Christine Wennersten

Linezolid resistance in a clinical isolate of Staphylococcus aureus.

The new oxazolidinone antimicrobial, linezolid, has been approved for the treatment of infections caused by various gram-positive bacteria, including meticillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE). Although instances of linezolid resistance in VRE have been reported, resistance has not been encountered among clinical isolates of S aureus. We have characterised an MRSA isolate resistant to linezolid that was recovered from a patient treated with this agent for dialysis-associated peritonitis.

Accessory Gene Regulator (agr) Locus in Geographically Diverse Staphylococcus aureus Isolates with Reduced Susceptibility to Vancomycin

ABSTRACT The majority of infections with glycopeptide intermediate-level resistant Staphylococcus aureus (GISA) originate in biomedical devices, suggesting a possible increased ability of these strains to produce biofilm. Loss of function of the accessory gene regulator (agr) of S. aureus has been suggested to confer an enhanced ability to bind to polystyrene. We studied agr in GISA, hetero-GISA, and related glycopeptide-susceptible S. aureus isolates. All GISA strains from diverse geographic origins belong to agr group II. All GISA strains were defective in agr function, as demonstrated by their inability to produce delta-hemolysin. Hetero-GISA isolate A5940 demonstrated a nonsense mutation in agrA that was not present in a pulsed-field gel electrophoresis-indistinguishable vancomycin-susceptible isolate from the same patient. Various other agr point mutations were noted in several clinical GISA and hetero-GISA isolates. A laboratory-generated agr-null strain demonstrated a small but reproducible increase in vancomycin heteroresistance after growth in vitro in subinhibitory concentrations of vancomycin. This was not seen in the isogenic agr group II parent strain in which agr was intact. The in vitro bactericidal activity of vancomycin was attenuated in the agr-null strain compared to the parent strain. These findings imply that compromised agr function is advantageous to clinical isolates of S. aureus toward the development of vancomycin heteroresistance, perhaps through the development of vancomycin tolerance.

Linezolid Resistance in Sequential Staphylococcus aureus Isolates Associated with a T2500A Mutation in the 23S rRNA Gene and Loss of a Single Copy of rRNA

Linezolid is an important therapeutic option for infections caused by resistant gram-positive bacteria. We report the characterization of sequential methicillin-resistant Staphylococcus aureus (MRSA) bloodstream isolates that developed resistance in a patient treated with a prolonged course of linezolid. Analysis of this series of clinical MRSA isolates detected, in the resistant isolates, the presence of a T2500A mutation in the domain V region of the 23S rRNA gene. In addition, the loss of a single copy of the 23S rRNA gene was found in 2 of the resistant isolates. As a result of these 2 factors, the proportion of mutant : wild-type 23S rRNA genes increased in association with an increase in the minimum inhibitory concentration of linezolid. The most recent isolate of this series was recovered 7 months after the patient discontinued linezolid and demonstrated reversion to a susceptible phenotype associated with a loss of the T2500A mutation.

Staphylococcus aureus Accessory Gene Regulator (agr) Group II: Is There a Relationship to the Development of Intermediate-Level Glycopeptide Resistance?

We previously determined that all 6 Staphylococcus aureus strains with confirmed intermediate-level resistance to glycopeptides (glycopeptide intermediate S. aureus [GISA]) from the United States that we tested belonged to accessory gene regulator (agr) group II. In the present study, we found that 56% of surveyed bloodstream methicillin-resistant S. aureus isolates (n = 148) at our hospital were agr group II, whereas only 24% of methicillin-susceptible S. aureus isolates (n = 33) were agr group II (P = .001). Population analysis of genetically engineered agr-null and parent wild-type strains of groups I, II, and IV revealed that, when agr function is lost, the agr group II knockout S. aureus was most likely to develop glycopeptide heteroresistance after growth in 1 microg/mL but not 16 microg/mL vancomycin. This strain was unique in showing decreased autolysis after growth in these conditions. This study suggests that some S. aureus strains have an intrinsic survival advantage under a glycopeptide selective pressure, which is possibly related to reduced autolysis after exposure to subinhibitory concentrations of glycopeptide.

Emergence of a Clinical Daptomycin-Resistant Staphylococcus aureus Isolate during Treatment of Methicillin-Resistant Staphylococcus aureus Bacteremia and Osteomyelitis

ABSTRACT The emergence of a clinically daptomycin-resistant Staphylococcus aureus isolate occurred during treatment of methicillin-resistant S. aureus bacteremia and probable vertebral osteomyelitis. The breakthrough isolate was indistinguishable from pretreatment daptomycin-susceptible isolates by pulsed-field gel electrophoresis. Daptomycin nonsusceptibility was confirmed by MIC and time-kill curve analyses.

Increasing resistance to beta-lactam antibiotics among clinical isolates of Enterococcus faecium: a 22-year review at one institution.

