Document Type: Final File

Authors

Department of Microbiology, School of Basic Sciences, Saveh Branch, Islamic Azad University, Saveh, Iran.

Abstract

Pseudomonas aeruginosa is a common cause of surgical-site infections and healthcare-associated infections in the bloodstream, and urinary tract. Iron oxide nanoparticles (IONPs) have shown, to possess antibacterial features. The nanoparticles' status as emerging therapeutic elements has motivated investigators to assess the effects of iron nanoparticles on the expression of TEM type beta-lactamase genes in P. aeruginosa. In this descriptive-analytic study, 60 clinical isolates of P. aeruginosa were isolated from burn wounds and respiratory excretions of Pasargad Research Laboratory of Tehran, Iran. All isolates were characterized using differential biochemical tests and confirmed samples as P. aeruginosa. Their genomic DNA was extracted and PCR reaction was performed to screen TEM-gene carrying isolates. Then MIC of IONPs against these strains was determined and finally, Real-time PCR performed to the determination of the expression of the TEM gene. Results showed that 8 isolates (13/33%) had the TEM beta-lactamase gene. The MIC and MBC of IONPs against P. aeruginosa strains were observed at 256 µg/mL or 125 µg/mL, while the MBC was determined at 500 µg/mL. In addition, statistical analysis of Real-time PCR data showed that there is a statistically significant difference between gene expression levels of IONPs treated isolates and non-treated ones. The results showed that TEM gene expression levels in two isolates treated with IONPs were 78% and 75% lower than untreated bacteria (P<0.001; r= 0.958). Our findings confirmed that IONPs are potential antibacterial agents and can be considered as promising treatments for recalcitrant P. aeruginosa infections.

Keywords

1.   Chalhoub H, Pletzer D, Weingart H, Braun Y, Tunney MM, Elborn JS, Rodriguez-Villalobos H, Plésiat P, Kahl BC, Denis O, Winterhalter M. Mechanisms of intrinsic resistance and acquired susceptibility of Pseudomonas aeruginosa isolated from cystic fibrosis patients to temocillin, a revived antibiotic. Sci. Rep. 7:40208; (2017).

2.   Chevalier S, Bouffartigues E, Bodilis J, Maillot O, Lesouhaitier O, Feuilloley MG, Orange N, Dufour A, Cornelis P. Structure, function and regulation of Pseudomonas aeruginosa porins. FEMS Microbiol. Rev. 41(5):698-722; (2017).

3.   Li X-Z, Plésiat P, Nikaido H. The challenge of efflux-mediated antibiotic resistance in Gram-negative bacteria. Clin. Microbiol. Rev. 28(2):337-418; (2015).

4.   Ventola CL. The antibiotic resistance crisis: part 1: causes and threats. Pharm & Therap. 40(4):277; (2015).

5.   Pasa O, Ozer B, Duran N, Inci M, Yula E. Beta-lactamase Enzymes of Clinical Pseudomonas aeruginosa Strains. West Indian Med. J. 65(1):40-5; (2016).

6.   Bell BG, Schellevis F, Stobberingh E, Goossens H, Pringle M. A systematic review and meta-analysis of the effects of antibiotic consumption on antibiotic resistance. BMC Infect. Dis. 14(1):13; (2014).

7.   Shanmugam K, Thyagarajan R, Katragadda R, Vajravelu L, Lakshmy A. Biofilm formation and Extended Spectrum Beta Lactamases (ESBL) producers among the gram negative bacteria causing Urinary tract infections. Int. J. of Med. Microb. and Trop. Dis. 3(3):86-90; (2017).

8.   Gellatly SL, Bains M, Breidenstein EB, Strehmel J, Reffuveille F, Taylor PK, Yeung AT, Overhage J, Hancock RE. Novel roles for two-component regulatory systems in cytotoxicity and virulence-related properties in Pseudomonas aeruginosa. AIMS Microbiol. 4(1):173-91; (2018).

9.   Oglesby AG, Farrow JM, Lee JH, Tomaras AP, Greenberg EP, Pesci EC, Vasil ML. The influence of iron on Pseudomonas aeruginosa physiology a regulatory link between iron and quorum sensing. J. Biol. Chem. 283(23):15558-67; (2008).

10. Ramezani Ali Akbari K, Abdi Ali A. Study of antimicrobial effects of several antibiotics and iron oxide nanoparticles on biofilm producing pseudomonas aeruginosa. NMJ. 4(1):37-43; (2017).

11. Lerner RN, Lu Q, Zeng H, Liu Y. The effects of biofilm on the transport of stabilized zerovalent iron nanoparticles in saturated porous media. Water Res. 46(4):975-85; (2012).

12. Fahmy HM, Mohamed FM, Marzouq MH, Mustafa AB, Alsoudi AM, Ali OA, Mohamed MA, Mahmoud FA. Review of Green Methods of Iron Nanoparticles Synthesis and Applications. J. Bionanosci. 14 (5):1-13; (2018).

13. De Vos P, Garrity GM. Bergey's manual of systematic bacteriology: Springer;p123-191; (2009).

14. Phillips K, McCallum N, Welch L. A comparison of methods for forensic DNA extraction: Chelex-100® and the QIAGEN DNA Investigator Kit (manual and automated). Forensic Sci. Int. 6(2):282-5; (2012).

15. Khatami M, Alijani H, Sharifi I, Sharifi F, Pourseyedi S, Kharazi S, Lima Nobre MA, Khatami M. Leishmanicidal activity of biogenic Fe3O4 nanoparticles. Sci. Pharm. 85(4):36; (2017).

