Detecting the Efflux Pumps Gene (mexB and oprM ) in MDR Pseudomonas Aeruginosa Isolated from Wound Infection

Wael Abbas Hameed  AL-Mhesin; Rana Mujahid Abdullah Abdullah; Okan Ürker

doi:10.30526/37.3.3447

Authors

Wael Abbas Hameed AL-Mhesin Department of Biology, Graduate School of Natural and Applied Sciences, Çankiri Karatekin University.
Rana Mujahid Abdullah Abdullah Department of Biology, College of Education for Pure Sciences (Ibn Al-Haitham), University of Baghdad, Baghdad, Iraq. https://orcid.org/0000-0001-6017-2005
Okan Ürker Department of Biology, Graduate School of Natural and Applied Sciences, Çankiri Karatekin University https://orcid.org/0000-0002-5103-7757

DOI:

https://doi.org/10.30526/37.3.3447

Keywords:

Keywords: Pseudomonas aeruginosa; Antibiotic resistance; Efflux pumps.

Abstract

The large illnesses caused by Pseudomonas aeruginosa and the more challenging therapy for antimicrobial resistance are both due to the organism's extensive spectrum of virulence factors and several mechanisms for antibiotic resistance. The aim of the study is to detect phenotypic and genotypic efflux pump genes in Pseudomonas aeruginosa. In the current study, 100 wound samples were collected from both sexes and different ages for a period from May 2022 to November 2022 from Teaching Laboratories Hospital/Medical City, Martyr Ghazi Hariri Hospital, and Baghdad Teaching Hospital. 19 isolates of Pseudomonas aeruginosa were taken from wound infections. The samples were cultured on cetrimide agar and MacConkey agar for diagnosis, and then they underwent several microscopic, phenotypic, and biochemical tests, where included Oxidase, Indole, Urease, Glucose and Triple sugar-iron test (TSI). Pseudomonas aeruginosa susceptibility to 11 antibiotics was tested using the disc diffusion method. It has been shown that Pseudomonas aeruginosa has multi-antibiotic resistance. It showed that the bacteria isolates are resistant to Levofloxim 42%, Ciprofloxacin 40%, Piperacillin tazobactam 35%, Amicacin 32%, Ceftazidime 31%, Gentamicin 31%, Cefepime 30%, Piperacillin 29%, Tobramycin 29%, Imipenem 17%, and Meropenem 16%. The ability of bacteria to produce efflux pumps was also studied using the Ethidium Bromide Agar Cartwheel Method, where it was found that 8/19 isolates (42%) were positive. The efflux pump genes mexB and oprM were studied. The results of this study showed that all isolates of Pseudomonas aeruginosa possess mexB at 100%. While the oprM increased by 57.8%

Author Biography

Okan Ürker, Department of Biology, Graduate School of Natural and Applied Sciences, Çankiri Karatekin University

.

References

Henry, R. Etymologia: Pseudomonas. Emerging Infectious Diseases 2012, 18 (8), 1241. DOI: 10.3201/eid1808.et1808.

Tayeb, L.A.; Ageron, E.; Grimont, F.; Grimont, P.A. Molecular phylogeny of the genus Pseudomonas based on rpoB sequences and application for the identification of isolates. Research in Microbiology 2005, 156(5),763-73. DOI: 10.1016/j.resmic.2005.02.009.

Pang, Z.; Raudonis, R.; Glick, B.R.; Lin, T.J.; Cheng, Z. (2019). Antibiotic resistance in Pseudomonas aeruginosa: mechanisms and alternative therapeutic strategies. Biotechnol Adv 37(1), 177-192. http://dx.doi.org/10.1016/j.biotechadv.2018.11.013

Al-Shwaikh, R.M.A.; AL-Shuwaikh A.M.A.; Alornaaouti, A.F. BOX-PCR Fingerprinting for molecular genotyping of P. aeruginosa. Asian Jr. of Microbiol. Biotech. Env. Sc. 2018, 20(3) : 2018 : 738-743

Diggle, S.P.; Whiteley, M. Microbe Profile: Pseudomonas aeruginosa: Opportunistic Pathogen and Lab Rat. Microbiol. (Reading England) 2020, 166(1), 30–33. http://dx.doi.org/10.1099/mic.0.000860

Rossi, E.; La Rosa, R.; Bartell, J.A.; Marvig, R.L.; Haagensen, J.A.J.; Sommer, L. M.; Molin, S.; Johansen, H.K. Pseudomonas aeruginosa adaptation and evolution in patients with cystic fibrosis. Nature reviews. Microbiology 2021, 19(5), 331–342. https://doi.org/10.1038/s41579-020-00477-5

Al-Saeedi, R., H., A.; Raheema, R.H. Molecular detection of some virulence genes In Pseudomonas aeruginosa clinical isolates. Indian Journal of Public Health Research & Development 2019, 10(4), 769-77.

