Generic placeholder image

Current Protein & Peptide Science

Author(s): Osman Albarri*, Manaf AlMatar*, Melda Meral Öcal and Fatih Köksal

DOI: 10.2174/1389203723666220630162920

DownloadDownload PDF Flyer Cite As
Overexpression of Efflux Pumps AcrAB and OqxAB Contributes to Ciprofloxacin Resistance in Clinical Isolates of K. pneumoniae

Page: [356 - 368] Pages: 13

  • * (Excluding Mailing and Handling)

Abstract

Background: Infection caused by multidrug-resistant K. pneumoniae is regarded as a severe public health concern worldwide, with most countries reporting an increase in fatality rates over time. Efflux pumps are significant determinants of acquired and/or intrinsic resistance in K. pneumoniae.

Objectives: Our aim is to explore efflux-mediated resistance mechanisms in K. pneumoniae by using quantitative real-time PCR in order to evaluate the expression of efflux pump genes (acrA, acrB, oqxA, and oqxB) and pump regulators (marA, soxS, and rarA).

Methods: Efflux pump inhibitor CCCP reduced MIC values of ciprofloxacin by 2 to 64-fold in 43/46 (93%) of MDR-K. pneumoniae isolates.

Results: Compared to the control strain (untreated one), our results demonstrated that acrA, acrB, oqxA, oqxB, marA, soxS, and rarA were overexpressed in 29 (63%), 24 (52%), 29 (63%), 24 (52%), 17 (37%), 16 (35%), and 16 (35%) of K. pneumoniae isolates, respectively. Additionally, a positive correlation was established between the expressions of acrAB and marA (r = 0.50, r = 0.45, respectively) and oqxAB and rarA (r = 0.462912, r = 0.519354, respectively).

Conclusion: Ciprofloxacin resistance was caused by overexpression of the efflux pump genes acrAB and oqxAB, as well as the transcriptional regulators marA, soxS, and rarA in clinical isolates of K. pneumonia.

Keywords: Klebsiella pneumoniae, efflux pumps, ciprofloxacin, CCCP, RT-PCR, transcriptional regulators.

