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Find video protocols related to scientific articles indexed in Pubmed.
Novel insight into antimicrobial resistance and sensitivity phenotypes associated to qac and norA genotypes in Staphylococcus aureus.
Microbiol. Res.
PUBLISHED: 06-13-2014
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Staphylococcus aureus strains harboring QacA, QacB, QacC, QacG transporters and norA promoter up-regulating mutations were characterized by phenotype microarray (PM), standard methods for susceptibility testing, and ethidium bromide efflux assays, in order to increase knowledge on phenotypes associated to efflux pumps and their substrates. PM data and standard susceptibility testing lead to the identification of new potential efflux targets, such as guanidine hydrochloride or 8-hydroxyquinoline for QacA and QacC pumps, respectively. The identification of compounds to which the presence of efflux pumps induced increased susceptibility opens new perspectives for potential adjunct anti-resistance treatment (i.e. strains bearing QacB transporters showed increased susceptibility to thioridazine, amitriptyline and orphenadrine). Although the tested isolates were characterized by high degree of heterogeneity, a hallmark of clinical isolates, direct ethidium bromide efflux assays were effective in highlighting differences in efflux efficiency among strains. These data add to characterization of substrate specificity in the different classes of staphylococcal multidrug efflux systems conferring specific substrate profiles and efflux features to each of them.
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Antimicrobial susceptibility monitoring of respiratory tract pathogens isolated from diseased cattle and pigs across Europe: the VetPath study.
Vet. Microbiol.
PUBLISHED: 04-06-2014
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VetPath is an ongoing pan-European antibiotic susceptibility monitoring programme collecting pathogens from diseased antimicrobial non-treated cattle, pigs and poultry. In the current study, 1001 isolates from cattle and pig respiratory tract infections were tested for their antimicrobial susceptibilities. Non-replicate lung samples or nasopharyngeal/nasal swabs were collected from animals with acute clinical signs in 11 countries during 2002-2006. Pasteurella multocida and Mannheimia haemolytica from cattle and P. multocida, Actinobacillus pleuropneumoniae and Streptococcus suis from pigs were isolated by standard methods. S. suis was also isolated from meningitis cases. MICs of 16 antibiotics were assessed centrally by broth microdilution following CLSI recommendations. Results were interpreted using CLSI breakpoints where available. P. multocida (231) and M. haemolytica (138) isolates were all susceptible to amoxicillin/clavulanic acid, ceftiofur, enrofloxacin and trimethoprim/sulfamethoxazole. Resistance to florfenicol and spectinomycin was 0.4% and 3.5% in P. multocida, respectively, and absent in M. haemolytica isolates. Tetracycline resistance was 5.7% and 14.6% for P. multocida and M. haemolytica. In pigs, 230 P. multocida, 220 A. pleuropneumoniae and 182 S. suis isolates were recovered. Resistance to amoxicillin/clavulanic acid, ceftiofur, enrofloxacin, florfenicol, tiamulin and tilmicosin was absent or <1%. Trimethoprim/sulfamethoxazole resistance was 3-6% and tetracycline resistance varied from 14.7% in A. pleuropneumoniae to 81.8% in S. suis. In conclusion, low resistance to antibiotics with defined clinical breakpoints, except for tetracycline, was observed among the major respiratory tract pathogens recovered from cattle and pigs. Since for approximately half of the antibiotics in this panel no CLSI-defined breakpoints were available, setting of the missing veterinary breakpoints is important.
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Epidemiology and antimicrobial susceptibility of Gram-negative aerobic bacteria causing intra-abdominal infections during 2010-2011.
