Novel Mechanism of Resistance to Oxazolidinones, Macrolides, and Chloramphenicol in Ribosomal Protein L4 of the Pneumococcus

Abstract

Two clinical Streptococcus pneumoniae isolates, identified as resistant to macrolides and chloramphenicol and nonsusceptible to linezolid, were found to contain 6-bp deletions in the gene encoding riboprotein L4. The gene transformed susceptible strain R6 so that it exhibited such resistance, with the transformants also showing a fitness cost. We demonstrate a novel bacterial mechanism of resistance to chloramphenicol and nonsusceptibility to linezolid.Macrolide resistance in Streptococcus pneumoniae is predominantly caused by acquisition of the erm(B) (target modification) (33) or mef(A) (drug efflux) (28) gene or a combination of these mechanisms (16). Mutations in 23S rRNA and riboproteins L4 and L22 have more recently been found to confer macrolide resistance (6,29,30). Nonsusceptibility to linezolid in laboratory-generated resistant Escherichia coli isolates (35) and enterococci (21) as well as in clinical isolates of methicillin-resistant Staphylococcus aureus (17, 32) and enterococci (4, 15) has been found to be conferred by mutations in domain V of 23S rRNA. To date, linezolid-nonsusceptible pneumococcal strains are extremely rare. Chloramphenicol resistance in the pneumococcus occurs by acquisition of the cat gene encoding chloramphenicol acetyltransferase (3,18). Chloramphenicol acetyltransferase acetylates chloramphenicol, resulting in derivatives that are unable to bind the ribosome (22,23). In this study, two clinical pneumococcal isolates with elevated macrolide, linezolid, and chloramphenicol MICs were identified and investigated for their resistance mechanisms.PU1071099 (PROTEKT surveillance study) (7) was isolated in Boston in 2001 from sputum of a 67-year-old, and TN33388 (ABCs program of the Centers for Disease Control) was isolated in Tennessee in 2003 from the blood of a 32-year-old who had been exposed to long-term azithromycin prophylaxis. Unencapsulated laboratory strain S. pneumoniae R6 was used in transformation studies. Pneumococci were routinely cultured at 37°C in 5% CO 2 on Mueller-Hinton agar supplemented with 5% horse blood. MICs were determined according to the CLSI

Genome sequencing of linezolid-resistant Streptococcus pneumoniae mutants reveals novel mechanisms of resistance

“…Instead, positions 1743 and 2503 were mutated and three copies of the domain V 23S rRNA had the A2503G mutation at 16 mg/mL, while the fourth copy was mutated when cells reached a MIC of 32 mg/mL (Supplemental Table 2). Small deletions in the 50S ribosomal protein L4 were observed in clinical strains of S. pneumoniae with decreased susceptibility to linezolid (Wolter et al 2005). It is salient to point out that in mutant 1974M2, whole-genome sequencing revealed that the L4 protein is mutated to G71R, a conserved region of L4.…”

“…In addition to mutations in the 23S rRNA, a 6-bp deletion in the ribosomal protein L4 has been described in two clinical isolates of S. pneumoniae that were resistant to chloramphenicol and not susceptible to linezolid (Wolter et al 2005). Recently, an integrated plasmid 23S rRNA methyltransferase coded by the cfr gene (for chloramphenicol-florfenicol resistance) was shown to confer resistance to linezolid in S. aureus (Toh et al 2007;Mendes et al 2008).…”

“…For example, a recent survey of U.S. isolates indicated that 25% of S. pneumoniae were resistant to at least two antibiotics, and a third of these isolates were resistant to at least four antibiotics (Thornsberry et al 2008), but all studies confirmed that linezolid is still very active against S. pneumoniae (Jones et al 2007). However, it was shown that linezolid-resistant strains (minimal inhibitory concentration [MIC] $ 8 mg/mL) of S. pneumoniae can be generated in vitro (Carsenti-Dellamonica et al 2005), that clinical S. pneumoniae strains not susceptible to linezolid (MIC $ 4 mg/mL) have been reported (Wolter et al 2005), and that one resistant clinical isolate of Streptococcus oralis has been described (Mutnick et al 2003). With the use of linezolid steadily increasing in the Western world, the potential for an increase in resistance to linezolid is likely to be inevitable.…”

Clinical and Microbiological Aspects of Linezolid Resistance Mediated by the cfr Gene Encoding a 23S rRNA Methyltransferase

“…The number of rRNA genes mutated depends on the dose and duration of linezolid exposure and has been shown to influence the level of linezolid resistance (3,15). Also, a linezolid-resistant isolate of S. pneumoniae was found to contain a 6-bp deletion in the gene encoding riboprotein L4 (30).…”

Cometabolic degradation of chloramphenicol via a meta-cleavage pathway in a microbial fuel cell and its microbial community

“…Therefore, the growth of E. coli and S. oneidensis was not markedly affected, which indicated that the antibacterial activity of CAP was largely eliminated by the MFC. CAP was first transformed to the CAPacetyl and AMCl2, which involved changes to its functional groups (Jiang et al, 2016;Wolter et al, 2005). Moreover, the aromatic structure of these two CAP metabolites was destroyed via a meta-cleavage pathway.…”