Optimum Growth Condition Of Acidithiobacillus Spp. And Its Response To Toxic Metal Stress And Biodiversity During Sludge Bioleaching

Bioleaching is an attractive approach for sludge treatment by which sludge-born metals can be removed and sludge dewaterability can be drastically improved. The major microorganisms responsible for bioleaching are Acidithiobacillus thiooxidans and Acidithiobacillus ferrooxidans. It is very important to understand well the optimum condition of Acidithiobacilli bacteria and its resistance to toxic heavy metals during bioleaching.In the present study, Acidithiobacillus thiooxidans TS6 and Acidithiobacillus ferrooxidans LX5 were studied by pure culture method under different culture conditions. It was found that optimum temperature and initial pH in medium for strain TS6 growth were 28℃and pH 2.0～6.0, respectively, as exhibiting that the sulfur-oxidizing activity was highest under these conditions. For strain LX5,the optimum growth temperature and pH in medium were 28℃～32℃and pH 2.0～2.5,respectively, in which Fe²⁺ was finally oxidized into Fe³⁺ within 48 hours. Total DNA of strain TS6 and strain LX5 were extracted and amplified for bacterial 16S rDNA gene sequences. The purified PCR product was sequenced and deposited in the GenBank database with accession number DQ067562 & DQ067563. Phylogenesis of two strains was built in term of the most close related bacteria found in RDP database.Strain TS6 and LX5 were studied for their resistance to Cr³⁺, Cu²⁺, Zn²⁺, and Ni²⁺ by pure culture method. TS6 could resist the attack of at least 1500 mg·L^-1of Cr³⁺, 400 mg·L^-1 of Cu²⁺, 1000 mg·L·L^-1 of Zn²⁺, and 250 mg·L^-1of Ni²⁺ in medium. Resistance of strain LX5 to C³⁺, Zn²⁺, and Ni²⁺ were also high, but its growth was restrained when concentration of Cu²⁺ reached to 200 mg·'L^-1. It was observed by electronic microscope that strain TS6 produced a great amount of extracellar polymer subtances (EPS) at a rate of 1467μg/10¹⁰ cells when strain TS6 grew in the medium with 1500 mg·L^-1of Cr³⁺. EPS produced by strain TS6 could enhanced its resistance to heavy metals and consequently caused rapid drop of media pH.An efficient method named by indirect approach for DNA extraction from sludge microorganism was also established in this study. DGGE and ERIC-PCR were used to investigate the change of bacterial community at different stages of sludge bioleaching. The results showed that strain TS6 existed throughout bioleaching period from initial fresh sludge with pH7-8 to bioleached sludge with pH 1-2.So the strain TS6 is the main functional bacterial in bioleaching system. The change of structural dynamics of microbial community was noticeably, some signature DNA band whose intensity was correlated to efficiency of sludge was found. The findings about these bands can thus be used to isolate the potentially functional microorganism in bioleaching system.