Bioremediation of Heavy Metals by a Novel Bacterial Strain Enterobacter cloacae and Its Antioxidant Enzyme Activity, Flocculant Production, and Protein Expression in Presence of Lead, Cadmium, and Nickel

  • Goutam BanerjeeEmail author
  • Shubhant Pandey
  • Arun Kumar Ray
  • Ravi Kumar


This investigation reported the isolation and characterization of a potent heavy metal accumulating bacterial strain Enterobacter cloacae B1 from polluted soil at Ghaziabad, India. The minimum inhibitory concentration of the selected bacterial strain was recorded to be 1100 ppm for lead, 900 ppm for cadmium, and 700 ppm for nickel. Bioaccumulation of lead by this bacterial strain was extremely high (95.25 %), followed by cadmium (64.17 %) and nickel (36.77 %). Antioxidant enzymes such as catalase (CAT) and superoxide dismutase (SOD) activity was determined in presence of lead, cadmium, and nickel at a concentration of 400 ppm. To monitor the physiological stress, malondialdehyde (MDA) level was also estimated. In order to optimize the flocculant production, bacterial strain E. cloacae B1 was cultured in specific medium at different incubation period (24 to 72 h), pH (6.0 to 9.0), and temperature (20 to 50 °C). It was observed that surfactant production was maximum at 72 h of incubation period (47.28 %), pH 8.0 (56.63 %), and temperature 40 °C (62.94 %). Protein expression profile in presence of these heavy metals was also interesting. Few proteins were noticed to be overexpressed in presence of these heavy metals.


Enterobacter cloacae 16S rRNA Minimum inhibitory concentration Heavy metal accumulation Antioxidant enzymes Flocculant production Protein expression 



We are grateful to Helix Biogenesis for providing laboratory facilities and financial supports.


