Within a susceptible wild-type population, a small fraction of cells, even <10(-9) , is not affected when challenged by an antimicrobial agent. This subpopulation has mutations that impede antimicrobial action, allowing their selection during clinical treatment. Emergence of resistance occurs in the frame of a selective compartment termed a mutant selection window (MSW). The lower margin corresponds to the minimum inhibitory concentration of the susceptible cells, whereas the upper boundary, named the mutant prevention concentration (MPC), restricts the growth of the entire population, including that of the resistant mutants. By combining pharmacokinetic/pharmacodynamic concepts and an MPC strategy, the selection of resistant mutants can be limited. Early treatment avoiding an increase of the inoculum size as well as a regimen restricting the time within the MSW can reduce the probability of emergence of the resistant mutants. Physiological and, possibly, genetic adaptation in biofilms and a high proportion of mutator clones that may arise during chronic infections influence the emergence of resistant mutants. Moreover, a resistant population can emerge in a specific selective compartment after acquiring a resistance trait by horizontal gene transfer, but this may also be avoided to some extent when the MPC is reached. Known linkage between antimicrobial use and resistance should encourage actions for the design of antimicrobial treatment regimens that minimize the emergence of resistance. Emergence of a resistant bacterial subpopulation within a susceptible wild-type population can be restricted with a regimen using an antibiotic dose that is sufficiently high to inhibit both susceptible and resistant bacteria.

During the COMParative Activity of Carbapenems Testing (COMPACT) surveillance study, 448 Pseudomonas aeruginosa clinical isolates were obtained from 16 Spanish hospitals. Nonsusceptibility (EUCAST breakpoints) to imipenem (35%), meropenem (33%), and/or doripenem (33%) was observed with 175 isolates (39%). Simultaneous resistance to these three drugs was observed with 126 of the 175 isolates (72%). Except for colistin, high resistance rates were observed among noncarbapenem antibiotics. Clonal relatedness was investigated by pulsed-field gel electrophoresis (PFGE) with SpeI, discriminating 68 patterns. Multilocus sequence typing (MLST) was performed on 84 isolates representing different PFGE types and all participating hospitals. Thirty-nine sequence types (STs) could be distinguished, and of these, ST175 (48 isolates, 10 hospitals), ST646 (16 isolates, 4 hospitals), ST532 (13 isolates, 3 hospitals), and ST111 (13 isolates, 7 hospitals) were the most frequently encountered. Minimum-spanning tree analysis confirmed a wide dissemination of different clones among participant hospitals, particularly ST175. PFGE pattern comparison within the four most frequent STs revealed that ST175 isolates were relatively uniform, while ST646, ST532, and ST111 isolates were highly diverse, with almost every isolate belonging to a unique pulsotype, even when originating from the same center. The population of carbapenem-nonsusceptible P. aeruginosa isolates from 16 hospitals is highly diverse, with one ST (ST175) representing a highly conserved clone disseminated in 10 of the 16 participant hospitals. This ST175 clone should be added to the list of P. aeruginosa clones at high risk for epidemic spread, such as the Liverpool, Manchester, and Melbourne clones previously found in cystic fibrosis patients and ST235 in the nosocomial setting.Antibiotic surveillance studies are necessary not only for the better understanding of bacterial epidemiology, but also for the design of control strategies for preventing bacterial resistance and establishing therapeutic guidelines. In this context, the COMParative Activity of Carbapenems Testing (COMPACT) multicenter study was performed in 16 European countries, including Spain (23). The main objectives of this surveillance study were to test the activity of currently used carbapenems against Pseudomonas aeruginosa clinical isolates and to detect carbapenem nonsusceptible isolates (12).In this multicenter study, Spanish authors concluded that percentages of carbapenem resistance among P. aeruginosa clinical isolates in their country (range of 21.4% to 32.6%) are very similar to those previously reported in other European countries. Doripenem was the most active drug against these resistant isolates (12). The presence of carbapenem-nonsusceptible P. aeruginosa isolates has been described worldwide, particularly in relation to outbreaks (1,5,6,17,18,25,28,30,31,37). Most of these studies focused on clinical isolates, and genetic relatedness was established using molecular typing tools based on macror...

Therapeutic regimens containing (-lactam antibiotics are selecting penicillin-resistant Streptococcus pneumoniae populations all over the world. The selective pressure after 4 h of exposure to different concentrations of amoxicillin, cefixime, cefuroxime, and cefotaxime for low-level or high-level penicillin-resistant S. pneumoniae was evaluated in an in vitro model with mixed populations with penicillin susceptibilities of 0.015, 0.5, 1, and 2 ,ug/ml. The antibiotic concentration selecting for low-level resistance strongly reduced the susceptible population. Increasing antibiotic concentrations tended to decrease the total proportion of penicillin-resistant bacteria because of reduced numbers of the low-level-resistant population. The antibiotic concentration selecting for high-level resistance produced fewer resistant populations, but most of the organisms selected represented high-level resistance. In general, amoxicillin was a good selector for the low-level-resistant population and a poor selector for high-level resistance; cefuroxime and cefotaxime were poor selectors for low-level resistance and better selectors than amoxicillin for high-level penicillin resistance. Cefixime was the best selector of low-level penicillin resistance. When only resistant populations were mixed, the strains with high-level resistance were selected even at low antibiotic concentrations. Determination of the effects of selective antibiotic concentrations on mixed cultures of bacteria expressing different antibiotic resistance levels may help researchers to understand the ecology and epidemiology of penicillin-resistant S. pneumoniae populations.

