Search tips
Search criteria

Results 1-25 (1140072)

Clipboard (0)

Related Articles

1.  Atomic Force Microscopy Investigation of the Morphology and Topography of Colistin-Heteroresistant Acinetobacter baumannii Strains as a Function of Growth Phase and in Response to Colistin Treatment▿  
Antimicrobial Agents and Chemotherapy  2009;53(12):4979-4986.
The prevalence of infections caused by multidrug-resistant gram-negative Acinetobacter baumannii strains and the lack of novel antibiotics under development are posing a global dilemma, forcing a resurgence of the last-line antibiotic colistin. Our aim was to use atomic force microscopy (AFM) to investigate the morphology and topography of paired colistin-susceptible and -resistant cells from colistin-heteroresistant A. baumannii strains as a function of bacterial growth phase and colistin exposure. An optimal AFM bacterial sample preparation protocol was established and applied to examine three paired strains. Images revealed rod-shaped colistin-susceptible cells (1.65 ± 0.27 μm by 0.98 ± 0.07 μm) at mid-logarithmic phase, in contrast to spherical colistin-resistant cells (1.03 ± 0.09 μm); the latter were also more diverse in appearance and exhibited a rougher surface topography (7.05 ± 1.3 nm versus 11.4 ± 2.5 nm for susceptible versus resistant, respectively). Cellular elongation up to ∼18 μm at stationary phase was more commonly observed in susceptible strains, although these “worm-like” cells were also observed occasionally in the resistant population. The effects of colistin exposure on the cell surface of colistin-susceptible and -resistant cells were found to be similar; topographical changes were minor in response to 0.5 μg/ml colistin; however, at 4 μg/ml colistin, a significant degree of surface disruption was detected. At 32 μg/ml colistin, cellular clumping and surface smoothening were evident. Our study has demonstrated for the first time substantial morphological and topographical differences between colistin-susceptible and -resistant cells from heteroresistant A. baumannii strains. These results contribute to an understanding of colistin action and resistance in regard to this problematic pathogen.
PMCID: PMC2786353  PMID: 19786595
2.  Different surface charge of colistin-susceptible and -resistant Acinetobacter baumannii cells measured with zeta potential as a function of growth phase and colistin treatment 
Electrostatic forces mediate the initial interaction between cationic colistin and Gram-negative bacterial cells. Lipopolysaccharide (LPS) loss mediates colistin resistance in some A. baumannii strains. Our aim was to determine the surface charge of colistin-susceptible and –resistant A. baumannii as a function of growth phase and in response to polymyxin treatment.
The zeta potential of A. baumannii ATCC 19606 and 10 clinical multidrug-resistant strains (MICs 0.5–2 mg/L) was assessed. Colistin-resistant derivatives (MIC >128 mg/L) of wild-type strains were selected in the presence of 10 mg/L colistin, including the LPS-deficient lpxA mutant, ATCC 19606R. To determine the contribution of LPS to surface charge, two complemented ATCC 19606R derivatives were examined, namely ATCC 19606R + lpxA (containing an intact lpxA gene) and ATCC 19606R + V (containing empty vector). Investigations were conducted as a function of growth phase and polymyxin treatment (1, 4 and 8 mg/L).
Wild-type cells exhibited a greater negative charge (−60.5 ± 2.36 to −26.2 ± 2.56 mV) thancolistin-resistant cells (−49.2 ± 3.09 to −19.1 ± 2.80 mV) at mid-log phase (ANOVA, P < 0.05). Opposing growth-phase trends were observed for both phenotypes: wild-type cells displayed reduced negative charge and colistin-resistant cells displayed increased negative charge at stationary compared with mid-logarithmic phase. Polymyxin exposure resulted in a concentration-dependent increase in zeta potential. Examination of ATCC 19606R and complemented strains supported the importance of LPS in determining surface charge, suggesting a potential mechanism of colistin resistance.
Zeta potential differences between A. baumannii phenotypes probably reflect compositional outer-membrane variations that impact the electrostatic component of colistin activity.
PMCID: PMC3001852  PMID: 21081544
physicochemical properties; Gram-negative; polymyxin
3.  Cell surface hydrophobicity of colistin-susceptible versus -resistant Acinetobacter baumannii determined by contact angles: methodological considerations and implications 
Journal of applied microbiology  2012;113(4):940-951.
Contact angle analysis of cell surface hydrophobicity (CSH) describes the tendency of a water droplet to spread across a lawn of filtered bacterial cells. Colistin-induced disruption of the Gram-negative outer membrane necessitates hydrophobic contacts with lipopolysaccharide (LPS). We aimed to characterize the CSH of Acinetobacter baumannii using contact angles, to provide insight into the mechanism of colistin resistance.
Contact angles were analysed for five paired colistin-susceptible and -resistant A. baumannii strains. Drainage of the water droplet through bacterial layers was demonstrated to influence results. Consequently, measurements were performed 0.66-sec after droplet deposition. Colistin-resistant cells exhibited lower contact angles (38.8±2.8° to 46.8±1.3°) compared to their paired-susceptible strains (40.7±3.0° to 48.0±1.4°; ANOVA; p<0.05). Contact angles increased at stationary phase (50.3±2.9° to 61.5±2.5° and 47.4±2.0° to 50.8±3.2°, susceptible and resistant, respectively, ANOVA; p<0.05), and in response to colistin 32-mgL−1 exposure (44.5±1.5° to 50.6±2.8° and 43.5±2.2° to 48.0±2.2°, susceptible and resistant, respectively; ANOVA; p<0.05). Analysis of complemented strains constructed with an intact lpxA gene, or empty vector, highlighted the contribution of LPS to CSH.
Compositional outer-membrane variations likely account for CSH differences between A. baumannii phenotypes, which influence the hydrophobic colistin-bacterium interaction.
