Abstract
The penicillin-binding protein (PBP) patterns of six strains of Bilophila wadsworthia were investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and subsequent fluorography of membrane preparations labelled with [3H]benzylpenicillin. The PBP profiles among the strains were similar; generally, seven to nine PBP-reactive bands could be visualized; their molecular weights ranged from 31 to 137 kDa. The relative affinities of the PBPs of four strains of B. wadsworthia for imipenem were examined and correlated with the morphological responses of the cells to imipenem. Morphological changes were examined by light and electron microscopies. Light microscopy revealed that at low concentrations (less than the MIC), imipenem induced the formation of rounded and bulging cells; rarely, elongation without filamentation was observed. In the presence of imipenem at the MIC, spheroplast formation was observed. Scanning and transmission electron microscopies revealed round forms together with larger, multilobate cells in the presence of subinhibitory concentrations of imipenem, suggesting that new growth sites were initiated while cell division was inhibited. Peeling of the outer membrane was also seen. Spheroplasts were very large (up to 30 microns in diameter) and stable in aqueous solution. Inhibition of the PBPs could be seen in the presence of low imipenem concentrations.
Highlights
The penicillin-binding protein (PBP) patterns of six strains of Biophila wadsworthia were investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and subsequent fluorography of membrane preparations labelled with 13H]benzylpenicillin
The PBP profiles found in the six B. wadsworthia strains tested were comparable to each other, and the molecular masses of the PBPs were fairly similar to those described in E. coli [21]
Inhibition of PBP 1 leads to spheroplasting and lysis, inhibition of PBP 2 results in spherical forms, and inhibition of PBP 3 causes inhibition of septum formation and initiation of new sites, resulting in filamentation of the cells [6, 20, 25]
Summary
The penicillin-binding protein (PBP) patterns of six strains of Biophila wadsworthia were investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and subsequent fluorography of membrane preparations labelled with 13H]benzylpenicillin. Light microscopy revealed that at low concentrations (less than the MIC), imipenem induced the formation of rounded and bulging cells; rarely, elongation without filamentation was observed. Scanning and transmission electron microscopies revealed round forms together with larger, multilobate cells in the presence of subinhibitory concentrations of imipenem, suggesting that new growth sites were initiated while cell division was inhibited. Inhibition of the PBPs could be seen in the presence of low imipenem concentrations. Our early studies [27] indicated significant resistance to ,B-lactamase-stable ,B-lactams, such as imipenem and cefoxitin, among strains of B. wadsworthia. The action of 3-lactam drugs on certain PBPs governs the morphological changes in bacteria seen in the presence of these antibiotics [6, 7, 18, 20]. The purpose of the study described here was to define the PBP patterns of B. wadsworthia, to describe the morphological changes that occur in the transition of the conventional growth to hazy growth upon exposure to imipenem, and to determine the role of PBPs in these changes
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