In vitro and In silico analysis of heavy metal-resistant bacteria from cooum river for bioremediation approaches

Main Article Content

Sugitha S
Abirami G

Keywords

Bioremediation, Metal tolerance, BLAST analysis, Ralstonia pickettii, Pseudomonas otitidis, Antibiotic resistance

Abstract

The world is facing a significant deficit in the quality and quantity of freshwater as a result of the contamination of rivers. Heavy metals are the most common pollutants in water bodies. In this study, the new discovery of bacterial strains from the Cooum river sample can be used to remove heavy metals and radioactive compounds from water sources through heavy metal bioremediation. The bacterial strains from the Cooum sample were isolated and identified using 16S rRNA gene sequencing and nucleotide BLAST analysis. Considering copper and lead as toxic metals, metal tolerance, Maximum Tolerance concentration (MTC), and antibiotic susceptibility tests were examined. Identification of genes associated with metal resistant mechanism and comparative analysis using In Silico studies. The results presented in this study support the concept that the two bacterial strains namely Ralstonia pickettii and Pseudomonas otitidis have significant bioremediation potential. In addition, a positive correlation has been found between metals and antibiotic resistance in the bacterial strains and the proteins and genes involved in the resistance of heavy metals were identified. This novel study paves a new way to remove toxic substances and heavy metals contaminated in water bodies. It leads researchers to identify genes that help to magnify the possibilities for microbial bioremediation.

Abstract 166 | pdf Downloads 198

References

1. R. N. Kumar, R. Solanki, and J. I. N. Kumar, “Seasonal variation in heavy metal contamination in water and sediments of river Sabarmati and Kharicut canal at Ahmedabad, Gujarat,” Environ Monit Assess, vol. 185, no. 1, pp. 359–368, Jan. 2013, doi: 10.1007/s10661-012-2558-4.
2. A. Ramadan, “Heavy Metal Pollution and Biomonitoring Plants in Lake Manzala, Egypt,” Pakistan Journal of Biological Sciences, vol. 6, no. 13, pp. 1108–1117, Jun. 2003, doi: 10.3923/pjbs.2003.1108.1117.
3. V. Masindi and K. L. Muedi, “Environmental Contamination by Heavy Metals,” in Heavy Metals, InTech, 2018. doi: 10.5772/intechopen.76082.
4. N. Shikazono, H. M. Zakir, and Y. Sudo, “Zinc contamination in river water and sediments at Taisyu Zn–Pb mine area, Tsushima Island, Japan,” J Geochem Explor, vol. 98, no. 3, pp. 80–88, Sep. 2008, doi: 10.1016/j.gexplo.2007.12.002.
5. W. Maret, “Zinc and Human Disease,” 2013, pp. 389–414. doi: 10.1007/978-94-007-7500-8_12.
6. L. Järup, “Hazards of heavy metal contamination,” Br Med Bull, vol. 68, no. 1, pp. 167–182, Dec. 2003, doi: 10.1093/bmb/ldg032.
7. Z. Hussain and K. M. M. Sheriff, “ Status of heavy metal concentrations in groundwater samples situated in and around on the bank of Cooum river at Chennai City, Tamil Nadu.,” Journal of Chemical and Pharmaceutical Research , vol. 5, pp. 73–77, 2013.
8. Finny and Abraham, “Variation of water quality across Cooum river in Chennai city,” vol. 2, p. 51, 2012.
9. Duraisamy and S. Latha, “Impact of pollution on marine environment -A case study of coastal Chennai ,” Indian J Sci Technol, vol. 4, no. 3, 2011.
10. T.R. Kalaivani and M.S. Dheenadayalan, “Seasonal fluctuation of Heavy Metal Pollution in Surface water,” International Research Journal of Environment Sciences, vol. 2, pp. 66–73, 2003.
