Eco-friendly synthesis of zinc oxide nanoparticles using Bacillus Subtilis, characterization and antibacterial potential against Staphylococcus aureus associated with cardiac catheterization

Document Type : Articles


Department of Biology, College of Sciences, University of AL-Qadisiyah, Iraq


Zinc Oxide nanoparticles (ZnONPs), which have well-known antimicrobial properties, are used extensively in various medical and general applications. In this analysis, 70-gram positive bacterial isolates were obtained from 100 patients using cardiac catheterization, with 54 Staphylococcus aureus and 16 other positive pathogenic bacteria. Accordingly, morphological, cultural and biochemical testes confirmed the results by VITEK 2 System. The synthesis of Zinc Oxide nanoparticles (ZnO NPs) was done using eco-friendly biological methods by Bacillus Subtilis filtrate which was identified and characterized by UV–Vis Spectrophotometer, SEM, AFM and FTIR, the pH value for the various of ZnONPS is about 7.1 and temperature 37 °C. Furthermore, the antibacterial efficacy of biological synthesized ZnO NPs against this isolated Staphylococcus aureus was determined. The results of SEM illustrated the morphology and sizes of ZnO NPs which are spherical and ovoid with the size range of 20-70 nm. The UV-Vis spectrum indicated the absorption bands of ZnO NPs at 378 nm. Antimicrobial susceptibility test was conducted for 54 isolates against 10 commonly-used antimicrobial agents using Kirby-Bauer disk diffusion method. The results of this study showed the highest rate of resistance against Amoxcillin/Clavulanic acid, Methicillin, tetracycline, Erythromycin and Azithromycin, and moderate resistance to Chloramphenicol. The synergistic effect of antibiotics (Amoxcillin / Clavulanic acid, Methicillin, tetracycline, Erythromycin, Azithromycin, Amikacine, penicillin G, Ampecilline, Trimethoprim\ sulphamethazole and Chloramphenicol) against Staphylococcus aureus was significantly increased in presence of ZnONPs compared to antibiotics only. Conclusion: ZnO NPs demonstrate a good synergistic effect with antibiotics, which can open avenues for a future combination therapy against pathogenic bacteria.

Graphical Abstract

Eco-friendly synthesis of zinc oxide nanoparticles using Bacillus Subtilis, characterization and antibacterial potential against Staphylococcus aureus associated with cardiac catheterization


