Neurotoxicity and Immunotoxicity of the Insecticide Isofenphos-methyl in Zebrafish (Danio rerio)

• Environmentally relevant IFP exposure disrupts embryonic development in zebrafish. • High-dose IFP exposure causes neurotoxicity, visual toxicity, and immunotoxicity. • IFP adversely affects CNS structure and neurogenesis. • IFP reduces the numbers of neutrophils and macrophages. • IFP may target multiple metabolic pathways. Isofenphos-methyl (IFP), an organophosphorus pesticide widely used in agriculture, has raised public health concerns because of its potential risks to human health. Although recent studies have linked IFP exposure to developmental toxicity, the underlying toxicological mechanisms remain insufficiently understood. To address this gap, this study employed a zebrafish model to investigate the developmental toxicity of IFP through an integrated approach combining behavioral, histopathological, biochemical, and transcriptomic analyses. Exposure to IFP from 2 to 96 hours post-fertilization (hpf) adversely affected early zebrafish embryonic development, with a calculated 96-h LC₅₀ of 6.407 mg/L. Embryos exposed to 0.1, 2, 4, 8 mg/L IFP diminished responsiveness to light/dark stimulation, and disrupted circadian rhythms. Moreover, IFP exposure impaired immune function, significantly reducing in macrophage and neutrophil numbers. Transcriptomic analysis revealed disruptions in neuroactive ligand–receptor interaction, leading to dysfunctional synaptic signalling and neuronal communication; and it also disturbed protein digestion and absorption, thereby limiting the availability of essential amino acid precursors (e.g., tryptophan and tyrosine) required for the synthesis of key neurotransmitters such as serotonin and dopamine (DA). These transcriptomic alterations were supported by altered neurotransmitter levels, brain mitochondrial damage, and histopathological abnormalities in both brain and ocular structures. RNA-sequencing results were further validated using quantitative real-time reverse transcription polymerase chain reaction (RT-qPCR), confirming the robustness of the data. Collectively, these findings demonstrate that IFP exerts pronounced neurotoxic and immunotoxic effects on developing zebrafish, underscoring potential risks during early developmental stages and the urgent need for stricter regulation of IFP contamination in aquatic environments.