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Multidrug-resistant Acinetobacter baumannii has emerged as a serious nosocomial pathogen in certain areas. In Brooklyn, New York, citywide surveillance revealed that ∼2 of every 3 isolates were resistant to carbapenem antibiotics. Genetic fingerprinting revealed that 2 strains accounted for 82% of these resistant isolates. Compared with carbapenem-susceptible isolates, carbapenem-resistant isolates had reduced expression of 47-, 44-, and 37-kDa outer-membrane proteins. No specific carbapenemase was found; however, carbapenem-resistant isolates expressed greater levels of a class C cephalosporinase. Although expression of penicillin-binding proteins varied among strains, no consistent pattern appeared to account for carbapenem resistance. An efflux pump, present in several strains, did not appear to contribute to carbapenem resistance. Clonal spread of carbapenem-resistant A. baumannii has occurred in hospitals in Brooklyn. The preliminary findings for a small number of strains suggest that diminished production of outer-membrane porins, together with increased expression of a class C cephalosporinase, appear to be important factors leading to carbapenem resistance in this region.

Isolates of serotype 23F Streptococcus pneumoniae with high levels of resistance to penicillin have been commonly recovered in Spain for more than a decade. Recently penicillin-resistant serotype 23F S. pneumoniae strains were also isolated from children attending a day-care center in Cleveland. A number of Spanish and Cleveland isolates were compared by electrophoretic analysis of penicillin-binding protein (PDP) profiles and DNA restriction endonuclease cleavage profiles of the PDP 2X and 2D genes amplified with the polymerase chain reaction and by multilocus enzyme electrophoresis. All strains were identical by these criteria. The findings demonstrate that the Spanish and Cleveland isolates are clonally related and suggest that this antibiotic resistant clone of serotype 23F S. pneumoniae has spread intercontinentally from Spain to the United States.

Surveillance for nasopharyngeal colonization with Streptococcus pneumoniae was maintained in a research day care center between 1985 and 1992. An outbreak of nasal carriage of a multidrug- resistant (MDR) serotype 23F organism occurred between May 1990 and December 1991 involving 14 of 52 children. Electrophoresis of penicillin-binding proteins (PBP) and pulsed-field gel electrophoresis (PFGE) of chromosomal DNA indicated that the MDR serotype 23F organism was closely related to a serotype 23F MDR clone that has been prevalent in Spain since the early 1980s. In June 1991, an MDR serotype 14 organism was isolated from a child who had previously carried the MDR serotype 23F strain. PFGE and PBP typing revealed that the MDR serotype 14 organism was very similar to the circulating MDR serotype 23F strain, suggesting serotype transformation. Dissemination of MDR pneumococcal strains and possibly spread of the MDR phenotype to additional serotypes may be facilitated in group day care.

Examination of several hundred penicillin-resistant clinical isolates of Streptococcus pneumoniae has revealed extensive strain-to-strain variation in the number and molecular size of penicillin-binding proteins (PBPs), This polymorphism has been used to classify resistant isolates into groups (PBP families) that share distinct electrophoretic profiles, We describe herein properties of four such PBP families: two from Spain (and/or Ohio) and one each from Hungary and Alaska. We have discovered that representative isolates assigned to each PBP family also share capsular serotype, antibiotic resistance pattern, pneumococcal surface protein A type, and multilocus enzyme genotype. The results demonstrate independent clonal origin for strains assigned to each PBP family. Each resistant clone occurs with uniquely high incidence within specific geographic areas.

β-Lactam antibiotics exert their antibacterial effects by inactivating the high-molecular-weight penicillin-binding proteins (PBPs) that are responsible for the final stages of peptidoglycan biosynthesis. The availability of the amino acid sequences of several low-molecular-weight PBPs, high-molecular-weight PBPs, and active-site serine β-lactamases has provided evidence that these groups of enzymes have a common, but distant, evolutionary origin. This view is strongly supported by the recent finding of a similarity in the three-dimensional structures of a low-molecular-weight PBP and class A β-lactamases. The high-molecular-weight PBPs of Escherichia coli are believed to possess an amino-terminal peptidoglycan transglycosylase domain and a carboxy-terminal penicillin-sensitive transpeptidase domain. These enzymes are inserted in the cytoplasmic membrane only at their amino termini, and water-soluble forms have been obtained that should be suitable for crystallization and X-ray analysis. Resistance to β-lactam antibiotics mediated by alterations of PBPs has been reported in some gram-negative bacteria. In isolates of Neisseria gonorrhoeae with chromosomally mediated resistance, penicillin-resistant PBPs have arisen from the introduction of multiple amino acid substitutions within the transpeptidase domain of the enzymes.