To identify any change in the antibiotic resistance of Enterococcus faecium, we examined the antibiotic susceptibilities of clinical strains (n = 84) isolated at one institution during the 22 years since 1968. A significant increase in resistance to penicillin was observed during the study period: the MICs of penicillin for 50 and 90% of isolates tested were 16 and 64 micrograms/ml, respectively, from 1969 to 1988 (n = 48; geometric mean MIC, 14 micrograms/ml) , whereas they were 256 and 512 micrograms/ml, respectively, from 1989 to 1990 (n = 36; geometric mean MIC, 123 micrograms/ml) (P less than 0.001). A comparable increase in resistance to ampicillin was also noted (P less than 0.001). No strains produced detectable beta-lactamase. In contrast, susceptibilities to vancomycin, teicoplanin, and ciprofloxacin remained stable. High-level resistance to gentamicin was observed in none of 48 isolates from 1969 to 1988, but was present in 22 of 36 strains (61%) from 1989 to 1990 (P less than 0.001) and was significantly associated with resistance (MIC, greater than or equal to 128 micrograms/ml) to penicillin (P less than 0.001). To assess the potential evolution of antibiotic resistance in this species, clinical isolates (n = 24) were compared with strains isolated in 1968 from a human population in the Solomon Islands that was never exposed to antibiotics. Solomon Island isolates were significantly more susceptible than all clinical strains to penicillin, ampicillin, and vancomycin (P less than 0.001 for each), but they exhibited no differences in susceptibility to teicoplanin or ciprofloxacin. The penicillin-binding affinity of penicillin-binding protein 5 (PBP 5) in penicillin-resistant clinical strains (MIC, 512 micrograms/ml) was notably lower than that in strains with more typical susceptibilities, suggesting an alteration in this PBP as a possible mechanism for increased penicillin resistance. Solomon Island strains most susceptible to penicillin demonstrated a prominent PBP 5* and the absence of PBP 5. These changes in the antibiotic resistance of E. faecium emphasize the importance of identifying this species in patients with serious enterococcal infections and the necessity of assessing its susceptibility to both beta-lactams and aminoglycosides if effective therapy is to be identified. Images

Linezolid Resistance in Staphylococcus aureus: Characterization and Stability of Resistant Phenotype

Linezolid is an important therapeutic option for treatment of infections caused by glycopeptide- and beta-lactam-resistant gram-positive organisms. Linezolid resistance is caused by mutations within the domain V region of the 23S ribosomal RNA (rRNA) gene, which is present in multiple copies in most bacteria. Among clinical Staphylococcus aureus isolates, there has been only 1 reported case of linezolid resistance. In the present study, this isolate was further characterized by determination of the number of mutant 23S rRNA copies, assessment of the stability of the resistant phenotype, and comparison of its growth characteristics with those of linezolid-susceptible S. aureus. All 5 copies of the 23S rRNA gene contained a G2576U mutation in the domain V region. After serial passage on antibiotic-free medium, the isolate maintained resistance to high concentrations of linezolid. Compared with 2 linezolid-susceptible S. aureus isolates, the linezolid-resistant S. aureus isolate demonstrated no significant differences in in vitro growth characteristics.

Antibiotic Synergism Against Listeria monocytogenes

The effectiveness of ampicillin, penicillin, streptomycin, and gentamicin against 20 strains of Listeria monocytogenes was studied in vitro. For all strains, the minimal bactericidal concentration (MBC) of both ampicillin and penicillin was much higher than the minimal inhibitory concentration (MIC). The MBC of both streptomycin and gentamicin was close to the MIC, but relatively high concentrations of these antibiotics were necessary to inhibit the growth of most of the strains of Listeria. The combination of penicillin plus streptomycin was synergistic against 19 of 20 strains and in the remaining strain produced enhanced killing (but of less magnitude than our criterion for synergism). Combinations of penicillin plus gentamicin, ampicillin plus streptomycin, and ampicillin plus gentamicin produced enhanced killing against all strains tested. No antagonism was observed when ampicillin or penicillin was combined with streptomycin or gentamicin.

In vitro activity of RP59500, an injectable streptogramin antibiotic, against vancomycin-resistant gram-positive organisms.

The in vitro activity of RP59500, a streptogramin antibiotic, against 146 clinical isolates of vancomycin-resistant gram-positive bacteria was examined. Five strains of the species Enterococcus casseliflavus and Enterococcus gallinarum, for which the MIC of vancomycin was 8 micrograms/ml, were also studied. Twenty-eight vancomycin-susceptible strains of Enterococcus faecalis and Enterococcus faecium were included for comparison. The drug was highly active against Leuconostoc spp., Lactobacillus spp., and Pediococcus spp. (MICs, < or = 2 micrograms/ml). RP59500 was more active against vancomycin-susceptible strains of E. faecium than E. faecalis (MICs for 90% of the strains [MIC90s], 1.0 versus 32 micrograms/ml). Vancomycin-resistant strains of E. faecalis were as resistant to RP59500 as vancomycin-susceptible strains (MIC90, 32 micrograms/ml), but some vancomycin-resistant E. faecium strains were relatively more resistant to the new agent (MIC90, 16; MIC range, 0.5 to 32 micrograms/ml) than were vancomycin-susceptible organisms of this species.

In Vitro Activities of the Glycylcycline GAR-936 against Gram-Positive Bacteria

ABSTRACT The in vitro activities of GAR-936, the 9-t-butylglycylamido derivative of minocycline, were compared with those of doxycycline, minocycline, and tetracycline against 527 gram-positive clinical isolates. GAR-936 inhibited all strains, including those resistant to other tetracyclines, at concentrations of ≤2 μg/ml, except two strains of JK diphtheroids for which the MIC was 4 μg/ml.