx 2, bmp4 and pax 6) seems to be impacted also at environmentally relevant concentrations. In a wider context, this study reveals several indications on the ability of antidepressants to affect non target animals occupying environments which may be contaminated by such compounds.

The effects of a therapeutic formalin bath on selected immunological and oxidative stress parameters in common carp (Cyprinus carpio).

Formalin is commonly used as a component of antiparasitic baths in fisheries. In this study the impact of this bath on the immune profile and oxidative stress parameters was evaluated. A formalin bath was prepared in the concentration of 185.3 mg L-1 (0.17 mL L-1) at a temperature of 20 °C. A total of 96 common carp Cyprinus carpio (Linnaeus, 1758) individuals were immersed in this bath for 60 min. The effects were monitored immediately, and then after 24, 48 h and 10 days following the treatment. The study revealed the most effects 10 days after the treatment, when we observed the decrease of lysozyme in skin mucus, the decrease of anti-inflammatory cytokine transforming growth factor beta in gill tissue and increase of interleukin 10 in cranial kidney tissue. The pro-inflammatory cytokine interleukin 1b showed an increase in gill tissue immediately after the bath and the increase in glutathione peroxidase in gill tissue was also observed 24 h and 10 days after bath treatment. The other investigated parameters did not show any significant changes. In conclusion, even though the formalin bath elevated some parameters as mentioned above, formalin used in the bath is probably safe as an antiparasitic treatment of fish.

Oxidative stress induced by fluoroquinolone enrofloxacin in zebrafish (Danio rerio) can be ameliorated after a prolonged exposure.

The purpose of our experiment was to evaluate the effect of enrofloxacin on biotransformation, oxidative stress and mRNA expression of related genes in fish as a non-target organisms. Zebrafish (Danio rerio) juveniles were treated with enrofloxacin at concentrations of 5, 10 and 500 µg/L for 14 days. A three-day-long test caused changes of catalytic activity of glutathione peroxidase and glutathione-S-transferase. Moreover, lipid peroxidation was observed at the highest concentration. No significant changes either in catalytic activity of antioxidant enzymes or elevated lipid peroxidation were observed from sampling day 7 on. mRNA expression of genes encoding antioxidant enzymes was also not affected by enrofloxacin after a 14-day exposure. This suggests the ability of D. rerio juveniles to adapt to enrofloxacin in a short time period. Moreover, enrofloxacin was not shown to affect collagen, cathepsin K, optic atrophy 1 and pyruvate kinase L/R mRNA expression in this study.

mRNA expression of antioxidant and biotransformation enzymes in zebrafish (Danio rerio) embryos after exposure to the tricyclic antidepressant amitriptyline.

Now-a-days, the occurrence of antidepressant residues in surface waters has become a major concern. Amitriptyline (AMI) has been described to treat depression and other disorders for decades. However, little is known about its effect on non-target organisms. The aim of this study was to assess the potential impact of AMI on the mRNA expression of antioxidant and detoxification enzymes during the early embryonic development of zebrafish (Danio rerio). Fertilized D. rerio embryos were exposed to AMI at concentrations of 300 ng/L and 30 µg/L and sampled 24, 48, 96, and 144 h post fertilization (hpf) to assess the mRNA expressions of cytochrome P450 1A1, glutathione-S-transferase, glutathione peroxidase, superoxide dismutase, and catalase. The time courses of the mRNA expressions of antioxidant and detoxification enzymes revealed characteristic changes during embryonic development causing generally transient changes post hatching; however, AMI did not cause any significant impact except in the case of CAT after 144 h, which was significantly upregulated by the AMI concentration of 30 µg/L. The results suggest that the antidepressant AMI causes only moderate to minor impacts on antioxidant and detoxification enzymes during early embryonic development of the non-target organism D. rerio and that CAT is the only biomarker affected by AMI.

The effect of silver nanoparticles and silver ions on zebrafish embryos (Danio rerio).

OBJECTIVES: The aim of this study was to establish and evaluate the mortality rate, hatching rate and observe the presence of sublethal changes in zebrafish embryos after exposure to silver ions and nanoparticles. METHODS: Tests were conducted on newly fertilized zebrafish embryos, according to the modified OECD guideline 236, using a semistatic method and 96 hour incubation time. Silver nitrate and two different silver nanoparticles, stabilized with 0.01% solution of maltose and gelatine in the first case, and stabilized with polyvinylpyrrolidone, in the latter, were tested. RESULTS: Significant differences in toxicity of tested substances were recorded. The value of 96hLC50 for silver nitrate was 58.44 µg/L. The value of 96hLC50, calculated for silver nanoparticles stabilized with 0.01% solution of maltose and gelatine, was nearly 100 times higher, 4.31 mg/L. The value 96hLC50 for silver nanoparticles stabilized with polyvinylpyrrolidone exceeded 100mg/L, occurrence of sublethal effects caused by silver nanoparticles stabilized with polyvinylpyrrolidone was insignificant in most of the exposition groups, but only in this substance caused decreased hatching rate. CONCLUSION: Properties of different silver nanoparticles play an important role in levels of their toxicity and predominant mechanisms of action. In general, silver nanoparticles are less toxic for Danio rerio embryos than silver ions.

Effects of waterborne antidepressants on non-target animals living in the aquatic environment: A review.

After application, antidepressants, like other pharmaceuticals, are excreted from human body in their native form or as metabolites and enter the aquatic environment via different pathways. As concentrations of antidepressant residues in water continue to increase, their effects on non-target animals are being discussed. The aim of this study is to summarize current knowledge about the effects of wateborne antidepressants on non-target animals living in surface waters - invertebrates, fish and amphibians. Selective serotonin reuptake inhibitors such as fluoxetine, sertraline, and citalopram have been found to effect behavior, reproduction, and development in both invertebrates and vertebrates. Venlafaxine, belonging to the group of selective serotonin-noradrenalin reuptake inhibitors, not only affected behavior but also showed the potential to reduce survival in fish. Tricylic antidepressants are known to have various side-effects when consumed by humans. Moreover, in fish, exposure resulted in a significant increase in mortality, developmental retardation, morphological anomalies, and pathological changes in brain, heart, and cranial and caudal kidney. In addition, changes in antioxidant enzyme activity as well as increased lipid peroxidation were observed, even at the lowest tested concentrations. According to current knowledge, antidepressants occuring in surface water are able to affect the behavior, reproduction, development, and survival of aquatic invertebrates and vertebrates.

Effects of glyphosate on early life stages: comparison between Cyprinus carpio and Danio rerio.

Glyphosate (N-(phosphonomethyl)glycine) is an active substance of many herbicides. According to literature studies, glyphosate residues and their metabolites have been commonly detected in surface waters and toxicological reports confirmed negative effects on living organisms. In this study, the acute embryo toxicity of glyphosate into two different fish species-common carp (Cyprinus carpio) and zebrafish (Danio rerio)-was investigated. Lethal endpoints, development disorder, and, in addition, other sublethal endpoints such as hatching rate, formation of somites, and development of eyes, spontaneous movement, heartbeat/blood circulation, pigmentation, and edema were recorded to indicate the mode of action of the toxic compound. Hatching retardation (p < 0.05) was observed in experimental groups of common carp exposed to glyphosate with significant statistical difference especially at the highest concentration after 72, 96, and 120 hpf. The significantly highest cumulative mortality at concentration of 50 mg/l was observed. In contrast, hatching stimulation was observed in embryos of zebrafish exposed to the highest concentration of glyphosate. The significantly highest cumulative mortality for zebrafish was observed only at concentration of 50 mg/l. Based on our results, early life stages of common carp are more sensitive in comparison to zebrafish to the toxic action of glyphosate.

Toxicity of naproxen sodium and its mixture with tramadol hydrochloride on fish early life stages.

Pharmaceuticals occur in water bodies as a consequence of their incomplete removal during waste water treatment processes. The occurence of pharmaceuticals in surface waters as well as their possible impact on aquatic vertebrates have received considerable attention in recent years. However, there is still a lack of informations on the chronic effects of widely used drugs as well as their possible mixture toxicity on non-target aquatic vertebrates as well as their possible mixture toxicity. The aim of this study was to assess the effects of naproxen sodium on early life stages of fish and evaluate its mixture toxicity with tramadol hydrochloride, which was assessed in our earlier study as a single substance. Two embryo-larval toxicity tests with common carp (Cyprinus carpio) were performed according to the OECD guideline 210 (Fish, Early-life Stage Toxicity Test) in order to assess the subchronic toxicity of naproxen sodium and tramadol hydrochlorid-naproxen sodium mixture at the concentrations of 10; 50; 100 and 200 µg/L. These experiments were conducted for 32 days. The subchronic exposure to naproxen sodium and naproxen sodium and tramadol hydrochloride mixture had a strong effect on the early life stages of common carp. Hatching, developmental rate, morphology, histopathology and, in the case of the naproxen sodium and tramadol hydrochloride mixture, mortality were influenced. The bioindicators of oxidative stress were also influenced. The LOEC was determined at 10 µg/L for both naproxen sodium and naproxen sodium and tramadol hydrochloride mixture.

Effects of selected tricyclic antidepressants on early-life stages of common carp (Cyprinus carpio).

Despite the fact that pharmaceuticals occur in surface water in low concentrations, they can still have adverse effect on aquatic biota, because these substances are designed to have a specific mode of action even at low concentrations. To our knowledge, only little is known about the long-term effect of tricyclic antidepressant residues on non-target animals, especially fish. Hence, the aim of this study was to assess the effect of subchronic exposure to selected tricyclic antidepressants - namely, amitriptyline, nortriptyline, and clomipramine and their mixture - on early-life stages of common carp (Cyprinus carpio). Two embryo-larval toxicity tests with common carp were performed, each taking 30 days. For the experiment, three tricyclic antidepressants as well as their mixtures were tested at three different concentrations - 10, 100 and 500 µg/L. Hatching and mortality were recorded twice a day. In addition, samples were taken at regular intervals in order to record developmental stage, morphometric and condition characteristics and morphological anomalies. At the end of the test, additional samples were taken for histopathological examination and also for the determination of antioxidant and biotransformation enzyme activity and lipid peroxidation and protein carbonylation. Long-term exposure resulted in a significant increase in mortality, developmental retardation, morphological anomalies, and pathological changes in brain, heart, and cranial and caudal kidney. In addition, changes in antioxidant enzyme activity as well as increased lipid peroxidation were observed, even at the lowest tested concentrations. Hence, environmentally relevant concentrations of TCAs have the potential to cause harmful effects on early-life stages of fish.

Effect of T-2 toxin-contaminated diet on common carp (Cyprinus carpio L.).

The T-2 toxin, a fungal metabolite produced by Fusarium molds, occurs in a range of agriculture products. Reduced availability of fish meal has led to increasing use of cereals as a source of protein in commercial aquaculture feeds, which has increased the potential for mycotoxin contamination. The purpose of this study was to investigate toxicity of T-2 toxin intake in common carp (Cyprinus carpio L.) using haematological, biochemical and immunological parameters and oxidative stress indices. In a four-week feeding trial, fish were fed a commercial diet with 5.3 mg/kg T-2 toxin added. Ingestion of contaminated diet did not lead to mortality of fish, probably due to lower feed intake. On the other hand, it significantly affected haematological variables such as haematocrit, haemoglobin, red blood cell counts leading to anemia and white blood cell counts leading to leukopenia due to lymphopenia. Plasma glucose concentration and alanine amino transferase activity showed a significant increase while triglycerides concentration decreased. Activity of ceruloplasmin was significantly decreased in plasma. Further, liver glutathione S-transferase activity was significantly increased and catalase activity decreased, in parallel with a significant increase in caudal kidney catalase activity and a decrease in glutathione peroxidase activity. Finally, lipid peroxidation (detected as malondialdehyde) was significantly increased in the liver and caudal kidney. Changes in non-specific immune response and cytokine levels in head kidney indicated immune system sensitivity to T-2 toxin. Overall, the results demonstrate that this feed-borne mycotoxin is able to induce anaemia and oxidative stress and cause changes in the immune response of common carp.

The Effects of Salicylic Acid on Juvenile Zebrafish Danio rerio Under Flow-Through Conditions.

The aquatic environment is becoming increasingly contaminated with pharmaceuticals. Salicylic acid (SA), which can be used individually or appear as a degradation product of the widely used acetylsalicylic acid was chosen for testing. Juvenile zebrafish Danio rerio were subjected to OECD test No. 215 (fish, juvenile growth test) with salicylic acid concentrations of 0.004; 0.04; 0.4; 4 and 40 mg/L. Specific growth rate (SGR), histological changes, and parameters of oxidative stress were evaluated. SA had no effects on histological changes, SGR, glutathione reductase, and lipid peroxidation. Increased catalytic activity of GPx was found at 0.04 mg/L compared to control, increased catalytic activity of catalase was found at 0.04 and 4 mg/L compared to control, and increased catalytic activity of glutathione-S-transferase was found at 0.004 and 0.04 mg/L compared to control (P < 0.05). Juvenile zebrafish turned out to be relatively insensitive to both environmentally relevant (0.004 mg/L) and higher concentrations of salicylic acid.

The effect of tramadol hydrochloride on early life stages of fish.

The aim of this study was to perform the fish embryo acute toxicity test (FET) on zebrafish (Danio rerio) and the early-life stage toxicity test on common carp (Cyprinus carpio) with tramadol hydrochloride. The FET was performed using the method inspired by the OECD guideline 236. Newly fertilized zebrafish eggs were exposed to tramadol hydrochloride at concentrations of 10; 50; 100 and 200µg/l for a period of 144h. An embryo-larval toxicity test on C. carpio was performed according to OECD guideline 210 also with tramadol hydrochloride at concentrations 10; 50; 100 and 200µg/l for a period of 32 days. Hatching was significantly influenced in both acute and subchronic toxicity assays. Subchronic exposure also influenced early ontogeny, both morphometric and condition characteristics and caused changes in antioxidant enzyme activity. The LOEC value was found to be 10µg/l tramadol hydrochloride.

Effects of therapeutic formalin bath on selected indices in fish.

OBJECTIVES: The aim of this study is to assess the effects of a treatment bath in a formalin solution on fish, focusing on haematological, biochemical and histopathological profiles. METHODS: A total of 96 common carps (Cyprinus carpio) were randomly assigned to eight groups. Four experimental groups were placed in the test solution for 60 minutes. The concentration of the formalin bath was 0.17 ml/l (38% formaldehyde), with the water temperature of 20°C. The effects of the bath were monitored immediately (E0) and 24 hours (E24h), 48 hours (E48h) and 10 days (E10d) after the bath. There was a control group (C0, C24h, C48h, C10d) for each of the experimental groups. RESULTS: Histopathological indices were strongly affected. Extensive changes were found on gill and skin immediately and also after 24 h, 48 h and 10 d of the formalin treatment bath. A plurality of mucinous elements was observed on the skin. Moreover, structural devastation of lamellas and numerous mucinous cells were observed on the gill. Changes were also found in haematological and biochemical indices. CONCLUSIONS: It can be concluded that after applying the treatment bath in the formalin solution, the monitored profiles were affected immediately after the bath and also after 10 days (histopathological changes of skin and gill). Such significant impact of therapeutic formalin bath should be considered in treatment of fish.

Effect of salicylic acid on early life stages of common carp (Cyprinus carpio).

Environmental concentrations of pharmaceutical residues are often low; nevertheless, they are designed to have biological effects at low doses. The aim of this study was to assess the effects of salicylic acid on the growth and development of common carp (Cyprinus carpio) early life stages with respect to antioxidant defence enzymes. An embryo-larval toxicity test lasting 34 days was performed according to OECD guidelines 210 (Fish, Early-life Stage Toxicity Test). The tested concentrations were 0.004, 0.04, 0.4, 4 and 20mg/l of salicylic acid. Hatching, early ontogeny, and both morphometric and condition characteristics were significantly influenced by subchronic exposure to salicylic acid. Also, changes in antioxidant enzyme activity and an increase in lipid peroxidation were observed. The LOEC value was found to be 0.004 mg/l salicylic acid. The results of our study confirm the suggestion that subchronic exposure to salicylic acid at environmental concentrations can have significant effects on aquatic vertebrates.

The effect of the fluoroquinolone norfloxacin on somatic indices and oxidative stress parameters in early stages of common carp (Cyprinus carpio L.).

OBJECTIVES: The aim of this study was to assess the effect of the subchronic exposure of early stages of common carp (Cyprinus carpio L.) to norfloxacin using morphometric data and oxidative stress parameters. METHODS: A subchronic toxicity test was performed on fertilized embryos of common carp according to the OECD Guidelines No. 210. Embryos were exposed to norfloxacin concentrations of 0.0001 (environmental), 0.1, 1.0, 5.0, and 10.0 mg.L(-1) for 34 days. RESULTS: At the end of the test (day 34), significant (p<0.05) stimulation of development was observed in all experimental groups, in contrast to the control. Significantly greater (p<0.01) total body length was also observed in the group exposed to 10.0 mg.L(-1) of norfloxacin compared to the control. A significant increase in the activity of glutathione S-transferase in all carp exposed to norfloxacin concentrations of 0.1 and 1.0 mg.L(-1) (p<0.01), and 5.0 mg.L(-1) (p<0.05) compared to control group was revealed. The activity of glutathione peroxidase was significantly lower (p<0.01) in experimental carp exposed to a norfloxacin concentration of 10.0 mg.L(-1). In experimental carp exposed to a norfloxacin concentration of 0.0001 mg.L(-1), a significant increase (p<0.05) in glutathione reductase activity was found. Significant (p<0.01) decreases in the content of thiobarbituric acid reactive substances in the groups exposed to norfloxacin concentrations of 1.0, 5.0, and 10.0 mg.L(-1) were revealed. CONCLUSION: From the results, we can conclude that norfloxacin has a negative impact on selected biochemical processes related to the production of reactive oxygen species in early-life stages of common carp.