Seleem, N.M., Abd El Latif, H. K., Shaldam, M. A.; El-Ganiny, A. Drugs with new lease of life as quorum sensing inhibitors: for combating MDR Acinetobacter baumannii infections. Eur. J. Clin. Microbiol. Infect. Dis. 2020, 39, 1687–1702. doi. http://dx.doi.org/10.1007/s10096-020-03882-z

Wang, M.; Zhao, L.; Wu, H.; Zhao, C.; Gong, Q.; Yu, W. Cladodionen is a potential quorum sensing inhibitor against Pseudomonas aeruginosa. Mar. Drugs 2020, 18, E205. http://dx.doi.org/10.3390/md18040205.

Foca, M.; Jakob , K.; Whittier, S.; Latta, P.D.; Factor, S.; Rubenstein, D.; Saiman, L. Endemic Pseudomonas aeruginosa infection in a neonatal intensive care unit. New England Journal of Medicine 2000, 343(10), 695-700.

Jodrá, V. M. ; Díaz-Agero Pérez , C. ; Sainz de Los Terreros Soler, L.; Saa Requejo, C.M.; Dacosta Ballesteros, D.; Quality Control Indicator Working Group. Results of the Spanish national nosocomial infection surveillance network (VICONOS) for surgery patients from January 1997 through December 2003. Am J Infect Control 2006, 34, 134–41. http://dx.doi.org/10.1016/j.ajic.2005.10.004

Krishnamoorthy, G.; Weeks, J.W.; Zhang, Z.; Chandler, C.E.; Xue, H.; Schweizer, H.P.; Ernst, R.K.; Zgurskaya, H.I. Efflux Pumps of Burkholderia thailandensis Control the Permeability Barrier of the Outer Membrane. Antimicrob. Agents Chemother 2019, 63, e00956-19. http://dx.doi.org/10.1128/AAC.00956-19

Kumar, S.; Lekshmi, M.; Parvathi, A.; Ojha, M.; Wenzel, N.; Varela, M.F. Functional and Structural Roles of the Major Facilitator Superfamily Bacterial Multidrug Efflux Pumps. Microorganisms 2020, 8, 266. http://dx.doi.org/10.3390/microorganisms8020266

Al-Saadi Z.H.; Abdullah. R.M. Phenotypic and molecular detection of Escherichia coli efflux pumps from UTI patients. Biochem. Cell. Arch. 2015, 19(Supplement 1), 2371-2376. DOI: 10.35124/bca.2019.19.S1.2371.

Venter, H.; Mowla, R.; Ohene-Agyei, T.; Ma, S. RND-type drug efflux pumps from Gram-negative bacteria: molecular mechanism and inhibition. Front. Microbiol. 2015, 6(377), 1-11. http://dx.doi.org/10.3389/fmicb.2015.00377.

Zhi Li, X.; Elkins, C.A.; Zgurskaya, H.I. Efflux-Mediated Antimicrobial Resistance in Bacteria Mechanisms, Regulation and Clinical Implications. International Publishing. Switzerland pp. 848, 2016.

Shao, X.; Xie, Y.; Zhang, Y.; Liu, J.; Ding, Y.; Wu, M.; Wang, X.; Deng, X. Novel therapeutic strategies for treating Pseudomonas aeruginosa infection. Expert Opinion On Drug Discovery 2020, 15(12), 1403–1423. https://doi.org/10.1080/17460441.2020.1803274 .

Levinson, W. Review of Medical Microbiology and Immunology. 14th ed. McGraw-Hill education, Inc. pp 821, 2016.

Hemraj, V.; Diksha, S.; Avneet, G. A Review on Commonly Used Biochemical Test for Bacteria. IJLS 2013, 1(1), 1-7.

Baron, E.J.; Finegold, S. M.; Peterson, I.L.R. Bailey and Scotts Diagnostic Microbiology. 9th ed. Mosby Company. Missouri, 2007.

CLSI. Performance standards for antimicrobial susceptibility testing. 28th ed. Clinical and Laboratory Standards Institute, Wayne, PA2018 (CLSI supplement M100), 2018.

MacFaddin J. F. Biochemical tests for Identification of Medical Bacteria. 4th ed., Waverly press, Inc., Baltimore, U.S.A, 2004.

Pourahmad, M.; Javadi, A.; Kamran, A.; Ataei, B.; Yaran, M.; Jharomi, A.S. A clinical debate concerning aminoglycoside resistance genes among Pseudomonas aeruginosa strains. Middle East Journal of Family Medicine 2018, 7(10), 172. http://dx.doi.org/10.5742/MEWFM.2018.93327

Al-Dahmoshi, H.O.M. Genotypic and Phenotypic Investigation of Alginate Biofilm Formation among Pseudomonas aeruginosa Isolated from Burn Victims in Babylon, Iraq. Ph.D. thesis, Babylon University, Science Faculty-Biology Department, Iraq, 2013.

Todar, K. Pseudomonas aeruginosa. Textbook of Bacteriology. Science Journal 2011, 304,.14219.

Leboffe, M.J. and Pierce, B.E. A photographic Atlas for the Microbiology Laboratory. 4th ed. Morton Publishing Company. pp96, 2011.