Graphical Abstract

[1]
Giske, C.G.; Monnet, D.L.; Cars, O.; Carmeli, Y. Clinical and economic impact of common mul-tidrug-resistant gram-negative bacilli. Antimicrob. Agents Chemother., 2008, 52(3), 813-821.
[http://dx.doi.org/10.1128/AAC.01169-07] [PMID: 18070961]
[2]
Osman, A.; Işıl, V.; Fatih, K. Microbial sidero-phores: Potential medicinal applications of the si-derophores. J. Biotechnol. Sci. Res, 2019, 6, 32.
[3]
AlMatar, M.; Albarri, O.; Makky, E.A.; Var, I.; Köksal, F. An overview of the activities of cefiderocol against sensitive and multidrug- re-sistant (MDR) bacteria. Mini Rev. Med. Chem., 2020, 20(18), 1908-1916.
[http://dx.doi.org/10.2174/1389557520666200818211405] [PMID: 32811410]
[4]
AlMatar, M.; Albarri, O.; Makky, E.A.; Var, I.; Köksal, F. A glance on the role of bacterial sidero-phore from the perspectives of medical and bio-technological approaches. Curr. Drug Targets, 2020, 21(13), 1326-1343.
[http://dx.doi.org/10.2174/1389450121666200621193018] [PMID: 32564749]
[5]
Livermore, D.M. Current epidemiology and growing resistance of gram-negative pathogens. Korean J. Intern. Med. (Korean. Assoc. Intern. Med.), 2012, 27(2), 128-142.
[http://dx.doi.org/10.3904/kjim.2012.27.2.128] [PMID: 22707882]
[6]
Okusu, H.; Ma, D.; Nikaido, H. AcrAB efflux pump plays a major role in the antibiotic resistance phenotype of Escherichia coli multiple-antibiotic-resistance (Mar) mutants. J. Bacteriol., 1996, 178(1), 306-308.
[http://dx.doi.org/10.1128/jb.178.1.306-308.1996] [PMID: 8550435]
[7]
Guillard, T.; de Jong, A.; Limelette, A.; Lebreil, A.L.; Madoux, J.; de Champs, C.; Group, C.S. Characterization of quinolone resistance mecha-nisms in enterobacteriaceae recovered from dis-eased companion animals in Europe. Vet. Microbiol., 2016, 194, 23-29.
[http://dx.doi.org/10.1016/j.vetmic.2015.11.033] [PMID: 26701806]
[8]
Osei Sekyere, J.; Amoako, D.G. Carbonyl cya-nide m-chlorophenylhydrazine (CCCP) reverses resistance to colistin, but not to carbapenems and tigecycline in multidrug-resistant Enterobacteri-aceae. Front. Microbiol., 2017, 8, 228.
[http://dx.doi.org/10.3389/fmicb.2017.00228] [PMID: 28261184]
[9]
AlMatar, M.; Albarri, O.; Makky, E.A.; Köksal, F. Efflux pump inhibitors: New updates. Pharmacol. Rep., 2021, 73(1), 1-16.
[http://dx.doi.org/10.1007/s43440-020-00160-9] [PMID: 32946075]
[10]
Jorgensen, J.H.; Ferraro, M.J.; Jorgensen, J.H.; Ferraro, M.J. Antimicrobial susceptibility testing: A review of general principles and contemporary practices. Clin. Infect. Dis., 2009, 49(11), 1749-1755.
[http://dx.doi.org/10.1086/647952] [PMID: 19857164]
[11]
Barry, A.L. An overview of the Clinical and Laboratory Standards Institute (CLSI) and its impact on antimicrobial susceptibility tests. In. Richard, S.; Lynn S.M.; Avery C.G. (Eds.), Antimicro-bial susceptibility testing protocols., 2007. CRC press
[12]
Dufour, P.; Colon, M. The tetrazolium reduc-tion method for assessing the viability of individu-al bacterial cells in aquatic environments: Im-provements, performance and applications. Hydrobiologia, 1992, 232(3), 211-218.
[http://dx.doi.org/10.1007/BF00013706]
[13]
Razavi, S.; Mirnejad, R.; Babapour, E. In-volvement of AcrAB and OqxAB efflux pumps in antimicrobial resistance of clinical isolates of Klebsiella pneumonia. J. Appl. Biotechnol. Reports, 2020, 7(4), 251.
[14]
AlMatar, M.; Var, I.; Kayar, B.; Köksal, F. Differential expression of resistant and efflux pump genes in MDR-TB isolates. Endocr. Metab. Immune Disorders-Drug Targets, 2020, 20(2), 271.
[http://dx.doi.org/10.2174/1871530319666191009153834]
