Overexpression of the adeB Gene in Clinical Isolates of Tigecycline-Nonsusceptible Acinetobacter baumannii without Insertion Mutations in adeRS

Abstract

Thirteen clinical isolates of multidrug-resistant Acinetobacter baumannii resistant to carbapenems (MRAB-C) with tigecycline nonsusceptibility were collected from individual patients in this study. None of the 13 isolates shared the same strain characteristics in molecular typing. All of them showed increased adeB transcription, as predicted. However, none of these tigecycline-nonsusceptible MRAB-C isolates were found to possess previously known adeRS mutations. Upregulation of adeB transcription may result from cross stimulation by other mechanisms. Multidrug-resistantAcinetobacter baumannii resistant to carbapenems (MRAB-C) has been increasingly reported worldwide, raising serious concerns about the limited antimicrobial treatment options (3, 6). Tigecycline (TGC) was therefore recommended for treating MRAB-C infections (4, 7, 13). However, overexpression of the adeABC efflux system regulated by the adeRS two-component system has been reported to result in tigecycline nonsusceptibility of A. baumannii (5,11,12,17,22). We noted more frequent isolation of TGC-nonsusceptible MRAB-C in clinics in Taiwan after the general recommendation of TGC usage against MRAB-C in March 2008. In this study, we investigated clinical characteristics, therapeutic management, and outcomes for patients infected with TGC-nonsusceptible MRAB-C. The mechanism of TGC nonsusceptibility of these clinical isolates was also explored.A total of 17 clinical isolates of MRAB-C were collected at the Tri-Service General Hospital (TSGH) in 2008 and 2009 and subjected to further studies. They were categorized into 3 groups: 4 TGC-susceptible MRAB-C isolates (group I), 4 TGC-intermediate MRAB-C isolates (group II), and 9 TGCresistant MRAB-C isolates (group III). These strains were collected from different individuals who had no overlapping stays in the same intensive care unit (ICU) or the same ward. A. baumannii ATCC 15151 was used as a control. All the isolates were identified using the Vitek 2 system (bioMérieux, Marcy l'Etoile, France), and identifications were confirmed by sequencing of the intergenic spacer (ITS) region of the 16S-23S rRNA genes (2). An A. baumannii isolate was defined as MRAB-C according to a previously described standard (3, 8). The MIC was determined by using the Vitek 2 system and Etest (AB Biodisk, Solna, Sweden). The U.S. Food and Drug Administration-recommended TGC susceptibility breakpoints for Enterobacteriaceae (susceptible, Ϲ2 mg/ml; intermediate, 4 mg/ml; resistant, м8 mg/ml) were used as MIC interpretation criteria. Mueller-Hinton (M-H) agar plates with or without 64 mg/liter 1-(1-naphthylmethyl)-piperazine (NMP; Steinheim, Germany), an efflux pump inhibitor, were used to verify the efflux pump effect on TGC resistance. All isolates were repeatedly investigated for their adeB transcripts by using the ABI Prism 7700 sequence detection system (Applied Biosystems, Warrington, United Kingdom) on three separate occasions. A cutoff threshold (CT) value was used to represent adeB transcripts quantitatively. The ⌬CT for a...

RND-Type Efflux Pumps in Multidrug-Resistant Clinical Isolates of Acinetobacter baumannii: Major Role for AdeABC Overexpression and AdeRS Mutations

“…Overexpression of AdeABC has been shown to confer reduced susceptibility to tigecycline (2,6,10,14,15,16). Thirteen of the 14 clinical isolates had a tigecycline MIC of Ն0.5 g/ml (Table 1) and were considered resistant (23).…”

“…An A 94 V substitution in AdeS was found in an A. baumannii isolate from a patient receiving tigecycline therapy that resulted in 6-fold adeB overexpression and an increase in the tigecycline MIC from 0.5 to 16 g/ml compared to the susceptible strain isolated previously from the same patient (14); however, because of the presence of additional single nucleotide polymorphisms (SNPs) between the two strains, the possibility of nonisogenic clinical isolates was raised (15). Lately, no AdeRS mutations were found in 13 unrelated MDR A. baumannii isolates with increased AdeABC expression and decreased tigecycline susceptibility (16).…”

First Occurrence of OXA-72-Producing Acinetobacter baumannii in Serbia

“…In addition, although no direct evidence of the role of the porins OmpA and OprD in antimicrobial resistance has been demonstrated, the observed changes may act on their functionality and, thus, on membrane permeability to several antimicrobial compounds. Similarly, we have analyzed the sequence of efflux pump regulatory genes, which are known to be involved in antibiotic resistance, namely, the AdeR/S two-component system and AdeN and AdeB, which regulate the AdeIJK and AdeAB efflux pumps, respectively (35)(36)(37). Although only a few substitutions were found in efflux pump regulatory genes, further investigations using reverse transcription-PCR will be necessary to correlate these substitutions with overexpression of these efflux pumps.…”

Antibiotic Resistance, Biofilms and Quorum Sensing in Acinetobacter Species

“…However, in the absence of an adeB gene knockout experiments, it is difficult to ascertain the overall contribution of the AdeABC efflux pump to tigecycline nonsusceptibility (Peleg et al, 2007;Hornsey et al, 2010). But, one recent study demonstrated that overexpression of the adeABC efflux pump resulted in tigecycline nonsusceptibility by quantizing transcripts of the adeB gene and demonstrating conversion of the tigecycline resistance pattern in the presence of an efflux pump inhibitor without any previously known mutation (Sun et al, 2010). When the isolates were analysed separately, there was an association between a higher MIC and elevated adeABC expression, although more isolates would need to be investigated to confirm this observation.…”