Ana Cristina Gales

Emerging Importance of Multidrug-Resistant Acinetobacter Species and Stenotrophomonas maltophilia as Pathogens in Seriously Ill Patients: Geographic Patterns, Epidemiological Features, and Trends in the SENTRY Antimicrobial Surveillance Program (1997–1999)

As part of the SENTRY Antimicrobial Surveillance Program, a total of 1078 Acinetobacter species and 842 Stenotrophomonas maltophilia isolates were collected between January 1997 and December 1999 from 5 geographic regions (Canada, the United States, Latin America, Europe, and the Asia-Pacific). The frequency of infections (by geographic region and body site), including those due to imipenem-resistant Acinetobacter species and trimethoprim-sulfamethoxazole (TMP-SMZ)-resistant S. maltophilia, was evaluated. The possibility of seasonal variations in bloodstream infections caused by Acinetobacter species was studied, as was the activity of several therapeutic antimicrobials against all strains. Acinetobacter species and S. maltophilia were most frequently associated with pulmonary infections, independent of the region evaluated. In contrast, patterns of antimicrobial resistance markedly varied among distinct geographic regions, especially for nosocomial isolates. Although the carbapenems were the most active antimicrobials against Acinetobacter species, nearly 11.0% of the nosocomial isolates were resistant to this drug group in both regions. TMP-SMZ, ticarcillin-clavulanic acid, gatifloxacin, and trovafloxacin were the only agents with consistent therapeutic activity against S. maltophilia isolates. Rates of resistance to TMP-SMZ ranged from 2% in Canada and Latin America to 10% in Europe. The geographic differences in resistance patterns among Acinetobacter species and S. maltophilia isolates observed in this study emphasize the importance of local surveillance in determining the most adequate therapy for acinetobacter and S. maltophilia infections and the possible clonal, epidemic nature of occurrence.

Characterization of Pseudomonas aeruginosa Isolates: Occurrence Rates, Antimicrobial Susceptibility Patterns, and Molecular Typing in the Global SENTRY Antimicrobial Surveillance Program, 1997–1999

During 1997-1999, a total of 70,067 isolates (6631 Pseudomonas aeruginosa isolates) were analyzed in the SENTRY program by geographic region and body site of infection. The respiratory tract was the most common source of P. aeruginosa. P. aeruginosa isolation rates increased during the study interval. Europe was the only region to show a significant decline in beta-lactam and aminoglycoside susceptibility rates. There was a reduction in the rates of susceptibility of Canadian isolates to imipenem and of Latin American isolates to meropenem. A total of 218 multidrug-resistant P. aeruginosa isolates (MDR-PSA; resistant to piperacillin, ceftazidime, imipenem, and gentamicin) were observed; MDR-PSA occurrence rates (percentages of all isolates) ranged from 8.2% (Latin America) to 0.9% (Canada). No antimicrobial inhibited >50% of MDR-PSA strains. Molecular characterization of selected, generally resistant strains was performed. Isolates showing unique ribogroups were found in Europe, Latin America, and the United States, but clonal spread was documented in several medical centers.

Contemporary Assessment of Antimicrobial Susceptibility Testing Methods for Polymyxin B and Colistin: Review of Available Interpretative Criteria and Quality Control Guidelines

ABSTRACT The emergence of infections caused by multidrug-resistantPseudomonas aeruginosa and Acinetobacter spp. has necessitated the search for alternative parenteral agents such as the polymyxins. The National Committee for Clinical Laboratory Standards (NCCLS) documents do not currently provide interpretative criteria for the testing of the polymyxins, colistin and polymyxin B. Therefore, an evaluation of the antimicrobial activity of colistin and polymyxin B was initiated using 200 bloodstream infection pathogens collected through the SENTRY Antimicrobial Surveillance Program. All susceptibility tests were performed according to the NCCLS recommendations. Polymyxin B and colistin displayed a nearly identical spectrum of activity, exhibiting excellent potency against P. aeruginosa (MIC90, 2 μg/ml) andAcinetobacter sp. (MIC90, 2 μg/ml). In contrast, they showed limited activity against some other nonfermentative bacilli such as Burkholderia cepacia(MIC90, ≥128 μg/ml). Excellent correlation was achieved between broth microdilution and agar dilution tests (r= 0.96 to 0.98); 94.3% of the results were ±1 log2dilution between the methods used for both compounds. At a resistance breakpoint of ≥4 μg/ml for both agents, unacceptable false-susceptible or very major errors were noted for colistin (5%) and polymyxin B (6%). Modified zone criteria for colistin (≤11 and ≥14 mm) and polymyxin B (≤10 and ≥14 mm) were suggested, but some degree of error persisted (≥3.5%). It is recommended that all susceptible disk diffusion results be confirmed by MIC tests using the preferred reference NCCLS method. The quality control (QC) ranges listed in the product package insert require an adjusted range by approximately 3 mm for both NCCLS gram-negative quality control strains. This evaluation of in vitro susceptibility test methods for the polymyxin class drugs confirmed continued serious testing error with the disk diffusion method, the possible need for breakpoint adjustments, and the recalculation of disk diffusion QC ranges. Clinical laboratories should exclusively use MIC methods to assist the therapeutic application of colistin or polymyxin B until disk diffusion test modifications are sanctioned and published by the NCCLS.

