Mechanisms of carbapenem resistance in Acinetobacter species and synergistic activities of antibiotics against carbapenem-resistant Acinetobacter species

Abstract

Acinetobacter calcoaceticus-A. baumannii complex is one of the most important opportunistic pathogens that cause nosocomial infection. Carbapenem resistance in Acinetobacter spp. has been increasingly reported worldwide. The objectives of this study were to investigate the prevalence and mechanisms of carbapenem resistance in Acinetobacter spp. Thai isolates and to determine the synergistic effects of antibiotic combinations against carbapenem-resistant Acinetobacter spp. The prevalences of carbapenem resistance in A. baumannii, A. pittii and A. nosocomialis were 86.6%, 26.3% and 22.7%, respectively. The most common carbapenemase gene in carbapenem-resistant A. baumannii, A. pittii and A. nosocomialis was blaOXA-23. The representative isolates carrying carbapenemase genes including 23 A. baumannii, 8 A. pittii and 5 A. nosocomialis were further investigated. The reduction of outer membrane proteins was observed in 9 A. baumannii and 4 A. pittii isolates, but was not detected in A. nosocomialis. The carO gene of A. baumannii AB58 and AB97 was disrupted by a novel insertion sequence, ISAba40. No overexpression of efflux pump was found in A. baumannii and A. nosocomialis, but was observed in 3 A. pittii isolates. The clonal study showed that the major clones of A. baumannii were ST195 and ST542, those of A. pittii were ST1178 and ST1425. A. nosocomialis belonged to a single clone, ST598. Analysis of antibiotic resistance islands demonstrated 4 subtypes of AbaR4 island. AbaR4 subtype carried blaOXA-23-like was found in A. baumannii and A. nosocomialis. The in vitro activities of antibiotic combinations including imipenem and meropenem plus amikacin, colistin or fosfomycin were evaluated. The best combination was imipenem plus fosfomycin which showed synergism in 65.2%, 62.5% and 80% of A. baumannii, A. pittii and A. nosocomialis, respectively. The synergism was confirmed by time-kill assay. The results were similar to checkerboard assay except that of A. baumannii AB250. All of time-kill tested isolates were resistant to fosfomycin, but the resistance mechanisms including MurA mutation, fosfomycin-modifying enzyme production, overexpression of efflux pump and cell wall recycling bypass pathway were not detected. The population analysis of Acinetobacter spp. showed subpopulations that had higher fosfomycin MICs. Tolerance to imipenem was found in A. baumannii AB250. The results indicate that imipenem tolerance may be associated with synergism when imipenem is combined with fosfomycin. The results showed that carbapenemase production is the major mechanism of carbapenem resistance in Acinetobacter spp. This is the first report of carbapenem-resistant A. pittii carrying blaOXA-58 and blaIMP-14a and the disruption of the carO gene by a novel ISAba40 in carbapenem-resistant A. baumannii. This study suggests that combination of imipenem and fosfomycin may become the effective combination therapy for treatment of carbapenem-resistant Acinetobacter spp. infection.