Document Type : Research Paper


1 Department of Microbiology, Falavarjan Branch, Islamic Azad University, Isfahan, Iran

2 Department of Pharmacology and Toxicology, School of Pharmacy, Hamadan University of Medical Sciences, Hamadan, Iran



Industrial activities present a significant threat to the environment and natural ecosystems like water and soil due to the release of toxic metals. This article primarily concentrates on the identification and isolation of bacteria, with the goal of effectively eliminating pollutants from industrial wastewater. In order to achieve this goal, the study was conducted to assess the ability of bacterial strains to tolerate copper (Cu) and zinc (Zn), as well as their antibiotic resistance and ability to tolerate elevated metal concentrations. The resistance of the isolates to various metals and antibiotics were assessed using the minimum inhibitory concentration (MIC) values and disc diffusion (DD) method, respectively. The technique of colony PCR was employed to determine the identity of the bacteria that were separated. Resistance to multiple antibiotics was assessed, including Penicillin, Sulfamethoxazole, Tetracycline, Erythromycin, Amoxicillin, Cefoxitin, Streptomycin, Chloramphenicol, Vancomycin, Gentamycin, Cephalothin, Rifampicin, and Novobiocin. In the current investigation, a total of 5 bacteria with a positive gram stain and 7 bacteria with a negative gram stain were identified. The study found that the effluent from the wastewater treatment plant in Razi industrial town showed resilience to copper ions, especially at a concentration of 7mM. The effluent wastewater from the refinery unit exhibited the greatest level of tolerance towards zinc, with a concentration as high as 6mM. The rise in copper and zinc levels in industrial wastewater treatment plants causes microorganisms to develop resistance to these heavy metals. The study of Gram-positive resistant bacteria conducted in this research focused on the examination of their susceptibility to zinc and copper. Notably, Staphylococcus hominis displayed resistance to a majority of the antibiotics evaluated. However, Kocuria rosea demonstrated sensitivity to all antibiotics. Agrobacterium fabrum exhibited susceptibility to all antibiotics as opposed to other Gram-negative bacteria resistant to zinc and copper. The findings of this study indicated that some strains displayed a degree of resistance to both antibiotics and heavy metals. The presence of heavy metals in bacteria isolated from a wastewater treatment plant exhibited the capability to restrict antibiotic resistance.


Alboghobeish, H., Tahmourespour, A. and Doudi, M. (2014) 'The study of Nickel Resistant Bacteria (NiRB) isolated from wastewaters polluted with different industrial sources', Journal of Environmental Health Science and Engineering, 12(44), pp.1-7. doi:
Alengebawy, A. et al. (2021) 'Heavy metals and pesticides toxicity in agricultural soil and plants: Ecological risks and human health implications', Toxics, 9(3), pp. 1-33. doi:
Aslam, B. et al. (2018) 'Antibiotic resistance: a rundown of a global crisis', Infect Drug Resist, 11, pp.1645-1658. doi:
Bischofberger, A.M. et al. (2020) 'Associations between sensitivity to antibiotics, disinfectants and heavy metals in natural, clinical and laboratory isolates of Escherichia coli', Environmental Microbiology, 22 (7), pp. 2664-2679. doi:
Briffa, j., Sinagra, E., and Blundell, R. (2020) 'Heavy metal pollution in the environment and their toxicological effects on humans, Heliyon, 6 (9), e04691. doi:
Bukowski, M. et al. (2019) 'Prevalence of antibiotic and heavy metal resistance determinants and virulence-related genetic elements in plasmids of Staphylococcus aureus, Frontiers in Microbiology, 10, 805. doi:
Callaway, T.R. et al. (2021) 'Alternatives to antibiotics: A symposium on the challenges and solutions for animal health and production', Antibiotics, 10 (5), pp. 1-15. doi:
Chandrangsu, P., Rensing, C., and Helmann, J.D. (2017) 'Metal homeostasis and resistance in bacteria', Nature Reviews Microbiology, 15 (6), pp. 338-350. doi: 10.1038/nrmicro.2017.15
Coque, T.M. et al. (2023) 'Antimicrobial resistance in the global health network: known unknowns and challenges for efficient responses in the 21st century', Microorganisms, 11 (2023). doi: 10.3390/microorganisms11041050
Das, T.K., and Poater, A. (2021) 'Review on the use of heavy metal deposits from water treatment waste towards catalytic chemical syntheses', International Journal of Molecular Sciences, 22 (24), 13383. doi:
Dickinson, A.W. et al. (2019) 'Heavy metal pollution and co-selection for antibiotic resistance: A microbial palaeontology approach, Environment International, 132, 105117. doi: 10.1016/j.envint.2019.105117
Dinu, L.A., Anghel, L., and Jurcoane, S. (2011) 'Isolation of heavy metal resistant strains from the battery manufactured polluted environment', Romanian Biotechnological Letters, 16 (6), pp. 102-106. Available at: (Accessed: 15 November 2023).