Survey of Bloodstream Infections Due to Gram-Negative Bacilli: Frequency of Occurrence and Antimicrobial Susceptibility of Isolates Collected in the United States,Canada, and Latin America for the SENTRY Antimicrobial Surveillance Program, 1997

During 1997, a total of 4,267 nosocomial and community-acquired bloodstream infections due to gram-negative organisms were reported from SENTRY hospitals in Canada (8 sites), the United States (30 sites), and Latin America (10 sites). Escherichia coli was the most common isolate (41% of all gram-negative isolates), followed by Klebsiella species (17.9%), Pseudomonas aeruginosa (10.6%), and Enterobacter species (9.4%). For all gram-negative isolates combined, the most active antimicrobials tested were meropenem, imipenem, and cefepime. The quinolones levofloxacin (MIC90, 2 microg/mL), ciprofloxacin (MIC90, 1 microg/mL), gatifloxacin (MIC90, 2 microg/mL), sparfloxacin (MIC90, 2 microg/mL), and trovafloxacin (MIC90, 2 microg/mL) were also active against most isolates. Bloodstream infection isolates from Latin America were uniformly more resistant to all classes of antimicrobial agents tested than were isolates from Canada or the United States. Resistance phenotypes consistent with extended-spectrum beta-lactamase production were also most common among E. coli and Klebsiella species from Latin America. Further investigation of the reasons for regional differences in resistance patterns is needed, as is ongoing surveillance to detect resistance trends and to guide antimicrobial use.

Worldwide diversity of Klebsiella pneumoniae that produce β-lactamase blaKPC-2 gene.

TOC summary: Clones harboring different plasmids with identical genetic structure could be the origin of worldwide spread.

Evaluation of a New Etest for Detecting Metallo-β-Lactamases in Routine Clinical Testing

ABSTRACT Several Etest (AB BIODISK, Solna, Sweden) gradient formats were developed for detection of metallo-β-lactamases based on the reduction of imipenem (IP) or ceftazidime (TZ) MICs in the presence of EDTA or 2-mercaptopropionic acid (MPA). The Etest metallo-β-lactamase (Etest MBL) strips consisted of a double-sided seven-dilution range of IP or TZ (4 to 256 μg/ml) and IP or TZ (1 to 64 μg/ml) overlaid with a constant concentration of EDTA or MPA. The prototype strips were evaluated on several agar media (brain heart infusion agar, Isosensitest agar, nutrient agar, and Mueller-Hinton agar for aerobes and brucella blood agar for anaerobes) with 138 challenge strains: Acinetobacter spp. (n = 9), Aeromonas spp. (n = 8), Chryseobacterium spp. (n = 28), Escherichia coli (n = 1), Klebsiella pneumoniae (n = 4), Pseudomonas aeruginosa (n = 14), Proteus mirabilis (n = 3), Serratia spp. (n = 10), Stenotrophomonas maltophilia (n = 43), Sphingobacterium spp. (n = 3), and Bacteroides fragilis group (n = 15). PCR analysis using specific primers for IMP-1, L1, CcrA, and blaB/C confirmed the presence of the metallo-β-lactamase genes. Enzyme assays were also performed with IP as an indicator substrate followed by EDTA inhibition profiles. EDTA was found to be a better inhibitor of metallo-β-lactamases, especially for anaerobes. IP was a better than TZ. Mueller-Hinton agar was the preferred medium, particularly when compared to Isosensitest agar, which frequently produced falsely low MICs for IP. Etest IP plus IP-EDTA with Mueller-Hinton agar had a sensitivity of 94% (79 of 84) and specificity of 95% (124 of 130). The Etest MBL strip appears to be an acceptable diagnostic reagent to detect metallo-β-lactamase phenotypes in the clinical microbiology laboratory.

Rapid Detection and Identification of Metallo-β-Lactamase-Encoding Genes by Multiplex Real-Time PCR Assay and Melt Curve Analysis

ABSTRACT Metallo-β-lactamase enzymes (MβL) are encoded by transferable genes, which appear to spread rapidly among gram-negative bacteria. The objective of this study was to develop a multiplex real-time PCR assay followed by a melt curve step for rapid detection and identification of genes encoding MβL-type enzymes based on the amplicon melting peak. The reference sequences of all genes encoding IMP and VIM types, SPM-1, GIM-1, and SIM-1 were downloaded from GenBank, and primers were designed to obtain amplicons showing different sizes and melting peak temperatures (Tm). The real-time PCR assay was able to detect all MβL-harboring clinical isolates, and the Tm-assigned genotypes were 100% coincident with previous sequencing results. This assay could be suitable for identification of MβL-producing gram-negative bacteria by molecular diagnostic laboratories.

