Volume 10, Issue 1 (3-2022)                   JoMMID 2022, 10(1): 36-41 | Back to browse issues page

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Sepehr A, Fereshteh S, Shahrokhi N. Detection of Efflux Pump Using Ethidium Bromide-Agar Cartwheel Method in Acinetobacter baumannii Clinical Isolates. JoMMID 2022; 10 (1) :36-41
URL: http://jommid.pasteur.ac.ir/article-1-410-en.html
Detection of Efflux Pump Using Ethidium Bromide-Agar Cartwheel Method in Acinetobacter baumannii Clinical Isolates
Amin Sepehr , Sepideh Fereshteh , Nader Shahrokhi
Molecular Biology Department, Pasteur Institute of Iran, Tehran, Iran
Abstract:   (1324 Views)
Introduction: In the past decade, multidrug-resistant Acinetobacter baumannii has become one of the most critical challenges in treating infected patients. The AdeABC efflux pump is the most important among the various resistance mechanisms. This pump can force various antibiotics and ethidium bromide out of the bacterial cell to the surrounding environment. Methods: In this study, nine A. baumannii clinical isolates were isolated and identified using different biochemicals (catalase, oxidase, TSI, hemolysis, growth at 44°C, and indole) and molecular (blaOxa51 gene) tests. Following the antibiogram test, the antibiotic resistance changes in the isolates in the presence and absence of efflux pump inhibitor (CCCP) were determined for tetracycline, and ciprofloxacin AdeABC efflux pump genes, including adeA, adeB, and adeC, were amplified by PCR. Finally, the presence of the AdeABC efflux pump was investigated using the agar ethidium-bromide cartwheel method (AEBCM). Results: According to the antibiogram test, all isolates were MDR. In the presence of efflux pump inhibitor, a reduced resistance for tetracycline was observed, but not for ciprofloxacin. The AdeABC efflux pump genes were detected in all isolates. An increase in the AdeABC pump activity in four isolates was confirmed using AEBCM. Conclusion: AEBCM, a fast and convenient tool for assessing the ethidium bromide secretion in various bacteria, provides a quick diagnosis and treatment of multidrug-resistant bacteria.
Keywords: Agar ethidium bromide cartwheel, AdeABC, Acinetobacter baumannii
Full-Text [PDF 2455 kb]   (1128 Downloads)    
Type of Study: Short communication | Subject: Anti-microbial agents, resistance and treatment protocols
Received: 2021/10/12 | Accepted: 2022/03/11 | Published: 2022/04/4
References
1. Khoshbayan A, Shariati A, Shahmoradi S, Baseri Z, Mozafari H. Prevalence and molecular mechanisms of colistin resistance in Acinetobacter baumannii clinical isolates in Tehran, Iran. Acta Microbiol Immunol Hung. 2021; 262-6. [DOI:10.1556/030.2021.01420]
2. Rajamohan G, V.B Srinivasan, W.A Gebreyes. Novel role of Acinetobacter baumannii RND efflux transporters in mediating decreased susceptibility to biocides. J Antimicrob Chemother. 2010; 65 (2): 228-32. [DOI:10.1093/jac/dkp427]
3. Abdi S, Ghotaslou R, Ganbarov K, Mobed A, Tanomand A, Yousefi M, et al. Acinetobacter baumannii efflux pumps and antibiotic resistance. Infect Drug Resist. 2020; 13: 423-34. [DOI:10.2147/IDR.S228089]
4. Xu C, S Bilya, W Xu. AdeABC efflux gene in Acinetobacter baumannii. New Microbes New Infect. 2019; 30: 100549. [DOI:10.1016/j.nmni.2019.100549]
5. Lari A.R, A Ardebili, A Hashemi. AdeR-AdeS mutations & overexpression of the AdeABC efflux system in ciprofloxacin-resistant Acinetobacter baumannii clinical isolates. Indian J Med Res. 2018; 147 (4): 413-21. [DOI:10.4103/ijmr.IJMR_644_16]
6. Verma P, V Tiwari. Targeting outer membrane protein component AdeC for the discovery of efflux pump inhibitor against AdeABC efflux pump of multidrug resistant Acinetobacter baumannii. Cell Biochem Biophys. 2018; 76 (3): 391-400. [DOI:10.1007/s12013-018-0846-5]
7. Shi Y, Xhaing H, Qingy J, Yunthig Y. Mechanism of eravacycline resistance in Acinetobacter baumannii mediated by a deletion mutation in the sensor kinase adeS, leading to elevated expression of the efflux pump AdeABC. Infect Genet Evol. 2020; 80: 104185. [DOI:10.1016/j.meegid.2020.104185]
8. Tegos P, Haynes G, JacobStrouse M, Khan T, Bologa G, Oprea I, et al. Microbial efflux pump inhibition: tactics and strategies. Curr Pharm Des. 2011; 17 (13): 1291-302. [DOI:10.2174/138161211795703726]
9. Pansook S, Incharoensakdi A, Phunpruch S. Effects of the photosystem II inhibitors CCCP and DCMU on hydrogen production by the unicellular halotolerant cyanobacterium Aphanothece halophytica. ScientificWorldJournal. 2019; 1030236. [DOI:10.1155/2019/1030236]
10. Richmond G, KL Chua, LJJoAC Piddock. Efflux in Acinetobacter baumannii can be determined by measuring accumulation of H33342 (bis-benzamide). J Antimicrob Chemother. 2013; 68 (7): 1594-600. [DOI:10.1093/jac/dkt052]
11. Martins M, Cusker M, Viveiros M, Couto I, Fanning S, Pagès J, et al. A simple method for assessment of MDR bacteria for over-expressed efflux pumps. Open Microbiol J. 2013; 7: 72-82. [DOI:10.2174/1874285801307010072]
12. Lin L, B-D Ling, X.-Z Li. Distribution of the multidrug efflux pump genes, adeABC, adeDE and adeIJK, and class 1 integron genes in multiple-antimicrobial-resistant clinical isolates of Acinetobacter baumannii-Acinetobacter calcoaceticus complex. Int J Antimicrob. 2009; 33 (1): 27-32. [DOI:10.1016/j.ijantimicag.2008.06.027]
13. Matthew J, Ellingtona J, Coelhoa J, Turtonb FM. Elaina W, Brown S, et al. Multiplex PCR for genes encoding prevalent OXA carbapenemases in Acinetobacter spp. Int J Antimicrob. 2006: 27 (4): 351-3. [DOI:10.1016/j.ijantimicag.2006.01.004]
14. Martins M, Cusker M, Viveiros M, Couto I, Fanning S, Pagès J, et al. Identification of efflux pump-mediated multidrug-resistant bacteria by the ethidium bromide-agar cartwheel method. In Vivo. 2011; 25 (2): 171-8.
