Comparative Study On Leaching Of Cu/Zn Sulphide By Mixed Microorganisms Before And After Mutation And Their Isolation In Pure Culture

Bioleaching is a complicated biochemical process in which a number of microbial genera are involved.Most of bioleaching microorganisms are chemoautotrophic.They grow slowly and have low efficiency in metal -leaching.How to improve the leaching efficiency is a problem of industrial applications.Up to date,most popular strategy is mutation of single microorganism isolated from mixed microorganisms of bioleaching.However,microorganisms for bioleaching industry are always in mixed form,and mostly,their pure culture may not be obtained. Therefore,there are limitations on mutation breed of single microorganism.In present study,microbial samples were collected from acid mine drainage and spring of Jiangxi,Yunnan,Guangdong and Taiwan province, mixed and enriched.To obtain higher growth rate and higher leaching efficiency-mixed cultures,the mixed microorganisms were mutated using several physical and chemical mutagens and their combinations. Furthermore,single strain isolation,characterization of 16S rDNA,and ecological analysis were performed for the screened mixed cultures.The results are as follows:The mixed cultures with higher concentrations of leaching Cu²⁺, designated A,B,C,D and E,were screened after mutation and leaching tests of chalcopyrite.â‘ The concentrations of leaching Cu²⁺for mixed culture A,B,C, D and E were increased by 61.40%,125.57%,1.45%,58.98%and 149.98%and the leaching rates of Cu²⁺in the cultures were correspondingly 17.56%,35.90%,0.41%,16.87%and 42.88%higher compared with the control,40 d after leaching in 9K medium containing 1%chalcopyrite pulp.The results indicate that Cu²⁺leaching activities of mixed organisms can be enhanced by physical and chemical mutations.â‘¡The amount of leaching Cu²⁺for mixed culture A,B,C,D and E was increased by 49.46%,71.68%,22.82%,44.85%and 97.57%, respectively,with 3.65%,5.29%,1.69%,3.31%and 7.20%higher than the control in Cu²⁺leaching rates,40 d after leaching in 9K medium each containing 10%Chalcopyrite pulp,again indicating that physical and chemical mutations may improve Cu²⁺leaching activities of mixed organisms.â‘¢When added with 20%chalcopyrite pulp and inoculated in 9K and Leathen mediums for 40 d,mixed culture E extracted 87.22%and 79.24%more Cu²⁺from chalcopyrite and had 3.6%and 3.59 higher Cu²⁺ leaching rates than the unmutated control,suggesting the acquired higher characteristics of Cu²⁺-leaching is stable.â‘£Mixed culture E was more resistant to Cu²⁺than the unmutated control,suggesting that it possesses potentials for industrial applications.⑤The growth of mixed culture A,B,C,D and E in 9k medium was 1-2 d higher than that of the unmutated control and the mixed cultures conserved on chalcopyrite grew 1-2 d faster than those not conserved on chalcopyrite.These results indicate that growth activity of leaching microorganisms can be kept by selective stress.After three rounds of leaching tests on sphalerite,mixed cultures B, D and E with higher efficiency of Zn²⁺-leaching were obtained from mixed microorganisms A,B,C,D and E mentioned above.â‘ Compared with the unmutated control,the concentrations of leaching Zn²⁺in 9K medium added with 1%sphalerite pulp for mixed cultures A,B,C,D and E were increased by 8.06%,85.97%,-2.39%,77.61%and 117.01%,respectively,and the leaching rates of Zn²⁺in the cultures were correspondingly 1.54%,16.43%,-0.46%,14.83%and 22.37%,indicating that mutation against mixed microorganisms may improve leaching effects.â‘¡Compared with the unmutated control,the concentrations of leaching Zn²⁺for mixed culture B,D and E inoculated in 9K medium with 10%sphalerite pulp were increased by 49.14%,44.86%and 54.77%, and the leaching rates of Zn²⁺in the cultures were correspondingly 2.27%,2.07%and 2.53%,,respectively.â‘¢When added with 20%sphalerite pulp and inoculated for 40 d, mixed culture B,D and E had 2.10%,1.59%and 2.24%higher Zn²⁺ leaching rates in 9K medium and 2.03%,1.89%and 2.56%higher Zn²⁺ leaching rates in Leathen medium than the unmutated control.â‘£Mixed culture B could adjust to 4%-16%pulp of sphalerite,and mixed culture E could adjust to 32%pulp of sphalerite.These results indicate that B and E are more tolerant to high concentrations of sphalerite pulp,suggesting that they have brighter prospects on industrial applications than D.Diversity of microbial clones of CK,CKCu,Bcu and BZn were compared in this section.The results showed that the microbial populations of the four samples were categorized into three groups,i.e, Leptospirillum ferriphilum,Acidithiobacillus ferrooxidans,Thiobacillus thiooxidant and Acidithiobaeillus caldus.With the culture conditions varying,some changes may occur in the population structures of the mixed cultures.However,in the present study,the dominant microorganisms of the bioleaching systems were still Thiobacillus thiooxidant and Acidithiobacillus caldus.The growth quantity of Acidithiobacillus ferrooxidan among mixed microoganisms is the smallest before mutation.After mutation,its quantity ratio in the culture of chalcopyrite and sphalerite is 20%and 29%respectively of all microorganisms and it propably plays an important role in bioleachingThe growth quantity of Thiobacillus thiooxidant among mixed microoganisms is small before mutation.After mutation,its quantity ratio in the culture of chalcopyrite and sphalerite is 69%and 57% respectively of all microorganisms and it propably plays an important role in bioleaching.â‘ A strain of Leptospirillum ferriphilum was isolated,and characterized by sequence analysis of 16S rDNA.Its optium growth temperature was 40℃,and its optium growth start pH was 1.5.â‘¡A strain of Acidithiobacillus caldus was isolated,and characterized by sequence analysis of 16S rDNA.Its optium growth temperature was 45℃,and its optium growth start pH was 1.8. â‘¢A strain of Sulfobacillus thermosulfidooxidans was isolated,and characterized by sequence analysis of 16S rDNA.Its optium growth temperature was 53℃,and its optium growth start pH was 2.0.â‘£No.1 strain of Acidithiobacillus ferrooxidans and No.2 strain of Thiobacillus thiooxidant were isolated.