Acinetobacter baumannii: An Emerging Multidrug-resistant Threat

Thomas D Gootz; Andrea Marra

Disclosures

Expert Rev Anti Infect Ther. 2008;6(3):309-325. 

In This Article

Acquired ß-lactamases of

In addition to the presence of the ADC and OXA-51 intrinsic ß-lactamases, similar to other Gram-negatives, A. baumannii can supplement its armamentarium with a number of acquired bla genes, including several Ambler class A serine ß-lactamases ( Table 2 ). Many variants of the TEM, SHV, VEB, PER and CTX-M enzymes are found occasionally in this organism, including many that are extended-spectrum ß-lactamases (ESBLs) with potent activity against third-generation cephalosporins.[81,82,83,84,85,86,87,88,89] Many of these isolates possess MDR phenotypes making it difficult to directly assess the role played by the ESBL alone in conferring ß-lactam resistance.[82] In one surveillance study from Italy, 31 out of 470 (6.6%) of clonally related A. baumannii encoded a TEM-92 ESBL with a pI of 5.9.[82] The blaTEM-92 gene was linked to a Tn3-like transposon (IS26) and the IS was inserted into the tnpR sequence of this gene.[82] The related clones were resistant to all ß-lactams tested, with the exception of the carbapenems. Factors in addition to bla TEM-92, which could have contributed to this extensive ß-lactam re-sistance, were not identified.

Other ESBL variants are not commonly found in Acinetobacters. For instance, despite their rapid dissemination in the Enterobacteriaceae across Europe,[85] the CTX-M ESBLs have only rarely been found in A. baumannii. One study from Argentina described the blaCTX-M-2 gene in a nosocomial isolate of A. baumanii.[90] The gene was part of a class 1 integron and was preceeded upstream by orf513; also in this integron were partial gene sequences for the metal efflux pump gene qacEL, sul1, blaOXA-2 and aac(6')-Ib aminoglycoside acetyltransferase.[90] Interspecies transfer of this integron was in evidence since it was also detected in isolates of Proteus mirabilis, Klebsiella pneumoniae, Enterobacter cloacae, Serratia marcescens and Salmonella spp. isolated from the same hospital.

The VEB and PER ESBLs are more commonly found in A. baumanni; PER-1 has been isolated in strains from Turkey, Korea and France; whereas, the VEB-1 enzyme has been reported in strains from France and Belgium.[87] Interestingly, in A. baumannii AYE isolated in France, the blaVEB-1 gene is part of an 86-kb genomic antibiotic-resistance island, which encodes 44 other antibiotic-resistance genes.[9] Clonal outbreaks of blaVEB-1 containing A. baumannii have been described in northern France in 2003-2004, where 53 hospitals reported 275 cases of infection.[84] The authors point out the difficulty in detecting the VEB phenotype by using cefepime or ceftazidime disks ± clavulanate, owing to the presence of the chromosomally encoded ADC ß-lactamase, which can be overexpressed in some strains. They found that the addition of 200 µg/ml cloxacillin in the agar medium inhibited the class C enzyme, allowing more pronounced reduction in the cephalosporin disk zone due to the inhibitory activity of clavulanate against the ESBL. This complexity may lead to an under-reporting of ESBLs in some surveillance studies.

The Ambler class D oxacillinases appear to be more widely distributed in clinical strains of A. baumannii.[69] The OXA carbapenemases found in Acinetobacter have been organized into either three or four subgroups based on their sequence divergence. One reference designates four subgroups consisting of OXA-23-like, OXA-24-like, OXA-51-like and OXA-58.[91] Unlike the blaOXA-51 enzyme, OXA variants, such as OXA-23 to OXA-27, and OXA-40, -49 and -58, can have substantial carbapenemase hydrolyzing activity depending on the isolate in which they are found ( Table 2 ).[91,92,93] Several studies have attempted to assess the role that OXA carbapenemases play in conferring carbapenem resistance in acinetobacters. The largest subgroup of the three carbapenemases retain the Ambler class D motif Y-G-N at positions 144-146, yet this does not appear to correlate with high carbapenemase activity. The OXA-58 carbapenemase is commonly found in A. baumannii isolates from Spain, Turkey, Greece, Austria, the UK, Argentina, Italy and the US military who have served in Iraq.[69] The blaOXA-58 and blaOXA-51 genes can contain, upstream of the structural gene, ISAba1, ISAba2, ISAba3 and IS18, a genetic fusion that probably upregulates enzyme expression.[69,94]