16. Seifi Mansour S, Ezzatzadeh E, Safarkar R. In vitro evaluation of its antimicrobial effect of the synthesized Fe3O4 nanoparticles using Persea Americana extract as a green approach on two standard strains. Asi. J. Gre. Chem. 3:353-65; (2019).

17. Turbett SE, Pierce VM. Overview of antibacterial susceptibility testing. UpToDate Retrieved January.;12:2019; (2017).

18. Espy MJ, Uhl JR, Sloan LM, Buckwalter SP, Jones MF, Vetter EA, Yao JD, Wengenack NL, Rosenblatt JE, Cockerill F3, Smith TF. Real-time PCR in clinical microbiology: applications for routine laboratory testing. Clin Microbiol. Rev. 19(1):165-256; (2006).

19. Rodríguez-Baño J, Gutiérrez-Gutiérrez B, Machuca I, Pascual A. Treatment of Infections Caused by Extended-Spectrum-Beta-Lactamase-, AmpC-, and Carbapenemase-Producing Enterobacteriaceae. Clin. Microbiol. Rev. 31(2):e00079-17; (2018).

20. Sharma VK, Johnson N, Cizmas L, McDonald TJ, Kim H. A review of the influence of treatment strategies on antibiotic resistant bacteria and antibiotic resistance genes. Chemosphere. 150:702-14; (2016).

21. Colque C, Orio AA, Feliziani S, Marvig R, Tobares A, Johansen H. Hypermutator Pseudomonas aeruginosa exploits multiple genetic pathways to develop multidrug resistance during long-term infections in the airways of cystic fibrosis patients. Bio. Rxiv. 8(3):80-89; (2019).

22. Cavallo J, Fabre R, Leblanc F, Nicolas-Chanoine M, Thabaut A. Antibiotic susceptibility and mechanisms of β-lactam resistance in 1310 strains of Pseudomonas aeruginosa: a French multicentre study (1996). J Antimicrob. Chemother. 46(1):133-6; (2000).

23. Bert F, Branger C, Lambert-Zechovsky N. Identification of PSE and OXA β-lactamase genes in Pseudomonas aeruginosa using PCR–restriction fragment length polymorphism. J. Antimicrob. Chemother. 50(1):11-8; (2002).

24. Elhariri M, Hamza D, Elhelw R, Dorgham SM. Extended-spectrum beta-lactamase-producing Pseudomonas aeruginosa in camel in Egypt: potential human hazard. Ann. Clin. Microbiol. Antimicrob. 16(1):21; (2017)..

25. Shojapour M, Shariati L, Karimi A, Zamanzad B. Prevalence of TEM-1 type beta-lactmase genes in Pseudomonas aeruginosa strains isolated from burn infections using Duplex PCR in Shahrekord, 2008. J. Arak. Univ. Med. Sci. 14(1); (2011).

 

26. Neupane, Bishnu Prasad, Dinesh Chaudhary, Sanjita Paudel, Sangita Timsina, Bipin Chapagain, Nirmala Jamarkattel, and Bishnu Raj Tiwari. Himalayan honey loaded iron oxide nanoparticles: synthesis, characterization and study of antioxidant and antimicrobial activities. Int. j. nanomed. 14:3533; (2019).

27. Khatami, Mehrdad, Hajar Q. Alijani, Mojtabi Haghighat, Mehdi Bamrovat, Sara Azhdari, Mohammad Ahmadian, Marcos Nobre, Mohammadreza Heidari, Mina Sarani, and Sanaz Khatami. Green Synthesis of Amorphous Iron Oxide Nanoparticles and their Antimicrobial Activity against Klebsiella pneumonia, Pseudomonas aeruginosa and Escherichia coli. Iran.  J. Biotechnol. 10:33-39; (2019).

28. Vasantharaj S, Sathiyavimal S, Senthilkumar P, LewisOscar F, Pugazhendhi A. Biosynthesis of iron oxide nanoparticles using leaf extract of Ruellia tuberosa: antimicrobial properties and their applications in photocatalytic degradation. J. Photochem. Photobiol. Biol. 192:74-82; (2019).

29. Khatami, M., Aflatoonian, M.R., Azizi, H., Mosazade, F., Hoshmand, A., Nobre, M.A.L., Poodineh, F.M., Khatami, M., Khraazi, S. and Mirzaeei, H. Evaluation of Antibacterial Activity of Iron Oxide Nanoparticles Against Escherichia coli. Practice. 18:19; (2017).

30. Masadeh, Majed M., Ghadah A. Karasneh, Mohammad A. Al-Akhras, Borhan A. Albiss, Khaled M. Aljarah, Sayer I. Al-Azzam, and Karem H. Alzoubi. Cerium oxide and iron oxide nanoparticles abolish the antibacterial activity of ciprofloxacin against gram positive and gram negative biofilm bacteria. Cytotechnology. 67(3):427-35; (2015).

31. Borcherding J, Baltrusaitis J, Chen H, Stebounova L, Wu CM, Rubasinghege G, Mudunkotuwa IA, Caraballo JC, Zabner J, Grassian VH, Comellas AP. Iron oxide nanoparticles induce Pseudomonas aeruginosa growth, induce biofilm formation, and inhibit antimicrobial peptide function. Envi. Sci: Nano. 1(2):123-32; (2014)..

32. Azam A, Ahmed AS, Oves M, Khan MS, Habib SS, Memic A. Antimicrobial activity of metal oxide nanoparticles against Gram-positive and Gram-negative bacteria: a comparative study. Int. j. nano. 7:6003; (2012).