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Bixafen-induced immunotoxicity in zebrafish: TLR4/NF-κB activation and DNA replication disruption as critical
Fei Fang1, Bo Peng1, Yanjuan Zhu1
1Shanghai Key Laboratory of Chemical Biology, School of Pharmacy, East China University of Science and Technology, Shanghai 200237, China.
Abstract:
Comprehensive understanding the toxic effects of pesticide with relative long half-life properties are always hot research topics in the world. In this study, the immunotoxic effects of Bixafen (BIX), a prominent Succinate Dehydrogenase Inhibitor (SDHI) fungicide, were systematically assessed at concentrations of 0.05, 0.1, and 0.2 μM, using a zebrafish (Danio rerio) larval model. Neutral red staining and confocal microscopy quantification showed that BIX exposure reduced immune cell numbers in a concentration-dependent manner. qPCR and Western blot validation revealed upregulated pro-inflammatory factors (IL-1β, IL-8, TNF-α) and TLR4 gene expression, while the lymphocyte marker gene rag1 was downregulated. Protein - level detection found significantly increased p-p65/p65 and TLR4 expression. Molecular docking experiments showed that BIX forms multiple interactions with the TLR4 protein. RNA-Seq analysis further confirmed that BIX interferes with DNA replication and metabolic pathways. Our results suggest that BIX may activate proinflammatory responses and disrupt immune cell homeostasis via the TLR4 signaling pathway, posing a potential immunotoxicity risk to non-target organisms. These findings provide insights into the molecular mechanisms of BIX-induced immunotoxicity and are critical for assessing the environmental safety and health risks associated with SDHI fungicides.
Insights
Bixafen (BIX), a fungicide, was found to harm zebrafish immune cells by disrupting the TLR4 pathway and increasing inflammation. This pesticide poses potential immunotoxicity risks to non-target organisms.
Area of Science:
- Environmental Toxicology
- Immunology
- Molecular Biology
Background:
- Pesticide toxicity, particularly from compounds with long half-lives, is a significant global research area.
- Succinate Dehydrogenase Inhibitor (SDHI) fungicides, like Bixafen (BIX), are widely used but their non-target effects require thorough investigation.
Purpose of the Study:
- To systematically evaluate the immunotoxic effects of Bixafen (BIX) on zebrafish larvae.
- To elucidate the molecular mechanisms underlying BIX-induced immunotoxicity, focusing on inflammatory pathways.
Main Methods:
- Zebrafish (Danio rerio) larvae were exposed to varying concentrations of BIX (0.05, 0.1, 0.2 μM).
- Immunotoxicity was assessed using neutral red staining, confocal microscopy, qPCR, Western blot, molecular docking, and RNA-Seq.
- Key indicators included immune cell counts, pro-inflammatory factors (IL-1β, IL-8, TNF-α), TLR4 signaling, lymphocyte markers (rag1), and DNA replication/metabolic pathways.
Main Results:
- BIX exposure reduced immune cell numbers in a dose-dependent manner.
- Upregulation of pro-inflammatory factors (IL-1β, IL-8, TNF-α) and TLR4 gene/protein expression was observed.
- Downregulation of the lymphocyte marker rag1 and evidence of BIX interacting with TLR4 were found.
- RNA-Seq indicated BIX interferes with DNA replication and metabolic pathways.
Conclusions:
- Bixafen (BIX) exhibits immunotoxicity in zebrafish, potentially by activating pro-inflammatory responses and disrupting immune cell homeostasis through the TLR4 signaling pathway.
- These findings highlight the potential immunotoxicity risk of BIX to non-target organisms.
- The study provides critical insights into the molecular mechanisms of BIX-induced immunotoxicity, aiding environmental safety and health risk assessments for SDHI fungicides.
