Methicillin-susceptible Strains Research Articles

Methicillin-Resistant Staphylococcal Colonization and Infection in a Long-Term Care Facility

To determine the natural history of colonization by methicillin-resistant Staphylococcus aureus (MRSA) among patients in a long-term care facility. We specifically sought to determine if MRSA colonization was predictive of subsequent infection. Cohort study. Long-term Veterans Affairs Medical Center. A total of 197 patients residing on two units were followed with regular surveillance cultures of the anterior nares. The development of staphylococcal infection. Thirty-two patients were persistent carriers of MRSA and 44 were persistent carriers of methicillin-susceptible strains (MSSA). Twenty-five percent of MRSA carriers had an episode of staphylococcal infection compared with 4% of MSSA carriers and 4.5% of non-carriers (P less than 0.01; relative risk 3.8; 95% CI, 2.0 to 6.4). The rate of development of infection among MRSA carriers was 15% for every 100 days of carriage. Using logistic regression analysis, persistent MRSA carriage was the most significant predictor of infection (P less than 0.001; odds ratio, 3.7). Seventy-three percent of all MRSA infections occurred among MRSA carriers. Isolates of MRSA from 7 patients were typed. Colonizing and infecting strains had the same phage type in all 7 patients and the same pattern of plasmid EcoRI restriction endonuclease fragments in 5 patients. Colonization of the anterior nares by MRSA predicts the development of staphylococcal infection in long-term care patients; most infections arise from endogenously carried strains. Colonization by MRSA indicates a significantly greater risk for infection than does colonization by MSSA. The results offer a theoretic rationale for reduction in MRSA infections by interventions aimed at eliminating the carrier state.

Daptomycin compared with teicoplanin and vancomycin for therapy of experimental Staphylococcus aureus endocarditis

The efficacies of daptomycin, teicoplanin, and vancomycin were compared in the therapy of experimental Staphylococcus aureus endocarditis. Rabbits infected with either of two methicillin-susceptible strains (SA-12871 or its moderately teicoplanin-resistant derivative SA-12873) or a methicillin-resistant S. aureus strain (MRSA-494) were treated with daptomycin, 8 mg/kg of body weight, every 8 h; teicoplanin, 12.5 mg/kg (low-dose teicoplanin [teicoplanin-LD], excluding MRSA-494) or 40 mg/kg (high-dose teicoplanin [teicoplanin-HD]) every 12 h; or vancomycin, 17.5 mg/kg every 6 h, for 4 days. Compared with no treatment daptomycin, teicoplamin-HD, and vancomycin significantly reduced bacterial counts of all test strains in vegetations and renal and splenic tissues (P less than 0.001). Teicoplanin-LD was equally effective against SA-12871 but failed against SA-12873, with three of six animals still being bacteremic at the end of therapy. For SA-12871, daptomycin was as effective as teicoplanin-HD and was superior to teicoplanin-LD and vancomycin (P = 0.02) in lowering vegetation bacterial counts. There were no differences between daptomycin, teicoplanin-HD, or vancomycin in the reduction of bacterial counts in tissues for any of the test strains. In rabbits infected with SA-12871, vegetations from 33% of teicoplanin-LD-treated, 6% of teicoplanin-HD-treated, and 13% of daptomycin-treated animals yielded organisms for which there were up to eightfold increases in the MICs. Resistance may have contributed to early death in one daptomycin-treated animal. No increases in the MICs for the test strain were detected in animals infected with SA-12873 or MRSA-494. We conclude that in this model and against these strains of S. aureus, daptomycin and teicoplanin-HD are as efficacious as vancomycin, but diminished susceptibility to both can develop during therapy.

Mathematical modeling of antimicrobial susceptibility data of Staphylococcus haemolyticus for 11 antimicrobial agents, including three experimental glycopeptides and an experimental lipoglycopeptide

Antimicrobial MIC data were obtained for 96 strains of Staphylococcus haemolyticus and the following 11 antimicrobial agents: methicillin, gentamicin, rifampin, fusidic acid, ciprofloxacin, vancomycin, teicoplanin; three experimental glycopeptides, MDL 62,873, MDL 62,208, and MDL 62,224; and an experimental lipoglycopeptide, ramoplanin. Resistance to methicillin and gentamicin was present in over 50% of the strains, although resistance to the other agents was present in less than 10%. It is shown how application of mathematical modeling techniques can add to the understanding of such MIC data. MICs of methicillin and gentamicin were highly correlated, suggesting that evolutionary pressures for development of resistance to these agents were similar. The structural relationships among the glycopeptides were accurately reflected in their spatial relationships within the model. MICs of ramoplanin were negatively correlated with MICs of some other antimicrobial agents, particularly gentamicin, suggesting that this agent is more active against gentamicin-resistant strains. Methicillin-resistant strains were more tightly clustered than were methicillin-susceptible strains, suggesting that methicillin-resistant strains were more closely related to each other than were methicillin-susceptible strains. Mathematical modeling techniques enable more detailed analysis of MIC data.

