Ni2+胁迫下嗜酸氧化亚铁硫杆菌亚铁氧化应激效应

Iron oxidation response of Acidithiobacillus ferrooxidans under Ni2+ stress

摘要:

【目的】嗜酸氧化亚铁硫杆菌(Acidithiobacillus ferrooxidans)作为湿法冶金的主要功能菌，其在重金属Ni²⁺胁迫下的亚铁氧化应激机制还不清楚。【方法】在两步法培养体系中，设计由低到高Ni²⁺胁迫浓度，利用分光光度法、电化学法和实时荧光定量PCR法等从宏观到微观探究Ni²⁺胁迫下A.ferrooxidans菌亚铁氧化应激效应。【结果】两步培养法体系下A.ferrooxidans菌能够耐受较高浓度的Ni²⁺毒性(≤30 g/L Ni²⁺)。当Ni²⁺胁迫浓度增加至40 g/L时，与对照组(不添加Ni²⁺胁迫)相比，A.ferrooxidans亚铁氧化率和速率显著降低(24 h的亚铁氧化抑制率约为59.4%)，亚铁氧化代谢相关的rus操纵子各功能基因的表达量也显著下调，尤其是基因cyc1(最高下调了16倍)和coxBACD(4个不同亚基最高下调2.7–7.4倍)；同时Fe²⁺氧化相关的胞外电子传递能力也降低，对照组(0 g/L Ni²⁺，29.0µA)最大还原峰电流值高于实验组(40 g/L Ni²⁺，24.5µA)。【结论】超高浓度Ni²⁺胁迫环境对A.ferrooxidans菌有毒性，导致该菌rus操纵子上各功能基因表达量下调，同时造成胞外电子传递速率降低，最终致使A.ferrooxidans菌亚铁氧化能力降低。研究结果将为含Ni固废高效生物浸出提供理论支持。

Abstract:

[Objective] Acidithiobacillus ferrooxidans is one of the key bacteria involved in the bioleaching of primary and secondary ores and solid wastes. In the bioleaching environments, high levels of heavy metals are one of the major challenges encountered by A. ferrooxidans. Resistance to heavy metals stress is essential for the adaptation of microorganisms to the extreme environments. However, the tolerance and iron oxidation response of A. ferrooxidans to Ni²⁺ stress remain unclear. [Methods] In this study, we employed a two-step culture system with the Ni²⁺concentration range designed within 0-40 g/L. The ferrous oxidation efficiency, expression of rus operon genes, and extracellular electron transfer of A. ferrooxidans exposed to Ni²⁺ stress were respectively investigated by phenanthroline spectrophotometry, real-time quantitative PCR, and cyclic voltammetry. [Results] A. ferrooxidans could tolerate Ni²⁺ stress at the concentration ≤30 g/L in the two-step culture system. However, 40 g/L Ni²⁺ significantly inhibited the ferrous iron oxidation activity of A. ferrooxidans, and the ferrous oxidation inhibition efficiency at the time point of 24 h was 59.4% that of the control (0 g/L Ni²⁺). Moreover, the high concentration (40 g/L) of Ni²⁺ decreased the transcriptional levels of rus operon genes. Especially, the transcriptional levels of cyc1 and coxBACD were maximally down-regulated by 16 folds and 2.7-7.4 folds, respectively. Meanwhile, the rate of extracellular electron transfer in A. ferrooxidans exposed to 40 g/L Ni²⁺ was decelerated to 24.5 µA in comparison to the control (0 g/L Ni²⁺, 29.0 µA). [Conclusion]High concentration (40 g/L) of Ni²⁺ down-regulated the expression of rus operon genes involved in Fe²⁺oxidation, inhibited the electron transport from Fe²⁺ oxidation, and finally slowed down ferrous oxidation activity. The findings provide theoretical support for the bioleaching of Ni-containing solid waste.