J Chemother
PUBLISHED: 02-20-2014
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The study for monitoring antimicrobial resistance trends (SMART) surveillance program monitors the epidemiology and trends in antibiotic resistance of intra-abdominal pathogens to currently used therapies. The current report describes such trends during 2010-2011. A total of 25?746 Gram-negative clinical isolates from intra-abdominal infections were collected and classified as hospital-associated (HA) if the hospital length of stay (LOS) at the time of specimen collection was ?48 hours, community-associated (CA) if LOS at the time of specimen collection was <48 hours, or unknown (no designation given by participating centre). A total of 92 different species were collected of which the most common was Escherichia coli: 39% of all isolates in North America to 55% in Africa. Klebsiella pneumoniae was the second most common pathogen: 11% of all isolates from Europe to 19% of all isolates from Asia. Isolates were from multiple intra-abdominal sources of which 32% were peritoneal fluid, 20% were intra-abdominal abscesses, and 16·5% were gall bladder infections. Isolates were further classified as HA (55% of all isolates), CA (39% of all isolates), or unknown (6% of all isolates). The most active antibiotics tested were imipenem, ertapenem, amikacin, and piperacillin-tazobactam. Resistance rates to all other antibiotics tested were high. Considering the current data set and high-level resistance of intra-abdominal pathogens to various antibiotics, further monitoring of the epidemiology of intra-abdominal infections and their susceptibility to antibiotics through SMART is warranted.
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Evaluation of the Clinical and Laboratory Standards Institute phenotypic confirmatory test to detect the presence of extended-spectrum ?-lactamases from 4005 Escherichia coli, Klebsiella oxytoca, Klebsiella pneumoniae and Proteus mirabilis isolates.
J. Med. Microbiol.
PUBLISHED: 01-29-2014
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A subset of Escherichia coli, Klebsiella oxytoca, Klebsiella pneumoniae and Proteus mirabilis isolates collected for the Study for Monitoring Antimicrobial Resistance Trends that were positive for the Clinical and Laboratory Standards Institute (CLSI) extended-spectrum ?-lactamase (ESBL) phenotypic confirmatory test (n?=?3245) or had an ertapenem MIC of ?0.5 µg ml(-1) (n?=?293), or both (n?=?467), were analysed for ESBL genes. Most ESBL phenotype E. coli or K. pneumoniae possessed an ESBL gene (95.8 and 88.4?%, respectively), and this was 93.1?% if carbapenem-non-susceptible K. pneumoniae were removed. This rate was lower for P. mirabilis (73.4?%) and K. oxytoca (62.5?%). Virtually all ESBL-positive isolates (99.5?%) were cefotaxime non-susceptible [CLSI or European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints)]. Fewer isolates (82?%) were ceftazidime non-susceptible (CLSI breakpoints). In addition, 21.1?% of E. coli, 25?% of K. oxytoca and 78.7?% of P. mirabilis isolates were ceftazidime susceptible but ESBL positive. This suggests that CLSI breakpoints for ceftazidime are too high to detect ESBLs. The lower EUCAST breakpoints detected ESBLs in E. coli and K. oxytoca better, but 59.6?% of ESBL-positive isolates of P. mirabilis were ceftazidime susceptible. For isolates with ertapenem MICs ?0.5 µg ml(-1), more accurate ESBL phenotype analysis was observed for E. coli and K. pneumoniae (sensitivity >95?% for both, specificity 94.4 and 54.1?%, respectively). If carbapenemase-positive K. pneumoniae were excluded, the specificity increased to 78?%. The positive predictive values for the ESBL phenotypic test with E. coli and K. pneumoniae were 97.6 and 81.8?%, respectively, and negative predictive values were 75.9 and 95.2?%, respectively. We therefore suggest that it would be prudent to confirm phenotypic ESBL-positive P. mirabilis, K. pneumoniae and K. oxytoca with molecular analysis.
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Evaluation of epidemiological cut-off values indicates that biocide resistant subpopulations are uncommon in natural isolates of clinically-relevant microorganisms.
