Innovative Transgenic Zebrafish Biosensor for Heavy Metal Detection

Abstract

Heavy metal contamination is an urgent environmental issue that poses a significant threat to human health and the ecosystem. To mitigate the adverse impacts of heavy metal pollution, the aim of this research was to develop genetically engineered zebrafish as biosensors, which offer a promising alternative for detecting heavy metal exposure, specifically Cd²⁺ and Zn²⁺. A novel heavy metal-sensitive gene construct metallothionine 2 promoter with DsRed reporter gene (mt2-DsRed2) was synthesized and integrated into zebrafish embryos using a Tol2 transposon transposase system with the transgenic zebrafish line subjected to biosensing applications for Cd²⁺ and Zn²⁺. The biosensor showed specific responses with linear correlation heavy metal concentration and DsRed fluorescence signal for Cd²⁺ and Zn²⁺ with (p < 0.01) a minimum detection limit of 4 ppb for each metal ion but not for the non-specific metal ion Ni²⁺, which makes it suitable for laboratory-based heavy metal assessment assays in the low heavy metal concentration ranges of 0-10 ppb. Additionally, the study investigated the toxicity of heavy metals on zebrafish early developmental stages applying a modified version of the OECD Fish Embryo Toxicity (FET-236) test. Accordingly, Cd²⁺, Zn²⁺, and Ni²⁺ exhibited no significant toxicity effects on zebrafish embryos within the low dose range of 2-10 ppb confirming the biocompatibility of the transgenic zebrafish biosensor for heavy metal sensing applications. Thus, the developed transgenic zebrafish line can accurately sense heavy metals Cd²⁺ and Zn²⁺ within the low dose range, making it a promising alternative laboratory assay for environmental monitoring and risk assessment.