Journal of Antimicrobial Chemotherapy (2000) 45, 797-802
© 2000 The British Society for Antimicrobial Chemotherapy
Activity of linezolid against Gram-positive cocci possessing genes conferring resistance to protein synthesis inhibitors
Laboratoire de Microbiologie, CHU de la côte de Nacre, Service de Microbiologie, Avenue de la côte de Nacre, 14033 Caen Cedex, France
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Abstract |
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 Top Abstract IntroductionMaterials and methodsResults and discussionReferences |
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Linezolid belongs to a new class of antimicrobials, the oxazolidinones, that act by inhibiting protein synthesis. To detect cross-resistance with other inhibitors of protein synthesis (chloramphenicol, macrolides, lincosamides, streptogramins, aminoglycosides and tetracyclines), the in vitro activity of linezolid was determined against isolates harbouring known genes conferring resistance to these antimicrobials. Neither the presence of modifying enzymes (LinA, LinA', LinB, Vgb, Vat, SatA, ANT(4') (4'')-I, AAC(6')-APH(2''), APHA-3 and Cat), nor the presence of an efflux mechanism (MsrA, MefE, MefA, MreA, Vga, TetK and TeL), nor the modification or protection of antimicrobial target (because of ribosomal methylases or TetM and TetO) affected linezolid activity as demonstrated by similar in vitro activity against resistant isolates and sensitive control isolates.
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Introduction |
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TopAbstractIntroductionMaterials and methodsResults and discussionReferences |
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Recent years have been marked by the increasing isolation of multidrug-resistant Gram-positive bacteria in clinical settings. As a result, new antimicrobials that possess unique mechanisms of action are urgently needed to treat with good efficacy infections caused by resistant pathogens.
Linezolid belongs to a new class of antimicrobial agents, the oxazolidinones, which are active against a variety of Gram-positive pathogenic bacteria, including methicillin-resistant isolates of Staphylococcus aureus and Staphylococcus epidermidis, vancomycin-resistant isolates of Enterococcus spp. and penicillin-resistant pneumococci.1–3 Early studies have demonstrated that this new agent exerts bacteriostatic effect by inhibiting protein synthesis.4 Linezolid binds to the 50S ribosomal subunit in a similar way to chloramphenicol, macrolides, lincosamides and streptogramins. The linezolid binding site is close to that of chloramphenicol and lincomycin, and linezolid competes with these two antimicrobials.5 However, in contrast to the latter two classes, oxazolidinones clearly have a distinct mechanism of action, as they do not have any effect on the peptidyl transferase activity but inhibit the formation of the initiation complex in bacterial translation systems.6
Many studies have shown that linezolid is active against multidrug-resistant organisms. However, mechanisms of resistance to protein synthesis inhibitors, such as aminoglycosides, chloramphenicol, macrolides and related antimicrobials or tetracyclines, have not been characterized at the genetic level in the isolates studied. Therefore, possible cross-resistance between linezolid and other protein synthesis inhibitors has not been systematically studied.
Three types of resistance mechanisms to antimicrobials that inhibit protein synthesis have been identified in clinical isolates of Gram-positive bacteria. For each type, several resistance genes have been described. The major ones are listed in Table I
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Modification of the ribosomal target remains the main mechanism of resistance against macrolides and related antimicrobials. N-6 dimethylation of a specific residue in 23S rRNA is conveyed by a family of methyltransferase enzymes encoded by the erm genes and yields crossresistance between macrolide, lincosamide and streptogramin B antimicrobials (MLSB phenotype).7,25 The tetM and tetO gene products have been found to associate with ribosomes, thus preventing tetracyclines from reaching their target.26,27
A second type of resistance is the drug inactivation conveyed by various enzymes including acetylases (aminoglycosides, chloramphenicol, streptogramins A), hydrolases (streptogramins B), nucleotidyltransferases (aminoglycosides, lincosamides) and phosphotransferases (aminoglycosides).
The third mechanism affects the rate of transport of streptogramins A, tetracyclines or macrolides across the cell membrane by active efflux.