In order to assess the role that OXA carbapenemases play in conferring carbapenem resistance, the blaOXA-23, blaOXA-40 and blaOXA-58 genes from three different clinical isolates of A. baumannii were cloned onto plasmid vectors and transformed into A. baumannii CIP70.10 and its point mutant, A. baumannii BM4547, which overexpresses the AdeABC efflux pump.[93] This study included three clinical strains (FER, CLA-1 and MAD) that were resistant to all ß-lactams tested, including imipenem and meropenem (summarized in Table 3 ). CLA-1 encoded blaOXA-40 and a knockout construct of this strain was made that resulted in lowered MICs. In a similar fashion, transfer of the blaOXA-23 carbapenemase from clinical strain FER to a cephalosporin- and carbapenem-susceptible A. baumannii CIP70.10 conferred resistance to cefepime and both carbapenems ( Table 3 ). Compared with the control strain CIP70.10, the transformant CIP70.10 (blaOXA-58) had elevated MICs to amoxicillin and the carbapenems, with little increase in MICs to cephalosporins.[93] The same blaOXA-58 plasmid construct transformed into A. baumannii BM4547 (overexpressing the efflux pump) significantly elevated the MICs to cefotaxime and cefepime, as well as increasing the MICs to imipenem and meropenem 16-fold. This particular set of strains illustrates the synergy obtained from both the OXA-58 carbapenemase and increased efflux in limiting access of a drug to its penicillin-binding protein (PBP) targets, elevating the MICs particularly to carbapenems. The broad-spectrum ß-lactam resistance of the transformants was comparable with that of the three clinical strains from which the oxa genes were cloned, suggesting that high-level resistance to the carbapenems is only clinically achievable when these carbapenemases are present with additional factors that reduce antibiotic accumulation in the periplasmic space, either through increased efflux or porin loss.[93]

Results from such detailed studies are valuable since the OXA-58, -23 and -24 carbapenemases are commonly found in A. baumannii strains that are resistant to carbapenems.[91,95,96,97] Recent epidemiological studies utilizing multiplex PCR methods have proven useful for documenting the dissemination of the OXA carbapenemase genes in clinical A. baumannii from the UK.[97,98] Outbreaks of MDR acinetobacters where these blaOXA genes are implicated have occurred in military patients treated at the Walter Reed Army Medical Hospital in the USA,[99] as well as in patients from Belgium,[100] Greece,[101] Italy,[102,103] Spain,[104] Argentina,[105] Australia,[106] Singapore[107] and China.[108] Interestingly, in several of these studies,[106,108] the structural gene for blaOXA was fused immediately upstream with ISAba1, a familiar arrangement implicated in overexpression of the carbapenemase in both A. baumannii and other Gram-negative bacilli.[109] In a study of such resistant acinetobacters from Australia, PCR primers were designed internal to the ISAba1 gene, including the immediate downstream region encoding the blaOXA genes.[106] All of the carbapenem-resistant and none of the carbapenem-susceptible A. baumannii generated an amplicon that, when sequenced, placed the IS adjacent to the blaOXA. In almost all strains examined, the 5'-end of the blaOXA gene was missing 7 bp (CTCTTTT), defining the point of insertion of the element.[106] It was of interest that many of the Australian MDR strains also contained the class I integron gene cassette array aacC1-orfP-orfP-orfQ-aadA1, which was not linked to the ISAba1-blaOXA fusion. This suggests that acquisition of multiple antibiotic-resistance determinants in the Acinetobacter genome favors the survival of such MDR strains under selective pressure of many different classes of antibiotics.

In addition to the Ambler class D carbapenemases, clear evidence exists that the Ambler class B metallo-ß-lactamases VIM, IMP and SIM are present in acinetobacters ( Table 2 ). The SIM-1 carbapenemase shares 64-69% identity with IMP-type enzymes and has been found to be encoded on integrons in isolates from Korea.[110] The genes encoding these metallo enzymes are not intrinsic to this species and evidence is mounting to indicate that the IMP, VIM, SPM-1 and GIM-1 class B enzyme genes are acquired from P. aeruginosa, which may act as a reservoir for these determinants.[111,112,113,114] Furthermore, A. baumannii clinical isolates encoding blaOXA genes together with a metallo-carbapenemase have been identified.[107,115,116] In a study from Singapore, of 44 A. baumanii-expressing carbapenemases, four isolates encoded both the blaOXA-58 and blaIMP-4, while 40 isolates encoded a blaOXA-23 only.[107] Several isolates encoded other blaOXA variants. These data suggest that the IMP-4 gene has disseminated in this population less extensively than in the OXA group.[107] All of the strains encoding both enzymes had imipenem MICs of at least 16 µg/ml, although strains encoding only the blaOXA-23 had comparable MICs to this agent. A very similar result was found when carbapenem-resistant A. baumannii from Shanghai and Hong Kong were characterized.[117]

In a recent surveillance study from Korea, acquired carbapenemase genes were detected in 136 out of 513 (26.5%) of carbapenem-resistant A. baumannii isolated between 2003 and 2004.[118]blaVIM-2 was the most commonly detected metallo-ß-lactamase gene in both A. baumannii and P. aeruginosa. Reports from Poland and Greece also indicate dissemination of the VIM-type gene, either present alone[115] or with the blaOXA-58 gene.[119] Given the propensity for these genes to be encoded on mobile genetic elements, such as transposons and integrons, continued dissemination of these genes has been followed by using multiplex PCR methodology.[116] To facilitate design of adequate PCR primers for multiplex methods, conserved regions of all the available VIM and IMP genes deposited in GenBank can easily be accessed.[162]

ß-lactamases have spread widely in populations of A. baumanii, including those that can encode carbapenem resistance, leaving few choices for effective therapy. The available evidence strongly suggests that additional mechanisms of resistance can be found in the same MDR strain, such as overexpressed efflux and outer membrane alterations that can augment the level of resistance to important antimicrobials. Future studies of MDR strains should also consider these additional resistance determinants in order to gain a clear picture of how MDR populations may acquire additional determinants.

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