Dixit, R. et al. (2015) 'Bioremediation of heavy metals from soil and aquatic environment: An overview of principles and criteria of fundamental processes', Sustainability, 7 (2), pp. 2189-2212. doi:
Edet, U.O., Bassey, I.U. and Joseph, A.P. (2023) 'Heavy metal co-resistance with antibiotics amongst bacteria isolates from an open dumpsite soil'. Heliyon, 9 (2), e13457. doi:
Escamilla-Rodríguez, A., Carlos-Hernández, S. and Díaz-Jiménez, L. (2021) 'Evidence of resistance of heavy metals from bacteria isolated from natural waters of a mining area in Mexico', Water, 13 (2021), 2766. doi:
Fashola, M.O., Ngole-Jeme, V.M., and Babalola, O.O. (2016) 'Heavy metal pollution from gold mines: Environmental effects and bacterial strategies for resistance', International Journal of Environmental Research and Public Health, 13 (11), 1047. doi:
Fawwaz Alfarras, A., Hamid Al-Fahdawi, M. and Albayaty, M.K. (2022) 'Heavy metal resistance ability of Pseudomonas species isolated from sludge and sewage in Iraq', Archives of Razi Institute journal, 77 (3), pp.1041-1047. doi:
Fillali, B.K. et al. (2000) 'Waste water bacterial isolates resistant to heavy metals and antibiotics', Current Microbiology, 41 (3), pp.151-156. doi:
Galetti, R. et al. (2019) 'Antibiotic resistance and heavy metal tolerance plasmids: the antimicrobial bulletproof properties of Escherichia fergusonii isolated from poultry', Infection and Drug Resistance, 12, pp.1029-1033. doi:
González Henao, S., and Ghneim-Herrera, T. (2021) 'Heavy Metals in Soils and the Remediation Potential of Bacteria Associated with the Plant Microbiome', Frontiers in Environmental Science, 9, 604216. doi:
Santo, C.E., Morais, P.V., and Grass, G. (2010) 'Isolation and characterization of bacteria resistant to metallic Copper surfaces', Applied and Environmental Microbiology, 76 (5), pp.1341-1348. doi:
Hammaini, A. et al. (2007) 'Biosorption of heavy metals by activated sludge and their desorption characteristics', Journal of Environmental Management, 84 (4), pp. 19-426. doi:
Heidarzadeh, M. et al. (2020) 'The effect of Typha Latifolia L. on heavy metals phytoremediation at the urban and industrial wastewater entrance to the Meighan wetland, Iran', Journal of Applied Research in Water and Wastewater, 7 (2), pp.167-171. doi:
Khaira, M.B., Yusuf, M.B., and Khan, F. (2022) 'Insights to antimicrobial resistance: heavy metals can inhibit antibiotic resistance in bacteria isolated from wastewater', Environmental Monitoring and Assessment, 194 (4), 252. doi:
Knapp, C.W. et al. (2017) 'Relationship between antibiotic resistance genes and metals in residential soil samples from Western Australia', Environmental Science and Pollution Research, 24, pp.2484–2494.