In this study, we have evaluated the activity of linezolid against various Gram-positive bacteria exhibiting the major mechanisms of resistance against inhibitors of protein synthesis characterized in clinical isolates.
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Materials and methods |
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TopAbstractIntroductionMaterials and methodsResults and discussionReferences |
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Bacterial isolates
For a majority of resistance gene classes, isogenic pairs of bacteria, differing only by the presence or absence of a known resistance determinant (transconjugants, transformants, transductants or cured derivatives) were used. When pairs were not available, a control isolate belonging to the same species but devoid of the resistance gene was used. Mutants constitutively resistant to macrolides were obtained from inducibly resistant isolates after selection on agar plates containing inhibitory concentrations of clindamycin, a non-inducer macrolide. Before testing the activity of linezolid, the presence of resistance genes in the isolate studied was checked by a specific PCR reaction.
Antimicrobial susceptibility
MICs of linezolid were determined by the agar dilution method according to NCCLS standards with Mueller– Hinton medium (bioMérieux, La Balme-les-Grottes, France).28 An inoculum of 104 cfu/spot was used. For streptococci, Mueller–Hinton medium was supplemented with 5% horse blood. The plates were incubated aerobically for 24 h at 37°C.
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Results and discussion |
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TopAbstractIntroductionMaterials and methodsResults and discussionReferences |
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As shown in Table II
, similar MICs of linezolid have been determined against isolates containing resistance genes affecting activity of protein synthesis inhibitors and those against control isolates. Thus, linezolid activity was not affected by the presence of modifying enzymes (LinA, LinA', LinB, Vgb, Vat, SatA, ANT(4') (4'')-I, AAC(6')-APH(2''), APHA-3 and Cat), or by the presence of active efflux systems (MsrA, MefE, MefA, MreA, Vga, TetK and TeL), or by the modification or protection of antimicrobial target (because of ribosomal methylases or TetM and TetO). Lack of activity of modifying enzymes studied against linezolid was not surprising. As linezolid belongs to a new antimicrobial family structurally unrelated to any other known family, drug inactivation by enzymes, activity of which is based on structure identification, was not expected to occur. By contrast, unrelated drugs can be substrates for efflux pumps that are responsible for multidrug resistance. In our study, none of the efflux pumps tested affect linezolid activity. Similarly, mechanisms that lead to modification of target site can confer cross-resistance to structurally unrelated antimicrobials if their binding sites are overlapping.7,25 This was not observed with MLSB resistance as a result of ribosomal methylation encoded by erm genes, although lincomycin and linezolid appeared to compete for their binding site. This is consistent with a different mechanism of action of the two molecules.6 Tetracyclines have a high affinity binding site at the central loop of domain V of 23S rRNA although binding to the 30S subunit and subsequent inhibition of the A-site are probably the major mode of action of this drug.39
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In conclusion, linezolid is a new antimicrobial agent that, by virtue of its novel structure and unique mechanism of action, is not cross-resistant with other antimicrobial classes, including inhibitors of protein synthesis that bind to the 50S ribosomal subunit.
Recent in vitro studies have shown that resistance to linezolid can appear by point mutation in the 23S rRNA domain V in S. aureus and Enterococcus faecalis probably at the binding site of this antimicrobial.40 Moreover, the same mutation was found in two clinical Enterococcus faecium isolates resistant to linezolid, which have emerged during linezolid therapy within the Linezolid Compassionate Use Program.41
Despite a possible but still very rare resistance by target mutation, this molecule could represent an advance in antimicrobial therapy, particularly for the problematic resistant organisms that are becoming widespread.
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Acknowledgments |
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We thank Patrice Courvalin, Ann Eady and Joyce Sutcliffe for the gift of isolates. This work was supported by a grant from Pharmacia–Upjohn (Kalamazoo, MI, USA).
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Notes |
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* Corresponding author. Tel: +33-2-31-06-45-72; Fax: +33-2-31-06-45-73; E-mail: leclercq-r{at}chu-caen.fr
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Received 9 August 1999; returned 7 December 1999; revised 7 January 2000; accepted 28 January 2000
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