15. Mortazavi SM, Farshadzadeh Z, Janabadi S, Musavi M, Shahi F, Moradi M, et al. Evaluating the frequency of carbapenem and aminoglycoside resistance genes among clinical isolates of Acinetobacter baumannii from Ahvaz, south-west Iran. New Microbes New Infect. 2020; 38: 100779. [DOI:10.1016/j.nmni.2020.100779]
16. Vázquez-López R, Solano-Gálvez SG, Vignon-Whaley JJJ, Vaamonde JAA, Alonzo LAP, Reséndiz AR, et al. Acinetobacter baumannii resistance: a real challenge for clinicians. Antibiotics. 2020; 9 (4): 205. [DOI:10.3390/antibiotics9040205]
17. Hatami R. The frequency of multidrug-resistance and extensively drug-resistant Acinetobacter baumannii in west of Iran. J Clin Microbiol Infect Dis. 2018; 1 (1): 4-8.
18. Tan TY, Lsy Ng, Poh K. Susceptibility testing of unconventional antibiotics against multiresistant Acinetobacter spp. by agar dilution and Vitek 2. Diagn Microbiol Infect Dis. 2007; 58 (3): 357-61. [DOI:10.1016/j.diagmicrobio.2007.02.008]
19. Nazari M, Azizi O, Solgi H, Feresheh H, Shokouhi Sh, Badmasi F. Emergence of carbapenem resistant Acinetobacter baumannii clonal complexes CC2 and CC10 among fecal carriages in an educational hospital. Int J Environ Health Res. 2021; 1-11. [DOI:10.1080/09603123.2021.1892036]
20. Honarmand Jahromy S, Ranjbar S, Pishkar L. Phenotypic Activity of Efflux Pumps by Carbonyl Cyanide M-Chlorophenyl Hydrazone (CCCP) and Mutations in GyrA and ParC Genes Among Ciprofloxacin-Resistant Acinetobacter baumannii Isolates. Jundishapur J Microbiol. 2020; 13 (4): e99435. [DOI:10.5812/jjm.99435]
21. Moazzen Z, Eslami G, Hashemi A. Efflux pump inhibitor carbonyl cyanide 3-chlorophenylhydrazone (CCCP) effect on the minimum inhibitory concentration of ciprofloxacin in acinetobacter baumannii strains. Arch Clin Infect Dis. 2018; 13 (2): e59644. [DOI:10.5812/archcid.59644]
22. Richet H, Fournier PE. Nosocomial infections caused by Acinetobacter baumannii a major threat worldwide. Infect Control Hosp Epidemiol. 2006; 27 (7): 645-6. [DOI:10.1086/505900]
23. Elshahat M, Aboelized E, Rahman RAE, Nabil M, Yassin A. Association between mutations in GYR A/B and PAR C/E genes of Acinetobacter Baumannii clinical isolates and Fluoroquinolones resistance. IMD. 2021; 2 (2): 343-51. [DOI:10.21608/mid.2021.64414.1127]
24. Coyne S, Guigon C, Courvalin P, Périchon B. Screening and quantification of the expression of antibiotic resistance genes in Acinetobacter baumannii with a microarray. Antimicrob Agents Chemother. 2010; 54 (1): 333-40. [DOI:10.1128/AAC.01037-09]
25. Coyne S, Rosenfeld N, Lambert T, Courvalin P, Périchon B. Overexpression of resistance-nodulation-cell division pump AdeFGH confers multidrug resistance in Acinetobacter baumannii. Antimicrob Agents Chemother. 2010; 54 (10): 4389-93. [DOI:10.1128/AAC.00155-10]
26. Yoon E-J, Courvalin P, Grillot-Courvalin C. RND-type efflux pumps in multidrug-resistant clinical isolates of Acinetobacter baumannii: major role for AdeABC overexpression and AdeRS mutations. Antimicrob Agents Chemother. 2013; 57 (7): 2989-95. [DOI:10.1128/AAC.02556-12]
27. Yang H, Huang L, Barnie PA, Su Z, Mi Z, Chen J, et al. Characterization and distribution of drug resistance associated β-lactamase, membrane porin and efflux pump genes in MDR A. baumannii isolated from Zhenjiang, China. Int J Clin Exp Med. 2015; 8 (9): 15393-402.
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This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.