Occurrence of a beta-lactam-inducible penicillin-binding protein in methicillin-resistant staphylococci

Abstract

The mechanism of methicillin resistance was investigated in methicillin-resistant staphylococci (MRS) and in variants which had lost methicillin resistance. Phase-contrast microscopy showed that cells swelled at low concentrations of 13-lactam antibiotics in both MRS and variants which had lost methicillin resistance. Cells of variants which had lost methicillin resistance were lysed easily when higher concentrations of antibiotic were used. In contrast, MRS cells remained swollen at even higher concentrations of antibiotics. Furthermore, bacterial growth was inhibited at antibiotic concentrations much lower than MICs for MRS. Examination of the penicillin-binding proteins (PBPs) in MRS revealed that a new PBP-2' (molecular weight, 74,000) was induced in large quantity by exposure to P-lactams. PBP-2' was produced constitutively in variants of MRS which had lost a penicillinase plasmid. The induction of PBP-2' by (-lactams was not detected in variants which had lost methicillin resistance. High concentrations of ,l-lactam were required for saturation of PBP-2'. The optimum antibiotic concentration for the induction of PBP-2' varied with the 3-lactam used as the inducer, and PBP-2' was produced in a larger amount at 32°C than at 370C. From these results, we suggest that the mechanism of methicillin resistance depends on the induction of PBP-2', which may function as a detour enzyme for PBP-2 or PBP-3 or may be a particular enzyme involved in peptidoglycan synthesis.Methicillin-resistant strains of Staphylococcus aureus were first reported in 1961 (19), soon after the introduction of methicillin into clinical use. Subsequently, it has been shown that these strains also are resistant to many penicillins and cephems other than methicillin (2,11,15). In addition, methicillin resistance (MR) has been found to be temperature dependent (1, 7, 12) and affected by pH (27), NaCl concentration (3,12), and inoculum size (12, 23). Although methicillip-resistant staphylococci (MRS) produce penicillinase (PCase), curing of the PCase plasmid does not reduce the level of MR (28).Four main penicillin binding proteins (PBPs) have been identified in S. aureus (20), and either one or both PBP-2 and PBP-3 have been suggested to be the lethal target(s) for P-lactam action (13). In MRS, production of altered PBPs with extremely low affinity for ,3-lactams (14,16,17) or an increase of an altered PBP-3 (6) has been described. However, the double-zone phenomenon occasionally detected on a disk diffusion susceptibility test of P-lactams, such as methicillin, nafcillin (24), or imipenem (4), for MRS cannot be explained by the decreased affinities of PBPs for ,-lactams. This phenomenon suggests the existence of another mechanism for MR.Although MRS had not been a major clinical problem in Japan, after the introduction of third-generation cephems in 1982 isolates of MRS increased. We isolated two variant types from our clinical isolates of MRS. One type retained MR, whereas PCase activity became negative, and the other type lost MR along with ...

Detection ofmecA, femA, andfemBgenes in clinical strains of staphylococci using polymerase chain reaction

“…Such strains, namely mecA-positive but probably non-PBP-2'-producing strains, have been detected previously and referred to as 'cryptically' methicillin-resistant strains [14]. The clinical problem posed by such strains is that during chemotherapy with beta-lactam antibiotics, production of PBP-2' may be induced, converting them into oxacillin-resistant strains as demonstrated in vitro [20]. For this reason, detection of mecA gene is also indispensable for precise differentiation of MRSA and the PCR method employed in this study will be a useful technique in clinical laboratories.…”

Methicillin-resistant staphylococci

“…1), termed PBP2a or PBP2' (assumed to be identical for the purposes of this review), that has a low binding affinity for beta-lactam antibiotics (22,67,80,82,160). Methicillin-resistant strains of coagulase-negative staphylococci also produce PBP2a (31,159).…”

Molecular Epidemiology of Methicillin-Resistant Staphylococcus aureus in Finland

“…The most clinically relevant and the most prevalent form of methicillin resistance is characterized by production of an additional penicillin binding protein, PBP2a (or PBP2') (26,228,275). PBP2a has probably evolved by recombination of a penicillinase gene and a PBP gene similar to PBP2 and PBP3 of Escherichia coli (255).…”