We studied the mechanism of resistance to imipenem in three clinical isolates of Pseudomonas aeruginosa. Two of these isolates arose from imipenem-susceptible strains isolated during therapy with imipenem and were associated with treatment failure. One of these two strains had previously been broadly resistant to β-lactams; the second acquired resistance to imipenem alone. One isolate of the third strain was resistant to imipenem but susceptible to other antipseudomonal β-lactams. No isolate contained β-lactamase activity capable of hydrolyzing imipenem at a detectable rate. Studies of the penicillin-binding proteins of all isolates revealed no differences in the number of proteins, molecular weight of, affinity for penicillin, or affinity for imipenem in any isolate. In each case the resistant isolate lacked one or more outer membrane proteins that were present in a susceptible isolate of the same strain. The observed alterations in outer membrane proteins may be associated with diminished permeability of the bacterial outer membrane to imipenem and may be the major factor responsible for resistance in these isolates.

β-lactams active against methicillin-resistant Staphylococcus aureus (MRSA) must resist penicillinase hydrolysis and bind penicillin-binding protein 2A (PBP 2A). Cefamandole might share these properties. When tested against 2 isogenic pairs of MRSA that produced or did not produce penicillinase, MICs of cefamandole (8–32 mg/L) were not affected by penicillinase, and cefamandole had a ⩾40 times greater PBP 2A affinity than did methicillin. In rats, constant serum levels of 100 mg/L cefamandole successfully treated experimental endocarditis due to penicillinase-negative isolates but failed against penicillinase-producing organisms. This suggested that penicillinase produced in infected vegetations might hydrolyze the drug. Indeed, cefamandole was slowly degraded by penicillinase in vitro. Moreover, its efficacy was restored by combination with sulbactam in vivo. Cefamandole also uniformly prevented MRSA endocarditis in prophylaxis experiments, a setting in which bacteria were not yet clustered in the vegetations. Thus, while cefamandole treatment was limited by penicillinase, the drug was still successful for prophylaxis of experimental MRSA endocarditis.

Enterococci are important nosocomial pathogens among which resistance to multiple antibiotics is being recognized with increasing frequency. We characterized three clinical isolates from three New York City hospitals that demonstrated concomitant resistance to vancomycin (one VanA, two VanB phenotypes) and high-level resistance to penicillin. Two Enterococcus faecium strains were intrinsically highly resistant to penicillin and showed very low affinity for penicillin of penicillin-binding protein 5. Unlike previously described glycopeptide-resistant enterococci, these strains were not hypersusceptible to β-lactam agents after vancomycin induction, and combinations of penicillin and vancomycin were not synergistic against them. A third isolate, Enterococcus faecalis, produced β-lactamase. Two of the three strains were also highly resistant to all aminoglycosides. Emergence of concomitant high-level resistance to multiple antibiotic classes among enterococci considerably narrows the therapeutic options for treatment of infections due to these opportunistic pathogens.

Penicillin-resistant strains of Streptococcus pneumoniae that are isolated with increasing frequency worldwide contain low-affinity penicillin-binding proteins (PBPs). The relatedness of PBPs from 55 resistant strains isolated on three continents wasinvestigatedby testing the reactivity of antibodies specific for PBP la or 2b and by comparing the PBP patterns. Seventeen patterns of antibody reactivity could be distinguished, 12 of which were specific to one isolate. Most strains, including all German and South African strains, had a unique PBP profile. A few groups of Spanish and Finnish isolates were identified where the strains within each group shared the same PBP profile, the same antigenic variants of PBPs la and 2b, and the same serogroup, suggesting that they represent different clones of S. pneumoniae. The results demonstrated highly variable pathways of resistance development and confirmed that resistant strains have emerged independently in different locations.

Previous studies have suggested that relatively penicillin-resistant (RPR) capsular group 9L strains in western Canada may be clonally related. To test this hypothesis, restriction fragment length polymorphisms (RFLPs) were examined using DNA probes for pspA and a newly recognized pneumococcal genetic element, IS1167. Penicillin-binding proteins (PBPs) and PBP genes from representative strains were also studied. All RPR type 9L strains demonstrated an identical RFLP when probed with IS1167, and 12 of 14 RPR strains had the same RFLP when examined with pspA. Amplification of pspA by polymerase chain reaction and restriction endonuclease digestion showed that the 9L strains had common DNA fragments not identified in any of the penicillin-susceptible strains. The 9L strains apparently have a low-affinity PBP 2B distinct from those of other capsular types. These data derived from new genetic markers and PBP analysis strongly support a clonal origin of RPR type 9L pneumococci of western Canada.