Description: We describe the clinical outcome of 17 patients with secondary Acinetobacter bacteremia whose isolates had a tigecycline MIC of ≤2 mg/liter and who received tigecycline within 2 days of bacteremia onset. The 14-day mortality rate of the tigecycline cohort was 41.2% (7/17), which was significantly higher than that of those receiving other appropriate antimicrobial agents (13.8%, 9/65; P = 0.018). However, the percentages of end-stage renal disease and congestive heart failure were higher in the tigecycline cohort. The efficacy of tigecycline was contingent upon the illness severity and bacterial species. Tigecycline should be applied cautiously for treatment of Acinetobacter bacteremia.

Description: Carbapenem-resistant Acinetobacter baumannii (CRAb) shelter cohabiting carbapenem-susceptible bacteria from carbapenem killing via extracellular release of carbapenem-hydrolyzing class D β-lactamases, including OXA-58. However, the mechanism of the extracellular release of OXA-58 has not been elucidated. In silico analysis predicted OXA-58 to be translocated to the periplasm via the Sec system. Using cell fractionation and Western blotting, OXA-58 with the signal peptide and C terminus deleted was not detected in the periplasmic and extracellular fractions. Overexpression of enhanced green fluorescent protein fused to the OXA-58 signal peptide led to its periplasmic translocation but not extracellular release, suggesting that OXA-58 is selectively released. The majority of the extracellular OXA-58 was associated with outer membrane vesicles (OMVs). The OMV-associated OXA-58 was detected only in a strain overexpressing OXA-58. The presence of OXA-58 in OMVs was confirmed by a carbapenem inactivation bioassay, proteomic analysis, and transmission electron microscopy. Imipenem treatment increased OMV formation and caused cell lysis, resulting in an increase in the OMV-associated and OMV-independent release of extracellular OXA-58. OMV-independent OXA-58 hydrolyzed nitrocefin more rapidly than OMV-associated OXA-58 but was more susceptible to proteinase K degradation. Rose bengal, an SecA inhibitor, inhibited the periplasmic translocation and OMV-associated release of OXA-58 and abolished the sheltering effect of CRAb. This study demonstrated that the majority of the extracellular OXA-58 is selectively released via OMVs after Sec-dependent periplasmic translocation. Addition of imipenem increased both OMV-associated and OMV-independent OXA-58, which may have different biological roles. SecA inhibitor could abolish the carbapenem-sheltering effect of CRAb.

Description: Minocycline-based combination therapy has been suggested to be a possible choice for the treatment of infections caused by minocycline-susceptible Acinetobacter baumannii , but its use for the treatment of infections caused by minocycline-resistant A. baumannii is not well established. In this study, we compared the efficacy of minocycline-based combination therapy (with colistin, cefoperazone-sulbactam, or meropenem) to that of colistin in combination with meropenem for the treatment of minocycline-resistant A. baumannii infection. From 2006 to 2010, 191 (17.6%) of 1,083 A. baumannii complex isolates not susceptible to minocycline from the Taiwan Surveillance of Antimicrobial Resistance program were collected. Four representative A. baumannii isolates resistant to minocycline, amikacin, ampicillin-sulbactam, ceftazidime, ciprofloxacin, cefepime, gentamicin, imipenem, levofloxacin, meropenem, and piperacillin-tazobactam were selected on the basis of the diversity of their pulsotypes, collection years, health care setting origins, and geographic areas of origination. All four isolates had tetB and overexpressed adeABC , as revealed by quantitative reverse transcription-PCR. Among all minocycline-based regimens, only the combination with colistin produced a fractional inhibitory concentration index comparable to that achieved with meropenem combined with colistin. Minocycline (4 or 16 μg/ml) in combination with colistin (0.5 μg/ml) also synergistically killed minocycline-resistant isolates in time-kill studies. Minocycline (50 mg/kg of body weight) in combination with colistin (10 mg/kg) significantly improved the survival of mice and reduced the number of bacteria present in the lungs of mice compared to the results of monotherapy. However, minocycline (16 μg/ml)-based therapy was not effective at reducing biofilm-associated bacteria at 24 or 48 h when its effectiveness was compared to that of colistin (0.5 μg/ml) and meropenem (8 μg/ml). The clinical use of minocycline in combination with colistin for the treatment of minocycline-resistant A. baumannii may warrant further investigation.