Binding Affinity for Penicillin-Binding Protein 2a Correlates with In Activity of -Lactan Antibiotics against Methicillin-Resistant Staphylococcus aureus

The beta-lactam antibiotics ticarcillin, nafcillin, imipenem, and ampicillin, which differ in antibacterial activity against methicillin-resistant strains of Staphylococcus aureus, were examined for affinity to penicillin-binding protein (PBP) 2a, which mediates methicillin resistance. The relative efficacy of each antibiotic was compared to vancomycin in a rabbit model of aortic valve endocarditis caused by either a methicillin-susceptible or methicillin-resistant strain of beta-lactamase-producing S. aureus. beta-lactamase inhibitors clavulanate and sulbactam were used in combination with ticarcillin and ampicillin, respectively. All beta-lactam antibiotics were effective against the susceptible strain. beta-lactam antibiotic activity in vitro and in vivo against the resistant strain correlated with its affinity for binding to PBP 2a. Lack of efficacy of beta-lactam antibiotics for the resistant strain was due to an inability to eradicate the resistant subpopulation of cells. Vancomycin was the most effective agent.

Increasing resistance of Staphylococcus aureus to ciprofloxacin

We demonstrated the marked emergence of resistance to ciprofloxacin among Staphylococcus arueus strains isolated at the Ann Arbor Veterans Administration Medical Center. All S. aureus isolates tested from 1984 to 1985 were susceptible, whereas 55.1% of methicillin-resistant and 2.5% of methicillin-susceptible strains from 1989 had high-level resistance to ciprofloxacin.

Binding of -Lactam Antibiotics to Penicillin-Binding Proteins in Methicillin-Resistant Staphylococcus aureus

Binding affinity of penicillin-binding proteins (PBP) of Staphylococcus aureus for several beta-lactam antibiotics was examined in a methicillin-susceptible strain and in two methicillin-resistant strains, one heterogeneous and one homogeneous, to determine relationships between PBP binding and expression of resistance. PBPs 1-4 had similar affinities in all three strains. PBP 2a affinities were similar in both resistant strains. For the susceptible strain and for the susceptible subpopulation of cells in the heterogeneous strain, growth inhibition correlated with binding to PBPs 1-3, suggesting that these PBPs mediate susceptibility in both. For resistant cells, growth inhibition correlated only with binding to PBP 2a, suggesting that this is the target in resistant cells. Determination of binding affinity for PBP 2a or the concentration that inhibits growth of the most resistant subpopulation of cells should be included in evaluation of beta-lactam antibiotics for potential activity against methicillin-resistant strains of S. aureus.

Correlation between esterase electrophoretic types and capsular polysaccharide types 5 and 8 among methicillin-susceptible and methicillin-resistant strains of Staphylococcus aureus

The relationship between capsular polysaccharide types 5 and 8 and esterase electrophoretic types (zymotypes) in 160 French clinical isolates of Staphylococcus aureus was studied. Methicillin-susceptible strains of capsular types 5 and 8 were represented by 11 zymotypes, indicating a high polymorphism. Methicillin-resistant strains were mainly distributed in only two distinct populations. The predominant population was represented by strains of zymotype 6 and capsular type 5, and the second population was represented by strains of zymotype 14 and capsular type 8.

Synergistic effect of quinolones and oxacillin on methicillin-resistant Staphylococcus species.