Important insight into the mechanism of colistin resistance has been provided. Greater consideration of contact angle mehodology is nescessary to ensure accurate analyses are performed.
PMCID: PMC3434258  PMID: 22574702
Antimicrobials; Lipopolysaccharide; Mechanism of Action
4.  Inhaled Colistin for Treatment of Pneumonia due to Colistin-Only-Susceptible Acinetobacter baumannii 
Yonsei Medical Journal  2013;55(1):118-125.
Colistin is used for the treatment of pneumonia associated with multidrug-resistant Acinetobacter baumannii and Pseudomonas aeruginosa. However, the best route of administration and dosage is not known. We report our experience with aerosolized colistin in twelve patients with pneumonia caused by colistin-only-susceptible (COS) A. baumannii.
Materials and Methods
We retrospectively reviewed patients' medical records who were treated with aerosolized colistin for the treatment of pneumonia.
Ten patients were treated only with aerosolized colistin inhalation and two patients received a 3-day course intravenous colistin, and then switched to colistin inhalation therapy. The median duration of aerosolized colistin therapy was 17 days (5-31 days). Four patients were treated only with aerosolized colistin, whereas 4 patients received concomitant glycopeptides, and 4 received concomitant levofloxacin or cefoperazone/sulbactam. At the end of the therapy, the clinical response rate and bacteriological clearance rate was 83% and 50%, respectively. Colistin-resistant strains were isolated from 3 patients after aerosolized colistin therapy; however, all of them showed favorable clinical response. The median interval between inhalation therapy and resistance was 7 days (range 5-19 days). Acute kidney injury developed in 3 patients. Two patients experienced Clostridium difficile associated diarrhea. One patient developed fever and skin rash after aerosolized colistin therapy. No patient developed neurotoxicity or bronchospasm.
Colistin inhalation therapy is deemed tolerable and safe, and could be beneficial as an adjuctive therapy for the management of pneumonia due to COS A. baumannii. However, the potential development of colistin resistance cannot be overlooked.
PMCID: PMC3874897  PMID: 24339296
Acinetobacter baumannii; colistin; pneumonia
5.  Heteroresistance to Colistin in Multidrug-Resistant Acinetobacter baumannii 
Multidrug-resistant Acinetobacter baumannii has emerged as a significant clinical problem worldwide and colistin is being used increasingly as “salvage” therapy. MICs of colistin against A. baumannii indicate its significant activity. However, resistance to colistin in A. baumannii has been reported recently. Clonotypes of 16 clinical A. baumannii isolates and ATCC 19606 were determined by pulsed-field gel electrophoresis (PFGE), and colistin MICs were measured. The time-kill kinetics of colistin against A. baumannii ATCC 19606 and clinical isolate 6 were investigated, and population analysis profiles (PAPs) were conducted. Resistance development was investigated by serial passaging with or without exposure to colistin. Five different PFGE banding patterns were found in the clinical isolates. MICs of colistin against all isolates were within 0.25 to 2 μg/ml. Colistin showed early concentration-dependent killing, but bacterial regrowth was observed at 24 h. PAPs revealed that heteroresistance to colistin occurred in 15 of the 16 clinical isolates. Subpopulations (<0.1% from inocula of 108 to 109 CFU/ml) of ATCC 19606, and most clinical isolates grew in the presence of colistin 3 to 10 μg/ml. Four successive passages of ATCC 19606 in broth containing colistin (up to 200 μg/ml) substantially increased the proportion of the resistant subpopulations able to grow in the presence of colistin at 10 μg/ml from 0.000023 to 100%; even after 16 passages in colistin-free broth, the proportion only decreased to 2.1%. This represents the first demonstration of heterogeneous colistin-resistant A. baumannii in “colistin-susceptible” clinical isolates. Our findings give a strong warning that colistin-resistant A. baumannii may be observed more frequently due to potential suboptimal dosage regimens recommended in the product information of some products of colistin methanesulfonate.
PMCID: PMC1563544  PMID: 16940086
6.  Biological Cost of Different Mechanisms of Colistin Resistance and Their Impact on Virulence in Acinetobacter baumannii 
Two mechanisms of resistance to colistin have been described in Acinetobacter baumannii. One involves complete loss of lipopolysaccharide (LPS), resulting from mutations in lpxA, lpxC, or lpxD, and the second is associated with phosphoethanolamine addition to LPS, mediated through mutations in pmrAB. In order to assess the clinical impacts of both resistance mechanisms, A. baumannii ATCC 19606 and its isogenic derivatives, AL1851 ΔlpxA, AL1852 ΔlpxD, AL1842 ΔlpxC, and ATCC 19606 pmrB, were analyzed for in vitro growth rate, in vitro and in vivo competitive growth, infection of A549 respiratory alveolar epithelial cells, virulence in the Caenorhabditis elegans model, and virulence in a systemic mouse infection model. The in vitro growth rate of the lpx mutants was clearly diminished; furthermore, in vitro and in vivo competitive-growth experiments revealed a reduction in fitness for both mutant types. Infection of A549 cells with ATCC 19606 or the pmrB mutant resulted in greater loss of viability than with lpx mutants. Finally, the lpx mutants were highly attenuated in both the C. elegans and mouse infection models, while the pmrB mutant was attenuated only in the C. elegans model. In summary, while colistin resistance in A. baumannii confers a clear selective advantage in the presence of colistin treatment, it causes a noticeable cost in terms of overall fitness and virulence, with a more striking reduction associated with LPS loss than with phosphoethanolamine addition. Therefore, we hypothesize that colistin resistance mediated by changes in pmrAB will be more likely to arise in clinical settings in patients treated with colistin.
PMCID: PMC3910726  PMID: 24189257
7.  Activities of colistin- and minocycline-based combinations against extensive drug resistant Acinetobacter baumannii isolates from intensive care unit patients 
BMC Infectious Diseases  2011;11:109.
