We have studied the mechanism by which b-lactam challenge leads to b-lactamase induction in Aeromonas hydrophila through transposon-insertion mutagenesis. Disruption of the DDcarboxypeptidases/endopeptidases, penicillin-binding protein 4 or BlrY leads to elevated monomer-disaccharide-pentapeptide levels in A. hydrophila peptidoglycan and concomitant overproduction of b-lactamase through activation of the BlrAB two-component regulatory system. During b-lactam challenge, monomer-disaccharide-pentapeptide levels increase proportionately with b-lactamase production and b-lactamase induction is inhibited by vancomycin, which binds muro-pentapeptides. Taken together, these data strongly suggest that the Aeromonas spp. blactamase regulatory sensor kinase, BlrB, responds to the concentration of monomerdisaccharide-pentapeptide in peptidoglycan.