Various combinations of antistaphylococcal antimicrobial agents have been tested against 17 selected Staphylococcus isolates, including methicillin-susceptible and methicillin-resistant strains of S. aureus and coagulase-negative Staphylococcus species. With the checkerboard technique the following combinations were tested: oxacillin-ofloxacin, oxacillin-temafloxacin, oxacillin-fleroxacin, vancomycin-fleroxacin, gentamicin-fleroxacin, and rifampin-fleroxacin. Against methicillin-resistant staphylococci the combination oxacillin-quinolone tested at 35 degrees C always showed a fractional inhibitory concentration (FIC) index of less than 0.75, which is interpreted as synergistic or additive. Equal or more synergistic effects were observed at 30 degrees C. In contrast, when methicillin-susceptible Staphylococcus species were tested, the FIC for the combination oxacillin-quinolone was always 1 or 2, which is considered to be indifferent. For the other mentioned combinations the FICs were also 1 or 2. Killing kinetics showed synergistic or additive bactericidal activity for the combination oxacillin-ofloxacin against methicillin-resistant Staphylococcus species, killing 1.5 to 2.8 log10 CFU more of these per ml than did the most active drug after 24 h of incubation. This difference was not observed for methicillin-susceptible strains. In vitro evidence for the potential clinical use of quinolones in treating infections due to methicillin-resistant staphylococci in combination with a beta-lactamase-resistant penicillin is provided.

New mechanism for methicillin resistance in Staphylococcus aureus: clinical isolates that lack the PBP 2a gene and contain normal penicillin-binding proteins with modified penicillin-binding capacity.

Seventeen clinical isolates of Staphylococcus aureus (from the United States and Europe) selected for low (borderline)-level methicillin resistance (MIC of methicillin, 2 to 4 micrograms/ml; MIC of oxacillin, 0.5 to 8 micrograms/ml) were examined for their mechanisms of resistance. Five strains were typical of heterogeneous S. aureus: they gave positive reactions with a DNA probe specific for mec and contained a small fraction (10(-6] of highly resistant cells (MIC, greater than 100 micrograms/ml). The rest of the 12 strains were homogeneous with respect to their methicillin resistance: the MIC of methicillin for all cells was 2 to 4 micrograms/ml, and no cells for which MICs were 50 micrograms/ml or higher were detectable (less than 10(-9]. None of these strains reacted with the mec-specific DNA probe. One representative strain of each group was characterized in more detail. Strain CDC-1, prototype of heterogeneous methicillin-resistant S. aureus, contained penicillin-binding protein (PBP) 2a; its DNA could transform a methicillin-susceptible and novobiocin-resistant recipient to methicillin resistance with ca. 35% linkage to Novr. Introduction of the "factor X" determinant (K. Murakami and A. Tomasz, J. Bacteriol. 171:874-879, 1989) converted strain CDC-1 to high, homogeneous resistance. Strain CDC-6, prototype of the second group of isolates, showed completely homogeneous MICs of methicillin, oxacillin, and cefotaxime. The strain contained modified "normal" PBPs: PBPs 1 and 2 showed low drug reactivity (and/or cellular amounts), and PBP 4 was present in elevated amounts. No PBP 2a could be detected. DNA isolated from strain CDC-6 could transform the methicillin-susceptible and novobiocin-resistant strain to methicillin resistance in a multistep fashion, but this resistance showed no genetic linkage to the Nov marker. We suggest that staphylococci with borderline resistance may contain at least three different classes of mechanism: heterogeneous, methicillin-resistant S. aureus, PBPs of modified drug reactivities, and the previously reported hyperproduction of beta-lactamase (L.K. McDougal and C. Thornsberry, J. Clin Microbiol. 23:832-839, 1986).

Lower autolytic activity in a homogeneous methicillin-resistant Staphylococcus aureus strain compared to derived heterogeneous-resistant and susceptible strains

It has been proposed that in addition to production of a penicillin-binding protein with low affinity for beta-lactam antibiotics, control of autolysin activity is involved in the mechanism of staphylococcal methicillin resistance. A homogeneous methicillin-resistant Staphylococcus aureus strain (DU4916) had lower rates of unstimulated, NaCl- and Triton X-100-stimulated autolysis, and daptomycin (LY146032)-induced lysis than a heterogeneous methicillin-resistant strain (DU4916-K7) and a methicillin-susceptible strain (DU4916S) derived from DU4916.

Lower autolytic activity in a homogeneous methicillin-resistant Staphylococcus aureus strain compared to derived heterogeneous-resistant and susceptible strains

It has been proposed that in addition to production of a penicillin-binding protein with low affinity for beta-lactam antibiotics, control of autolysin activity is involved in the mechanism of staphylococcal methicillin resistance. A homogeneous methicillin-resistant Staphylococcus aureus strain (DU4916) had lower rates of unstimulated, NaCl- and Triton X-100-stimulated autolysis, and daptomycin (LY146032)-induced lysis than a heterogeneous methicillin-resistant strain (DU4916-K7) and a methicillin-susceptible strain (DU4916S) derived from DU4916.