Extensive drug resistance of Acinetobacter baumannii is a serious problem in the clinical setting. It is therefore important to find active antibiotic combinations that could be effective in the treatment of infections caused by this problematic 'superbug'. In this study, we analyzed the in vitro activities of three colistin-based combinations and a minocycline-based combination against clinically isolated extensive drug resistant Acinetobacter baumannii (XDR-AB) strains.
Fourteen XDR-AB clinical isolates were collected. The clonotypes were determined by polymerase chain reaction-based fingerprinting. Susceptibility testing was carried out according to the standards of the Clinical and Laboratory Standards Institute. Activities of drug combinations were investigated against four selected strains and analyzed by mean survival time over 12 hours (MST12 h) in a time-kill study.
The time-kill studies indicated that the minimum inhibitory concentration (MIC) of colistin (0.5 or 0.25 μg/mL) completely killed all strains at 2 to 4 hours, but 0.5×MIC colistin showed no bactericidal activity. Meropenem (8 μg/mL), minocycline (1 μg/mL) or rifampicin (0.06 μg/mL) did not show bactericidal activity. However, combinations of colistin at 0.5×MIC (0.25 or 0.125 μg/mL) with each of the above were synergistic and shown bactericidal activities against all test isolates. A combination of meropenem (16 μg/mL) with minocycline (0.5×MIC, 4 or 2 μg/mL) was synergitic to all test isolates, but neither showed bactericidal activity alone. The MST12 h values of drug combinations (either colistin- or minocycline-based combinations) were significantly shorter than those of the single drugs (p < 0.01).
This study indicates that combinations of colistin/meropenem, colistin/rifampicin, colistin/minocycline and minocycline/meropenem are synergistic in vitro against XDR-AB strains.
PMCID: PMC3098177  PMID: 21521536
8.  fAUC/MIC is the most predictive pharmacokinetic/pharmacodynamic index of colistin against Acinetobacter baumannii in murine thigh and lung infection models 
To elucidate the pharmacokinetic/pharmacodynamic (PK/PD) index that predicts colistin efficacy against Acinetobacter baumannii in neutropenic murine thigh and lung infection models, and to determine the extent of the emergence of resistance in vivo to colistin.
PK/PD of colistin was studied in thigh and lung infection models against A. baumannii ATCC 19606 and two multidrug-resistant clinical isolates (two of the three strains were colistin heteroresistant). Dose fractionation studies were conducted over a daily dose range of 1–160 mg/kg colistin sulphate. Bacterial burden in tissues was measured at 24 h. Non-linear least squares regression analyses were employed to determine the PK/PD index (fAUC/MIC, fCmax/MIC or fT>MIC) best correlating with the efficacy of colistin in each model. Real-time population analysis profiles were conducted for tissue samples to monitor the emergence of resistance.
The fAUC/MIC was the PK/PD index that correlated best with efficacy in both thigh (R2 = 0.90) and lung (R2 = 0.80) infection models. The fAUC/MIC targets required to achieve stasis and 1 log kill against the three strains were 1.89–7.41 and 6.98–13.6 in the thigh infection model, respectively, while the corresponding values were 1.57–6.52 and 8.18–42.1 in the lung infection model. Amplification of colistin-resistant subpopulations was revealed for all strains in both models after 24 h colistin treatment.
This study indicates the importance of achieving adequate time-averaged exposure to colistin and defined target fAUC/MIC values for various magnitudes of kill. Amplification of resistant subpopulations indicates the importance of investigating rational combinations with colistin. The results will facilitate efforts to optimize colistin use in humans.
PMCID: PMC2920176  PMID: 20573659
Gram-negative bacteria; emergence of resistance; population analysis profiles; heteroresistance
9.  Rapid Determination of Colistin Resistance in Clinical Strains of Acinetobacter baumannii by Use of the Micromax Assay 
Journal of Clinical Microbiology  2013;51(11):3675-3682.
Colistin is an old antibiotic which has been used as a therapeutic option for carbapenem- and multidrug-resistant Gram-negative bacteria, like Acinetobacter baumannii. This pathogen produces life-threatening infections, mainly in patients admitted to intensive care units. Rapid detection of resistance to colistin may improve patient outcomes and prevent the spread of resistance. For this purpose, Micromax technology was evaluated in four isogenic A. baumannii strains with known mechanisms of resistance to colistin and in 66 isolates (50 susceptible and 16 resistant). Two parameters were determined, DNA fragmentation and cell wall damage. To assess DNA fragmentation, cells trapped in a microgel were incubated with a lysing solution to remove the cell wall, and the released nucleoids were visualized under fluorescence microscopy. Fragmented DNA was observed as spots that diffuse from the nucleoid. To assess cell wall integrity, cells were incubated with a lysis solution which removes only weakened cell walls, resulting in nucleoid release exclusively in affected cells. A dose-response relationship was demonstrated between colistin concentrations and the percentages of bacteria with DNA fragmentation and cell wall damage, antibiotic effects that were delayed and less frequent in resistant strains. Receiver operating characteristic (ROC) curves demonstrated that both DNA fragmentation and cell wall damage were excellent parameters for identifying resistant strains. Obtaining ≤11% of bacteria with cell wall damage after incubation with 0.5 μg/ml colistin identified resistant strains of A. baumannii with 100% sensitivity and 96% specificity. Results were obtained in 3 h 30 min. This is a simple, rapid, and accurate assay for detecting colistin resistance in A. baumannii, with strong potential value in critical clinical situations.
