Enrichment,Isolation And Characterization Of Bacteria Oxidizing As(?) And Research Of Oxidation Efficiency For As(?) In Aqueous Environment

During this research,the samples were collected at the tailings and surrounding areas of the Aotao Shan mine in the Shexingshan town of Shuangfeng County,Loudi County,Hunan Province.The arsenic pollution was studied from these samples,in the meantime those also were used for the screening,separation and purification of As(III)-oxidizing microorganisms.In order to obtain the As(III)-oxidizing becteria for the remediation of acid mine wastewater especially for As(III)contaminated water.The main results are as follows:(1)Most of the soil samples were acidic according to the results of physicochemical property,the value of p H ranged between 2.82 to 5.60.The degree of heavy metal contaminations of each sample were different,and all the samples had various levels of arsenic pollution.Among these samples,No.5 and No.7 were the most serious arsenic contaminations,which were 373mg/kg and 119mg/kg respectively.After the cultivation of seven samples with autotrophic/heterotrophic culture conditions,the p H value in 3.0,5.0 and 6.0 under various acidic conditions,only 5 samples(No.1,No.2,No.5,No.6 and No.7)showed the positive results according to PDC(pyrrolidine dithiocarbamate)test.With all these positive samples for As(III)oxidation,No.1,No.5 and No.7 contained As(III)-oxidizing microorganisms living in both autotrophic and heterotrophic culture conditions,the As(III)-oxidizing microorganisms in No.2 and No.6 could only survive in heterotrophic culture conditions.The number of As(III)-oxidizing bacteria in the No.5 sample(p H value was 3.0)got the most,also with the highest proportion which was more than 10% in the total number.(2)The samples were incubated with autotrophic and heterotrophic conditions at different p H values of 3.0,5.0 and 6.0 respectively.A total of 8 different enriched samples were subjected to 3-4 times of separation and purification experiments.After PDC test for As(III)oxidization under the condition of p H 3.0,7 different single bacterial colonies were obtained from the sample No.1 with heterotrophic culture,5 different single bacterial colonies were obtained from the sample No.3 with autotrophic and heterotrophic cultures.Under the condition of p H 5.0,one single bacterial colony was obtained from the sample No.6 with heterotrophic culture.At the p H 6.0,65 different single bacterial colonies were obtained from the autotrophic culture and 84 colonies were obtained from heterotrophic culture in the sample No.7.Therefore,162 As(III)-oxidizing bacteria colonies were obtained from 5 arsenic-contaminated soil samples.(3)The DNA was analyzed by PCR-SSCP(Polymerase Chain Reaction and Single-Strand Conformation Polymorphism).From the results,there were 3 files were different from each other under the condition of p H 3.0,which means at least 3 different As(III)-oxidizing bacteria strains were obtained.As well,at least one different As(III)-oxidizing bacteria strain was obtained under the condition of p H 5.0 and 8 different As(III)-oxidizing bacteria strains were obtained at p H 6.0.In summary,at least 12 As(III)-oxidizing bacteria strains belonging to different species were obtained from both autotrophic and heterotrophic culture conditions.For the further analysis and identification of all of the above As(III)-oxidizing bacteria strains and the exploration of low cost additional carbon sources,and the engineering applications,will be in the follow-up study to continue to complete.(4)The growth rates of autotrophic As(III)-oxidizing strains and heterotrophic As(III)-oxidizing strains in disparate samples were different at p H 3.0,5.0 and 6.0 respectively.The growth rate of the strains was relatively fast under heterotrophic culture conditions.In the autotrophic condition,although the growth rate of the becteria was not positively correlated with p H value,the fastest growth rate among the strains was the one at p H 6.0.The concentration of As(III)ranged from 100 to 500 mg/L conditions,these growth conditions of two As(III)-oxidizing strains both were good: under 100mg/L arsenic concentration conditions,the vast majority of bacteria maintained activity and growed well;very few As(III)-oxidizing strain died even in 500 mg/L concentration.For the application of two As(III)-oxidizing strains from sample No.7,inoculated these strains into artificial contaminated arsenic wastewater ranging 100 to 500 mg/L concentration.The oxidation efficiency of As(III)was also very high,the oxidation efficiency is 100% when the concentration of arsenic is 100 and 200mg/L,and the oxidation efficiency could reach 50% even at 500mg/L.