[15]
Schmittgen, T.D.; Livak, K.J. Analyzing real-time PCR data by the comparative C(T) method. Nat. Protoc., 2008, 3(6), 1101-1108.
[http://dx.doi.org/10.1038/nprot.2008.73] [PMID: 18546601]
[16]
Livak, K.J.; Schmittgen, T.D. Analysis of rela-tive gene expression data using real-time quantita-tive PCR and the 2− ΔΔCT method. Methods, 2001, 25(4), 402.
[17]
Huet, A.A.; Raygada, J.L.; Mendiratta, K.; Seo, S.M.; Kaatz, G.W. Multidrug efflux pump overexpression in Staphylococcus aureus after sin-gle and multiple in vitro exposures to biocides and dyes. Microbiology, 2008, 154(Pt 10), 3144-3153.
[http://dx.doi.org/10.1099/mic.0.2008/021188-0] [PMID: 18832320]
[18]
Rodrigues, L.; Machado, D.; Couto, I.; Am-aral, L.; Viveiros, M. Contribution of efflux activi-ty to isoniazid resistance in the Mycobacterium tu-berculosis complex. Infect. Genet. Evol., 2012, 12(4), 695-700.
[http://dx.doi.org/10.1016/j.meegid.2011.08.009] [PMID: 21871582]
[19]
Zheng, J.X.; Lin, Z.W.; Sun, X.; Lin, W.H.; Chen, Z.; Wu, Y.; Qi, G.B.; Deng, Q.W.; Qu, D.; Yu, Z.J. Overexpression of OqxAB and MacAB efflux pumps contributes to eravacycline resistance and heteroresistance in clinical isolates of Klebsiel-la pneumoniae. Emerg. Microbes Infect., 2018, 7(1), 139.
[http://dx.doi.org/10.1038/s41426-018-0141-y] [PMID: 30068997]
[20]
Heiat, M.; Rezaeimehr, M.; Moghaddam, M.; Ranjbar, R.; Najafi, A. Molecular genetic analysis of quinolone resistance-determining region of DNA Gyrase-A in fluoroquinolones resistant Klebsiella pneumoniae based on GenBank data and reported studies. Mol. Gen. Microbiol. Virol., 2014, 29(4), 211-215.
[http://dx.doi.org/10.3103/S0891416814040041]
[21]
Roodsari, M.R.; Fallah, F.; Taherpour, A.; Vala, M.H.; Hashemi, A. Carbapenem-resistant bacteria and laboratory detection methods. Arch. Pediatr. Infect. Dis., 2014, 2(1), 188.
[22]
Bialek-Davenet, S.; Lavigne, J-P.; Guyot, K.; Mayer, N.; Tournebize, R.; Brisse, S.; Leflon-Guibout, V.; Nicolas-Chanoine, M-H. Differential contribution of AcrAB and OqxAB efflux pumps to multidrug resistance and virulence in Klebsiella pneumoniae. J. Antimicrob. Chemother., 2015, 70(1), 81-88.
[http://dx.doi.org/10.1093/jac/dku340] [PMID: 25193085]
[23]
Gabr, B.M.; Zamzam, A.S.A.; Eisa, E.A.; El-Baradey, G.F.; Eldeen, M.A.S. Detection of oqxA and oqxB efflux pump genes among nosocomial coliform bacilli: An observational cross-sectional study. J. Acute Dis., 2021, 10(3), 117.
[http://dx.doi.org/10.4103/2221-6189.316676]
[24]
Park, Y.; Choi, Q.; Kwon, G.C.; Koo, S.H. Molecular epidemiology and mechanisms of tigecycline resistance in carbapenem-resistant Klebsiella pneumoniae isolates. J. Clin. Lab. Anal., 2020, 34(12), e23506.
[http://dx.doi.org/10.1002/jcla.23506] [PMID: 32815626]
[25]
Khoshnood, S.; Heidary, M.; Hashemi, A.; Shahi, F.; Saki, M.; Kouhsari, E.; Eslami, G.; Gou-darzi, H. Involvement of the AcrAB efflux pump in ciprofloxacin resistance in clinical Klebsiella pneumoniae isolates. Infect. Disorders-Drug Tar-gets, 2021, 21(4), 564.
[http://dx.doi.org/10.2174/1871526520999200905121220]
[26]
Lee, Y-J.; Huang, C-H.; Ilsan, N.A.; Lee, I-H.; Huang, T-W. Molecular epidemiology and characterization of carbapenem-resistant Klebsiella pneumoniae isolated from urine at a teaching hos-pital in Taiwan. Microorganisms, 2021, 9(2), 271.
[http://dx.doi.org/10.3390/microorganisms9020271] [PMID: 33525554]
[27]
Elgendy, S.G.; Abdel Hameed, M.R.; El-Mokhtar, M.A. Tigecycline resistance among Klebsiella pneumoniae isolated from febrile neu-tropenic patients. J. Med. Microbiol., 2018, 67(7), 972-975.
