Microbial Diversity During The Remediation Of Cadmium-Polluted Soil Of Mining Area And Quantitative Monitoring Of Engineering Bacteria

Cadmium is a heavy metal pollutant that has large toxicity to organisms. The activity of indigenous soil microbial community is usually inhibited by high levels of cadmium. This tested soil was collected from the mining area of Daye Country, Hubei province, China. The content of Copper and Cadmium in the soil all exceeds the standard. The soil is mildly polluted by Copper and severely polluted by Cadmium. In the previous studies, the contaminated soil was bioremediated with the application of straw and engineered-bacterium. This study evaluated the influence of rice straw (5.2,10.3and23.2t/ha) and an engineered-bacterium X4/pCIM on the soil microbial community by Biolog and DGGE to detect the variation of microbial diversity of soil during the remediation. Real-time PCR was applied to quantitative monitoring of the bacterial strain X4/pCIM. The main results are outlined as follows:1. Biolog analysis revealed that the soil microbial community composition changed after combined application of the rice straw and engineered bacteria. But it has different effects on soil rhizosphere microorganisms and non-rhizosphere microorganisms. Using low-dose straw increased the physiological activity of the non-rhizosphere soil microbial communities and inhibited the physiological activity of the rhizosphere soil microbial communities. In contrast, the addition of the high-dose straw inhibited the physiological activity of the non-rhizosphere soil microbial communities and increased the physiological activity of the rhizosphere soil microbial communities. Using medium-dose straw both inhibited the physiological activity of the non-rhizosphere soil microbial communities and the physiological activity of the rhizosphere soil microbial communities. By contrast with the application of straw, the joint application of rice straw and engineered bacteria significantly improved the the physiological activity of the non-rhizosphere soil microbial communities and markedly inhibited the physiological activity of the rhizosphere soil microbial communities. Accoding to the results on the analysis of the diversity index obtained from Biolog data, using low-dose straw enhanced the richness and evenness of the non-rhizosphere soil microbial communities. On the contrary, the addition of the high-dose straw reduced the richness and evenness of the non-rhizosphere soil microbial communities and improved the richness and evenness of the rhizosphere soil microbial communities. Using medium-dose straw inhibited the richness and evenness of both the non-rhizosphere and the rhizosphere soil microbial communities. The joint application of low-dose or medium dose rice straw and engineered bacteria improved the richness and evenness of the non-rhizosphere soil microbial communities and inhibited the richness and evenness of the rhizosphere soil microbial communities. The joint application of high-dose rice straw and engineered bacteria both increased the richness and evenness of the non-rhizosphere or rhizosphere soil microbial communities. As the results of analysis on functional diversity, the straw had no significant effect on the carbon source utilization efficiency of non-rhizosphere microbial community. The joint application of high-dose rice straw and engineered bacteria influenced the carbon source utilization efficiency of non-rhizosphere microbial community. Using low-dose or high-dose straw and the joint application of low-dose rice straw and engineered bacteria changed the carbon source utilization efficiency of rhizosphere microbial community. These results demonstrated that the carbon sources that rhizosphere soil microbes use shifted.2. DGGE showed that the dominate microorganisms are uncultured microorganisms in the tested soil. In the DGGE patterns, band6-2,3-3,7-2,10-1,9-1,7-1,3-1were uncultured Acidobacteria bacteria, their percentage is58.33%. Band10-2belonged to uncultured gamma proteobacterium, its proportion is8.33%. Band3-2,6-1were uncultured soil bacteria, their percentage is16.67%. Band4-2was Enterobacter sp., with a percentage of8.33%. Band4-1was Geobacillus toebi, its proportion is8.33%. Gene sequence3-3had a distinct band in all the tested soil, it was an excellent cadmium-resistant bacteria.3. Real-time PCR results showed that the number of X4/pCIM in the soil is approximately10CFU/g soil after120days. In all non-rhizosphere samples, X4/pCIM achieved the best colonization in the sample RS2+B. The number of X4/pCIM in the sample was2797±176CFU/g soil. The number of X4/pCIM in sample RS1+B and RS3+B were2483±210CFU/g soil and1870±109CFU/g soil, respectively. There were significant differences (p<0.05) in all non-rhizosphere samples. Using medium-dose straw significantly improved the ability of colonization of X4/pCIM in the non-rhizosphere soil. The addition of high-dose straw significantly inhibited the ability of colonization of X4/pCIM in the non-rhizosphere soil. In all rhizosphere samples, X4/pCIM achieved the best colonization in the sample RS3+BR, its number is3967±247CFU/g soil. The number of X4/pCIM in sample RS1+BR and RS2+BR was1639±190CFU/g soil and750 ±56CFU/g soil, respectively. There were also significant differences (p<0.05) in all rhizosphere samples. The addition of low-dose or high-dose straw improved the ability of colonization of X4/pCIM in the rhizosphere soil, but using medium-dose straw inhibited the ability of colonization of X4/pCIM in the rhizosphere soil.In summary, using rice straw and engineered bacteria can alter the soil microbial community composition. The influence of rice straw and engineered bacteria on the microbial diversity was different with the change of the applied straw dose. There was no apparent law.