Baek Seongeun, Seo Jinbeom, Yun Taegwan, Kim Jin, Shin YuJin, Choi Jiwoo, Chang JuOae, Kim Inseo, Yang Yung-Hun, Kim Wooseong, Lee Wonsik
College of Pharmacy, Graduate School of Pharmaceutical Sciences, Ewha Womans University, Seoul 03760, Republic of Korea.
School of Pharmacy, Sungkyunkwan University, Suwon 16419, Republic of Korea.
Ecotoxicol Environ Saf. 2025 Mar 15;293:118014. doi: 10.1016/j.ecoenv.2025.118014. Epub 2025 Mar 12.
Bacterial persisters are dormant phenotypic variants that are tolerant to antibiotics, contributing to treatment failure and the emergence of antimicrobial resistance. Although the formation of persisters has been extensively studied in regards to bacterial infections and treatment, such as antibiotic exposure or intracellular survival within macrophages, the role of environmental stressors in persister formation remains largely unexplored. In this study, we investigate the role of environmental heavy metals, specifically arsenic (As), cadmium (Cd), and mercury (Hg), in promoting persister cell formation in Staphylococcus aureus and Escherichia coli. Log-phase cultures were exposed to heavy metals (5 mM As, 1.25 mM Cd, 4 µM Hg for S. aureus; 12.5 mM As, 2 mM Cd, and 15 µM Hg for E. coli) for 0.5 h to induce persister cells. We observed that exposure to these metals induced persister cell formation, confirmed by intracellular ATP levels through microscopy and luciferase assays, as well as by reactive oxygen species (ROS) levels using carboxy-H2DCFDA. Short-term heavy metal exposure strongly depleted intracellular ATP while generating ROS. Moreover, we observed enhanced expression of genes involved in the SOS response, including recA, umuC, dinB, rexA, rexB, sulA, rpoS, and soxR, as measured by qPCR. This response was likely induced by elevated ROS levels following heavy metal exposure. Furthermore, we demonstrate that heavy metal-induced bacterial persisters exhibited a substantially increased emergence of antibiotic resistance, as shown by ciprofloxacin resistance developing in the presence of heavy metals. Therefore, our results clearly demonstrate that heavy metals can induce persister cells by depleting cellular ATP and generating ROS, and these bacterial responses to heavy metals substantially contribute to antibiotic resistance. These findings highlight the intricate relationship between environmental heavy metals, bacterial persister formation, and antibiotic resistance, emphasizing the need for a "One Health" strategy to address the growing antibiotic resistance crisis.
细菌持留菌是对抗生素具有耐受性的休眠表型变体，会导致治疗失败和抗菌药物耐药性的出现。尽管关于细菌感染和治疗（如抗生素暴露或在巨噬细胞内的细胞内存活）过程中持留菌的形成已得到广泛研究，但环境应激源在持留菌形成中的作用仍很大程度上未被探索。在本研究中，我们调查了环境重金属，特别是砷（As）、镉（Cd）和汞（Hg），在促进金黄色葡萄球菌和大肠杆菌中持留菌细胞形成方面的作用。对数期培养物暴露于重金属（金黄色葡萄球菌为5 mM As、1.25 mM Cd、4 μM Hg；大肠杆菌为12.5 mM As、2 mM Cd和15 μM Hg）0.5小时以诱导持留菌细胞。我们观察到，暴露于这些金属会诱导持留菌细胞形成，这通过显微镜检查和荧光素酶测定的细胞内ATP水平以及使用羧基-H2DCFDA的活性氧（ROS）水平得到证实。短期重金属暴露会强烈消耗细胞内ATP，同时产生活性氧。此外，通过qPCR检测，我们观察到参与SOS反应的基因（包括recA、umuC、dinB、rexA、rexB、sulA、rpoS和soxR）的表达增强。这种反应可能是由重金属暴露后活性氧水平升高诱导的。此外，我们证明重金属诱导的细菌持留菌表现出抗生素耐药性的显著增加，如在重金属存在下环丙沙星耐药性的产生所示。因此，我们的结果清楚地表明，重金属可通过消耗细胞内ATP和产生活性氧来诱导持留菌细胞，并且这些细菌对重金属的反应极大地促进了抗生素耐药性。这些发现突出了环境重金属、细菌持留菌形成和抗生素耐药性之间的复杂关系，强调了采取“同一健康”策略来应对日益严重的抗生素耐药性危机的必要性。