Widespread quinolone resistance among methicillin-resistant Staphylococcus aureus isolates in a general hospital.

Ofloxacin and ciprofloxacin resistance (MIC, greater than 4 micrograms/ml) was encountered in 45 of 50 clinical isolates of methicillin-resistant Staphylococcus aureus. None of 20 methicillin-susceptible strains was resistant to the quinolones (P less than 10(-6). Quinolone-susceptible and -resistant isolates did not differ with respect to culture source or bacteriophage type. The future usefulness of quinolones for S. aureus infection may be limited.

In vitro interaction between rifampin and clindamycin against pathogenic coagulase-negative staphylococci.

The MICs and MBCs for 90% of strains tested (MIC90 and MBC90, respectively) of rifampin for 75 clinical isolates of pathogenic coagulase-negative staphylococci (PCNS) were 0.03 and 0.25 microgram/ml, respectively, while the MIC90 and MBC90 of clindamycin were both greater than 25 micrograms/ml. Although no synergy between rifampicin and clindamycin was found among the 15 strains studied by the checkerboard method, 6 of 12 selected strains showed synergy by the kill-curve method. No antagonism was observed by either method. All 30 strains rapidly developed resistance to rifampin in vitro, and this could be prevented by the simultaneous presence of 1.0 microgram of clindamycin per ml in the 24 methicillin-susceptible PCNS strains. The synergy between rifampin and clindamycin observed in vitro for some strains of PCNS, together with the prevention of emergence of resistance to rifampin by clindamycin, suggests that this antibiotic combination may be useful for the treatment of infections caused by methicillin-susceptible PCNS.

The antibacterial activity of meropenem in combination with gentamicin or vancomycin

The kinetics of bacterial killing by meropenem alone and in combination with gentamicin (for Pseudomonas aeruginosa) and vancomycin (for Staphylococcus aureus) were studied for two strains of each species. Against the two strains of P. aeruginosa, meropenem at concentrations up to 4 x MIC was rapidly bactericidal--but regrowth occurred by 24 h. The addition of half the MIC of gentamicin to the MIC of meropenem led to a more rapid decline of the colony count and to the prevention of regrowth of the strain which was gentamicin-susceptible. Similar results were obtained for a methicillin-susceptible strain of S. aureus when vancomycin was added at a concentration of half the MIC. A methicillin-resistant strain also was killed by a combination of vancomycin at half the MIC plus meropenem at the MIC. The study showed that the killing action of meropenem against P. aeruginosa and staphylococci is enhanced by the addition of gentamicin or vancomycin respectively.

Rapid detection (4 h) of methicillin-resistant Staphylococcus aureus by a bioluminescence method.

A 4-h bioluminescence method for methicillin susceptibility determination was compared with reference methods. Of the Staphylococcus aureus strains tested, 80 were methicillin resistant, 180 were methicillin susceptible, and 10 were borderline susceptible. There was 100% correlation between bioluminescence and reference methods for methicillin-susceptible and methicillin-resistant strains. All borderline-susceptible strains were identified as methicillin resistant by bioluminescence.

Effects of growth of methicillin-resistant and -susceptible Staphylococcus aureus in the presence of beta-lactams on peptidoglycan structure and susceptibility to lytic enzymes.

Growth of methicillin-resistant Staphylococcus aureus DU4916 in the presence of methicillin yielded crude cell walls that showed an increased rate of autolysis and purified cell walls (PCW) and peptidoglycan (PG) that had increased susceptibilities to autolysin extracted with LiCl and to lysozyme. The PG of cells grown in the presence of methicillin had markedly decreased cross-linking and O acetylation. Growth of the methicillin-susceptible strain H in the presence of subinhibitory concentrations of cefoxitin, a specific inhibitor of penicillin-binding protein (PBP) 4, caused a substantial decrease in PG cross-linking and O acetylation and increased susceptibilities of PCW and PG to LiCl-extracted autolysin and to lysozyme. Strain DU4916 cells grown in the presence of methicillin did not show an increased rate of autolysis or an increased susceptibility to vancomycin- or D-cycloserine-induced lysis, even though their PG was hypo-cross-linked. This implies that the potential for increased autolysis is controlled in intact cells and that this regulation may be involved in the methicillin resistance phenomenon. Growth of the methicillin-susceptible strain DU4916S in the presence of methicillin yielded PCW and PG that showed small increases in susceptibilities to LiCl-extracted autolysin and to lysozyme and a small decrease in PG cross-linking. Comparison of the PBPs of a penicillinase-nonproducing derivative of strain DU4916 (DU4916-K7) with those of strain DU4916S in intact cells and isolated membranes revealed that PBPs 1 to 4 had similar high beta-lactam antibiotic affinities in both strains and identified an additional PBP, PBP2(1), with low beta-lactam affinity in the methicillin-resistant strain DU4916-K7. The low degree of cross-linking of PG in strain DU4916 cells grown with methicillin was probably due mainly to inhibition of the secondary cross-linking function of PBP 4.