Adekunle, O.C.; Mustapha, A., Odewale, G.; Ojedele, R.O. Detection of antibiotic resistance genes among multiple drug resistant Pseudomonas aeruginosa strains isolated from clinical sources in selected health institutions in Kwara State. Research Journal of Health Sciences 2022, 10(1), 40-48. http://dx.doi.org/10.4314/rejhs.v10i1.5.

Jalil, M. B.; Abdul-Hussein, Z. R.; Al-Hmudi, H. A. Isolation and identification of multi drug resistant biofilm producer with burn wound infection in Basra province/ Iraq. IJDR, 2017, 7(11), 1-5.

Al-Shwaikh, R.M.A.; Alornaaouti, A.F. Detection of tox A gene in Pseudomonas aeruginosa that isolates from different clinical cases by using PCR . Ibn Al-Haitham Journal for Pure and Applied Science. 2017, Special Issue, 26-30. http://dx.doi.org/10.30526/2017.IHSCICONF.1767

Al-Azzawi , S.N.; Abdullah R.M. Study of the resistance of P. aeruginosa isolated from wounds and burns for some disinfects and antiseptic from some Baghdad hospitals. Journal of Pharmaceutical Sciences and Research 2018, 10(6), 1481-1484.

Obaid, S.A.; Al-Shwaikh, R.M. Evaluation the Efficacy of Bacteriophage against Pseudomonas aeruginosa Isolated from Wound and Burn Infections. Pakistan Journal of Medical & Health Sciences 2022, 16(04), APR 2022. http://dx.doi.org/10.53350/pjmhs22164440

Al-Arnaouti, A.F.M. Study of genotyping and some virulence factors for Pseudomonas aeruginosa bacteria. MSc. thesis, College of Education for Pure Sciences, Ibn Al-Haytham, University of Baghdad, 123 pp, 2015.

Peymani, A.; Naserpour-Farivar, T.; Zare, E.; Azarhoosh, K.H. Distribution of blaTEM, blaSHV, and blaCTX-M genes among ESBL producing Pseudomonas aeruginosa isolated from Qazvin and Tehran hospitals. Iran. Journal of Preventive Medicine and Hygiene 2017, 58(2), 155.

Al-Azzawi, S.N.A. Molecular study of the resistance of Pseudomonas aeruginosa isolated from wounds and burns treated with antiseptics. Master's thesis, College of Education for Pure Sciences, Ibn Al-Haitham, University of Baghdad, 2018.

Abdullah R.M. A Study the Effect of TiO2 Nanoparticles Combination with Antibiotics and Plant extracts Against Some Gram Negative Bacteria. Baghdad Science Journal 2016, 13(3), 425-434. http://dx.doi.org/10.21123/bsj.2016.13.3.0425

Helmy, O.M.; Kashef, M.T. Different phenotypic and molecular mechanisms associated with multidrug resistance in Gram-negative clinical isolates from Egypt. Infection and Drug Resistance 2016, 10, 479-498. http://dx.doi.org/10.2147/IDR.S147192.

Abbas, H.A.; El-Ganiny, A.M.; Kamel, H.A. Phenotypic and genotypic detection of antibiotic resistance of Pseudomonas aeruginosa isolated from urinary tract infections. African Health Sciences 2018, 18(1), 11-21. http://dx.doi.org/10.4314/ahs.v18i1.3.

Aghayan, S.S.; Kalalian Mogadam, H. Fazli, M. Darban-Sarokhalil, D. Khoramrooz, S.S.; Jabalameli, F.; Yaslianifard, S.; Mirzaii, M. (2017). The Effects of Berberine and Palmatine on Efflux Pumps Inhibition with Different Gene Patterns in Pseudomonas aeruginosa Isolated from Burn Infections. Avicenna J Med Biotechnol. 2017, 9(1), 2-7.

Murugan, N.; MalathiJ Therese, K.L.; Madhavan, H. N. Application of six multiplex PCR’s among 200 clinical isolates of Pseudomonas aeruginosa for the detection of 20 drug resistance encoding genes. The Kaohsiung Journal of Medical Sciences 2018, 34(2), 79–88,. http://dx.doi.org/10.1016/j.kjms.2017.09.010.

Abdallah A.L.; El Azawy, D.S.H. ; Mohammed, H.A. and El Maghraby, H. M. Expression of Mex AB-Opr M efflux pump system and meropenem resistance in Pseudomonas aeruginosa isolated from surgical intensive care unit. Microbes and Infectoious Diseases 2021, 2(4), 781-789. http://dx.doi.org/10.21608/mid.2021.92720.1187.

Tsutsumi, K. Yonehara, R.; Ishizaka-Ikeda, E.; Miyazaki, N.; Maeda, S.; Iwasaki, K.; Nakagawa, A.; Yamashita, E. Structures of the wild-type MexAB–OprM tripartite pump reveal its complex formation and drug efflux mechanism. Nature communications 2019, 10, 1–10. http://dx.doi.org/10.1038/s41467-019-09463-9.