Browse > Article
http://dx.doi.org/10.4014/jmb.0906.06026

Structure and Diversity of Arsenic-Resistant Bacteria in an Old Tin Mine Area of Thailand  

Jareonmit, Pechrada (Department of Soil Science, Kasetsart University)
Sajjaphan, Kannika (Department of Soil Science, Kasetsart University)
Sadowsky, Michael J. (Department of Soil, Water, and Climate, and Bio Technology Institute, University of Minnesota)
Publication Information
Journal of Microbiology and Biotechnology / v.20, no.1, 2010 , pp. 169-178 More about this Journal
Abstract
The microbial community structure in Thailand soils contaminated with low and high levels of arsenic was determined by denaturing gradient gel electrophoresis. Band pattern analysis indicated that the bacterial community was not significantly different in the two soils. Phylogenetic analysis obtained by excising and sequencing six bands indicated that the soils were dominated by Arthrobacter koreensis and $\beta$-Proteobacteria. Two hundred and sixty-two bacterial isolates were obtained from arsenic-contaminated soils. The majority of the As-resistant isolates were Gramnegative bacteria. MIC studies indicated that all of the tested bacteria had greater resistance to arsenate than arsenite. Some strains were capable of growing in medium containing up to 1,500 mg/l arsenite and arsenate. Correlations analysis of resistance patterns of arsenite resistance indicated that the isolated bacteria could be categorized into 13 groups, with a maximum similarity value of 100%. All strains were also evaluated for resistance to eight antibiotics. The antibiotic resistance patterns divided the strains into 100 unique groups, indicating that the strains were very diverse. Isolates from each antibiotic resistance group were characterized in more detail by using the repetitive extragenic palindromic-PCR (rep-PCR) DNA fingerprinting technique with ERIC primers. The PCR products were analyzed by agarose gel electrophoresis. The genetic relatedness of 100 bacterial fingerprints, determined by using the Pearson product-moment similarity coefficient, showed that the isolates could be divided into four clusters, with similarity values ranging from 5-99%. Although many isolates were genetically diverse, others were clonal in nature. Additionally, the arsenic-resistant isolates were examined for the presence of arsenic resistance (ars) genes by using PCR, and 30% of the isolates were found to carry an arsenate reductase encoded by the arsC gene.
Keywords
Denaturing gradient gel electophoresis (DGGE); arsenic-resistant bacteria; repetitive extragenic palindromic-PCR (rep-PCR); DNA fingerprinting;
Citations & Related Records

Times Cited By Web Of Science : 4  (Related Records In Web of Science)
연도 인용수 순위
  • Reference
1 Chang, J. S., Y. H. Kim, and K. W. Kim. 2008. The ars genotype characterization of arsenic-resistant bacteria from arsenic-contaminated gold-silver mines in the Republic of Korea. Appl. Microbiol. Biotechnol. 80: 155-165.   DOI   ScienceOn
2 Bennett, R. L. and M. H. Malamy. 1970. Arsenate resistant mutants of Escherichia coli and phosphate transport. Biochem. Biophys. Res. Commun. 40: 496-503.   DOI   ScienceOn
3 Butcher, B. G., S. M. Deane, and D. E. Rawlings. 2000. The chromosomal arsenic resistance genes of Thiobacillus ferrooxidans have an unusual arrangement and confer increased arsenic and antimony resistance to Escherichia coli. Appl. Environ. Microbiol. 66: 1826-1833.   DOI   ScienceOn
4 Gelsomino, A., A. C. Keijzer-Wolters, G. Cacco, and J. D. Van Elsas. 1999. Assessment of bacterial community structure in soil by polymerase chain reaction and denaturing gradient gel electrophoresis. J. Microbiol. Methods 38: 1-15.   DOI
5 Santini, J. M. and R. N. Vanden Hoven. 2004. Molybdenum-containing arsenite oxidase of the chemolithoautotrophic arsenite oxidizer NT-26. J. Bacteriol. 186: 1614-1619.   DOI   ScienceOn
6 Amacher, M. C. 1996. Nickel, cadmium and lead, pp. 739-768. In D. L. Spark, A. L. Page, P. A. Helmke, R. H. Loeppert, P. N. Soltanpour, M. A. Takatabai, C. T. Johnson, and M. E. Summer (eds.). Method of Soil Analysis Part 3: Chemical Methods. Soil Science Society of America Inc., Wisconsin, WI.