PMCID: PMC3889750  PMID: 23985913
10.  Activities of Vancomycin-Containing Regimens against Colistin-Resistant Acinetobacter baumannii Clinical Strains 
Treatment of infections due to extensively drug-resistant (XDR) Acinetobacter baumannii often involves the use of antimicrobial agents in combination. Various combinations of agents have been proposed, with colistin serving as the backbone in many of them. Recent data suggest that glycopeptides, in particular vancomycin, may have unique activity against laboratory-adapted and clinical strains of A. baumannii, alone and in combination with colistin. The aim of the present study was to test this approach against three unique colistin-resistant A. baumannii clinical strains using combinations of vancomycin (VAN), colistin (COL), and doripenem (DOR). All three strains possessed the signature phosphoethanolamine modification of the lipid A moiety associated with colistin resistance and unique amino acid changes in the PmrAB two-component signal transduction system not observed in colistin-susceptible strains. In checkerboard assays, synergy (defined as a fractional inhibitory concentration index [FICI] of ≤0.5) was observed between COL and VAN for all three strains tested and between COL and DOR in two strains. In time-kill assays, the combinations of COL-DOR, COL-VAN, and COL-DOR-VAN resulted in complete killing of colistin-resistant A. baumannii in 1, 2, and all 3 strains, respectively. In the Galleria mellonella moth model of infection, the combinations of DOR-VAN and COL-DOR-VAN led to significantly increased survival of the larvae, compared with other combinations and monotherapy. These findings suggest that regimens containing vancomycin may confer therapeutic benefit for infection due to colistin-resistant A. baumannii.
PMCID: PMC3632926  PMID: 23422916
11.  Growth Retardation, Reduced Invasiveness, and Impaired Colistin-Mediated Cell Death Associated with Colistin Resistance Development in Acinetobacter baumannii 
Two colistin-susceptible/colistin-resistant (Cols/Colr) pairs of Acinetobacter baumannii strains assigned to international clone 2, which is prevalent worldwide, were sequentially recovered from two patients after prolonged colistin administration. Compared with the respective Cols isolates (Ab248 and Ab299, both having a colistin MIC of 0.5 μg/ml), both Colr isolates (Ab249 and Ab347, with colistin MICs of 128 and 32 μg/ml, respectively) significantly overexpressed pmrCAB genes, had single-amino-acid shifts in the PmrB protein, and exhibited significantly slower growth. The Colr isolate Ab347, tested by proteomic analysis in comparison with its Cols counterpart Ab299, underexpressed the proteins CsuA/B and C from the csu operon (which is necessary for biofilm formation). This isolate also underexpressed aconitase B and different enzymes involved in the oxidative stress response (KatE catalase, superoxide dismutase, and alkyl hydroperoxide reductase), suggesting a reduced response to reactive oxygen species (ROS) and, consequently, impaired colistin-mediated cell death through hydroxyl radical production. Cols isolates that were indistinguishable by macrorestriction analysis from Ab299 caused six sequential bloodstream infections, and isolates indistinguishable from Ab248 caused severe soft tissue infection, while Colr isolates indistinguishable from Ab347 and Ab249 were mainly colonizers. In particular, a Cols isolate identical to Ab299 was still invading the bloodstream 90 days after the colonization of this patient by Colr isolates. These observations indicate considerably lower invasiveness of A. baumannii clinical isolates following the development of colistin resistance.
PMCID: PMC3910856  PMID: 24247145
12.  Clinical Use of Colistin Induces Cross-Resistance to Host Antimicrobials in Acinetobacter baumannii 
mBio  2013;4(3):e00021-13.
The alarming rise in antibiotic resistance has led to an increase in patient mortality and health care costs. This problem is compounded by the absence of new antibiotics close to regulatory approval. Acinetobacter baumannii is a human pathogen that causes infections primarily in patients in intensive care units (ICUs) and is highly antibiotic resistant. Colistin is one of the last-line antibiotics for treating A. baumannii infections; however, colistin-resistant strains are becoming increasingly common. This cationic antibiotic attacks negatively charged bacterial membranes in a manner similar to that seen with cationic antimicrobials of the innate immune system. We therefore set out to determine if the increasing use of colistin, and emergence of colistin-resistant strains, is concomitant with the generation of cross-resistance to host cationic antimicrobials. We found that there is indeed a positive correlation between resistance to colistin and resistance to the host antimicrobials LL-37 and lysozyme among clinical isolates. Importantly, isolates obtained before and after treatment of individual patients demonstrated that colistin use correlated with increased resistance to cationic host antimicrobials. These data reveal the overlooked risk of inducing cross-resistance to host antimicrobials when treating patients with colistin as a last-line antibiotic.
Increased use of the cationic antibiotic colistin to treat multidrug-resistant Acinetobacter baumannii has led to the development of colistin-resistant strains. Here we report that treatment of patients with colistin can induce not only increased resistance to colistin but also resistance to host cationic antimicrobials. This worrisome finding likely represents an example of a broader trend observed in other bacteria against which colistin is used therapeutically such as Pseudomonas aeruginosa and Klebsiella pneumoniae. Furthermore, these data suggest that the possible future use of an array of cationic antimicrobial peptides in development as therapeutics may have unintended negative consequences, eventually leading to the generation of hypervirulent strains that are resistant to innate host defenses. The potential for the induction of cross-resistance to innate immune antimicrobials should be considered during the development of new therapeutics.
PMCID: PMC3663567  PMID: 23695834
13.  Resistance to Colistin in Acinetobacter baumannii Associated with Mutations in the PmrAB Two-Component System▿  
The mechanism of colistin resistance (Colr) in Acinetobacter baumannii was studied by selecting in vitro Colr derivatives of the multidrug-resistant A. baumannii isolate AB0057 and the drug-susceptible strain ATCC 17978, using escalating concentrations of colistin in liquid culture. DNA sequencing identified mutations in genes encoding the two-component system proteins PmrA and/or PmrB in each strain and in a Colr clinical isolate. A colistin-susceptible revertant of one Colr mutant strain, obtained following serial passage in the absence of colistin selection, carried a partial deletion of pmrB. Growth of AB0057 and ATCC 17978 at pH 5.5 increased the colistin MIC and conferred protection from killing by colistin in a 1-hour survival assay. Growth in ferric chloride [Fe(III)] conferred a small protective effect. Expression of pmrA was increased in Colr mutants, but not at a low pH, suggesting that additional regulatory factors remain to be discovered.
