Effects of inactivation of d,d-transpeptidases of Acinetobacter baumannii on bacterial growth and susceptibility to β-lactam antibiotics.

Resistance to β-lactams, the most used antibiotics worldwide, constitutes the major problem for treatment of bacterial infections. In the nosocomial pathogen Acinetobacter baumannii, β-lactamase-mediated resistance to the family of β-lactam antibiotics, carbapenems, has resulted in the selection and dissemination of multidrug-resistant isolates, which often cause infections characterized by high mortality rates. There is thus an urgent demand for new β-lactamase-resistant antibiotics that also inhibit their targets, penicillin-binding proteins (PBPs). As some PBPs are indispensable for biosynthesis of the bacterial cell wall and survival, we evaluated their importance for growth of A. baumannii by performing gene inactivation studies of d,d-transpeptidase domains of high-molecular mass (HMM) PBPs individually and in combination with one another. We showed that PBP3 is essential for A. baumannii survival, as deletion mutants of this d,d-transpeptidase were not viable. Inactivation of PBP1a resulted in partial cell lysis and retardation of bacterial growth, and these effects were further enhanced by additional inactivation of PBP2 but not PBP1b. Susceptibility to β-lactam antibiotics increased 4-8-fold for the A. baumannii PBP1a/PBP1b/PBP2 triple mutant and 2-4-fold for all remaining mutants. Analysis of peptidoglycan structure revealed a significant change in the muropeptide composition of the triple mutant and demonstrated that lack of d,d-transpeptidase activity of PBP1a, PBP1b, and PBP2 is compensated by an increase in l,d-transpeptidase-mediated crosslinking activity of LdtJ. Overall, our data showed that in addition to essential PBP3, simultaneous inhibition of PBP1a and PBP2 or PBPs in combination with LdtJ could represent potential strategies for design of novel drugs against A. baumannii.