  1. Abassi, N. A., & Kushad, M. M. (1998). Active oxygen-scavenging enzymes activities in developing apple flowers and fruits. Scientia Horticulturae, 74, 183–194.CrossRefGoogle Scholar
  2. Aebi, H. (1984). Catalase in vitro. Methods in Enzymology, 105, 121–126.CrossRefGoogle Scholar
  3. Ahemad, M. (2012). Implication of bacterial resistance against heavy metals in bioremediation: a review. Institute of Integrative Omics and Applied Biotechnology Journal, 3(3), 39–46.Google Scholar
  4. Ahemad, M., & Malik, A. (2011). Bioaccumulation of heavy metals by zinc resistant bacteria isolated from agricultural soils and irrigated with wastewater. Bacteriology Journal, 2, 12–21.CrossRefGoogle Scholar
  5. Bailly, C., Leymarie, J., Lehner, A., Rousseau, S., Come, D., & Corbineau, F. (2004). Catalase activity and expression in developing sunflower seeds as related to drying. Journal of Experimental Botany, 55, 475–483.CrossRefGoogle Scholar
  6. Banerjee, G., Ray, A. K., Askarian, F., & Ringø, E. (2013). Characterization and identification of enzyme-producing autochthonous bacteria from the gastrointestinal tract of two Indian air-breathing fish. Beneficial Microbes, 4, 277–284.CrossRefGoogle Scholar
  7. Batta, N., Subudhi, S., Lal, B., & Devi, A. (2013). Isolation of lead tolerant novel species, Acinetobacter sp. TL-3: assessment of bioflocculating activity. Indian Journal of Experimental Biology, 51, 1004–1011.Google Scholar
  8. Bernhoft, R. A. (2013). Cadmium toxicity and treatment. The Scientific World Journal. doi: 10.1155/2013/394652.Google Scholar
  9. Beveridge, T. J., & Doyle, R. J. (1989). Metal ions and bacteria. New York: Wiley.Google Scholar
  10. Bodour, A. A., Drees, K. P., & Maier, R. M. (2003). Distribution of biosurfactant-producing bacteria in undisturbed and contaminated arid southwestern soils. Applied and Environmental Microbiology, 69(6), 3280–3287.CrossRefGoogle Scholar
  11. Bruins, M. R., Kapil, S., & Oehme, F. W. (2000). Microbial resistance to metals in the environment. Ecotoxicology and Environmental Safety, 45, 198–207.CrossRefGoogle Scholar
  12. Brüske-Hohlfeld, I. (2009). Environmental and occupational risk factors for lung cancer. Methods in Molecular Biology, 472, 3–23.CrossRefGoogle Scholar
  13. Chen, C. Y., & Lin, T. H. (1998). Nickel toxicity to human term placenta: in vitro study on lipid per oxidation. Journal of Toxicology and Environmental Health Part A, 54, 37–47.CrossRefGoogle Scholar
  14. Choudhary, M., Jetley, U. K., Khan, M. A., Zutshi, S., & Fatma, T. (2007). Effect of heavy metal stress on Proline, Malondialdehyde and Superoxide Dismutase activity in the Cyanobacterium spirulina platensis-S5. Ecotoxicology and Environmental Safety, 66, 204–209.CrossRefGoogle Scholar
  15. Deeb, B. E., & Altalhi, A. D. (2009). Degradative plasmid and heavy metal resistance plasmid naturally coexist in phenol and cyanide assimilating bacteria. American Journal of Biochemistry and Biotechnology, 5(2), 84–93.CrossRefGoogle Scholar
  16. Desai, J. D., & Banat, I. M. (1997). Microbial production of surfactants and their commercial potential. Microbiology and Molecular Biology Reviews, 61, 47–64.Google Scholar
  17. Dhail, S. (2013). Microbial enhanced oil recovery using potent biosurfactant produced by Pseudomonas sp. Arabian sea, Mumbai. Journal of Petroleum and Gas Engineering, 4(3), 57–60.Google Scholar
  18. Draper, H. H., & Hadley, M. (1990). Malondialdehyde determination as index of lipid peroxidation. Methods in Enzymology, 186, 421–423.CrossRefGoogle Scholar
  19. Etelvina, M. A. P. F., Ana Isabel, G. L., & Sofia Isabel, A. P. (2005). Cadmium tolerance plasticity in Rhizobium leguminoserum bv. Viciae: glutathione as a detoxifying agent. Canadian Journal of Microbiology, 51, 7–14.CrossRefGoogle Scholar
  20. Ewing, J. F., & Janero, D. R. (1995). Microplate superoxide dismutase assay employing a nonenzymatic superoxide generator. Analytical Biochemistry, 232, 243–248.CrossRefGoogle Scholar
  21. Fietcher, A. (1992). Biosurfactant: moving toward industrial application. Trends in Biotechnology, 10, 208–217.Google Scholar
  22. Georgiou, G., Lin, S. C., & Sharma, M. M. (1992). Surface active compounds from microorganisms. Biotechnology, 10(1), 60–65.CrossRefGoogle Scholar
  23. Goldstein, G. W. (1992). Neurological concepts of lead poisoning in children. Pediatric Annals, 21(6), 384–388.CrossRefGoogle Scholar
  24. Gupta, U. C., & Gupta, S. C. (1998). Trace element toxicity relationships to crop production and livestock and human health: implications for management. Communications in Soil Science and Plant Analysis, 29, 1491–1522.CrossRefGoogle Scholar
  25. Hassanshahian, M. (2014). Isolation and characterization of biosurfactant producing bacteria from Persian Gulf (Bushehr provenance), Marine Pollution Bulletin (
  26. Hookoom, M., & Puchooa, D. (2013). Isolation and identification of heavy metal tolerant bacteria from industrial and agricultural areas in Mauritius. Current Research in Microbiology and Biotechnology, 1(3), 119–123.Google Scholar