PMCID: PMC2737849  PMID: 19528270
14.  Unique Structural Modifications Are Present in the Lipopolysaccharide from Colistin-Resistant Strains of Acinetobacter baumannii 
Antimicrobial Agents and Chemotherapy  2013;57(10):4831-4840.
Acinetobacter baumannii is a nosocomial opportunistic pathogen that can cause severe infections, including hospital-acquired pneumonia, wound infections, and sepsis. Multidrug-resistant (MDR) strains are prevalent, further complicating patient treatment. Due to the increase in MDR strains, the cationic antimicrobial peptide colistin has been used to treat A. baumannii infections. Colistin-resistant strains of A. baumannii with alterations to the lipid A component of lipopolysaccharide (LPS) have been reported; specifically, the lipid A structure was shown to be hepta-acylated with a phosphoethanolamine (pEtN) modification present on one of the terminal phosphate residues. Using a tandem mass spectrometry platform, we provide definitive evidence that the lipid A isolated from colistin-resistant A. baumannii MAC204 LPS contains a novel structure corresponding to a diphosphoryl hepta-acylated lipid A structure with both pEtN and galactosamine (GalN) modifications. To correlate our structural studies with clinically relevant samples, we characterized colistin-susceptible and -resistant isolates obtained from patients. These results demonstrated that the clinical colistin-resistant isolate had the same pEtN and GalN modifications as those seen in the laboratory-adapted A. baumannii strain MAC204. In summary, this work has shown complete structure characterization including the accurate assignment of acylation, phosphorylation, and glycosylation of lipid A from A. baumannii, which are important for resistance to colistin.
PMCID: PMC3811424  PMID: 23877686
15.  Post Neurosurgical Meningitis due to Colistin Heteroresistant Acinetobacter baumannii 
Recently Acinetobacter baumannii isolates have emerged as a problematic infectious agent that causes meningitis in neurosurgical patients. Colistin has been used successfully for the treatment of A. baumannii meningitis but colistin resistant isolates have been reported worldwide.
Case Presentation:
Two isolates of A. baumannii were cultured during a five-day period from cerebrospinal fluid (CSF) samples of a 20-year-old man with a gunshot trauma in the abdomen, which had exited from his back. Antimicrobial susceptibility tests of isolates were performed. Multiplex PCR was performed for detection of blaOXA-23-like, blaOXA-24-like and blaOXA-58-like genes. Metallo-β-lactamase genes such as: blaVIM, blaIMP, blaSPM and blaNDM were sought by singleplex PCR. In order to evaluate the genetic relationship, two isolates were examined by the repetitive extragenic palindromic-polymerase chain reaction (REP_PCR) method.
E-test results showed that the isolates were sensitive to colistin and tigecycline with minimum inhibitory concentration of (MIC) 0.25 µg/mL and 1.5 µg/mL, respectively. Secondly the isolates were resistant to colistin with MIC > 256 µg/mL but remained sensitive to tigecycline with MIC 1.5 µg/mL. On the basis of the multiplex PCR, both of the isolates were positive for blaOXA-23-like. Other investigated genes such as blaOXA-24-like, blaOXA-58-like, blaVIM, blaIMP, blaSPM and blaNDM were negative. REP-PCR results showed that two isolates were derived from a single strain and both were the same. The results of our study revealed that the firs isolate of A. baumannii was colistin heteroresistant and was changed to completely resistant during therapy. Diagnosis and treatment of A. baumannii meningitis is very important and to avoid treatment failure we suggest that all A. baumannii isolates obtained from CSF should be evaluated properly for colistin heteroresistance.
PMCID: PMC4295316  PMID: 25632326
Acinetobacter baumannii; Meningitis; Colistin; Neurosurgery
16.  Comparative Evaluation of the VITEK 2, Disk Diffusion, Etest, Broth Microdilution, and Agar Dilution Susceptibility Testing Methods for Colistin in Clinical Isolates, Including Heteroresistant Enterobacter cloacae and Acinetobacter baumannii Strains▿  
Antimicrobial Agents and Chemotherapy  2007;51(10):3726-3730.
Increasing antibiotic resistance in gram-negative bacteria has recently renewed interest in colistin as a therapeutic option. The increasing use of colistin necessitates the availability of rapid and reliable methods for colistin susceptibility testing. We compared seven methods of colistin susceptibility testing (disk diffusion, agar dilution on Mueller-Hinton [MH] and Isosensitest agar, Etest on MH and Isosensitest agar, broth microdilution, and VITEK 2) on 102 clinical isolates collected from patient materials during a selective digestive decontamination or selective oral decontamination trial in an intensive-care unit. Disk diffusion is an unreliable method to measure susceptibility to colistin. High error rates and low levels of reproducibility were observed in the disk diffusion test. The colistin Etest, agar dilution, and the VITEK 2 showed a high level of agreement with the broth microdilution reference method. Heteroresistance for colistin was observed in six Enterobacter cloacae isolates and in one Acinetobacter baumannii isolate. This is the first report of heteroresistance to colistin in E. cloacae isolates. Resistance to colistin in these isolates seemed to be induced upon exposure to colistin rather than being caused by stable mutations. Heteroresistant isolates could be detected in the broth microdilution, agar dilution, Etest, or disk diffusion test. The VITEK 2 displayed low sensitivity in the detection of heteroresistant subpopulations of E. cloacae. The VITEK 2 colistin susceptibility test can therefore be considered to be a reliable tool to determine susceptibility to colistin in isolates of genera that are known not to exhibit resistant subpopulations. In isolates of genera known to (occasionally) exhibit heteroresistance, an alternative susceptibility testing method capable of detecting heteroresistance should be used.
