耐酸砷还原菌Bacillus sp.strain P3-23的分离鉴定及其对高砷土壤砷释放影响

Isolation and Identification of an Acid-tolerant Arsenate-reducing Bacterium Bacillus sp.strain P3-23 and Its Role in the Release of Arsenic from High-arsenic Soil

砷还原微生物在原生高砷地下水形成中起关键作用,研究其对不同环境因素改变的响应以及对砷迁移与转化的影响是十分必要的。从石门土壤中分离得到一株耐酸砷还原菌,研究其形态和生理生化特征,通过分子生物学和微生物学手段对其进行系统分类和生理生化特性鉴定,并使用微生物学方法在不同温度、pH值和电子供体条件下进行培养,探究其对环境因素波动的适应能力,检测菌株对高砷土壤砷释放的影响。实验结果表明:该耐酸砷还原菌为芽孢杆菌属(Bacillus)成员,故命名为Bacillus sp.strain P3-23(以下简称P3-23);菌株P3-23最适宜的生长温度为30℃,在pH值为3.5~7.5范围内均可生长且具有砷还原能力,能够利用乳酸钠、丙酮酸钠、柠檬酸钠、酵母味素、丙三醇、葡萄糖、蔗糖为电子供体;菌株P3-23能够在72 h内完全还原2.0 mM As(Ⅴ)且菌液浓度呈上升趋势,具有强耐砷能力,40.0 mM砷存在条件下仍能生长;菌株P3-23促进土壤中砷释放能力较强,砷形态分析显示释放的可溶性砷中As^(3+)占比达83.3%以上。菌株P3-23的分离不仅丰富了人们对砷还原微生物的认识,也说明砷还原微生物对不同环境因素胁迫具有相应的应答机制,预示着其可能存在于极端环境中。对不同环境中砷还原微生物进行研究,在完善砷的生物地球化学循环模式的同时有助于砷污染机制及砷修复生物手段的探索。

Arsenate-reducing microorganisms play an important role in the formation of primary high-arsenic groundwater.It is necessary to study their responses to environmental factors’changing and their effects on migration and transformation of arsenic.We isolate a single acid-tolerant arsenate-reducing bacteria from arsenic contaminated soil of Shimen,analyze its morphological,physiological and biochemical characteristics,and classify it and conduct physiological and biochemical characteristics identification by means of molecular biology and microbiology approach,and culture it in different temperatures and pH values and di-fferent kinds of electronic donor by using the microbiological method to explore the adaption ability to environmental factors’changing and to detect the effect of the strain on the arsenic release of high-arsenic soil.The results show that the acid-tolerant arsenate-reducing bacteria is an arsenate-reducing bacterium.It belongs to Bacillus,and thus is proposed as Bacillus sp.strain P3-23.Its optimum temperature is 30℃and it can grow under pH 3.5~7.5.P3-23 can utilize lactate,pyruvate,citrate,yeast extract,glycerol,glucose and sucrose as solo electron donor.2 mM As(Ⅴ)can be reduced completely within 72 hours in the presence of P3-23 and the bacteria has a tendency of growing.Strain P3-23 is capable of tolerating extremely high concentrations of arsenic.It can thrive when it is exposed to 40.0 mM As(Ⅴ).Strain P3-23 catalyzes dissolution of arsenic from high-arsenic soil under anaerobic condition,in which As3^(+)accounts for more than 83.3%of the total arsenic.The results broaden our horizons about arsenate-reducing bacteria,showing that arsenate-reducing bacteria can overcome different environmental stresses,which indicates that it may exist in different extreme environments.Study on arsenate-reducing bacterium is helpful to perfect the biogeochemistry cycle of arsenic as well as exploring of the arsenic pollution mechanism and the biological arsenic remediation means.