Larsson, D.G.J., and Flach, C.F. (2022) 'Antibiotic resistance in the environment', Nature Reviews Microbiology, 20 (5), pp. 257-269. doi:
Li, F.J. et al. (2022) 'Pollution, sources, and human health risk assessment of heavy metals in urban areas around industrialization and Urbanization-Northwest China', Chemosphere, 308 (Part 2), 136396. doi:
Li, L.G., Xia, Y., and Zhang, T. (2017) 'Co-occurrence of antibiotic and metal resistance genes revealed in complete genome collection', ISME J, 11, pp. 651-662. doi:
Long, Z. et al. (2021) 'Effect of different industrial activities on soil heavy metal pollution, ecological risk, and health risk, Environmental Monitoring and Assessment, 193 (1), 20. doi:
Marzan, L.W. et al. (2017) 'Isolation and biochemical characterization of heavy-metal resistant bacteria from tannery effluent in Chittagong city, Bangladesh: Bioremediation viewpoint', The Egyptian Journal of Aquatic Research, 43 (1), pp. 65-74. doi:
Najar, I.N., et al. (2022) 'Coexistence of heavy metal tolerance and antibiotic resistance in thermophilic bacteria belonging to genus Geobacillus', Frontiers in Microbiology, 13, 914037. doi:
Nasrazadani, A., Tahmourespour, A., and Hoodaji, M. (2010) 'Determination of bacteria resistance threshold to Lead, Zinc and Cadmium in three industrial wastewater samples', Journal of Environmental Studies, 36 (56), pp. 25-27. doi:
Nies, D.H. (1992) 'Resistance to cadmium, cobalt, zinc and nickel microbes', Plasmid, 27 (1), pp. 17-28. doi:
Nouri, M., and Montazer Faraj, A. (2022) 'Monitoring and assessment of water quality in Tehran city using physicochemical and microbial indexes', Journal of Applied Research in Water and Wastewater, 9 (2), pp.121-124. doi:
Pande, V. et al. (2022) 'Microbial interventions in bioremediation of heavy metal contaminants in Agroecosystem', Frontiers in Microbiology, 13, 824084. doi:
Pham, V.H.T. et al. (2022) 'Bacterial Biosorbents, an Efficient Heavy Metals Green Clean-Up Strategy: Prospects, Challenges, and Opportunities', Microorganisms, 10 (3), 610. doi: 10.3390/microorganisms10030610
Rahimzadeh Torabi, L. et al. (2021)’ Isolation, characterization, and effectiveness of bacteriophage Pɸ-Bw-Ab against XDR Acinetobacter baumannii isolated from nosocomial burn wound infection. Iranian Journal of Basic Medical Science, 24(9): pp. 1254-1263. doi:
Rajbanshi, A. (2008) 'Study on heavy metal resistant bacteria in Guheswori Sewage treatment plant', Our Nature, 6, pp. 52-57. doi: v6i1.1655
Igiri, B.E. et al. (2018) 'Toxicity and bioremediation of heavy metals contaminated ecosystem from tannery wastewater: A review', Journal of Toxicology, 2018, 2568038. doi:
Russell, A.D., and Chopra, I. (1990) Understanding antibacterial action and resistance. 2nd edn. New York: Ellis Horwood.
Sabry, S.A., Ghozian, H.A., and Abou-Zeid, D.M. (1997) 'Metal tolerance and antibiotic resistance patterns of a bacterial population isolated from sea water', Journal of Applied Microbiology, 82 (2), pp. 245-252. doi: 1997.tb02858.x
Sadeghabady, Z., Doudi, M., and Rahimzadeh Torabi, L. (2020) 'Isolation and molecular recognition of heavy metal and antibiotic resistant bacteria in the inlet and outlet sewage of Ahvaz refinery, Iran', Microbiology, Metabolites and Biotechnology, 3 (1), pp. 41-52. doi:
Salehi, M., Akhtari, M., and Akhavan Sepahi, A. (2017) 'Evaluation of resistance to heavy metals of zinc sulfate and cadmium sulphate in Escherichia coli strain isolated from surface waters of Tehran city using seial dilution method in pipe', New Cellular and Molecular Biotechnology Journal, 20 (7), pp. 85-89. doi:
Shahsanaei Goneirani, M., Doudi, M., and Ahadi, A.M. (2016) 'The study of simultaneous resistance to heavy metals and antibiotics in resistant bacteria to silver and cadmium isolated from the wastewater', International Journal of Molecular and Clinical Microbiology, 6 (2016), pp. 643-651. Available at: sup (Accessed: 15 November 2023).
Sharma, P. (2022) 'Role and significance of biofilm-forming microbes in phytoremediation - A review', Environmental Technology & Innovation, 25, 102182. doi:
Silver, S., and Ji, G. (1994) 'Newer systems for bacterial resistances to toxic heavy metals', Environmental Health Perspectives, 102 (suppl3), pp.107-13. doi:
Sinegani, A.S., and Younessi, N. (2017) 'Antibiotic resistance of bacteria isolated from heavy metalpolluted soils with different land uses', Journal of Global Antimicrobial Resistance, 10, pp.247-255. doi:
Spain, A., and Alm, E. (2003) 'Implication of microbial heavy metal tolerance in the environment', Reviews in Undergraduate Research, 2, pp. 1-6. Available at: (Accessed: 15 November 2023).