Contemporary activity of colistin and polymyxin B against a worldwide collection of Gram-negative pathogens: results from the SENTRY Antimicrobial Surveillance Program (2006–09)

OBJECTIVES To comparatively evaluate the antimicrobial activities of colistin and polymyxin B with those of other antimicrobials against a worldwide collection of 40 625 Gram-negative bacilli. METHODS Antimicrobial susceptibility testing was performed and interpreted using the CLSI broth microdilution method except for colistin against Enterobacteriaceae. RESULTS The polymyxins showed potent in vitro activities (MIC₉₀, ≤ 0.5-1 mg/L) against this large collection of clinical isolates, with very low resistance rates (< 0.1%-1.5%). Resistance to the polymyxins remained stable among organisms tested except for Klebsiella spp. isolates collected from the Asia-Pacific and Latin American regions, where a trend towards greater resistance was observed (P  ≤  0.05). In addition, an important reduction in imipenem susceptibility among Acinetobacter spp. and Klebsiella spp. was demonstrated in most geographical regions. CONCLUSIONS Although the polymyxins showed excellent in vitro activity against the vast majority of Gram-negative bacilli evaluated, a trend to greater resistance was observed in the Asia-Pacific and Latin American regions. Therefore, the clinical use of polymyxins must be cautious and surveillance monitored.

Multidrug-resistant Pseudomonas aeruginosa and Acinetobacter baumannii: resistance mechanisms and implications for therapy

Pseudomonas aeruginosa and Acinetobacter baumannii are major nosocomial pathogens worldwide. Both are intrinsically resistant to many drugs and are able to become resistant to virtually any antimicrobial agent. An increasing prevalence of infections caused by multidrug-resistant (MDR) isolates has been reported in many countries. The resistance mechanisms of P. aeruginosa and A. baumannii include the production of β-lactamases, efflux pumps, and target-site or outer membrane modifications. Resistance to multiple drugs is usually the result of the combination of different mechanisms in a single isolate or the action of a single potent resistance mechanism. There are many challenges in the treatment of MDR P. aeruginosa and A. baumannii, especially considering the absence of new antimicrobials in the drug-development pipeline. In this review, we present the major resistance mechanisms of P. aeruginosa and A. baumannii, and discuss how they can affect antimicrobial therapy, considering recent clinical, microbiological, pharmacokinetic and pharmacodynamic findings of the main drugs used to treat MDR isolates.

Antimicrobial activity and spectrum of the new glycylcycline, GAR-936 tested against 1,203 recent clinical bacterial isolates

The in vitro activity of GAR-936, a new semisynthetic glycylcycline, was evaluated in comparison with two tetracyclines and several other antimicrobial agents. A total of 1,203 recent clinical isolates were tested by reference broth or agar dilution methods. Among the members of the family Enterobacteriaceae, GAR-936 was generally two- to four-fold more active than minocycline, and two- to 16-fold more active than tetracycline. All enteric bacilli MIC90 results were < or = 4 microg/mL; the exception being Proteus mirabilis and indole-positive Proteae (> or = 8 microg/mL). GAR-936 demonstrated excellent activity against all gram-positive cocci with 90% of the penicillin-resistant Streptococcus pneumoniae isolates inhibited at 0.03 microg/ml, while the same isolates had a MIC90 of 8 and > 8 microg/mL for minocycline and tetracycline, respectively. All Enterococcus spp., including vancomycin-resistant isolates, were inhibited at 0.25 microg/mL of GAR-936 (MIC90, 0.12 or 0.25 microg/mL). Although GAR-936 (MIC50, 0.25 microg/mL) was two-fold less active than minocycline (MIC50, 0.12 microg/mL) against oxacillin-resistant Staphylococcus aureus, all isolates were inhibited at < or = 0.25 microg/mL. GAR-936 demonstrated good activity against nonfermentative bacteria such as Acinetobacter spp. (MIC90, 2 microg/ml) and Stenotrophomonas maltophilia (MIC90, 4 microg/mL), but the compound exhibited only modest activity against Pseudomonas aeruginosa (MIC50, 8 microg/mL). Haemophilus influenzae (MIC90, 1-2 microg/mL), Moraxella catarrhalis (MIC90, 0.12 microg/mL), and various Neisseria spp. (MIC90, 0.12-0.5 microg/mL) were susceptible to GAR-936. These results indicate that GAR-936 has potent in vitro activity against a wide range of clinically important pathogenic bacteria, and that several gram-positive and -negative isolates resistant to older tetracyclines and other drug classes remain susceptible to GAR-936, the newest glycylcycline candidate for clinical use.