7 Cai, J., K. Salmon, and M. S. DuBow. 1998. A chromosomal ars operon homologue of Pseudomonas aeruginosa confers increased resistance to arsenic and antimony in Escherichia coli. Microbiology 144: 2705-2713.   DOI   ScienceOn
8 Cervantes, C., G. Ji, J. L. Ramirez, and S. Silver. 1994. Resistance to arsenic compounds in microorganisms. FEMS Microbiol. Rev. 15: 355-367.
9 Da Costa, E. W. B. 1972. Variation in the toxicity of arsenic compounds to microorganisms and the suppression of the inhibitory effects of phosphate. Appl. Environ. Microbiol. 23: 46-53.
10 Ehrlich, H. L. 1996. Geomicrobial interactions with arsenic and antimony, pp. 276-293. In H. L. Ehrlich (ed.), Geomicrobiology, 3rd Ed. Marcel Dekker Inc., New York, NY.
11 Ford, T., J. Jay, A. Patel, M. Kile, P. Prommasith, T. Galloway, R. Sanger, K. Smith, and M. Depledge. 2005. Use of ecotoxicological tools to evaluate the health of New Bedford Harbor sediments: A microbial biomarker approach. Environ. Health Perspect. 113: 186-191.
12 Fordyce, F., M. Williams, A. Paijitprapapon, and P. Charoenchaisri. 1995. Hydrogeochemistry of arsenic in an area of chronic mining-related arsenic, Ron Phibun District, Nakhon si Thammarat Province, Thailand: Preliminary results. BGS Technical Report WC/94/79R.
13 Morases, S. R., R. B. Goncalves, C. Mouton, L. Seldin, M. C. S. Ferreira, and R. M. C. P. Domingues. 2000. Use of rep-PCR to define genetic relatedness among Bacteroides fragilis strains. J. Med. Microbiol. 49: 279-284.
14 Oden, K. L., T. B. Gladysheva, and B. P. Rosen. 1994. Arsenate reduction mediated by the plasmid-encoded ArsC protein is coupled to glutathione. Mol. Microbiol. 12: 301-306.   DOI   ScienceOn
15 Pontius, F., K. G. Brown, and C. J. Chen. 1994. Health implications of arsenic in drinking water. J. Am. Water Works Assoc. 86: 52-63.
16 Rosenstein, R., A. Peschel, B. Wieland, and F. Gotz. 1992. Expression and regulation of the antimonite, arsenite, and arsenate resistance operon of Staphylococcus xylosus plasmid pSX267. J. Bacteriol. 174: 3676-3683.
17 Saltikov, C. W. and B. H. Olson. 2002. Homology of Escherichia coli R773 arsA, arsB, and arsC genes in arsenic-resistant bacteria isolated from raw sewage and arsenic-enriched creek waters. Appl. Environ. Microbiol. 68: 280-288.   DOI   ScienceOn
18 Santini, J. M., L. I. Sly, R. D. Schnagl, and J. M. Macy. 2000. A new chemolithotrophic arsenite-oxidising bacterium isolated from a goldmine: Phylogenetic, physiological and preliminary biochemical studies. Appl. Environ. Microbiol. 66: 92-97.   DOI   ScienceOn
19 Silver, S. and L. T. Phung. 1996. Bacterial heavy metal resistance: New surprises. Annu. Rev. Microbiol. 50: 753-789.   DOI   ScienceOn
20 Silver, S. and L. T. Phung. 2005. Genes and enzymes involved in bacterial oxidation and reduction of inorganic arsenic. Appl. Environ. Microbiol. 71: 599-608.   DOI   ScienceOn
21 Williams, M. 1997. Mining-related arsenic hazards: Thailand case study. Technical Report WC/97/490. British Geological Survey.
22 Butcher, B. G. and D. E. Rawlings. 2002. The divergent chromosomal ars operon of Acidithiobacillus ferrooxidans is regulated by an atypical ArsR protein. Microbiology 148: 3983-3992.
23 Cindy, H. N., V. Torsvik, and L. Ovreas. 2000. Soil community analysis using DGGE of 16S rDNA polymerase chain reaction products. Soil Sci. Soc. Am. J. 64: 1382-1388.   DOI
24 Nakatsu, C. H. 2007. Soil microbial community analysis using denaturing gradient gel electrophoresis. Soil Sci. Soc. Am. J. 71: 562-571.   DOI
25 Stewart, J. W. B. and J. R. Bettany. 1982. Mercury, pp. 367-384. In A. L. Page (ed.). Method of Soil Analysis Part 2: Chemical and Microbiological Properties, 2nd Ed. Soil Science Society of America Inc., Wisconsin, WI.
