耐多种重金属细菌的筛选、鉴定和酶活测定

Isolation,Identification and Enzyme Activity Determination of Heavy Metal Tolerant Bacteria

重金属污染严重危害土壤生态环境质量。通过筛选耐多种金属离子的细菌可为复合型重金属污染地区的生态环境修复提供微生物菌种资源。该研究从腾格里沙漠生物土壤结皮中进行菌种初筛,并在重金属(W^(6+)、Mn^(2+)、As^(3+)、Ni^(2+)、Mg^(2+))最低胁迫浓度下进行复筛。根据细菌在培养基上的生长状况,得到能够耐受5种重金属的2个菌株,16S rRNA序列比对结果表明分别为炭疽芽孢杆菌(Bacillus anthracis L3)和贝莱斯芽孢杆菌(Bacillus velezensis L11)。菌株L3和L11对Mn^(2+)、Ni^(2+)、As^(3+)、Mg^(2+)、W^(6+)5种重金属的耐受范围分别为200~2000 mg/L、10~40 mg/L、100~300 mg/L、10~800 mmoL/L、10~400 mmol/L和200~2000 mg/L、10~40 mg/L、100~200 mg/L、10~700 mmoL/L、10~180 mmol/L,且均对Pb^(2+)、Fe^(3+)、K^(+)、Na^(+)具有耐受性。在不同重金属高浓度胁迫下,扫描电镜结果表明L3菌体对不同重金属胁迫呈现出不同的形态变化。在As^(3+)的胁迫下,L11的菌体表面由粗糙变得饱满光滑,L3和L11菌体的黏附性和聚集性有所提高,可能是由于分泌了胞外聚合物。生长曲线测定结果表明2株菌的生长在不同重金属高浓度胁迫下均受到抑制,且L3和L11对Mg^(2+)的耐受性显著高于其他离子。酶活测定结果表明2株菌的MDA酶活性均与对照组无显著性差异,仅有Ni^(2+)胁迫下L3和Mn^(2+)胁迫下的L11的SOD酶活性与对照组有显著性差异。不同浓度梯度重金属胁迫条件下,株菌L3和L11对不同重金属的适应性和稳定性有差异。该研究获得的L3和L11均具有优良的修复土壤重金属污染的潜在价值,但对不同重金属的解毒机制还需深入研究。

Heavy metal pollution seriously endangers the soil ecological environment quality.Screening bacteria resistant to multiple metal ions can provide microbial strain resources for the restoration of the ecological environment in complex heavy metal contaminated areas.In this paper,a preliminary screening of strains from biological soil crusts was conducted in the Tengger Desert,and re-screened them under the lowest stress concentration of heavy metals(W^(6+),Mn^(2+),As^(3+),Ni^(2+),Mg^(2+)).According to the growth of the bacteria on the medium,two strains that can tolerate five heavy metals were obtained.The 16S rRNA phylogenetic results showed that they were Bacillus anthracis L3 and Bacillus velezensis L11,respectively.The tolerance ranges of strain L3 and L11 to 5 heavy metals of Mn^(2+),Ni^(2+),As^(3+),Mg^(2+),W^(6+)were 200~2000 mg/L,10~40 mg/L,100~300 mg/L,10~800 mmol/L,10~400 mmol/L and 200~2000 mg/L,10~40 mg/L,100~200 mg/L,10~700 mmol/L,10~180 mmol/L,respectively.And strain L3 and L11 were also tolerant to Pb^(2+),Fe^(3+),K^(+),Na^(+).Under high concentration stress of different heavy metals,the scanning electron microscopy results showed that the L3 strain showed different morphological changes to different heavy metal stresses.Under the stress of As^(3+),the surface of L11 cells changed from rough to full and smooth,and the viscosity and aggregation of L3 and L11 cells increased,which may be due to the secretion of extracellular polymers.The results of growth curve determination showed that the growth of the two strains was inhibited under different high concentration stresses of heavy metals,and the tolerance of L3 and L11 to Mg^(2+)was significantly higher than that of other ions.The results of enzyme activity determination showed that the MDA enzyme activities of the two strains were not significantly different from those of the control group.Only the SOD enzyme activities of L3 under Ni^(2+)stress and L11 under Mn^(2+)stress were significantly different from those of the control group.Under different concentration gradients of heavy metal stress,strains L3 and L11 have different adaptability and stability to different heavy metals.Both L3 and L11 obtained in this study have excellent potential value in remediation of heavy metal pollution in soil,but the detoxification mechanism of different heavy metals needs to be studied in depth.