Antibiotics are widely used to improve human and animal health and treat infections. Antibiotics are often used in livestock farms and fisheries to prevent diseases and promote growth. Recently, there has been increasing interest in the presence of antibiotics in aquatic environments. Low levels of antibiotic components are frequently detected in surface water, seawater, groundwater, and even drinking water. Antibiotics are consistently and continually discharged into the natural environment as parent molecules or metabolites, which are usually soluble and bioactive, and this results in a pseudo and persistent pollution. The effects of environmental antibiotic toxicity on non-target organisms, especially aquatic organisms, have become an increasing concern. Although antibiotics have been detected worldwide, their ecological and developmental effects have been poorly investigated, particularly in non-target organisms. This review describes the toxicity and underlying mechanism of antibiotic contamination in aquatic organisms, including the effects on vertebrate development. A considerable number of antibiotic effects on aquatic organisms have been investigated using acute toxicity assays, but only very little is known about the long-term effects. Aquatic photosynthetic autotrophs, such as Pseudokirchneriella subcapitata, Anabaena flos-aquae, and Lemna minor, were previously used for antibiotic toxicity tests because of low cost, simple operation, and high sensitivity. Certain antibiotics show a different degree of potency in algal toxicity tests on the basis of different test algae. Antibiotics inhibit protein synthesis, chloroplast development, and photosynthesis, ultimately leading to growth inhibition; some organisms exhibit growth stimulation at certain antibiotic concentrations. Daphnia magna and other aquatic invertebrates have also been used for checking the toxicity priority of antibiotics. When investigating the acute effect of antibiotics (e.g., growth inhibition), concentrations in standard laboratory organisms are usually about two or three orders of magnitude higher than the maximal concentrations in the aquatic environment, resulting in the underestimation of antibiotic hazards. Vertebrate organisms show a promising potential for chronic toxicity and potentially subtle effects of antibiotics, particularly on biochemical processes and molecular targets. The adverse developmental effects of macrolides, tetracyclines, sulfonamides, quinolones, and other antibiotic groups have been evaluated in aquatic vertebrates such as Danio rerio and Xenopus tropicalis. In acute toxicity tests, low levels of antibiotics have systematic teratogenic effects on fish. The effects of antibiotics on oxidative stress enzymes and cytochrome P450 have been investigated. Cytotoxicity, neurotoxicity, and genotoxicity have been observed for certain antibiotic amounts. However, there are no firm conclusions regarding the chronic toxicity of antibiotics at environmentally relevant levels because of the lack of long-term exposure studies. Herein, future perspectives and challenges of antibiotic toxicology were discussed. Researchers should pay more attention to the following points: chronic toxicity and potentially subtle effects of environmentally relevant antibiotics on vertebrates; effects of toxicity on biochemical processes and mode of action; combined toxicity of antibiotics and other antibiotics, metabolites, and heavy metals; and environmental factors such as temperature and pH.