11. J. Azariah and Azariah.H, “Let the fishes live in Cooum,” vol. 3, pp. 12–19, 1987.
12. N. S. Elangovan and M. Dharmendirakumar, “Assessment of Groundwater Quality along the Cooum River, Chennai, Tamil Nadu, India,” JChem, vol. 2013, pp. 1–10, 2013, doi: 10.1155/2013/672372.
13. S. Aishwarya et al., “Structural, functional, resistome and pathogenicity profiling of the Cooum river,” Microb Pathog, vol. 158, p. 105048, Sep. 2021, doi: 10.1016/j.micpath.2021.105048.
14. P. A. Sahayaraj and K. Ayyadurai, “Bioaccumulation of lead in milk of buffaloes from Cooum River Belt in Chennai.,” J Environ Biol, vol. 30, no. 5, pp. 651–4, Sep. 2009.
15. Dhamodharan, S. Abinandan, U. Aravind, G. P. Ganapathy, and S. Shanthakumar, “Distribution of Metal Contamination and Risk Indices Assessment of Surface Sediments from Cooum River, Chennai, India,” Int J Environ Res, vol. 13, no. 5, pp. 853–860, Oct. 2019, doi: 10.1007/s41742-019-00222-8.
16. M. J. BUNCH, “Soft Systems Methodology and the Ecosystem Approach: A System Study of the Cooum River and Environs in Chennai, India,” Environ Manage, vol. 31, no. 2, pp. 182–197, Feb. 2003, doi: 10.1007/s00267-002-2721-8.
17. E. Vanchhawng and S. Jayaraj, “GENOTOXIC POTENTIAL OF CHANNA PUNCTATUS ON RIVER COOUM, CHENNAI,” 2012.
18. F. Masood and A. Malik, “Current Aspects of Metal Resistant Bacteria in Bioremediation: From Genes to Ecosystem,” in Management of Microbial Resources in the Environment, Dordrecht: Springer Netherlands, 2013, pp. 289–311. doi: 10.1007/978-94-007-5931-2_11.
19. M. R. Bruins, S. Kapil, and F. W. Oehme, “Microbial Resistance to Metals in the Environment,” Ecotoxicol Environ Saf, vol. 45, no. 3, pp. 198–207, Mar. 2000, doi: 10.1006/eesa.1999.1860.
20. T. Wadia and S. K. Jain, “Isolation, Screening and Identification of Lipase Producing Fungi from Oil Contaminated Soil of Shani Mandir Ujjain,” Int J Curr Microbiol Appl Sci, vol. 6, no. 7, pp. 1872–1878, Jul. 2017, doi: 10.20546/ijcmas.2017.607.223.
21. B. Neeta, V. Maansi, and S. B. Harpreet, “Characterization of heavy metal (cadmium and nickle) tolerant Gram negative enteric bacteria from polluted Yamuna River, Delhi,” Afr J Microbiol Res, vol. 10, no. 5, pp. 127–137, Feb. 2016, doi: 10.5897/AJMR2015.7769.
22. H. Hussein and H. Moawad, “Isolation and characterization of Pseudomonas resistant to heavy metals contaminants ,” vol. 7, pp. 11–22, 2003.
23. K. Ghaima, A. Mohamed, and W. Yehia, “ Resistance and bioadsorption of Cadmium by Pseudomonas aeruginosa isolated from
agricultural soil,” International Journal of Applied Environmental Sciences, vol. 12, 2017.
24. B. Yamina, B. Tahar, and F. Marie Laure, “Isolation and screening of heavy metal resistant bacteria from wastewater: a study of heavy metal co-resistance and antibiotics resistance,” Water Science and Technology, vol. 66, no. 10, pp. 2041–2048, Nov. 2012, doi: 10.2166/wst.2012.355.
25. L. W. Marzan, M. Hossain, S. A. Mina, Y. Akter, and A. M. M. A. Chowdhury, “Isolation and biochemical characterization of heavy-metal resistant bacteria from tannery effluent in Chittagong city, Bangladesh: Bioremediation viewpoint,” The Egyptian Journal of Aquatic Research, vol. 43, no. 1, pp. 65–74, Mar. 2017, doi: 10.1016/j.ejar.2016.11.002.
