Acute Exposure to Environmentally Relevant Concentrations of Ciprofloxacin and Levonadifloxacin Alters Behavior, Organ Health, and Stress Response in Adult Zebrafish

ABSTRACT

Antibiotic pollution in aquatic systems is an emerging global concern, but the sublethal effects of acute exposure on aquatic vertebrates are poorly understood. This study examined the acute toxicity by exposing adult zebrafish to three concentrations (1, 5, and 10 mg/L) of ciprofloxacin (CIP) and levonadifloxacin (LND) for 96 h. Behavioral, histological, biochemical, and transcriptional changes were assessed. In the novel tank‐dive test, both antibiotics induced concentration‐ and time‐dependent anxiogenic behaviors, such as reduced exploration, decreased total distance traveled, and less time in the upper zone. Histopathological analysis showed progressive tissue damage beginning in the gill epithelium and spreading to the intestine and muscle. Overall, lesion severity increased with higher concentrations and was consistently higher in CIP‐exposed fish. Antioxidant enzyme activity exhibited significant changes in superoxide dismutase, catalase, and glutathione peroxidase 1. Early increase in enzyme levels at 48 h coincided with reduced transcription of sod1 , cat1 , and gpx1a . At 96 h, transcription levels increased while protein levels remained stable. Pathway analysis grouped these genes within interconnected oxidative stress networks rather than cell death pathways. Overall, the results indicate that exposure to both fluoroquinolones for 96 h causes a staged oxidative stress response, along with behavioral disruptions and tissue damage. CIP caused stronger immediate biological effects than LND at the same concentrations, although both antibiotics disturbed organismal homeostasis at sublethal levels. These findings highlight the ecological importance of short‐term antibiotic contamination and demonstrate the value of multiple endpoints for detecting early toxic effects in aquatic organisms.