PLoS ONE
PUBLISHED: 01-01-2014
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To date there are no clear criteria to determine whether a microbe is susceptible to biocides or not. As a starting point for distinguishing between wild-type and resistant organisms, we set out to determine the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) distributions for four common biocides; triclosan, benzalkonium chloride, chlorhexidine and sodium hypochlorite for 3319 clinical isolates, with a particular focus on Staphylococcus aureus (N?=?1635) and Salmonella spp. (N?=?901) but also including Escherichia coli (N?=?368), Candida albicans (N?=?200), Klebsiella pneumoniae (N?=?60), Enterobacter spp. (N?=?54), Enterococcus faecium (N?=?53), and Enterococcus faecalis (N?=?56). From these data epidemiological cut-off values (ECOFFs) are proposed. As would be expected, MBCs were higher than MICs for all biocides. In most cases both values followed a normal distribution. Bimodal distributions, indicating the existence of biocide resistant subpopulations were observed for Enterobacter chlorhexidine susceptibility (both MICs and MBCs) and the susceptibility to triclosan of Enterobacter (MBC), E. coli (MBC and MIC) and S. aureus (MBC and MIC). There is a concern on the potential selection of antibiotic resistance by biocides. Our results indicate however that resistance to biocides and, hence any potential association with antibiotic resistance, is uncommon in natural populations of clinically relevant microorganisms.
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A Review of Ten Years of the Study for Monitoring Antimicrobial Resistance Trends (SMART) from 2002 to 2011.
Pharmaceuticals (Basel)
PUBLISHED: 09-29-2013
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Surveillance of antimicrobial agent resistance provides important information to guide microbiologists and infectious disease specialists understanding of the control and the spread of resistance mechanisms within the local environment. Continued monitoring of antimicrobial resistance patterns in the community and in local hospital environments is essential to guide effective empiric therapy. The Study for Monitoring Antimicrobial Resistance Trends (SMART) has monitored the in vitro susceptibility patterns of clinical Gram-negative bacilli to antimicrobial agents collected worldwide from intra-abdominal infections since 2002 and urinary tract infections since 2009. Resistance trends, with a particular focus on carbapenem resistance and the rate of extended-spectrum ?-lactamases (ESBLs), were analyzed. Isolates from intra-abdominal infections (n = 92,086) and urinary-tract infections (n = 24,705) were collected and tested using Clinical and Laboratory Standards Institute methods. This review presents carbapenem susceptibility and ESBL rates over ten years of SMART study analysis, including key publications during this period. The SMART study has proved to be a valuable resource in determining pathogen prevalence and antibiotic susceptibility over the last ten years and continues to provide evidence for regulatory susceptibility breakpoints and clinical decision making.
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Evaluation of reduced susceptibility to quaternary ammonium compounds and bisbiguanides in clinical isolates and laboratory-generated mutants of Staphylococcus aureus.
Antimicrob. Agents Chemother.
PUBLISHED: 05-13-2013
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The MICs and minimum bactericidal concentrations (MBCs) for the biocides benzalkonium chloride and chlorhexidine were determined against 1,602 clinical isolates of Staphylococcus aureus. Both compounds showed unimodal MIC and MBC distributions (2 and 4 or 8 mg/liter, respectively) with no apparent subpopulation with reduced susceptibility. To investigate further, all isolates were screened for qac genes, and 39 of these also had the promoter region of the NorA multidrug-resistant (MDR) efflux pump sequenced. The presence of qacA, qacB, qacC, and qacG genes increased the mode MIC, but not MBC, to benzalkonium chloride, while only qacA and qacB increased the chlorhexidine mode MIC. Isolates with a wild-type norA promoter or mutations in the norA promoter had similar biocide MIC distributions; notably, not all clinical isolates with norA mutations were resistant to fluoroquinolones. In vitro efflux mutants could be readily selected with ethidium bromide and acriflavine. Multiple passages were necessary to select mutants with biocides, but these mutants showed phenotypes comparable to those of mutants selected by dyes. All mutants showed changes in the promoter region of norA, but these were distinct from this region of the clinical isolates. Still, none of the in vitro mutants displayed fitness defects in a killing assay in Galleria mellonella larvae. In conclusion, our data provide an in-depth comparative overview on efflux in S. aureus mutants and clinical isolates, showing also that plasmid-encoded efflux pumps did not affect bactericidal activity of biocides. In addition, current in vitro tests appear not to be suitable for predicting levels of resistance that are clinically relevant.