26. Rajbanshi, “Study on Heavy Metal Resistant Bacteria in Guheswori Sewage Treatment Plant,” Our Nature, vol. 6, no. 1, pp. 52–57, Mar. 2009, doi: 10.3126/on.v6i1.1655.
27. M. U. Mustapha and N. Halimoon, “Screening and Isolation of Heavy Metal Tolerant Bacteria in Industrial Effluent,” Procedia Environ Sci, vol. 30, pp. 33–37, 2015, doi: 10.1016/j.proenv.2015.10.006.
28. L. Ouyang et al., “A study on the nitrogen removal efficacy of bacterium Acinetobacter tandoii MZ-5 from a contaminated river of Shenzhen, Guangdong Province, China,” Bioresour Technol, vol. 315, p. 123888, Nov. 2020, doi: 10.1016/j.biortech.2020.123888.
29. C. Jiang, X. Sheng, M. Qian, and Q. Wang, “Isolation and characterization of a heavy metal-resistant Burkholderia sp. from heavy metal-contaminated paddy field soil and its potential in promoting plant growth and heavy metal accumulation in metal-polluted soil,” Chemosphere, vol. 72, no. 2, pp. 157–164, May 2008, doi: 10.1016/j.chemosphere.2008.02.006.
30. G. Roeselers et al., “Evidence for a core gut microbiota in the zebrafish,” ISME J, vol. 5, no. 10, pp. 1595–1608, Oct. 2011, doi: 10.1038/ismej.2011.38.
31. K. Sunda and R. Vidya, “High Chromium Tolerant Bacterial Strains from Palar River Basin: Impact ofTannery Pollution,” Research Journal of Environmental and Earth Sciences, vol. 2, pp. 112–117, 2010.
32. Wiegand, K. Hilpert, and R. E. W. Hancock, “Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances,” Nat Protoc, vol. 3, no. 2, pp. 163–175, Feb. 2008, doi: 10.1038/nprot.2007.521.
33. S. Abbasiliasi and T. I. Azmi, “Antimicrobial Activity and Antibiotic Sensitivity of Three Isolates of Lactic Acid Bacteria From Fermented Fish Product, Budu,” Malays J Microbiol, vol. 5, pp. 33–37, 2009.
34. C. Edward Raja And G. S. Selvam, “Isolation, Identification And Characterization Of Heavy Metal Resistant Bacteria From Sewage,” International joint symposium on Geodisaster prevention and Geoenvironment in Asia, 2009.
35. D. R. VanDevanter, J. M. van Dalfsen, J. L. Burns, and N. Mayer-Hamblett, “In Vitro Antibiotic Susceptibility of Initial Pseudomonas aeruginosa Isolates From United States Cystic Fibrosis Patients,” J Pediatric Infect Dis Soc, vol. 4, no. 2, pp. 151–154, Jun. 2015, doi: 10.1093/jpids/pit052.
36. S. ben MILOUD et al., “First Description of Various Bacteria Resistant to Heavy Metals and Antibiotics Isolated from Polluted Sites in Tunisia,” Pol J Microbiol, vol. 70, no. 2, pp. 161–174, Jun. 2021, doi: 10.33073/pjm-2021-012.
37. Z.-H. Cui et al., “Rapid Screening of Essential Oils as Substances Which Enhance Antibiotic Activity Using a Modified Well Diffusion Method,” Antibiotics, vol. 10, no. 4, p. 463, Apr. 2021, doi: 10.3390/antibiotics10040463.
38. W. M. N. H. Kumari, S. Thiruchittampalam, M. S. S. Weerasinghe, N. V. Chandrasekharan, and C. D. Wijayarathna, “Characterization of a Bacillus megaterium strain with metal bioremediation potential and In silico discovery of novel cadmium binding motifs in the regulator, CadC,” Appl Microbiol Biotechnol, vol. 105, no. 6, pp. 2573–2586, Mar. 2021, doi: 10.1007/s00253-021-11193-2.