26 Ji, G., E. A. Garber, L. G. Armes, C.-M. Chen, J. A. Fuchs, and S. Silver. 1994. Arsenate reductase of Staphylococcus aureus plasmid pI258. Biochemistry 33: 7294-7299.   DOI   ScienceOn
27 Silver, S. and M. Walderhaug. 1992. Gene regulation of plasmid and chromosome determined inorganic ion transport in bacteria. Microbiol. Rev. 56: 1-33.
28 Williams, M., F. Fordyce, A. Paijitprapapon, and P. Charoenchaisri. 1996. Arsenic contamination in surface drainage and groundwater in part of the southeast Asian tin belt, Nakhon Si Thammarat, Southern Thailand. Environ. Geol. 27: 16-33.   DOI   ScienceOn
29 Becker, J. M., T. Parkin, H. N. Cindy, D. W. Jayson, and A. Konopka. 2006. Bacterial activity, community structure and centimeter-scale spatial heterogeneity in contaminated soil. Microbiol. Ecol. 51: 220-231.   DOI   ScienceOn
30 Anderson, C. R. and G. M. Cook. 2004. Isolation and characterization of arsenate-reducing bacteria from arsenic-contaminated sites in New Zealand. Curr. Microbiol. 48: 341-347.   DOI   ScienceOn
31 Dombek, P. E., L. K. Johnson, S. T. Zimmerley, and M. J. Sadowsky. 2000. Use of repetitive DNA sequences and the PCR to differentiate Escherichia coli isolates from human and animal sources. Appl. Environ. Microbiol. 66: 2572-2577.   DOI   ScienceOn
32 Loynachan, T. E. 2002. Laboratory Manual for Agronomy: Soil Microbial Ecology. Iowa State University, Iowa.
33 Sato, T. and Y. Kobayashi. 1998. The ars operon in the skin element of Bacillus subtilis confers resistance to arsenate and arsenite. J. Bacteriol. 180: 1655-1661.
34 Abdrashitova, S. A., G. G. Abdullina, and A. N. Ilialetdinov. 1986. Role of arsenites in lipid peroxidation in Pseudomonas putida cells oxidizing arsenite. Mikrobiologiya 55: 212-216.
35 Nelson, D. W. and L. E. Sommers. 1982. Total carbon, organic carbon and organic matter, pp. 570-572. In A. L. Page (ed.). Method of Soil Analysis Part 2: Chemical and Microbiological Properties, 2nd Ed. American Society Agronomy Inc., Wisconsin. WI.
36 Tisa, L. S. and B. P. Rosen. 1990. Molecular characterization of an anion pump: The ArsB protein is the membrane anchor for the ArsA protein. J. Biol. Chem. 265: 190-194.
37 Zwart, G. and J. Bok. 2002. Protocol DGGE. Available at .
38 Chen, C. M., T. K. Misra, S. Silver, and B. P. Rosen. 1986. Nucleotide sequence of the structural genes for an anion pump: The plasmid-encoded arsenical resistance operon. J. Biol. Chem. 261: 15030-15038.
39 Lopez-Maury, L., F. J. Florencio, and J. C. Reyes. 2003. Arsenic sensing and resistance system in the cyanobacterium Synechocystis sp. strain PCC 6803. J. Bacteriol. 185: 5363-5371.   DOI   ScienceOn
40 Anderson, G. L., J. Williams, and R. Hille. 1992. The purification and characterisation of the arsenite oxidase from Alcaligenes faecalis, a molybdenum-containing hydroxylase. J. Biol. Chem. 267: 23674-23682.
41 Muller, D., D. Lievremont, D. D. Simeonova, J. C. Hubert, and M. C. Lett. 2003. Arsenite oxidase aox genes from a metal-resistant ${\beta}$-Proteobacterium. J. Bacteriol. 185: 135-141.   DOI   ScienceOn
42 Suzuki, K., N. Wakao, T. Kimura, K. Sakka, and K. Ohmiya. 1998. Expression and regulation of the arsenic resistance operon of Acidiphilium multivorum AIU 301 plasmid pKW301 in Escherichia coli. Appl. Environ. Microbiol. 64: 411-418.
