Overproduction of efflux pumps caused reduced susceptibility to carbapenem under consecutive imipenem-selected stress in Acinetobacter baumannii

Infection and Drug Resistance Dovepress open access to scientific and medical research O ri g i n a l R e s e a r c h Infection and Drug Resistance downloaded from https://www.dovepress.com/ by 54.37.117.73 on 13-Jul-2018 For personal use only. Open Access Full Text Article Overproduction of efflux pumps caused reduced susceptibility to carbapenem under consecutive imipenem-selected stress in Acinetobacter baumannii This article was published in the following Dove Press journal: Infection and Drug Resistance Yanpeng Zhang 1,2 Zhuocheng Li 2 Xiaolong He 1 Fanglin Ding 2 Weiqing Wu 2 Yong Luo 2 Bing Fan 2 Hong Cao 1 1 Department of Microbiology, Guangdong Provincial Key Laboratory of Tropical Disease Research, School of Public Health, Southern Medical University, Guangzhou, 510515, China; 2 Laboratory Department of the First Affiliated Hospital of Shenzhen University, Shenzhen, 518000, China Purpose: Acinetobacter baumannii is an important pathogen in the nosocomial infections worldwide. Combining with carbapenemases, efflux pumps and outer membrane proteins (OMPs) have been thought to affect the development of carbapenem resistance in A. baumannii. This study aimed to investigate the contributions of different efflux pumps and OMPs in developing carbapenem resistance in a clinical isolate of A. baumannii and reveal the possible mechanism of overproduction of main efflux pumps. Patients and methods: In this study, an imipenem-susceptible clinical isolate was identified as A. baumannii and named SZE. Several common carbapenemases were detected by polymerase chain reaction (PCR). Imipenem-selected mutants were selected from SZE by serial subcultivations on Mueller–Hinton agar, and the minimum inhibitory concentration (MIC) was detected. Gene expressions of four families of efflux pumps, five OMPs, and blaOXA-51 were determined by reverse transcription quantitative PCR, and comparisons were made between SZE strain and the imipenem-selected mutants. The adeRS system in SZE and its mutant was sequenced and aligned. Results: Under consecutive imipenem-selected stress, the MIC to imipenem increased gradually from 0.125 μg/mL to 8 μg/mL. The effect of resistance inducement was almost neutralized when treated with an efflux pump inhibitor. The expression of efflux pumps, adeB, adeG, and adeJ, was increased by 6.9-, 4.0-, and 2.1-fold in mutants, respectively, compared to SZE. A single mutation (G to A) at position 58 was detected in the regulatory adeRS system and possibly upregulated the adeB expression, and then affected the carbapenem resistance in A. baumannii strains. Conclusion: In conclusion, under consecutive imipenem-selected stress in vitro, A. baumannii strain evolved the ability to reduce susceptibility to a variety of antimicrobials by overproduction of efflux pumps. Especially, the resistance-nodulation-cell division super family and a nucleotide mutant in adeRS regulating system caused the overexpression of adeABC. Keywords: Acinetobacter baumannii, efflux pump, multidrug resistance, imipenem, outer membrane protein Introduction Correspondence: Hong Cao Department of Microbiology, Guangdong Provincial Key Laboratory of Tropical Disease Research, School of Public Health, Southern Medical University, No 1023-1063, Sha Tai Road, Guangzhou, 510515, China Tel +86 20 6164 8723 Fax +86 20 6164 8307 Email 457 submit your manuscript | www.dovepress.com Infection and Drug Resistance 2018:11 457–467 Dovepress © 2018 Zhang et al. This work is published and licensed by Dove Medical Press Limited. The full terms of this license are available at https://www.dovepress.com/terms. php and incorporate the Creative Commons Attribution – Non Commercial (unported, v3.0) License (http://creativecommons.org/licenses/by-nc/3.0/). By accessing the work you hereby accept the Terms. Non-commercial uses of the work are permitted without any further permission from Dove Medical Press Limited, provided the work is properly attributed. For permission for commercial use of this work, please see paragraphs 4.2 and 5 of our Terms (https://www.dovepress.com/terms.php). http://dx.doi.org/10.2147/IDR.S151423 Powered by TCPDF (www.tcpdf.org) Acinetobacter baumannii is a Gram-negative nonfermentative coccobacillus. In recent years, this opportunistic pathogen has emerged as one of the main causes of hospital-acquired infections, such as ventilator-associated pneumonia, urinary tract infections, bloodstream infections, and surgical wound infections.1 The risk factors associated with A. baumannii infections include invasive medical procedure, mechanical ventilation, immune suppression, burns, and trauma. Additionally, A. baumannii is one of the most frequently isolated clinical pathogens in China.2 Carbapenems, such as imipenem and meropenem, either alone or in combination with other antibiotics Infection and Drug Resistance downloaded from https://www.dovepress.com/ by 54.37.117.73 on 13-Jul-2018 For personal use only. Zhang et al are currently the most effective therapeutic options for A. baumannii infections. However, carbapenem-resistant clinical isolates of A. baumannii have notably increased in recent years, and have resulted in a delay in treatment. It is significant to elucidate the molecular mechanism underlying the resistance of A. baumannii to carbapenem for solving the prevailing issue. A. baumannii has a plastic genome, which contributes to the acquisition and dissemination of multiple resistance mechanisms. These genome plasticity mechanisms include mutations and insertions of mobile elements, such as plasmids, integrons, transposons, and resistant islands.1 Moreover, the overexpression of intrinsic carbapenemase genes, enzymatic degradation, and modification of the target site decreased permeability, and efflux pumps are common possible mechanisms for the acquisition of resistance. In clinical isolates, chromosomally located resistance island and plasmids carrying different antimicrobial resistance determinants have shown an outstanding ability to rapid evolution of resistance when subjected to the pressure of new antimicrobials.3 Multidrug-resistant Acinetobacter spp. can acquire antimicrobial agent resistance genes via class 1 integrons.4,5 Acquired or endogenous carbapenemase activity, together with decreased outer membrane permeability and overproduction of efflux pumps, constitutes the causes of carbapenem resistance in A. baumannii clinical strains.1,6 Commonly, there are two intrinsic b-lactam hydrolyzing enzymes, AmpC and OXA-51-like, in A. baumannii. Yet these two enzymes show only weak hydrolysis to carbapenems.7 Located upstream of the position of blaOXA-51-like gene, the insertion sequence element ISAba1 has been reported to upregulate the expression of the blaOXA-51-like gene, and accordingly conferred resistance to carbapenem antimicrobials in A. baumannii.8 Main carbapenemases, including Ambler Class D enzymes (also known as oxacillinses, such as OXA-23li (...truncated)