PMCID: PMC2043258  PMID: 17646414
17.  In Vitro Pharmacodynamic Properties of Colistin and Colistin Methanesulfonate against Pseudomonas aeruginosa Isolates from Patients with Cystic Fibrosis 
The in vitro pharmacodynamic properties of colistin and colistin methanesulfonate were investigated by studying the MICs, time-kill kinetics, and postantibiotic effect (PAE) against mucoid and nonmucoid strains of Pseudomonas aeruginosa isolated from patients with cystic fibrosis. Twenty-three clinical strains, including multiresistant strains, and one type strain were selected for MIC determination. Eleven strains were resistant; MICs for these strains were >128 mg/liter. For the susceptible strains, MICs of colistin ranged from 1 to 4 mg/liter, while the MICs of colistin methanesulfonate were significantly higher and ranged from 4 to 16 mg/liter. The time-kill kinetics were investigated with three strains at drug concentrations ranging from 0.5 to 64 times the MIC. Colistin showed extremely rapid killing, resulting in complete elimination at the highest concentrations within 5 min, while colistin methanesulfonate killed more slowly, requiring a concentration of 16 times the MIC to achieve complete killing within 24 h. Colistin exhibited a significant PAE of 2 to 3 h at 16 times the MIC against the three strains after 15 min of exposure. For colistin methanesulfonate, PAEs were shorter at the concentrations tested. Colistin methanesulfonate had lower overall bactericidal and postantibiotic activities than colistin, even when adjusted for differences in MICs. Our data suggest that doses of colistin methanesulfonate higher than the recommended 2 to 3 mg/kg of body weight every 12 h may be required for the effective treatment of P. aeruginosa infections in cystic fibrosis patients.
PMCID: PMC90373  PMID: 11181360
18.  Synergistic Activity of Colistin and Rifampin Combination against Multidrug-Resistant Acinetobacter baumannii in an In Vitro Pharmacokinetic/Pharmacodynamic Model 
Combination therapy may be required for multidrug-resistant (MDR) Acinetobacter baumannii. This study systematically investigated bacterial killing and emergence of colistin resistance with colistin and rifampin combinations against MDR A. baumannii. Studies were conducted over 72 h in an in vitro pharmacokinetic (PK)/pharmacodynamic (PD) model at inocula of ∼106 and ∼108 CFU/ml using two MDR clinical isolates of A. baumannii, FADDI-AB030 (colistin susceptible) and FADDI-AB156 (colistin resistant). Three combination regimens achieving clinically relevant concentrations (constant colistin concentration of 0.5, 2, or 5 mg/liter and a rifampin maximum concentration [Cmax] of 5 mg/liter every 24 hours; half-life, 3 h) were investigated. Microbiological response was measured by serial bacterial counts. Population analysis profiles assessed emergence of colistin resistance. Against both isolates, combinations resulted in substantially greater killing at the low inoculum; combinations containing 2 and 5 mg/liter colistin increased killing at the high inoculum. Combinations were additive or synergistic at 6, 24, 48, and 72 h with all colistin concentrations against FADDI-AB030 and FADDI-AB156 in, respectively, 8 and 11 of 12 cases (i.e., all 3 combinations) at the 106-CFU/ml inoculum and 8 and 7 of 8 cases with the 2- and 5-mg/liter colistin regimens at the 108-CFU/ml inoculum. For FADDI-AB156, killing by the combination was ∼2.5 to 7.5 and ∼2.5 to 5 log10 CFU/ml greater at the low inoculum (all colistin concentrations) and high inoculum (2 and 5 mg/liter colistin), respectively. Emergence of colistin-resistant subpopulations was completely suppressed in the colistin-susceptible isolate with all combinations at both inocula. Our study provides important information for optimizing colistin-rifampin combinations against colistin-susceptible and -resistant MDR A. baumannii.
PMCID: PMC3719722  PMID: 23716052
19.  Aerosolized colistin for the treatment of nosocomial pneumonia due to multidrug-resistant Gram-negative bacteria in patients without cystic fibrosis 
Critical Care  2005;9(1):R53-R59.
The clinical and economic consequences of the emergence of multidrug-resistant Gram-negative bacteria in the intensive care unit (ICU) setting, combined with the high mortality rate among patients with nosocomial pneumonia, have stimulated a search for alternative therapeutic options to treat such infections. The use of adjunctive therapy with aerosolized colistin represents one of these. There is extensive experience with use of aerosolized colistin by patients with cystic fibrosis, but there is a lack of data regarding the use of aerosolized colistin in patients without cystic fibrosis.
We conducted the present study to assess the safety and effectiveness of aerosolized colistin as an adjunct to intravenous antimicrobial therapy for treatment of Gram-negative nosocomial pneumonia. We retrospectively reviewed the medical records of patients hospitalized in a 450-bed tertiary care hospital during the period from October 2000 to January 2004, and who received aerosolized colistin as adjunctive therapy for multidrug-resistant pneumonia.
Eight patients received aerosolized colistin. All patients had been admitted to the ICU, with mean Acute Physiological and Chronic Health Evaluation II scores on the day of ICU admission and on day 1 of aerosolized colistin administration of 14.6 and 17.1, respectively. Six of the eight patients had ventilator-associated pneumonia. The responsible pathogens were Acinetobacter baumannii (in seven out of eight cases) and Pseudomonas aeruginosa (in one out of eight cases) strains. Half of the isolated pathogens were sensitive only to colistin. The daily dose of aerosolized colistin ranged from 1.5 to 6 million IU (divided into three or four doses), and the mean duration of administration was 10.5 days. Seven out of eight patients received concomitant intravenous treatment with colistin or other antimicrobial agents. The pneumonia was observed to respond to treatment in seven out of eight patients (four were cured and three improved [they were transferred to another facility]). One patient deteriorated and died from septic shock and multiple organ failure. Aerosolized colistin was well tolerated by all patients; no bronchoconstriction or chest tightness was reported.