  27. Jiménez-Ortega, V., Barquilla, P. C., Fernández-Mateos, P., Cardinali, D. P., & Esquifino, A. I. (2012). Cadmium as an endocrine disruptor: correlation with anterior pituitary redox and circadian clock mechanisms and prevention by melatonin. Free Radical Biology and Medicine, 53(12), 2287–2297.CrossRefGoogle Scholar
  28. Johncy-Rani, M., Hemambika, B., Hemapriya, J., & Rajeshkannan, V. (2010). Comparative assessment of heavy metal removal by immobilized and dead bacterial cells: a biosorption approach. Global Journal of Environmental Research, 4(1), 23–30.Google Scholar
  29. Kafilzadeh, F., Afrough, R., Johari, H., & Tahery, Y. (2012). Range determination for resistance/tolerance and growth kinetic of indigenous bacteria isolated from lead contaminated soils near gas stations (Iran). European Journal of Experimental Biology, 2(1), 62–69.Google Scholar
  30. Kasprzak, K. S., Bal, W., & Karaczyn, A. A. (2003). The role of chromatin damage in nickel-induced carcinogenesis. A review of recent developments. Journal of Environmental Monitoring, 5, 183–187.CrossRefGoogle Scholar
  31. Kowall, N. W., Pendleury, W. W., & Kessler, J. B. (1989). Aluminium-induced neurofibrillary degeneration affects a subset of neurons in rabbit cerebral cortex, basal forebrain and upper brain stem. Neuroscience, 29, 329–337.CrossRefGoogle Scholar
  32. Lenaèrtovaè, V., Holovskaè, K., & Javorsky, P. (1998). The influence of mercury on the antioxidant enzyme activity of rumen bacteria Streptococcus bovis and Selenomonas ruminantium. FEMS Microbiology Ecology, 27, 319–325.CrossRefGoogle Scholar
  33. Master, C., Multhaup, G., & Simms, G. (1985). Neuronal origin of a cerebral amyloid: neurofibrillary tangles of Alzheimer’s disease contain the same protein as the amyloid of plaque cores and blood vessels. EMBO Journal, 4, 2757–2763.Google Scholar
  34. Mondal, S., Roy, T., & Ray, A. K. (2010). Characterization and identification of enzyme-producing bacteria isolated from the digestive tract of bata, Labeo bata. Journal of the World Aquaculture Society, 41, 369–377.CrossRefGoogle Scholar
  35. Nath, S., Deb, B., & Sharma, I. (2012). Isolation and characterization of cadmium and lead resistant bacteria. Global Advance Research Journal of Microbiology, 1(11), 194–198.Google Scholar
  36. Nies, D. (1999). Microbial heavy-metal resistance. Applied Microbiology and Biotechnology, 51, 730–750.CrossRefGoogle Scholar
  37. Pandey, S., Saha, P., Biswas, S., & Maiti, T. K. (2011). Characterization of two metals resistant Bacillus strains isolated from slag deposal site at Burnpur, India. Journal of Environmental Biology, 32, 773–779.Google Scholar
  38. Pandey, S., Barai, P. K., & Maiti, T. K. (2013). Influence of heavy metals on the activity of antioxidant enzymes in the metal resistant strains of Ochrobactrum and Bacillus sp. Journal of Environmental Biology, 34, 1033–1037.Google Scholar
  39. Poornima, M., Kumar, R. S., & Thomas, P. D. (2014). Isolation and molecular characterization of bacterial Strains from tannery effluent and reduction of chromium. International Journal of Current Microbiology and Applied Science, 3(4), 530–538.Google Scholar
  40. Ren, W. X., Li, P. J., Geng, Y., & Li, X. J. (2009). Biological leaching of heavy metals from a contaminated soil by Aspergillus niger. Journal of Hazardous Materials, 167(1–3), 164–169.CrossRefGoogle Scholar
  41. Schalk, I. J., Hannauer, M., & Braud, A. (2011). New roles of bacterial siderophores in metal transport and tolerance. Environmental Microbiology, 13, 2844–2854.CrossRefGoogle Scholar
  42. Selvi, A. T., Anjugam, E., Devi, R. A., Madhan, B., Kannappan, S., & Chandrasekaran, B. (2012). Isolation and characterization of bacteria from tannery effluent treatment plant and their tolerance to heavy metals and antibiotics. Asian Journal of Experimental Biological Sciences, 3(1), 34–41.Google Scholar
  43. Singh, A. K., Dhanjal, S., & Cameotra, S. S. (2014). Surfactin restores and enhances swarming motility under heavy metal stress. Colloids and Surfaces B: Biointerfaces, 116, 26–31.CrossRefGoogle Scholar
  44. Switala, J., & Leuwen, P. C. (2002). Diversity of properties among catalases. Archives of Biochemistry and Biophysics, 401, 145–154.CrossRefGoogle Scholar
  45. Thyssen, J. P., Gawkrodger, D. J., White, I. R., Julander, A., Menné, T., & Lidén, C. (2013). Coin exposure may cause allergic nickel dermatitis: a review. Contact Dermatitis, 68, 3–14.CrossRefGoogle Scholar
  46. Ugbenyen, A. M., & Okoh, A. I. (2013). Characteristics of a bioflocculant produced by a consortium of Cobetia and Bacillus species and its application in the treatment of waste waters. Water SA, 40(1), 139–1144.Google Scholar
  47. World Health Organization. (1995). Inorganic lead. Geneva: Environmental Health Criteria. No. 165.Google Scholar

Copyright information

© Springer International Publishing Switzerland 2015

Authors and Affiliations

  • Goutam Banerjee
    • 1
    Email author
  • Shubhant Pandey
    • 2
  • Arun Kumar Ray
    • 1
  • Ravi Kumar
    • 3
  1. 1.Department of ZoologyVisva-Bharati UniversitySantiniketanIndia
  2. 2.National Institute of Science Education and ResearchBhucneswarIndia
  3. 3.Helix BiogenesisNoidaIndia

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