Treatment Of Wastewater Containing Cr(Ⅵ) With Bacillus Thuringiensis

As a result of inappropriate waste-disposal of industries, significant amounts of chromium has been discharged into environment, causing serious environmental pollution. Reduction of Cr(VI) to Cr(III) is a potential useful process for remediation of Cr(VI)-affected environment. Bacillus thuringiensis (Bt) is an ideal material to removal Cr(VI) pollution with the merits of none pathogenicity and strong Cr(VI)-reducing capacity. Hence, 76 Bt strains form our lab were studied to reduce Cr(VI). The results showed Bt strains BRC-HZM7 and BRC-XQ15 could reduce chromium (VI) from 50 mg/L to <0.5 mg/L in 24 h, which reached Chinese standards for wastewater emission. It indicates that Bt has a great capacity for Cr(VI) reduction.To further characterize the factors affecting Cr(VI) reduction, a genome sequencing Bt 407 Cry- was exploited for its Cr(VI) reduction ability. The affecting factors, including initial Cr(VI) concentrations, initial cell concentrations, pH, temperatures, glucose and other heavy metals, on Cr(VI)-reduction were described. The results showed that the optimum inoculated cell concentration was 1%, the optimum initial pH value was 9.0 and the optimum temperature was 35oC. The addition of Mn2+, Mo2+, Cu2+, Ni2+and glucose promoted chromium-reduction while the reduction was inhibited by Zn2+ and Co2+. Bt 407 Cry- can reduce Cr(VI) from 50 mg/L to <0.5 mg/L in 36 h under the optimal condition, which reached Chinese standards for wastewater emission. After optimum, maximum reduction of Cr(VI) under the optimum condition (pH 9.0 and 35 oC) was 9.25 times what it was before optimum.By genome sequence analysis, Bt 407 Cry- was found to contain some Cr(VI) reduction genes, which may be responsible for the observed rapid chromate reduction. Furthermore, a chrA gene encoding a putative chromate transporter conferring chromate resistance was also identified. Genes regulating Cr(VI) reduction were further investigated by mini-Tn10 transposon mutation. The vector pIC333 was transformed into Bt 407 Cry- and generated a library of Bt 407 Cry- random insertion mutants, which capacity of 1500. We selected 9 mutants which Cr(VI)-reducing capacity are extremely higher than that of Bt 407 Cry- (p<0.01), were designated as Bt 407-Cr5, Bt 407-Cr15, Bt407-Cr22, Bt407-Cr90, Bt407-Cr108, Bt407-Cr215, Bt 407-Cr259, Bt 407-Cr279 and Bt 407-Cr281. Total DNA of the mutants were digested with HindⅢcompletely, respectively. Then the digested DNA was self-ligated overnight at 16℃and transformed into competent Escherichia coli TG1 cells. The insert sites were all defined as conjunction protein gene. Furthermore, the results of phenotype analysis of the 9 mutants showed that the growth curve of Bt 407 Cry- was not different from the mutants. This indicated the difference of Cr(VI)-reducing capacity between Bt 407 Cry- and its mutants was not because of the amount of bacteria. And the results of total Cr analysis revealed that a maximum of 166 mg/L Cr(VI) was reduced in 24 h, whereas the values of total Cr was almost the same throughout the period of Cr(VI)-reduction. This suggested the main mode of action of Bt 407 Cry- was reduction.