39. P. C. Bull and D. W. Cox, “Wilson disease and Menkes disease: new handles on heavy-metal transport,” Trends in Genetics, vol. 10, no. 7, pp. 246–252, Jul. 1994, doi: 10.1016/0168-9525(94)90172-4.
40. K. D. Pruitt, T. Tatusova, and D. R. Maglott, “NCBI reference sequences (RefSeq): a curated non-redundant sequence database of genomes, transcripts and proteins,” Nucleic Acids Res, vol. 35, no. Database, pp. D61–D65, Jan. 2007, doi: 10.1093/nar/gkl842.
41. R. Khan, G.-S. Park, S. Asaf, S.-J. Hong, B. K. Jung, and J.-H. Shin, “Complete genome analysis of Serratia marcescens RSC-14: A plant growth-promoting bacterium that alleviates cadmium stress in host plants,” PLoS One, vol. 12, no. 2, p. e0171534, Feb. 2017, doi: 10.1371/journal.pone.0171534.
42. O. Mackenzie and G. Grigoryan, “Protein structural motifs in prediction and design,” Curr
Opin Struct Biol, vol. 44, pp. 161–167, Jun. 2017, doi: 10.1016/j.sbi.2017.03.012.
43. Klonowska et al., “Novel heavy metal resistance gene clusters are present in the genome of Cupriavidus neocaledonicus STM 6070, a new species of Mimosa pudica microsymbiont isolated from heavy-metal-rich mining site soil,” BMC Genomics, vol. 21, no. 1, p. 214, Dec. 2020, doi: 10.1186/s12864-020-6623-z.
44. Pal, J. Bengtsson-Palme, C. Rensing, E. Kristiansson, and D. G. J. Larsson, “BacMet: antibacterial biocide and metal resistance genes database,” Nucleic Acids Res, vol. 42, no. D1, pp. D737–D743, Jan. 2014, doi: 10.1093/nar/gkt1252.
45. Liu and M. Pop, “ARDB--Antibiotic Resistance Genes Database,” Nucleic Acids Res, vol. 37, no. Database, pp. D443–D447, Jan. 2009, doi: 10.1093/nar/gkn656.
46. S. Kumar, M. Nei, J. Dudley, and K. Tamura, “MEGA: A biologist-centric software for evolutionary analysis of DNA and protein sequences,” Brief Bioinform, vol. 9, no. 4, pp. 299–306, Mar. 2008, doi: 10.1093/bib/bbn017.
47. Saitou N and Nei M, “The neighbor-joining method: a new method for reconstructing phylogenetic trees.,” Mol Biol Evol, Jul. 1987, doi: 10.1093/oxfordjournals.molbev.a040454.
48. Bergey DH and Holt JG, “Bergey’s Manual of Determinative Bacteriology,” no. 9, 2000.
49. Kalsoom et al., “Isolation and screening of chromium resistant bacteria from industrial waste for bioremediation purposes,” Brazilian Journal of Biology, vol. 83, 2023, doi: 10.1590/1519-6984.242536.
50. M. M. Kabir, A. N. M. Fakhruddin, M. A. Z. Chowdhury, Md. K. Pramanik, and Z. Fardous, “Isolation and characterization of chromium(VI)-reducing bacteria from tannery effluents and solid wastes,” World J Microbiol Biotechnol, vol. 34, no. 9, p. 126, Sep. 2018, doi: 10.1007/s11274-018-2510-z.
51. M. RYAN, J. PEMBROKE, and C. ADLEY, “Ralstonia pickettii: a persistent Gram-negative nosocomial infectious organism,” Journal of Hospital Infection, vol. 62, no. 3, pp. 278–284, Mar. 2006, doi: 10.1016/j.jhin.2005.08.015.