The emergence of Tn4401-bla(KPC-3) within a pKPN3/4-like plasmid and its novel association with ST384 and ST388 K. pneumoniae clones in Spain is reported. Although bla(KPC-3) has been scarcely reported in Europe, the location of this Tn4401a in a widespread K. pneumoniae plasmid supports the possibility of broader future dissemination.

During the 2 year period following the emergence of non-ST258 KPC-3-producing K. pneumoniae isolates in our institution, the blaKPC-3 and blaKPC-2 genes efficiently penetrated other Enterobacteriaceae lineages. Non-ST258 K. pneumoniae isolates were mostly responsible for the dissemination of KPC enzymes, producing a complex epidemiological picture.

Inhaled administration of tobramycin assures high concentrations in cystic fibrotic lungs, improving the therapeutic ratio over that of parenteral tobramycin levels, particularly against Pseudomonas aeruginosa. Conventional Clinical and Laboratory Standards Institute (CLSI; formerly National Committee for Clinical Laboratory Standards) breakpoints only consider parenteral levels and do not take into account these high antimicrobial concentrations. The Spanish Antibiogram Committee (The MENSURA Group) has tentatively defined specific breakpoint values for inhaled tobramycin when testing P. aeruginosa isolates from cystic fibrosis (CF) patients (susceptible, <64 g/ml; resistant, >128 g/ml). The antimicrobial susceptibilities of 206 prospectively collected CF P. aeruginosa isolates were determined by the reference agar dilution method. For tobramycin, the performance of high range tobramycin Etest strips (AB Biodisk, Solna, Sweden) and conventional tobramycin disks were assessed with the same collection. Applying MENSURA proposed breakpoints, 95.1% of the strains were categorized as susceptible to tobramycin, either using agar dilution or Etest high-range strips (99% categorical agreement between both methods). With CLSI breakpoints, susceptibility rates decreased to 79.1 and 81.1% for agar dilution and Etest strips, respectively (83.5% categorical agreement). Minor, major, and very major errors for Etest strips (CLSI criteria) were 13.6, 1.2, and 14.8%, respectively. Upon applying the new proposed criteria for inhaled tobramycin, only one major and one very major error were observed with Etest strips. Whenever inhaled tobramycin is considered for therapy, we suggest that P. aeruginosa strains from CF patients categorized as intermediate or resistant to tobramycin according to the CLSI criteria should be retested with high-range Etest strips and recategorized using MENSURA interpretive criteria. CLSI breakpoints should still be followed when intravenous tobramycin is used in CF patients, particularly during the course of exacerbations.

Ten unique changes in the 590-641 region of PBP2B and no specific murM alleles were found in S. pneumoniae strains isolated in Spain with an amoxicillin MIC>or=8 mg/L (MICs from 6 to 12 mg/L by 1 mg/L step dilution). In addition, the presence of specific mutations in PBP2B seems to play a key role in the presence of different murM alleles in these amoxicillin-resistant pneumococcal strains.

Susceptibility results with low reproducibility by the same or different methods have been observed for metallo-beta-lactamase (MBL)-producing Enterobacteriaceae. Eighteen VIM-1-producing Klebsiella pneumoniae isolates (one per patient) belonging to a single epidemic clone in our hospital (2005 to 2008) but with different susceptibilities to carbapenems were studied. Imipenem MICs ranged from 8 to >128 mg/liter by standard CLSI microdilution, from <1 to >8 mg/liter by the semiautomatic Wider system, and from 0.75 to >32 mg/liter by Etest. Meropenem MICs ranged from 0.5 to 128, <1 to >8, and 0.38 to >32 mg/liter, respectively. Ertapenem MICs by CLSI microdilution and Etest ranged from 1 to 64 and 0.75 to >32 mg/liter, respectively. The rates of essential agreement (؎1 log 2 dilution) for imipenem and meropenem MICs between the Wider system and the reference microdilution method were 45% and 49%, respectively. Those between Etest and the reference microdilution method for imipenem, meropenem, and ertapenem MICs were 33%, 67%, and 84%. The rates of very major errors for the Wider system and Etest were 33% and 28% for imipenem and 25% and 75% for meropenem, respectively. Low MIC reproducibility was observed even when the same inoculum was used (differences up to 4-fold dilutions). Heteroresistance was suspected due to the presence of colonies in the Etest inhibition zone. It was confirmed by population analysis profiles of 4 isolates displaying different imipenem MICs, with the exception of an OmpK36-porin-deficient isolate that homogeneously expressed carbapenem resistance (MIC, >128 mg/liter). Low carbapenem MIC reproducibility could be due to the presence of resistant subpopulations and variable expression of the resistance mechanisms. Since carbapenem MICs are not good markers of MBL production, reliable and reproducible phenotypic methods are needed to detect the presence of this mechanism.