43 Johnson, L. K., M. B. Brown, E. A. Carruthers, J. A. Ferguson, P. E. Dombek, and M. J. Sadowsky. 2004. Sample size, library composition and genotypic diversity among natural populations of Escherichia coli from different animals influence accuracy of determining sources of fecal pollution. Appl. Environ. Microbiol. 70: 4478-4485.   DOI   ScienceOn
44 Diorio, C., J. Cai, J. Marmor, R. Shinder, and M. S. DuBow. 1995. An Escherichia coli chromosomal ars operon homolog is functional in arsenic detoxification and is conserved in Gramnegative bacteria. J. Bacteriol. 177: 2050-2056.
45 Gladysheva, T. B., K. L. Oden, and B. P. Rosen. 1994. Properties of the arsenate reductase of plasmid R773. Biochemistry 33: 7288-7293.   DOI   ScienceOn
46 Macy, J. M., K. Nunan, K. D. Hagen, D. R. Dixon, P. J. Harbour, M. Cahill, and L. I. Sly. 1996. Chrysiogenes arsenatis gen. nov., sp. nov., a new arsenate respiring bacterium isolated from gold mine wastewater. Int. J. Syst. Bacteriol. 46: 1153-1157.   DOI   ScienceOn
47 Fournier, P. E., D. Vallenet, V. Barbe, S. Audic, H. Ogata, L. Poirel, et al. 2006. Comparative genomics of multidrug resistance in Acinetobacter baumannii. PLOS Genet. 2: e7.   DOI
48 Kaur, P. and B. P. Rosen. 1992. Plasmid-encoded resistance to arsenic and antimony. Plasmid 28: 29-40.
49 Torsvik, V., R. Sorheim, and J. Goksoyr. 1996. Total bacterial diversity in soil and sediment communities: A review. J. Ind. Microbiol. 17: 170-178.   DOI
50 Cullen, W. R. and K. J. Reimer. 1898. Arsenic speciation in the environment. Chem. Rev. 89: 713-764.
51 Gee, G. W. and J. W. Bauder. 1986. Particle-size analysis, pp. 399-404. In A. Klute (ed.). Method of Soil Analysis Part 1: Physical and Mineralogical Methods, 2nd Ed. American Society Agronomy Inc., Wisconsin, WI.
52 Sun, Y., E. A. Polishchuk, U. Radoja, and W. R. Cullen. 2004. Identification and quantification of arsC genes in environmental samples by using realtime PCR. J. Microbiol. Methods 58: 335-349.   DOI   ScienceOn
53 Quinn, J. P. and G. McMullan. 1995. Carbon-arsenic bond cleavage by a newly isolated Gram-negative bacterium, strain ASV2. Microbiology 141: 721-725.   DOI
54 Gans, J., M. Wolinsky, and J. Dunbar. 2005. Computational improvements reveal great bacterial diversity and high metal toxicity in soil. Science 309: 1387-1390.   DOI   ScienceOn
55 Ji, G. and S. Silver. 1992. Reduction of arsenate to arsenite by the ArsC protein of the arsenic resistance operon of Staphylococcus aureus plasmid pI258. Proc. Natl. Acad. Sci. U.S.A. 89: 9474-9478.   DOI   ScienceOn
56 Newman, D. K., E. K. Kennedy, J. D. Coates, D. Ahmann, D. Ellis, J. D. R. Lovley, and F. M. M. Morel. 1997. Dissimilatory arsenate and sulfate reduction in Desulfotomaculum auripigmentum sp. nov. Arch. Microbiol. 168: 380-388.   DOI   ScienceOn
57 Tamaki, S. and W. T. Frankenberger. 1992. Environmental biochemistry of arsenic. Rev. Environ. Contam. Toxicol. 124: 79-110.   DOI
58 Ahmann, D., A. L. Roberts, L. R. Krumholz, and F. M. M. Morel. 1994. Microbe grows by reducing arsenic. Nature 370: 750.
59 Ellis, P. J., T. Conrads, R. Hille, and P. Kuhn. 2001. Crystal structure of the 100 kDa arsenite oxidase from Alcaligenes faecalis in two crystal forms at 1.64 ${\AA}$ and 2.03 ${\AA}$. Structure 9: 125-132.   DOI
60 Macur, R. E., C. R. Jackson, L. M. Botero, T. R. McDermott, and W. P. Inskeep. 2004. Bacterial populations associated with the oxidation and reduction of arsenic in an unsaturated soil. Environ. Sci. Technol. 38: 104-111.   DOI   ScienceOn