[1] Beyth, N.; Houri-Haddad, Y.; Domb, A.; Khan, W. and Hazan, R. Alternative Antimicrobial Approach: Nano-Antimicrobial Materials. Evidence-Based Complementary and Alternative Medicine. 16 (2015) 246012.
[2] Jalal, R.; Goharshadi, E.K.; Abareshi, M.; Moosavi, M.; Yousefi, A. and Nancarrow, P.  ZnO nanofluids: green synthesis,characterization, and antibacterial activity. Mater. Chem. Phys. 121(1) (2010) 198–120.
[3] Jones, N.; Ray, B.; Ranjit, K. T.; and Manna, A. C.  Antibacterial activityof ZnO nanoparticle suspensions on a broad spectrum ofmicroorganisms. FEMS Microbiology Letters, 279(1) (2008) 71–76.
[4] Abdulrahman, N.B., Nssaif, Z.M. . Antimicrobial Activity of Zinc Oxide, titanium Dioxide and Silver Nanoparticles Against Mithicillin-Resistant Staphylococcus aureus Isolates. Tikrit J. Pure Sci., 21(3) (2016) 49-53.      
[5] Ahmadi Shadmehri, A., Namvar, F., Miri, H., Yaghmaei, P., Nakhaei Moghaddam, M. . Assessment of antioxidant and antibacterial activities of Zinc Oxide nanoparticles, Graphene and Graphene decorated by Zinc Oxide nanoparticles. Inter. J. Nano Dimension 10 (2019) 350–358.                                        
[6] Heer, A.S.K. Mansooria, S.M. and Chamria, N. Biosynthesis andcharacterization of Zno nanoparticles using ficus religiosa leavesextract. World J. Phrma. Res., 6 (10) (2017), 818-826.
[7] Baskar, G.; Chandhuru, J.; Fahad, K. S.; and Praveen, A.S.  MycologicalSynthesis, Characterization and Antifungal Activity of Zinc Oxide Nanoparticles. Asian Pharma Press, 3(4) (2013) 142–146.
[8] Kasemets, K.; Ivask, A.; Dubourguier, H.C. and Kahru, A. Toxicity of nanoparticles of ZnO, CuO and TiO2 to yeast Saccharomycescerevisiae. Toxicol. In Vitro, 23(6) (2009) 1116–1122.
[9] Kulkarni, S. S. and Shirsat, M. D. Optical and Structural Properties of Zinc Oxide Nanoparticles. IJARPS. 2(1) (2015) 14-18.
[10] Li, M.; Zhu, L. and Lin, D. Toxicity of ZnO nanoparticles to Escherichiacoli: mechanism and the influence of medium components. Environ.Sci. Technol. 45(5) (2011) 1977–1983.
[11] Alok D, Vyom S Toxicity assessment of nanomaterials: methodsand challenges. Anal. Bioanal. Chem. 398(2) (2010) 589-605.
[12] Shantikumar N, Abhilash S, VVDivya R, Deepthy M, Seema N, ManzoorK,  Satish  R  Role  of  size  scale  of  ZnO  nanoparticles  andmicroparticles on toxicity toward bacteria and osteoblast cancer cellsJ. Mater. Sci: Mater. Med. 20 (2009) 235–241.
[13] Toshiaki  O,  Osamu  Y,  Yasuhiro  I,  Zenbe-e  N  Antibacterial activity of ZnO powder with crystallographic orientation. J. Mater. Sci. Mater. Med. 19(3) (2008) 1407-1412.
[14] Zhongbing H,  Xu Zh, Danhong Y, Guangfu Y, Xiaoming L, Yunqing K,Yadong  Yao, Di  Huang,  Baoqing  H  Toxicological  effect of ZnO nanoparticles based on bacteria. Langmuir, 24(8) (2008) 4140–4144.
[15] Al-Aawadi, KK. Antibiotic profile and molecular characterization of Staphylococcus aureus isolated from Tonsillitis patients at Thi-Qar Province. Master thesis. College of  Sciences Thi-Qar University, Iraq (2014).                                                                
[16] Al-Dahbi, A M. and Al-Mathkhury, H J. Distribution of Methicillin Resistant Staphylococcus aureus in Iraqi patients and healthcare worker. Iraqi. J. Sci., 54 (2013) 293-300.
[17] Alikhani, A., et al., Minimal inhibitory concentration of microorganisms causing surgical site infection in referral hospitals in North of Iran, 2011-2012. Caspian J. Internal Med. 6(1) (2015) 34-39.                                                                                                   
[18] Al-Mussawi, AA Detection of Staphylococcus aureus and Methicillin  Resistant Staphylococcus aureus (MRSA) from human clinical specimens using conventional  biochemical testes and chromogenic  media. Indian J. Epp. Res. 4(2) (2014) 7-9.                                 
[19] Asghar, AH Molecular characterization of Methicillin-Resistant Staphylococcus aureus isolated from tertiary care Hospitals. Pak. J. Med. Sci., 30(4) (2014)  698-702.
[20] MacFaddin, J F. Biochemical tests for identification of medical bacteria. (3rd.ed). Lippinocott Williams &Wilkins, USA, (2000) 555-565.                                      
[21] Selvarajan E, Mohanasrinivasan V. Biosynthesis and characterization of ZnO nanoparticles using Lactobacillus plantarumVITES07. Mater. Lett. 112 (2013) 180-182.
[22] Wang D, Cui L, Chang X, Guan D. Biosynthesis and characterization of zinc oxide nanoparticles from Artemisia annua andinvestigate their effect on proliferation, osteogenic differentiationand mineralization in human osteoblast-like MG-63 Cells. J Photochem Photobiol B. 202 (2020) 111652.