Description: The rate of recovery of carbapenem-resistant Acinetobacter baumannii (CRAB) isolates has increased significantly in recent decades in Taiwan. This study investigated the molecular epidemiology of CRAB with a focus on the mechanisms of resistance and spread in isolates with bla OXA-23-like or bla OXA-24-like . All 555 CRAB isolates in our multicenter collection, which were recovered from 2002 to 2010, were tested for the presence of class A, B, and D carbapenemase genes. All isolates with bla OXA-23-like or bla OXA-24-like were subjected to pulsed-field gel electrophoresis, and 82 isolates (60 isolates with bla OXA-23-like and 22 isolates with bla OXA-24-like ) were selected for multilocus sequence typing to determine the sequence type (ST) and clonal group (CG) and for detection of additional β-lactamase and aminoglycoside resistance genes. The flanking regions of carbapenem and aminoglycoside resistance genes were identified by PCR mapping and sequencing. The localization of bla OXA was determined by S1 nuclease and I-CeuI assays. The numbers of CRAB isolates carrying bla OXA-23-like or bla OXA-24-like , especially those carrying bla OXA-23-like , increased significantly from 2008 onward. The bla OXA-23-like gene was carried by antibiotic resistance genomic island 1 (AbGRI1)-type structures located on plasmids and/or the chromosome in isolates of different STs (CG92 and novel CG786), whereas bla OXA-24-like was carried on plasmids in CRAB isolates of limited STs (CG92). No class A or B carbapenemase genes were identified. Multiple aminoglycoside resistance genes coexisted in CRAB. Tn 6180 -borne armA was found in 74 (90.2%) CRAB isolates, and 58 (70.7%) isolates had Tn 6179 upstream, constituting AbGRI3. bla TEM was present in 38 (46.3%) of the CRAB isolates tested, with 35 (92.1%) isolates containing bla TEM in AbGRI2-type structures, and 61% of ampC genes had IS Aba1 upstream. We conclude that the dissemination and spread of a few dominant lineages of CRAB containing various resistance island structures occurred in Taiwan.

Description: The Clinical and Laboratory Standards Institute (CLSI) and the European Committee on Antimicrobial Susceptibility Testing (EUCAST) offer different recommendations for carbapenem MIC susceptibility breakpoints for Acinetobacter species. In addition, the clinical efficacy of the intermediate category remains uncertain. This study was designed to determine the optimal predictive breakpoints based on the survival of patients with Acinetobacter bacteremia treated with a carbapenem. We analyzed the 30-day mortality rates of 224 adults who received initial carbapenem monotherapy for the treatment of Acinetobacter bacteremia at 4 medical centers over a 5-year period, according to the carbapenem MICs of the initial isolates. The 30-day mortality was about 2-fold greater in patients whose isolates had carbapenem MICs of ≥8 mg/liter than in those with isolates with MICs of ≤4 mg/liter. The differences were significant by bivariate analysis (53.1% [60/113] versus 25.2% [28/111], respectively; P 〈 0.001) and on survival analysis by the log rank test ( P 〈 0.001). Classification and regression tree analysis revealed a split between MICs of 4 and 8 mg/liter and predicted the same difference in mortality, with a P value of 〈0.001. Carbapenem treatment for Acinetobacter bacteremia caused by isolates with carbapenem MICs of ≥8 mg/liter was an independent predictor of 30-day mortality (odds ratio, 4.218; 95% confidence interval, 2.213 to 8.039; P 〈 0.001). This study revealed that patients with Acinetobacter bacteremia treated with a carbapenem had a more favorable outcome when the carbapenem MICs of their isolates were ≤4 mg/liter than those with MICs of ≥8 mg/liter.

