A resistance mechanism to Cephem antibiotics in Staphylococcus aureus

Abstract

The mechanism of the resistance to cephem-antibiotics was studied with two clinical isolates of Staphylococcus aureus, JS1 and JS2. Both strains were confirmed to produce β-lactamase possessing penicillinase-activity but no apparent cephalosporinase-activity. Non-β-lactamase-producing subclones, JS11 and JS21, obtained by curing the β-lactamase-encoding plasmid through cultivating JS1 and JS2 at 44C overnight, were as resistant to cephemantibiotics as the parent strains. This suggests that the resistance to cephem-antibiotics is not related to penicillinase.
Fluorography of penicillin-binding proteins (PBPs) of JS11 and JS21 indicated that the intrinsically cephem-resistant strains possessed a specific 78 kilodalton (Kd) protein (PBP2') besides PBP1 (81Kd), PBP2 (77Kd), PBP3 (74Kd), and PBP4 (44Kd) proteins. The latter four PBPs were shown also in the respective sensitive strains, JS111 and JS211, obtained by 10 times successive subculturing of JS11 and JS21 in drug-free medium. The 78Kd protein hardly bound to ¹⁴C-penicillin G at a concentration of 0.01mM, Whereas the other PBPs, PBP1, PBP2, PBP3, and PBP4, in either the resistant parents or sensitive strains bound abundantly to it. A competitive binding study using non-radioactive cefoxitin and PCG revealed that less than 30% of the 78Kd protein (PBP2') was saturated with cefoxitin at a concentration of 25μg/ml, whereas 84% each of the other PBPs was saturated.
Almost the same amounts of ¹⁴C-penicillin G bound to each of PBPs of the intrinsically resistant staphylococci with either the membrane fraction or whole live cells, indicating that no penetration barrier exists in the intrinsically cephem-resistant S. aureus.
It was concluded that the two cephemresistant clinical isolates S. aureus possess a specific 78Kd PBP that catalyzes crosslinking of murein even in the presence of cephem-antibiotics because of its low affinity to the drugs.