Seven open reading frames, annotated as potential penicillin-binding-protein-encoding genes (imoO441, lmo0540, Imo1438, lmo1892, Imo2039, Imo2229, and lmo2754), were targeted for insertional mutagenesis in Listeria monocytogenes EGDe. These genes were found to contribute in various degrees to beta-lactam resistance, cell morphology, or the virulence potential of this organism.