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The use of machine learning methodologies to analyse antibiotic and biocide susceptibility in Staphylococcus aureus.
PLoS ONE
PUBLISHED: 02-19-2013
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The rise of antibiotic resistance in pathogenic bacteria is a significant problem for the treatment of infectious diseases. Resistance is usually selected by the antibiotic itself; however, biocides might also co-select for resistance to antibiotics. Although resistance to biocides is poorly defined, different in vitro studies have shown that mutants presenting low susceptibility to biocides also have reduced susceptibility to antibiotics. However, studies with natural bacterial isolates are more limited and there are no clear conclusions as to whether the use of biocides results in the development of multidrug resistant bacteria.
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The MUT056399 inhibitor of FabI is a new antistaphylococcal compound.
Antimicrob. Agents Chemother.
PUBLISHED: 08-08-2011
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MUT056399 is a highly potent new inhibitor of the FabI enzyme of both Staphylococcus aureus and Escherichia coli. In vitro, MUT056399 was very active against S. aureus strains, including methicillin-susceptible S. aureus (MSSA), methicillin-resistant S. aureus (MRSA), linezolid-resistant, and multidrug-resistant strains, with MIC(90)s between 0.03 and 0.12 ?g/ml. MUT056399 was also active against coagulase-negative staphylococci, with MIC(90)s between 0.12 and 4 ?g/ml. The antibacterial spectrum is consistent with specific FabI inhibition with no activity against bacteria using FabK but activity against FabI-containing Gram-negative bacilli. In vitro, resistant clones of S. aureus were obtained at a low frequency. All of the resistant clones analyzed were found to contain mutations in the fabI gene. In vivo, MUT056399, administered subcutaneously, protected mice from a lethal systemic infection induced by MSSA, MRSA, and vancomycin-intermediate S. aureus strains (50% effective doses ranging from 19.3 mg/kg/day to 49.6 mg/kg/day). In the nonneutropenic murine thigh infection model, the same treatment with MUT056399 reduced the bacterial multiplication of MSSA and MRSA in the thighs of immunocompetent mice. These properties support MUT056399 as a very promising candidate for a novel drug to treat severe staphylococcal infections.
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Comparative activity of ceftobiprole against Gram-positive and Gram-negative isolates from Europe and the Middle East: the CLASS study.
J. Antimicrob. Chemother.
PUBLISHED: 03-29-2011
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to assess the in vitro activity of ceftobiprole and comparators against a recent collection of Gram-positive and Gram-negative pathogens, in order to detect potential changes in susceptibility patterns, and to evaluate the Etest assay for ceftobiprole susceptibility testing.
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Comparative activity of carbapenem testing: the COMPACT study.
J. Antimicrob. Chemother.
PUBLISHED: 03-08-2011
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Doripenem is a new carbapenem recently introduced into Europe. The COMParative Activity of Carbapenem Testing (COMPACT) study compared the susceptibility of common Gram-negative bacilli causing serious infections in hospitalized patients with doripenem, imipenem and meropenem.
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Activity of the new cephalosporin ceftaroline against bacteraemia isolates from patients with community-acquired pneumonia.
