In vitro selection of mutants of Streptococcus pneumoniae resistant to macrolides and linezolid: relationship with susceptibility to penicillin G or macrolides

299

195

Linezolid is an oxazolidinone antibiotic in clinical use for the treatment of serious infections of resistant Gram-positive bacteria. It inhibits protein synthesis by binding to the peptidyl transferase center on the ribosome. Almost all known resistance mechanisms involve small alterations to the linezolid binding site, so this review will therefore focus on the various changes that can adversely affect drug binding and confer resistance. High-resolution structures of linezolid bound to the 50S ribosomal subunit show that it binds in a deep cleft that is surrounded by 23S rRNA nucleotides. Mutation of 23S rRNA has for some time been established as a linezolid resistance mechanism. Although ribosomal proteins L3 and L4 are located further away from the bound drug, mutations in specific regions of these proteins are increasingly being associated with linezolid resistance. However, very little evidence has been presented to confirm this. Furthermore, recent findings on the Cfr methyltransferase underscore the modification of 23S rRNA as a highly effective and transferable form of linezolid resistance. On a positive note, detailed knowledge of the linezolid binding site has facilitated the design of a new generation of oxazolidinones that show improved properties against the known resistance mechanisms. LINEZOLID

Section: Resistance Caused By 23s Rrna Mutationsmentioning

confidence: 99%

Resistance to Linezolid Caused by Modifications at Its Binding Site on the Ribosome

Long

Vester

2012

299

195

“…This study and a growing number of other reports (7,17,32) show that oxazolidinone resistance mechanisms are not limited to 23S rRNA mutations. L3 mutations, in addition to some recently described oxazolidinone resistance determinants, including inactivation of an endogenous ribosomal methyltransferase and enhanced drug efflux (7), may help to explain some of the numerous reports of LZD r strains with unknown resistance mechanisms (3,9,10,15,25).…”

Section: Fig 1 Structural Analysis Of Ribosomal Mutations In Clinicmentioning

confidence: 99%

Mutations in Ribosomal Protein L3 Are Associated with Oxazolidinone Resistance in Staphylococci of Clinical Origin

Locke

Hilgers

Shaw

2009

115

Following recent reports of ribosomal protein L3 mutations in laboratory-derived linezolid-resistant (LZD r ) Staphylococcus aureus, we investigated whether similar mutations were present in LZD r staphylococci of clinical origin. Sequence analysis of a variety of LZD r isolates revealed two L3 mutations, ⌬Ser145 (S. aureus NRS127) and Ala157Arg (Staphylococcus epidermidis 1653059), both occurring proximal to the oxazolidinone binding site in the peptidyl transferase center. The oxazolidinone torezolid maintained a >8-fold potency advantage over linezolid for both strains.

“…tions in 23S rRNA and the ribosomal proteins L3 (28,29) and L4 (9, 51) have been associated with linezolid resistance but without substantial documentation of the cause-effect relationship. The 23S rRNA mutations reported sometimes correspond to those found in laboratory selection experiments, but in others cases there is no genetic evidence relating the specific 23S rRNA mutations to the linezolid resistance phenotype (3,9,27,41).…”

mentioning

confidence: 90%

Mutations in 23S rRNA at the Peptidyl Transferase Center and Their Relationship to Linezolid Binding and Cross-Resistance

Long

Munck

Andersen

et al. 2010

The oxazolidinone antibiotic linezolid targets the peptidyl transferase center (PTC) on the bacterial ribosome. Thirteen single and four double 23S rRNA mutations were introduced into a Mycobacterium smegmatis strain with a single rRNA operon. Converting bacterial base identity by single mutations at positions 2032, 2453, and 2499 to human cytosolic base identity did not confer significantly reduced susceptibility to linezolid. The largest decrease in linezolid susceptibility for any of the introduced single mutations was observed with the G2576U mutation at a position that is 7.9 Å from linezolid. Smaller decreases were observed with the A2503G, U2504G, and G2505A mutations at nucleotides proximal to linezolid, showing that the degree of resistance conferred is not simply inversely proportional to the nucleotide-drug distance. The double mutations G2032A-C2499A, G2032A-U2504G, C2055A-U2504G, and C2055A-A2572U had remarkable synergistic effects on linezolid resistance relative to the effects of the corresponding single mutations. This study emphasizes that effects of rRNA mutations at the PTC are organism dependent. Moreover, the data show a nonpredictable crossresistance pattern between linezolid, chloramphenicol, clindamycin, and valnemulin. The data underscore the significance of mutations at distal nucleotides, either alone or in combination with other mutated nucleotides, in contributing to linezolid resistance.The bacterial ribosome is an important target for many clinically relevant antibiotics, including linezolid, which is effective in the treatment of infections with various multidrug-resistant Gram-positive cocci, especially methicillin-resistant Staphylococcus aureus and vancomycin-resistant enterococci (26). Several lines of evidence have established that oxazolidinones bind to the peptidyl transferase center (PTC) on the large ribosomal subunit. The evidence includes competitive binding studies with chloramphenicol and lincomycin (25), the clustering of single rRNA resistance mutations in the peptidyl transferase loop (22), in vivo cross-linking experiments (4, 23), and crystal structures of linezolid-50S subunit complexes (archaeal 50S complex [20] and bacterial 50S complex [49]). These structures show that linezolid binds to the A-site portion of the PTC (Fig. 1A) overlapping the site of the aminoacyl moiety of A-sitebound tRNA (20,49). The linezolid binding pocket is lined by eight 23S rRNA nucleotides (marked by wedges in Fig. 1B) that are highly conserved among the three domains of life (49), and the drug is not in direct contact with any ribosomal proteins. Despite this high conservation, indirect evidence suggests that linezolid affects bacterial and mitochondrial ribosomes but not cytosolic ribosomes (23,34). It has been proposed that the specificity may be determined by nucleotides a bit further away from the binding site acting through the nearby conserved nucleotides (23).The linezolid resistance mechanisms characterized thus far involve alteration at or around the PTC by modification...