铬污染土壤修复过程中土壤细菌群落多样性的RFLP分析

RFLP analysis bacterial communities of remedied chromium-contaminated soil by different methods

【目的】对污染土壤修复过程中土壤细菌群落多样性的变化进行研究。【方法】以淹水培养后的模拟铬污染土壤为供试材料,通过直接提取土壤中总细菌DNA,利用细菌专一引物克隆细菌16S rDNA片段,分别建立克隆文库。利用PCR-RFLP技术,分析比较了土壤淹水10 d(对照,S1)、添加Cr(Ⅵ)淹水10 d(S2)、添加Cr(Ⅵ)和Fe(OH)3淹水10 d(S3)及20 d(S4)4个处理中土壤细菌群落的变化。【结果】用专一引物克隆细菌16S rDNA片段,分别建立了克隆文库;用限制性内切酶RsaⅠ进行细菌16S rDNA PCR-RFLP分析,分别得到123,120,97和69个酶切类型,库容值分别为54.92%,55.43%,65.33%和76.60%;Shannon-Wiener指数、Gini指数、物种丰富度指数(dMa)和物种均匀度指数(Jgi)均表现为S1>S2>S3>S4,以上4个指数的变异系数分别为11.51%,1.84%,23.64%和1.55%;基于细菌多样性参数的聚类分析结果,将对照S1和添加Cr(Ⅵ)处理的S2归于一类,而2个添加Fe处理的土壤S3和S4聚为一类。【结论】经过10 d淹水处理,土壤中添加的Cr(Ⅵ)有98%以上可发生转化,土壤残留的Cr(Ⅵ)含量基本达到稳定。添加铬矿渣处理(S2)的土壤细菌群落多样性和对照(S1)基本一致,添加氧化铁的2个处理(S3和S4)的细菌群落多样性基本接近,但低于S1和S2;添加氧化铁的处理(S3和S4)中,均出现了明显的优势细菌群落,其分别属于荧光假单胞菌属和不动杆菌属。

[Objective] The changes of soil bacterial community were studied during the process of soil remediation. [Method] The simulation Cr（Ⅵ） contaminated soil was used as test material,which had been treated by waterlogging for 10 days （S2）,adding ferrihydrite and waterlogging for 10 days （S3） and 20 days （S4）. As control,the nature soil waterlogged for 10 clays （S1） was used. Total genomic DNA of bacteria was extracted directly from soil treatments. The 16S rDNAs were amplified by PCR with the bacterial universal primers 63F/1387R. [Result] Using RFLP four clone libraries were analyzed based on patterns of restriction endonuclease Rsa Ⅰ , the OUT of four samples was 123,120,97 and 69 respectively. And the coverage was 54.92% ,55.43% ,65.33% and 76.60%. The bacterial diversity indexes of Shannon-Wiener,Gini and species richness index dMr , evenness index Jgi represented an identical order S1 〉 S2 〉 S3〉 S4. The C. V. of four bacterial diversity indexes was 11.51%, 1.84% ,23.64% and 1.55%. Based on the bacterial diversity index clustering analysis,treatments of S1 and S2 were classified into the same group, while S3 and S4 showed similarity to some degree. [Conclusion] After 10d anaerobic treatment by waterlogging,the chromium-contaminated was almost reduced, and the bacterial diversity level was similar between S1 and S2,which was higher than that of S3 and S4. Dominant bacteria which were helpful to eliminate heavy metal contamination and had Fe （Ⅲ）-reducing potential were found in S3 and S4.