[http://dx.doi.org/10.1099/jmm.0.000770] [PMID: 29799385]
[28]
Li, J.; Xu, Q.; Ogurek, S.; Li, Z.; Wang, P.; Xie, Q.; Sheng, Z.; Wang, M. Efflux pump acrab confers decreased susceptibility to piperacillin-tazobactam and ceftolozane-tazobactam in tigecy-cline-non-susceptible Klebsiella pneumoniae. Infect. Drug Resist., 2020, 13, 4309-4319.
[http://dx.doi.org/10.2147/IDR.S279020] [PMID: 33273833]
[29]
Blanco, P.; Hernando-Amado, S.; Reales-Calderon, J.A.; Corona, F.; Lira, F.; Alcalde-Rico, M.; Bernardini, A.; Sanchez, M.B.; Martinez, J.L. Bacterial multidrug efflux pumps: Much more than antibiotic resistance determinants. Microorganisms, 2016, 4(1), 14.
[http://dx.doi.org/10.3390/microorganisms4010014] [PMID: 27681908]
[30]
Correia, S.; Poeta, P.; Hébraud, M.; Capelo, J.L.; Igrejas, G. Mechanisms of quinolone action and resistance: Where do we stand? J. Med. Microbiol., 2017, 66(5), 551-559.
[http://dx.doi.org/10.1099/jmm.0.000475] [PMID: 28504927]
[31]
Chen, Y.; Hu, D.; Zhang, Q.; Liao, X-P.; Liu, Y-H.; Sun, J. Efflux pump overexpression contrib-utes to tigecycline heteroresistance in Salmonella enterica serovar Typhimurium. Front. Cell. Infect. Microbiol., 2017, 7, 37.
[http://dx.doi.org/10.3389/fcimb.2017.00037] [PMID: 28261566]
[32]
Jomehzadeh, N.; Ahmadi, K.; Bahmanshiri, M.A. Investigation of plasmid-mediated quinolone resistance genes among clinical isolates of Klebsiel-la pneumoniae in southwest Iran. J. Clin. Lab. Anal., 2022, e24342.
[http://dx.doi.org/10.1002/jcla.24342] [PMID: 35293043]
[33]
Szabo, O.; Kocsis, B.; Szabo, N.; Kristof, K.; Szabo, D. Contribution of OqxAB efflux pump in selection of fluoroquinolone-resistant Klebsiella pneumoniae. Can. J. Infect. Dis. Med. Microbiol., 2018, 2018, 4271638.
[http://dx.doi.org/10.1155/2018/4271638]
[34]
Chetri, S.; Das, B.J.; Bhowmik, D.; Chanda, D.D.; Chakravarty, A.; Bhattacharjee, A. Tran-scriptional response of mar, sox and rob regulon against concentration gradient carbapenem stress within Escherichia coli isolated from hospital ac-quired infection. BMC Res. Notes, 2020, 13(1), 168.
[http://dx.doi.org/10.1186/s13104-020-04999-2] [PMID: 32192538]
[35]
Zhang, Q.; Lin, L.; Pan, Y.; Chen, J. Charac-terization of tigecycline heteroresistant Klebsiella pneumoniae clinical isolates from a Chinese ter-tiary care teaching hospital. Front. Microbiol., 2021, 12, 671153.
[http://dx.doi.org/10.3389/fmicb.2021.671153] [PMID: 34413834]
[36]
Veleba, M.; De Majumdar, S.; Hornsey, M.; Woodford, N.; Schneiders, T. Genetic characteri-zation of tigecycline resistance in clinical isolates of Enterobacter cloacae and Enterobacter aero-genes. J. Antimicrob. Chemother., 2013, 68(5), 1011-1018.
[http://dx.doi.org/10.1093/jac/dks530] [PMID: 23349441]
[37]
Jiménez-Castellanos, J-C.; Wan Ahmad Kamil, W.N.I.; Cheung, C.H.P.; Tobin, M.S.; Brown, J.; Isaac, S.G.; Heesom, K.J.; Schneiders, T.; Avison, M.B. Comparative effects of overpro-ducing the AraC-type transcriptional regulators MarA, SoxS, RarA and RamA on antimicrobial drug susceptibility in Klebsiella pneumoniae. J. Antimicrob. Chemother., 2016, 71(7), 1820-1825.
[http://dx.doi.org/10.1093/jac/dkw088] [PMID: 27029850]
[38]
Zhong, X.; Xu, H.; Chen, D.; Zhou, H.; Hu, X.; Cheng, G. First emergence of acrAB and oqx-AB mediated tigecycline resistance in clinical iso-lates of Klebsiella pneumoniae pre-dating the use of tigecycline in a Chinese hospital. PLoS One, 2014, 9(12), e115185.
[http://dx.doi.org/10.1371/journal.pone.0115185] [PMID: 25503276]