Description: Breakthrough Acinetobacter bacteremia during carbapenem therapy is not uncommon, and it creates therapeutic dilemmas for clinicians. This study was conducted to evaluate the clinical and microbiological characteristics of breakthrough Acinetobacter bacteremia during carbapenem therapy and to assess the efficacy of various antimicrobial therapies. We analyzed 100 adults who developed breakthrough Acinetobacter bacteremia during carbapenem therapy at 4 medical centers over a 6-year period. Their 30-day mortality rate was 57.0%, and the carbapenem resistance rate of their isolates was 87.0%. Among patients with carbapenem-resistant Acinetobacter bacteremia, breakthrough bacteremia during carbapenem therapy was associated with a significantly higher 14-day mortality (51.7% versus 37.4%, respectively; P = 0.025 by bivariate analysis) and a higher 30-day mortality ( P = 0.037 by log rank test of survival analysis) than in the nonbreakthrough group. For the treatment of breakthrough Acinetobacter bacteremia during carbapenem therapy, tigecycline-based therapy was associated with a significantly higher 30-day mortality (80.0%) than those with continued carbapenem therapy (52.5%) and colistin-based therapy (57.9%) by survival analysis ( P = 0.047 and 0.045 by log rank test, respectively). Cox regression controlling for confounders, including severity of illness indices, demonstrated that treatment with tigecycline-based therapy for breakthrough Acinetobacter bacteremia was an independent predictor of 30-day mortality (hazard ratio, 3.659; 95% confidence interval, 1.794 to 7.465; P 〈 0.001). Patients with breakthrough Acinetobacter bacteremia during carbapenem therapy posed a high mortality rate. Tigecycline should be used cautiously for the treatment of breakthrough Acinetobacter bacteremia that develops during carbapenem therapy.

Description: The role of carbapenem-resistant Acinetobacter baumannii (CRAb) in polymicrobial infection remains elusive. Having observed the ability of CRAb to shelter other susceptible bacteria from carbapenem killing, we sought to determine the factors contributing to this sheltering effect by transforming different recombinant plasmids into recipient A. baumannii cells. The sheltering effects of CRAb were reproduced in recipient A. baumannii cells that highly expressed carbapenem-hydrolyzing class D β-lactamases (CHDLs) through their associated strong promoter. With the use of Western blot analysis and a bioassay, the highly expressed CHDLs were found to be extracellularly released and led to hydrolysis of carbapenem. The level of extracellular CHDLs increased after challenge with a higher concentration of CHDL substrates, such as carbapenem and ticarcillin. This increased CHDL may, in part, be attributed to cell lysis, as indicated by the presence of extracellular gyrase. In the planktonic condition, the sheltering effect for the cocultured susceptible bacteria might represent an indirect and passive effect of the CRAb self-defense mechanism, because coculture with the susceptible pathogen did not augment the amount of the extracellular CHDLs. Polymicrobial infection caused by CRAb and a susceptible counterpart exerted higher pathogenicity than monomicrobial infection caused by either pathogen alone in mice receiving carbapenem therapy. This study demonstrated that CHDL-producing CRAb appears to provide a sheltering effect for carbapenem-susceptible pathogens via the extracellular release of CHDLs and, by this mechanism, can enhance the pathogenesis of polymicrobial infection in the presence of carbapenem therapy.

Description: Acinetobacter baumannii biofilms are difficult to eradicate. We investigated the effects of meropenem (2 mg/liter), imipenem (2 mg/liter), sulbactam (4 mg/liter), colistin (2 mg/liter), and tigecycline (2 mg/liter), alone or in combination, on biofilm-embedded carbapenem-resistant and carbapenem-susceptible A. baumannii (CRAb and CSAb, respectively) cells, as well as on the architecture of the biofilms. A. baumannii ATCC 15151 (Ab15151) and its OXA-82-overproducing transformant, along with two clinical CSAb and two clinical CRAb isolates of differing clonalities, were used. The minimal bactericidal concentrations for biofilm-embedded cells of the six tested isolates were 〉50-fold those of their planktonic cells. When used individually, meropenem exhibited a higher killing effect than the other four antimicrobials on biofilm-embedded CSAb cells in the colony biofilm assay. For two clinical CRAb isolates, meropenem plus sulbactam or sulbactam plus tigecycline showed 〉100-fold the bactericidal effect exhibited by these agents used alone after 48 h of treatment. The effect of antimicrobials on the architecture of Ab15151 biofilm emitting green fluorescence was determined by confocal laser scanning microscopy using COMSTAT software. Significant decreases in the maximum biofilm thickness were observed after exposure to meropenem and imipenem. Meropenem plus sulbactam significantly decreased the biomass and mean thickness and increased the roughness coefficient of biofilms, but sulbactam plus tigecycline only decreased the maximum and mean biofilm thickness compared to any of these agents used alone. Meropenem was active against biofilm-embedded CSAb, whereas meropenem plus sulbactam exhibited synergism against biofilm-embedded CRAb and caused significantly more damage to the biofilm architecture than did any of the agents used alone.