Aerosolized colistin may be a beneficial adjunctive treatment in the management of nosocomial pneumonia (ventilator associated or not) due to multidrug-resistant Gram-negative bacteria.
PMCID: PMC1065114  PMID: 15693967
apnea; bronchoconstriction; colistin; inhaled; nosocomial pneumonia
20.  In Vivo Efficacy of Glycopeptide-Colistin Combination Therapies in a Galleria mellonella Model of Acinetobacter baumannii Infection ▿ 
The treatment of Acinetobacter baumannii infections poses a significant clinical challenge, with isolates resistant to all commonly used agents increasingly being reported. With few new agents in the pipeline, clinicians are increasingly turning to combinations of antimicrobials in the hope that they may act synergistically together. In this study we assessed the activities of two glycopeptide-colistin combinations both in vitro and using a Galleria mellonella caterpillar model of A. baumannii infection. In checkerboard assays both vancomycin and teicoplanin were highly active against susceptible and multidrug-resistant strains of A. baumannii when combined with colistin (fractional inhibitory concentration [FIC] of <0.25). Treatment of G. mellonella caterpillars infected with lethal doses of A. baumannii resulted in significantly enhanced survival rates when either vancomycin or teicoplanin was given with colistin compared to colistin treatment alone (P < 0.05). This effect was most marked when vancomycin was the glycopeptide administered, although this agent was also highly effective as monotherapy, possibly through an immunomodulatory action on the G. mellonella response to A. baumannii infection. This work suggests that glycopeptide-colistin combinations are highly active against A. baumannii both in vitro and in a simple animal model of infection. They should be considered further as potential treatments for difficult-to-treat A. baumannii infections.
PMCID: PMC3122470  PMID: 21502628
21.  Activity of Cecropin A-Melittin Hybrid Peptides against Colistin-Resistant Clinical Strains of Acinetobacter baumannii: Molecular Basis for the Differential Mechanisms of Action 
Acinetobacter baumannii has successfully developed resistance against all common antibiotics, including colistin (polymyxin E), the last universally active drug against this pathogen. The possible widespread distribution of colistin-resistant A. baumannii strains may create an alarming clinical situation. In a previous work, we reported differences in lethal mechanisms between polymyxin B (PXB) and the cecropin A-melittin (CA-M) hybrid peptide CA(1-8)M(1-18) (KWKLFKKIGIGAVLKVLTTGLPALIS-NH2) on colistin-susceptible strains (J. M. Saugar, T. Alarcón, S. López-Hernández, M. López-Brea, D. Andreu, and L. Rivas, Antimicrob. Agents Chemother. 46:875-878, 2002). We now demonstrate that CA(1-8)M(1-18) and three short analogues, namely CA(1-7)M(2-9) (KWKLFKKIGAVLKVL-NH2), its Nα-octanoyl derivative (Oct-KWKLFKKIGAVLKVL-NH2), and CA(1-7)M(5-9) (KWKLLKKIGAVLKVL-NH2) are active against two colistin-resistant clinical strains. In vitro, resistance to colistin sulfate was targeted to the outer membrane, as spheroplasts were equally lysed by a given peptide, regardless of their respective level of colistin resistance. The CA-M hybrids were more efficient than colistin in displacing lipopolysaccharide-bound dansyl-polymyxin B from colistin-resistant but not from colistin-susceptible strains. Similar improved performance of the CA-M hybrids in permeation of the inner membrane was observed, regardless of the resistance pattern of the strain. These results argue in favor of a possible use of CA-M peptides, and by extension other antimicrobial peptides with similar features, as alternative chemotherapy in colistin-resistant Acinetobacter infections.
PMCID: PMC1426946  PMID: 16569836
22.  In Vitro Synergy of Colistin Combinations against Colistin-Resistant Acinetobacter baumannii, Pseudomonas aeruginosa, and Klebsiella pneumoniae Isolates 
Colistin resistance, although uncommon, is increasingly being reported among Gram-negative clinical pathogens, and an understanding of its impact on the activity of antimicrobials is now evolving. We evaluated the potential for synergy of colistin plus trimethoprim, trimethoprim-sulfamethoxazole (1/19 ratio), or vancomycin against 12 isolates of Acinetobacter baumannii (n = 4), Pseudomonas aeruginosa (n = 4), and Klebsiella pneumoniae (n = 4). The strains included six multidrug-resistant clinical isolates, K. pneumoniae ATCC 700603, A. baumannii ATCC 19606, P. aeruginosa ATCC 27853, and their colistin-resistant derivatives (KPm1, ABm1, and PAm1, respectively). Antimicrobial susceptibilities were assessed by broth microdilution and population analysis profiles. The potential for synergy of colistin combinations was evaluated using a checkerboard assay, as well as static time-kill experiments at 0.5× and 0.25× MIC. The MIC ranges of vancomycin, trimethoprim, and trimethoprim-sulfamethoxazole (1/19) were ≥128, 4 to ≥128, and 2/38 to >128/2,432 μg/ml, respectively. Colistin resistance demonstrated little impact on vancomycin, trimethoprim, or trimethoprim-sulfamethoxazole MIC values. Isolates with subpopulations heterogeneously resistant to colistin were observed to various degrees in all tested isolates. In time-kill assays, all tested combinations were synergistic against KPm1 at 0.25× MIC and 0.5× MIC and ABm1 and PAm1 at 0.5× MIC. In contrast, none of the tested combinations demonstrated synergy against any colistin-susceptible P. aeruginosa isolates and clinical strains of K. pneumoniae isolates. Only colistin plus trimethoprim or trimethoprim-sulfamethoxazole was synergistic and bactericidal at 0.5× MIC against K. pneumoniae ATCC 700603. Colistin resistance seems to promote the in vitro activity of unconventional colistin combinations. Additional experiments are warranted to understand the clinical significance of these observations.