52. M. P. Ryan, J. T. Pembroke, and C. C. Adley, “Ralstonia pickettii in environmental biotechnology: potential and applications,” J Appl Microbiol, vol. 103, no. 4, pp. 754–764, Oct. 2007, doi: 10.1111/j.1365-2672.2007.03361.x.
53. M. Venkateswar Reddy, G. N. Nikhil, S. Venkata Mohan, Y. V. Swamy, and P. N. Sarma, “Pseudomonas otitidis as a potential biocatalystfor polyhydroxyalkanoates (PHA) synthesis using synthetic wastewater and acidogenic effluents,” Bioresour Technol, vol. 123, pp. 471–479, Nov. 2012, doi: 10.1016/j.biortech.2012.07.077.
54. L. L. Clark, J. J. Dajcs, C. H. McLean, J. G. Bartell, and D. W. Stroman, “Pseudomonas otitidis sp. nov., isolated from patients with otic infections,” Int J Syst Evol Microbiol, vol. 56, no. 4, pp. 709–714, Apr. 2006, doi: 10.1099/ijs.0.63753-0.
55. K. Ghaima, A. Mohamed, W. Yehia, A. Meshhdany, and A. Abdulhassan, “Resistance and bioadsorption of Cadmium by Pseudomonas aeruginosa isolated from agricultural soil,” International Journal of Applied Environmental Sciences, vol. 12, Jan. 2017.
56. Baker-Austin, M. S. Wright, R. Stepanauskas, and J. V. McArthur, “Co-selection of antibiotic and metal resistance,” Trends Microbiol, vol. 14, no. 4, pp. 176–182, Apr. 2006, doi: 10.1016/j.tim.2006.02.006.
57. S. H. A. Hassan, R. N. N. Abskharon, S. M. F. Gad El-Rab, and A. A. M. Shoreit, “Isolation, characterization of heavy metal resistant strain ofPseudomonas aeruginosa isolated from polluted sites in Assiut city, Egypt,” J Basic Microbiol, vol. 48, no. 3, pp. 168–176, Jun. 2008, doi: 10.1002/jobm.200700338.
58. G. O. Oyetibo, M. O. Ilori, S. A. Adebusoye, O. S. Obayori, and O. O. Amund, “Bacteria with dual resistance to elevated concentrations of heavy metals and antibiotics in Nigerian contaminated systems,” Environ Monit Assess, vol. 168, no. 1–4, pp. 305–314, Sep. 2010, doi: 10.1007/s10661-009-1114-3.
59. M. Virieux-Petit, F. Hammer-Dedet, F. Aujoulat, E. Jumas-Bilak, and S. Romano-Bertrand, “From Copper Tolerance to Resistance in Pseudomonas aeruginosa towards Patho-Adaptation and Hospital Success,” Genes (Basel), vol. 13, no. 2, p. 301, Feb. 2022, doi: 10.3390/genes13020301.
60. Permina, A. Kazakov, O. Kalinina, and M. Gelfand, “Comparative genomics of regulation of heavy metal resistance in Eubacteria,” BMC Microbiol, vol. 6, no. 1, p. 49, Dec. 2006, doi: 10.1186/1471-2180-6-49.
61. T. Coenye, J. Goris, P. de Vos, P. Vandamme, and J. J. LiPuma, “Classification of Ralstonia pickettii-like isolates from the environment and clinical samples as Ralstonia insidiosa sp. nov.,” Int J Syst Evol Microbiol, vol. 53, no. 4, pp. 1075–1080, Jul. 2003, doi: 10.1099/ijs.0.02555-0.
62. J. Giner-Lamia, L. López-Maury, J. C. Reyes, and F. J. Florencio, “The CopRS Two-Component
System Is Responsible for Resistance to Copper in the Cyanobacterium Synechocystis sp. PCC 6803 ,” Plant Physiol, vol. 159, no. 4, pp. 1806–1818, Aug. 2012, doi: 10.1104/pp.112.200659.
63. Yang et al., “Biosequestration via cooperative binding of copper by Ralstonia pickettii,” Environ Technol, vol. 31, no. 8–9, pp. 1045–1060, Jul. 2010, doi: 10.1080/09593330.2010.487290.