[23] Kumar, H. and Rani, R. Structural and Optical Characterization of ZnONanoparticles Synthesized by Microemulsion Route. Inter. Lett. Chem. Phys. Astronomy, 14 (2013) 26-36.
[24] Mohan KK, Mandal BK, Appala NE, Sinha M, Siva KK,Sreedhara RP. Synthesis and characterisation of flower shapedzinc oxide nanostructures and its antimicrobial activity. SpectrochimActa A Mol Biomol Spectrosc. 15 (2013) 256-259.
[25] Pati, R.; Mehta, R.K.; Mohanty, S.; Padhi, A.; Sengupta, M.; Vaseeharan, B. Topical application of zinc oxide nanoparticles reducesbacterial skin infection in mice and exhibits antibacterial activity byinducing oxidative stress response and cell membrane disintegration inmacrophages. Nanomedicine., 10(6) (2014) 1195-208.
[26] Płaza, G. A.; Chojniak, J. and Banat, I. M. Biosurfactant MediatedBiosynthesis of Selected Metallic Nanoparticles. Int. J. Mol. Sci., 15 (2014) 13720-13737.
[27] Lingling  Z,  Yunhong  J,  Yulong  D,  Malcolm  P,  David  Y  .Investigation  into  the  antibacterial  behavior  of  suspensions  of ZnOnanoparticles (ZnO nanofluids). J. Nanoparticle Res. 9(3) (2006) 479-489.
[28] Mohsen J,  Zahra B. Protein  nanoparticle: A unique  system  as drug delivery vehicles. Afr. J. Biotechnol., 7(25) (2008) 4926-4934.
[29] K. Mandava, K. Kadimcharla, N. R. Keesara, S. N. Fatima, P. Bommena, U. R. Batchu, ‘Green Synthesis of Stable Copper Nanoparticles and Synergistic Activity with Antibiotics’, Indian J. Pharm. Sci. 79(5) (2017) 695 –700.
[30]Bhave PP, Kartikeyan S, Ramteerthakar MN, Patil NR. Bacteriological study of surgical site infections in a tertiary care hospital at Miraj, Maharashtra state, India. Int. J. Res. Med. Sci. 4(7) (2016) 2630-2635.
[31] Blauwet, L. A., & Redberg, R. F. The role of sex-specific results reporting in cardiovascular disease. Cardiology in review, 15(6) (2007) 275-278.
[32] Zavareh, MS. Tohidi, M. Sabouri A. Infectious and coronary artery disease. ARYA Atheroscler. 12 (1) (2016) 41-49.
[33] Bouza, E.; Muñoz, P.; López-Rodrígues ,J.; Jesús Pérez ,M.; Rincón, C.;Martín Rabadán, P.; Sánchez, C.& Bastida, EA needleless closed system device (CLAVE) protects from intravascular catheter tip and hub. J. Hosp. Infect., 54( 2003) 279-287.
[34] Levy, S.B. and Marshall, B. Antibacterial resistance worldwide: causes,challenges and responses, Nat. Med., 10 (2004) S122-S129.
[35] Organization, W.H. Antimicrobial resistance: global report on surveillance, World Health Organization. (2014).
[36] Li, X., Xu, H., Chen, Z. and Chen, G. Biosynthesis of Nanoparticles bymicroorganisms and there application. J. Nanomat. 8 (2011) 1-16.
[37] Mukherjee, A.; Sadiq, M. I.; Prathna, T. C. and Chandrasckaran, N. Antimicrobial activity of aluminum oxide nanoparticles for potential clinical applications. Communicating Res. Technological Adv. (2011). 245–251.
[38] Rizwan W, Nagendra KK, Akhilesh KV, Anurag M, Hwang IH, You-Bing Y,  Hyung-Shik  Sh, Young-Soon  K  Fabrication  and  growthmechanism  of  ZnO  nanostructures  and  their  cytotoxic  effect  onhuman  brain  tumor  U87,  cervical  cancer  HeLa,  and  normal  HEKcells. J. Biol. Inorg. Chem., 16(3) (2010b) 431-442.
[39] Siddiqi, K.S., Rahman, A., Tajuddin, Husen, A. Properties of Zinc Oxide Nanoparticles and Their Activity Against Microbes. Nanoscale Res. Lett., 13 (2018) 141-149.
[40] Kulkarni, S. S. and Shirsat, M. D. Optical and Structural Properties of Zinc Oxide Nanoparticles. IJARPS 2(1) (2015)  14-18.     
[41] Pavani, K. V, Balakrishna, K. and Cheemarla, N. R. Biosynthesis of Zinc Nanoparticles by Aspergillus species, 5(1) (2011) 27–36.
[42] Reddy  KM,  Kevin  F,  Jason  B,  Denise  GW,  Cory  H,  Alex  P. Selective  toxicity  of  zinc  oxide  nanoparticles  to  prokaryotic  and  eukaryotic systems. J. Appl. Phys. Lett., 90(21) (2007) 1-3.
[43] Khalil, M. A. E. F.; Sonbol, F. I.; Mohamed, A. F. B. and Ali, S. S. Comparative study of virulence factors among ESβL-producing and nonproducing Pseudomonas aeruginosa clinical isolates. Tur. J. Med. Sci. 45 (2015) 60-69.
[44] Slomberg, D.L., Lu, Y., Broadnax, A.D., Hunter, R.A., Carpenter, A.W., Schoenfisch, M.H. Role of size and shape on biofilm eradication for nitric oxide-releasing silica nanoparticles. ACS Appl. Mater. Inter. 5(19) (2013) 9322–9329.
[45] Kasemets, K.; Ivask, A.; Dubourguier, H.C. and Kahru, A. Toxicity of nanoparticles of ZnO, CuO and TiO2 to yeast Saccharomyces cerevisiae. Toxicol. In Vitro, 23(6) (2009) 1116–1122.
Volume 12, Issue 2
June 2022
Pages 159-168
  • Receive Date: 10 October 2021
  • Revise Date: 15 April 2022
  • Accept Date: 22 April 2022
  • First Publish Date: 01 June 2022