Int. J. Antimicrob. Agents
PUBLISHED: 02-08-2009
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The activity of ceftaroline, a novel cephalosporin, was evaluated against 1337 isolates from patients with bacteraemic community-acquired pneumonia (CAP) requiring hospitalisation (including 119 Haemophilus influenzae, 9 Moraxella catarrhalis, 164 Staphylococcus aureus, 1007 Streptococcus pneumoniae and 38 Streptococcus pyogenes). Minimum inhibitory concentrations (MICs) were determined by broth microdilution according to Clinical and Laboratory Standards Institute (CLSI) guidelines, and susceptibility category assessments were made using CLSI or US Food and Drug Administration (FDA) breakpoints. Ceftaroline MICs were < or = 0.008-0.06 mg/L against H. influenzae, 0.25-2mg/L against methicillin-resistant S. aureus (MRSA), 0.06-1mg/L against methicillin-susceptible S. aureus, 0.015-0.5mg/L against M. catarrhalis and < or = 0.008-0.5mg/L against S. pneumoniae, and all S. pyogenes isolates had ceftaroline MICs < or = 0.008mg/L. Ceftaroline was more active than ceftriaxone or cefepime both against MRSA and penicillin-resistant pneumococci. More than 90% of MRSA isolates were resistant to clarithromycin and levofloxacin but were susceptible to linezolid or tigecycline. A high rate of clarithromycin resistance was also observed in the pneumococci. Ceftaroline was very active in vitro against all CAP isolates, including MRSA and penicillin-non-susceptible pneumococci, in contrast to the other beta-lactams tested. These data confirm ceftaroline as a new cephalosporin with enhanced anti-Gram-positive activity and suggest that ceftaroline has the potential to be a useful new agent in the treatment of CAP-associated bacteraemic infections.
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Activity of oritavancin against methicillin-resistant staphylococci, vancomycin-resistant enterococci and ?-haemolytic streptococci collected from western European countries in 2011.
J. Antimicrob. Chemother.
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To determine the activity of oritavancin against methicillin-resistant staphylococci, vancomycin-resistant enterococci (VRE) and ?-haemolytic streptococci recently isolated from acute bacterial skin and skin structure infections or bacteraemia in western Europe.
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In vitro activity of ceftaroline and comparator antimicrobials against European and Middle East isolates from complicated skin and skin-structure infections collected in 2008-2009.
Int. J. Antimicrob. Agents
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The activities of ceftaroline, the active metabolite of the pro-drug ceftaroline fosamil, a novel anti-meticillin-resistant staphylococcal cephalosporin, and nine comparators were determined against surveillance isolates collected in 2008-2009. Over 3000 isolates associated with complicated skin and skin-structure infections (cSSSIs) were collected from 106 centres in 19 countries. MICs were determined using CLSI broth microdilution methodology. Clonal relatedness of meticillin-resistant Staphylococcus aureus (MRSA) with raised ceftaroline MICs (2 mg/L) was assessed by MLST, PFGE and mec typing. The presence of Panton-Valentine leukocidin in these isolates was also determined. Ceftaroline was active against 500 MRSA and 479 meticillin-susceptible S. aureus, with MIC(50/90) values of 0.5/2 mg/L and 0.25/0.25 mg/L, respectively. For coagulase-negative staphylococci (CoNS), the ceftaroline MIC(50/90) values for meticillin-resistant strains (n=159) were the same as those seen for MRSA. Meticillin-susceptible CoNS (n=113) had the same MIC(90) as that seen with S. aureus, but the MIC(50) was lower at 0.06 mg/L. Ceftaroline was also active against ?-haemolytic streptococci (n=526; MIC(50/90)=0.004/0.015 mg/L), other streptococci (n=75; 0.015/0.06 mg/L), common Enterobacteriaceae (n=897; 0.25/?128 mg/L) and Enterococcus faecalis (n=329; 1/16 mg/L). Those MRSA with ceftaroline MICs of 2mg/L were found to be from four clonal groups associated with the country of origin. These data confirm the broad-spectrum in vitro activity of ceftaroline against cSSSI pathogens. Ceftaroline is unique among clinically available cephalosporins, having good in vitro activity against MRSA and meticillin-resistant CoNS and moderate activity against Gram-negative bacteria.
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A novel resistance mechanism to triclosan that suggests horizontal gene transfer and demonstrates a potential selective pressure for reduced biocide susceptibility in clinical strains of Staphylococcus aureus.