64. B. Borremans, J. L. Hobman, A. Provoost, N. L. Brown, and D. van der Lelie, “Cloning and Functional Analysis of the pbr Lead Resistance Determinant of Ralstonia metallidurans CH34,” J Bacteriol, vol. 183, no. 19, pp. 5651–5658, Oct. 2001, doi: 10.1128/JB.183.19.5651-5658.2001.
65. M. C. Thaller et al., “Metallo-β-Lactamase Production by Pseudomonas otitidis : a Species-Related Trait,” Antimicrob Agents Chemother, vol. 55, no. 1, pp. 118–123, Jan. 2011, doi: 10.1128/AAC.01062-10.
66. K. M. Sta Ana, J. Madriaga, and M. P. Espino, “β-Lactam antibiotics and antibiotic resistance in Asian lakes and rivers: An overview of contamination, sources and detection methods,” Environmental Pollution, vol. 275, p. 116624, Apr. 2021, doi: 10.1016/j.envpol.2021.116624.
67. C. Zheng, Y. Li, L. Nie, L. Qian, L. Cai, and J. Liu, “Transcriptional and Functional Studies of a Cd(II)/Pb(II)-Responsive Transcriptional Regulator(CmtR) from Acidithiobacillus ferrooxidans ATCC 23270,” Curr Microbiol, vol. 65, no. 2, pp. 117–121, Aug. 2012, doi: 10.1007/s00284-012-0117-4.
68. Meletis, “Carbapenem resistance: overview of the problem and future perspectives,” Ther Adv Infect Dis, vol. 3, no. 1, pp. 15–21, Feb. 2016, doi: 10.1177/2049936115621709.
69. C. Rensing and G. Grass, “Escherichia coli mechanisms of copper homeostasis in a changing environment,” FEMS Microbiol Rev, vol. 27, no. 2–3, pp. 197–213, Jun. 2003, doi: 10.1016/S0168-6445(03)00049-4.
70. K. Bondarczuk and Z. Piotrowska-Seget, “Molecular basis of active copper resistance mechanisms in Gram-negative bacteria,” Cell Biol Toxicol, vol. 29, no. 6, pp. 397–405, Dec. 2013, doi: 10.1007/s10565-013-9262-1.
71. M. R. B. Binet and R. K. Poole, “Cd(II), Pb(II) and Zn(II) ions regulate expression of the metal-transporting P-type ATPase ZntA in Escherichia coli,” FEBS Lett, vol. 473, no. 1, pp. 67–70, May 2000, doi: 10.1016/S0014-5793(00)01509-X.
72. L.-G. Li, Y. Xia, and T. Zhang, “Co-occurrence of antibiotic and metal resistance genes revealed in complete genome collection,” ISME J, vol. 11,no. 3, pp. 651–662, Mar. 2017, doi: 10.1038/ismej.2016.155.
73. J. Chen, J. Li, H. Zhang, W. Shi, and Y. Liu, “Bacterial Heavy-Metal and Antibiotic Resistance Genes in a Copper Tailing Dam Area in Northern China,” Front Microbiol, vol. 10, Aug. 2019, doi: 10.3389/fmicb.2019.01916.
74. W. R. Pearson, G. Robins, and T. Zhang, “Generalized neighbor-joining: more reliable phylogenetic tree reconstruction,” Mol Biol Evol, vol. 16, no. 6, pp. 806–816, Jun. 1999, doi: 10.1093/oxfordjournals.molbev.a026165.
75. M. Nei, “Phylogenetic Analysis In Molecular Evolutionary Genetics,” Annu Rev Genet, vol. 30, no. 1, pp. 371–403, Dec. 1996, doi: 10.1146/annurev.genet.30.1.371.
76. V. Masindi and K. L. Muedi, “Environmental Contamination by Heavy Metals,” in Heavy Metals, InTech, 2018. doi: 10.5772/intechopen.76082.