Chronic neonicotinoid exposure disrupts survival, development, digestive enzymes, and gut microbiome in honeybee...

Honeybee queens (Apis mellifera L.) are essential for colony sustainability, most study mainly focused on the acute toxicity of chemical insecticides. This study investigated the chronic effects of three prevalent neonicotinoids on honeybee queen larvae development, digestive physiology, and gut microbiota. It was determined that exposure to elevated concentrations (10-40 mg/L) significantly increased larval mortality (up to 41.70%), which reduced both capping and emergence rates, with imidacloprid causing a 43.80% decline in emergence at 40 mg/L. Chronic exposure (1-25 mg/L) to clothianidin notably decreased birth weight by 11.40% and altered thoracic and abdominal morphometrics. Moreover, imidacloprid and clothianidin suppressed amylase activity by up to 94%, while acetamiprid enhanced it by nearly 60%. Additionally, 16S rRNA gene sequencing revealed significant shifts in gut microbiota composition, characterized by increased abundance of Firmicutes and decreased Bacteroidota, despite minor changes in overall diversity. Functional predictions indicated alterations in carbohydrate metabolism, amino acid metabolism, and membrane transport pathways. These findings demonstrate that chronic neonicotinoid exposure disrupts metabolic and microbial homeostasis in honeybee queen larvae, posing a serious threat to honeybee colony fitness, which is important and provides essential evidence for ecological risk assessment and the development of safer pesticide application strategies in apiculture and crop protection.