Int. J. Antimicrob. Agents
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The widely used biocide triclosan selectively targets FabI, the NADH-dependent trans-2-enoyl-acyl carrier protein reductase, which is an important target for narrow-spectrum antimicrobial drug development. In relation to the growing concern about biocide resistance, we compared in vitro mutants and clinical isolates of Staphylococcus aureus with reduced triclosan susceptibility. Clinical isolates of S. aureus as well as laboratory-generated mutants were assayed for minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) phenotypes and genotypes related to reduced triclosan susceptibility. A potential epidemiological cut-off (ECOFF) MBC of >4 mg/L was observed for triclosan in clinical isolates of S. aureus. These showed significantly lower MICs and higher MBCs than laboratory mutants. These groups of strains also had few similarities in the triclosan resistance mechanism. Molecular analysis identified novel resistance mechanisms linked to the presence of an additional sh-fabI allele derived from Staphylococcus haemolyticus. The lack of predictive value of in-vitro-selected mutations for clinical isolates indicates that laboratory tests in the present form appear to be of limited value. More importantly, detection of sh-fabI as a novel resistance mechanism with high potential for horizontal gene transfer demonstrates for the first time that a biocide could exert a selective pressure able to drive the spread of a resistance determinant in a human pathogen.
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Activity of ceftaroline against serotyped Streptococcus pneumoniae isolates from Europe and South Africa associated with community-acquired bacterial pneumonia (2007-08).
J. Antimicrob. Chemother.
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To determine the activity of ceftaroline (the active metabolite of the prodrug ceftaroline fosamil), a new cephalosporin recently approved in the USA for the treatment of community-acquired bacterial pneumonia, against serotyped Streptococcus pneumoniae causing community-acquired bacterial pneumonia from Europe and South Africa.
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Comparative Activity of Carbapenem Testing (COMPACT) study in Germany.
Int. J. Antimicrob. Agents
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The aim of this study was to determine the current susceptibility of hospital isolates of contemporary Gram-negative pathogens to the carbapenems doripenem, imipenem and meropenem. Between May and October 2008, seven centres in Germany were invited to collect and submit Pseudomonas aeruginosa, Enterobacteriaceae and other Gram-negative bacterial Intensive Care Unit (ICU)/non-ICU isolates from patients with complicated intra-abdominal infections (cIAIs), bloodstream infections (BSIs) or nosocomial pneumonia (NP). Susceptibility was determined at each centre by Etest. A central laboratory performed species confirmation as well as limited susceptibility and quality control testing. In total, 363 isolates were collected, comprising 46.0% Enterobacteriaceae, 45.2% P. aeruginosa, 4.7% Acinetobacter spp. and 4.1% other Gram-negatives. Most isolates (47.9%) were collected from NP, 32.8% were from cIAIs and 19.3% from BSIs; 57.3% were obtained from ICU patients. The MIC(90) values (minimum inhibitory concentration for 90% of the isolates) for doripenem, meropenem and imipenem were, respectively, 4, 16 and 32 mg/L against P. aeruginosa, 0.06, 0.06 and 0.5mg/L against Enterobacteriaceae and ? 64 mg/L for each carbapenem against other Gram-negative isolates. Using European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints, 81.1%, 75.6% and 79.3% of P. aeruginosa were susceptible to doripenem, imipenem and meropenem, respectively. Against all pathogens combined, MIC(90) values for ICU versus non-ICU isolates, respectively, were 4 mg/L vs. 1mg/L for doripenem, 8 mg/L vs. 1mg/L for meropenem and ? 64 mg/L vs. 8 mg/L for imipenem. Doripenem showed comparable activity against P. aeruginosa from patients with BSIs, cIAIs or NP. Similar findings were observed for Enterobacteriaceae and other Gram-negatives, including Acinetobacter spp. Doripenem generally showed similar or slightly better activity than meropenem and better activity than imipenem against Gram-negative pathogens collected in Germany.
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