Screening method for rapid detection of methicillin-resistant (heteroresistant) Staphylococcus aureus.

A broth screening method was developed to detect methicillin-resistant (heteroresistant) strains of Staphylococcus aureus from primary plates. Lyophilized vials containing cation-supplemented Mueller-Hinton broth, 2% NaCl, and 10 micrograms of methicillin and 6 micrograms of oxacillin per ml were hydrated and tested with 129 methicillin-resistant and 35 methicillin-susceptible strains of S. aureus. With the addition of a tetrazolium indicator after 5 h of incubation, 96.9% of resistant strains were detected. No false-positive results occurred.

Pefloxacin therapy for experimental endocarditis caused by methicillin-susceptible or methicillin-resistant strains of Staphylococcus aureus.

The therapeutic efficacy of pefloxacin in experimental endocarditis caused by methicillin-susceptible or methicillin-resistant Staphylococcus aureus was evaluated. In rabbits infected with a methicillin-susceptible strain, 4 days of pefloxacin therapy significantly reduced both the number of bacteria per gram of vegetation and the mortality rate compared with untreated controls, and pefloxacin was equivalent to cephalothin. Pefloxacin was also as effective as vancomycin in reducing vegetation titers and mortality rate in animals with endocarditis caused by a methicillin-resistant strain. These results suggest that pefloxacin may be an effective agent in the therapy of serious infections caused by either methicillin-susceptible or -resistant strains of S. aureus.

In vitro activities of rifapentine and rifampin, alone and in combination with six other antibiotics, against methicillin-susceptible and methicillin-resistant staphylococci of different species.

The antistaphylococcal activity of rifapentine, a new rifamycin SV derivative, was evaluated in vitro and compared with that of rifampin. A total of 313 staphylococcal strains freshly isolated from clinical material and including representatives of all currently recognized Staphylococcus species of human origin were used. The susceptibility to methicillin of all the test strains was determined preliminarily. Despite minor differences with some species, the MICs of rifapentine were found to be substantially similar to those of rifampin. Methicillin-resistant strains of all species were most resistant to rifapentine and rifampin than were their methicillin-susceptible counterparts. For most strains tested, the MBCs of both rifamycins exceeded by twofold the respective MICs. Both the checkerboard dilution and time-kill methods were used to determine the interactions of rifapentine or rifampin with six different antibiotics: cefamandole, vancomycin, teicoplanin, gentamicin, erythromycin, and fusidic acid. No significant differences between the two rifamycins in the combinations were observed against either methicillin-susceptible or methicillin-resistant strains. Minor differences were noted depending on the second antibiotic tested or the staphylococcal species examined. Antagonism was never observed, and indifference was the prevalent response. Cases of synergism were observed occasionally with the checkerboard method and slightly more often with the time-kill method.

Teicoplanin versus nafcillin and vancomycin in the treatment of experimental endocarditis caused by methicillin-susceptible or -resistant Staphylococcus aureus.

In rabbits with experimentally induced endocarditis, the efficacy of teicoplanin compared favorably both with that of nafcillin for infection by a methicillin-susceptible strain of Staphylococcus aureus and with that of vancomycin for infection by a methicillin-resistant strain of S. aureus. In a 4-day treatment regimen, teicoplanin was as effective as either nafcillin or vancomycin in eliminating organisms from aortic valve vegetations in the respective infection. In a 10-day regimen for methicillin-resistant S. aureus endocarditis, both teicoplanin and vancomycin sterilized the vegetations of some rabbits, but the relapse rate was high for both. These results justify further investigation into the role of teicoplanin for the treatment of serious infections caused by S. aureus.