Tahmourespour, A., and Kasra Kermanshahi, R. (2007) 'Determination of adaptation to heavy metals in bacteria resistant to industrial effluents', Water and Wastewater Journal, 61, pp. 53-59. Available at: (Accessed: 15 November 2023).
Tahmourespour, A. (2021) 'Heavy metals and antibiotic co-resistance in bacterial isolates of industrial effluents', Journal of Water and Wastewater, 32 (5), pp. 12-20. doi:
Tchounwou, P.B. et al. (2012) 'Heavy metal toxicity and the environment', Experientia supplementum, 101, pp. 133-64. doi:
Teitzel, G.M., and Persek, M.R. (2003) 'Heavy metal resistance of biofilm and Planktonic Pseudomonas aeruginosa, Applied and Environmental Microbiology, 69 (4), pp. 2313–2320. doi:
Timmis, K. et al. (2017), 'The contribution of microbial biotechnology to economic growth and employment creation, Microbial Biotechnology, 10 (5), pp. 1137-1144. doi:
Tripathi, M. et al. (2023) 'Microbial biosorbent for remediation of dyes and heavy metals pollution: A green strategy for sustainable environment', Frontiers in Microbiology, 14, 1168954. doi:
Türkmen, D. et al. (2022) 'Heavy metal ions removal from wastewater using cryogels, A review', Frontiers in Sustainability, 3, 765592. doi:
Tytła, M. (2019) 'Assessment of heavy metal pollution and potential ecological risk in sewage sludge from municipal wastewater treatment plant located in the most industrialized region in poland-case study', International Journal of Environmental Research and Public Health, 16 (13), 2430. doi:
Uroko, R., and Njoku, O. (2021) 'Ecological and human health risks assessment of heavy metals in vegetables grown in palm oil mill effluents irrigated farmland', Journal of Applied Research in Water and Wastewater, 8 (2), pp. 140-149. doi:
Vats, P., Kaur, U.J., and Rishi, P. (2022) 'Heavy metal-induced selection and proliferation of antibiotic resistance, A review', Journal of Applied Microbiology, 132 (6), pp. 4058-4076. doi:
Verma, T. et al. (2001) 'Chromat tolerant bacteria isolated from tannery effluent', Bioresource Technology, 78 (1), pp. 31-35. doi:
Vidu, R. et al. (2023) 'Removal of heavy metals from wastewaters: A challenge from current treatment methods to nanotechnology applications', Toxics, 8 (4), 101. doi:
Wales, A.D., and Davies, R.H. (2015) 'Co-Selection of Resistance to Antibiotics, Biocides and Heavy Metals, and Its Relevance to Foodborne Pathogens', Antibiotics (Basel), 4 (4), pp. 567-604. doi:
Weisburg, W.G. et al. (1991) '16S ribosomal DNA amplification for phylogenetic study', Journal Bacteriolgy, 173 (2), pp. 697-703. doi:
Xu, Y. et al. (2017) 'Effect of the selective pressure of sub-lethal level of heavy metals on the fate and distribution of ARGs in the catchment scale', Environmental Pollution, 220 (PtB), pp. 900-908. doi:
Yazdankhah, S., Skjerve, E., and Wasteson, Y. (2018) 'Antimicrobial resistance due to the content of potentially toxic metals in soil and fertilizing products', Microbial Ecology in Health and Disease, 29 (1), 1548248. doi:
Ybarra, G.R. and Webb, R. (1999) 'Effects of divalent metal cations and resistance mechanisms of the Cyanobacterium synechococcus sp Strain OCC 7942', Journal of Hazardous Substance Research, 2, pp. 1-9. doi:
Yektamanesh, H., Doudi, M., and Emami, Z. (2021) 'Isolation, molecular identification, and bioremediation activity of chromium-resistant bacteria collected from waste waters in leather tanning plants of Khuzestan, Iran', Biological Journal of Microorganism, 10 (38), pp. 71-84. doi:
Yosofi, Y., Almasi, A., and Mousavi, S.A. (2017) 'Studying the efficiency of anaerobic stabilization pond on removing BOD and COD and changing the sulfur compounds in wastewater of Oil Refine', Journal of Applied Research in Water and Wastewater, 4 (1), pp. 331-333. doi: