Environmental Toxicological Study And Risk Assessment Of Tetracycline And Cu In Teleost Embryos

With the rapid development of intensive farming and aquaculture activities, pharmaceuticals and metals are often used as feed additives to promote efficient production. Tetracycline antibiotics and copper are widely used in aquaculture activities. The wide usage and poor removal of pharmaceuticals from the wastewater lead to the high concentration of pharmaceuticals in the water environment. In recent years, sewage water, surface water, groundwater, and even drinking water were detected in the presence of antibiotics. High concentrations of antibiotics can induce toxicity to aquatic organisms and threaten the safety of the aquatic ecosystem. Long-term exposure to pharmaceuticals, such as antibiotics can induce drug resistant in organisms, posing a threat to human health and food safety. There is a need to understand the environmental and human health effects of these emerging contaminants in aquatic environment.In this study, the impacts of tetracycline and copper in aquatic organisms were investigated in the developing zebrafish embryos. Furthermore, the joint toxicity of tetracycline and copper ions were studied.The toxicity of tetracycline in zebrafish embryos were conducted via acute exposure scheme. Acute exposure results showed that the 96 h exposure of tetracycline has no obvious effect on the survival of developing zebrafish embryo, but significantly inhibits the growth and development of zebrafish embryos. The exposed larvae displayed developmental delay phenotypes, including hatching delay, shorter body length, increased yolk sac area and uninflated swim bladder. The 96 h-EC10 of tetracycline to zebrafish embryos is 3.16 μg/L, which is below the environment concentration of tetracycline (please fill in the common range of environmental concentrations here). At molecular level, we further investigated the toxicity mechanisms by examing the gene expression profiles of SOD and CAT, together with the detection of ROS in exposed zebrafish larvae. These results showed that tetracycline exposure resulted in oxidative damage to zebrafish embryos. AO staining results showed that the exposure to tetracycline causes apoptosis in the heart and tail of zebrafish larvae. Gene expression study showed that exposure to tetracycline induces ROS which causes apoptosis in the zebrafish larvae, and the results also indicate that caspase-dependent apoptotic pathway may greatly contribute to tetracycline-induced apoptosis in the early-life stages of zebrafish. In addition, we investigated the effects of tetracycline on marker genes related to resistance mechanisms and gene regulating drug biotransformation. The results suggest that tetracycline could induce resistance to pharmaceuticals and Cytochrome P450s that are involved in the biotransformation of tetracycline in zebrafish larvae.Toxicology effects and underlying mechanisms of Cu in zebrafish embryos were studied using similar approaches. Exposure to copper ions induced evident lethal and teratogenic effects in zebrafish embryos, the LC10 and EC10 of 96 h exposure is 2.5μg/L and 0.1 μg/L respectively. Exposure to Cu led to the development delay of embryos, hatching inhibition, yolk sac absorption, shortened body length, and abnormal development of head and swimming bladder. After that, ICP-MS, AO staining and RT-PCR methods were used to determine the toxic mechanisms of Cu. In vivo uptake rate of Cu decreased with increased concentration and the enrichment in head area is more efficient than that in the trunk area. RT-PCR data revealed that the exposure to low concentration of Cu disturbed neuro development and heart development on zebrafish embryos, but did not affect the MT gene expression. AO staining data also showed that exposure to Cu caused apoptosis in the heart area and there was a small increased amount of apoptotic cells in the tail of exposed zebrafish larvae.Lastly, the joint toxicity of tetracycline and copper ions were studied in developing zebrafish embryos. The 96 h exposure results showed tetracycline can reduce the toxicity of Cu in zebrafish embryos. ICP-MS data showed that tetracycline can significantly reduce the in vivo total Cu content in zebrafish larvae. It is proposed that a complex is formed during the interaction on joint exposure, and the formation of the complex reduced the toxicity of Cu in exposed embryos. These results suggest that the joint interaction between tetracycline and copper is antagonism.In summary, the study showed that tetracycline can obviously inhibit the growth and development of zebrafish embryo via oxidative damage and apoptosis. And Cu induced developmental toxicity and cardiac toxicity to zebrafish embryos through apoptosis and the change of the gene expression at environmental relevant concentrations. In additional, co-exposure of Cu with other environmental contaminants such as tetracycline can change the bioavailability and the associated toxicity of Cu in aquatic organisms. This study provides toxicological information for the establishment of environmental standards and the ecological risk assessment of antibiotics and heavy metals in aquatic environment.