PMCID: PMC3421898  PMID: 22751540
23.  Detection of colistin sensitivity in clinical isolates of Acinetobacter baumannii in Iran 
Nosocomial infection caused by Acinetobacter baumannii has emerged as a serious problem world-wide. Finding the suitable drug is an important priority. The aim of this study was to determine colistin (polymyxin E) resistance in clinical isolates of A. baumannii from intensive care units (ICUs) of Al Zahra Hospital.
Materials and Methods:
Sixty isolates of A. baumannii from patients hospitalized in ICU (Al Zahra Hospital, Isfahan University of Medical Sciences [IUMS]) were studied. All isolates of A. baumannii were tested for colistin susceptibility by Eopsilometer test (E-test).
Of the 60 isolates 57, (95%) were multidrug resistant (MDR) and 76.6% (46/60) were highly resistant. The rate of colistin resistant with the E-test method was 11.6% (7/60).
As the frequency of resistance to colistin is low, it can be used as an easily available drug for treatment of MDR A. baumannii strains, which are susceptible to colistin.
PMCID: PMC4078383  PMID: 25002899
Acinetobacter baumannii; colistin resistance; multidrug resistant organism
24.  Efficacy of Rifampin and Its Combinations with Imipenem, Sulbactam, and Colistin in Experimental Models of Infection Caused by Imipenem-Resistant Acinetobacter baumannii▿  
There are currently no defined optimal therapies available for multidrug-resistant (MDR) Acinetobacter baumannii infections. We evaluated the efficacy of rifampin, imipenem, sulbactam, colistin, and their combinations against MDR A. baumannii in experimental pneumonia and meningitis models. The bactericidal in vitro activities of rifampin, imipenem, sulbactam, colistin, and their combinations were tested using time-kill curves. Murine pneumonia and rabbit meningitis models were evaluated using the A. baummnnii strain Ab1327 (with MICs for rifampin, imipenem, sulbactam, and colistin of 4, 32, 32, and 0.5 mg/liter, respectively). Mice were treated with the four antimicrobials and their combinations. For the meningitis model, the efficacies of colistin, rifampin and its combinations with imipenem, sulbactam, or colistin, and of imipenem plus sulbactam were assayed. In the pneumonia model, compared to the control group, (i) rifampin alone, (ii) rifampin along with imipenem, sulbactam, or colistin, (iii) colistin, or (iv) imipenem plus sulbactam significantly reduced lung bacterial concentrations (10.6 ± 0.27 [controls] versus 3.05 ± 1.91, 2.07 ± 1.82, 2.41 ± 1.37, 3.4 ± 3.07, 6.82 ± 3.4, and 4.22 ± 2.72 log10 CFU/g, respectively [means ± standard deviations]), increased sterile blood cultures (0% versus 78.6%, 100%, 93.3%, 93.8%, 73.3%, and 50%), and improved survival (0% versus 71.4%, 60%, 46.7%, 43.8%, 40%, and 85.7%). In the meningitis model rifampin alone or rifampin plus colistin reduced cerebrospinal fluid bacterial counts (−2.6 and −4.4 log10 CFU/ml). Rifampin in monotherapy or with imipenem, sulbactam, or colistin showed efficacy against MDR A. baumannii in experimental models of pneumonia and meningitis. Imipenem or sulbactam may be appropriate for combined treatment when using rifampin.
PMCID: PMC2825983  PMID: 20047914
25.  In vitro activity of tigecycline in combination with various antimicrobials against multidrug resistant Acinetobacter baumannii 
Infections sustained by multidrug-resistant (MDR) and pan-resistant Acinetobacter baumannii have become a challenging problem in Intensive Care Units. Tigecycline provided new hope for the treatment of MDR A. baumannii infections, but isolates showing reduced susceptibility have emerged in many countries, further limiting the therapeutic options. Empirical combination therapy has become a common practice to treat patients infected with MDR A. baumannii, in spite of the limited microbiological and clinical evidence supporting its efficacy. Here, the in vitro interaction of tigecycline with seven commonly used anti-Acinetobacter drugs has been assessed.
Twenty-two MDR A. baumannii isolates from Intensive Care Unit (ICU) patients and two reference strains for the European clonal lineages I and II (including 3, 15 and 6 isolates that were resistant, intermediate and susceptible to tigecycline, respectively) were tested. Antimicrobial agents were: tigecycline, levofloxacin, piperacillin-tazobactam, amikacin, imipenem, rifampicin, ampicillin-sulbactam, and colistin. MICs were determined by the broth microdilution method. Antibiotic interactions were determined by chequerboard and time-kill assays. Only antibiotic combinations showing synergism or antagonism in both chequerboard and time-kill assays were accepted as authentic synergistic or antagonistic interactions, respectively.
Considering all antimicrobials in combination with tigecycline, chequerboard analysis showed 5.9% synergy, 85.7% indifference, and 8.3% antagonism. Tigecycline showed synergism with levofloxacin (4 strains; 16.6%), amikacin (2 strains; 8.3%), imipenem (2 strains; 8.3%) and colistin (2 strains; 8.3%). Antagonism was observed for the tigecycline/piperacillin-tazobactam combination (8 strains; 33.3%). Synergism was detected only among tigecycline non-susceptible strains. Time-kill assays confirmed the synergistic interaction between tigecycline and levofloxacin, amikacin, imipenem and colistin for 5 of 7 selected isolates. No antagonism was confirmed by time-kill assays.
This study demonstrates the in vitro synergistic activity of tigecycline in combination with colistin, levofloxacin, amikacin and imipenem against five tigecycline non-susceptible A. baumannii strains, opening the way to a more rationale clinical assessment of novel combination therapies to combat infections caused by MDR and pan-resistant A. baumannii.
PMCID: PMC2693502  PMID: 19460166

Results 1-25 (1140072)