Carvacrol derived from <i>Origanum vulgare</i> oil was evaluated for its acaricidal efficacy and mechanism of action against <i>Haemaphysalis longicornis</i>, a primary vector of severe fever with a thrombocytopenia syndrome. Essential oils extracted from <i>O. vulgare</i> leaves cultivated in Germany, Albania, and Iran were analyzed. Among them, the German oil exhibited the highest acaricidal potency due to its elevated carvacrol content (83.38%). Carvacrol was isolated and its identity was confirmed using GC/MS, NMR, and FT-IR analyses. Carvacrol demonstrated significant toxicity across all developmental stages of <i>H. longicornis</i>, with LC₅₀ values of 3.47, 8.21, and 15.27 μg/cm³ for larvae, nymphs, and adults, respectively-representing 4.4-, 3.5-, and 3.2-fold higher potency compared to diethyltoluamide. The acaricidal mechanism of carvacrol involved multiple pathways: (i) inhibition of acetylcholinesterase activity by up to 85.4%, leading to neural disruption; (ii) suppression of cytochrome P450 monooxygenases (47.4% reduction), α-esterase (24.8% reduction), and β-esterase (28.6% reduction); and (iii) disruption of neural signaling pathways critical for survival. Chemical analysis confirmed carvacrol as the predominant active component in <i>O. vulgare</i> oil, with additional contributions from thymol and thymoquinone. Although its concentration in Iranian oil was lower, thymoquinone demonstrated the highest acaricidal potency (4.98 μg/cm³). Nevertheless, the abundance and superior efficacy of carvacrol establish it as the primary component contributing to the oil's bioactivity. This study demonstrates that carvacrol is a promising eco-friendly alternative to synthetic acaricides for the control of <i>H. longicornis</i>. Its ability to inhibit acetylcholinesterase activity and suppress detoxifying enzymes suggests potential for overcoming resistance mechanisms associated with synthetic chemicals. Further studies should focus on optimizing the formulations and applying them in the field to improve efficacy.