Mitochondrial Gene Expression as a Novel Biomarker for Detecting and Discriminating Neurotoxic Pesticide Exposure in Ramulus phyllodeus (Chen & He, 2008)

This study investigates the mitochondrial transcriptomic responses of Ramulus phyllodeus (Chen & He, 2008); Phasmatodea: Phasmatidae) to acute exposure to four widely used neurotoxic insecticides: chlorpyrifos, cyfluthrin, emamectin benzoate, and acetamiprid. Using quantitative real-time PCR (qRT-PCR), we quantified transcriptional changes in 10 mitochondrial protein-coding genes, which showed significant transcriptional changes (p < 0.05) when the insect was exposed to four commonly used pesticides (each at a concentration of 5 μg/L) for 24 h. Exposure to chlorpyrifos induced significant upregulation of ND2 (2.08 ± 0.048) and ND5 (1.38 ± 0.15). Cyfluthrin triggered coordinated upregulation across seven genes: ND1 (1.71 ± 0.07), ND2 (2.33 ± 0.38), ND3 (1.74 ± 0.25), ND5 (1.65 ± 0.38), COX1 (2.91 ± 0.40), COX3 (1.69 ± 0.18), and Cytb (2.81 ± 0.53). Emamectin benzoate induced the upregulation of ND1 (1.98 ± 0.21), ND2 (3.04 ± 0.41), ND3 (1.82 ± 0.26), ND4 (2.79 ± 0.64), COX1 (2.36 ± 0.34), ATP6 (3.26 ± 0.61), and Cytb (2.39 ± 0.81). Acetamiprid induced more selective upregulation, affecting only ND1 (1.67 ± 0.18), ND4 (1.43 ± 0.16), and ND5 (1.66 ± 0.10). Critically, each insecticide elicited a distinct, non-overlapping transcriptional signature, defined by both the identity and magnitude of responsive genes, indicating compound-specific modulation of mitochondrial gene expression. Notably, no gene exhibited significant downregulation under any single-compound treatment, and all differentially expressed genes were upregulated exclusively in response to individual pesticides. This absence of transcriptional suppression suggests that these neurotoxicants converge on shared upstream stress-response pathways that preferentially activate mitochondrial biogenesis or compensatory transcription, rather than inducing global transcriptional repression. Collectively, these findings establish mitochondrial protein-coding genes in R. phyllodeus as sensitive, mechanistically grounded molecular sentinels for neurotoxic pesticide exposure. The compound-specific transcriptional profiles further suggest potential utility in multiplex detection strategies for environmental monitoring, enabling discrimination among individual residues.