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- Title
Substitutions in PBP3 confer resistance to both ampicillin and extended-spectrum cephalosporins in Haemophilus parainfluenzae as revealed by site-directed mutagenesis and gene recombinants.
- Authors
Wienholtz, Nanna H.; Barut, Aynur; Nørskov-Lauritsen, Niels
- Abstract
<bold>Objectives: </bold>To determine the association of amino acid substitutions in PBP3 with β-lactam susceptibility in Haemophilus parainfluenzae.<bold>Methods: </bold>Single and multiple amino acid mutations at positions 385, 511 and 526 were introduced into PBP3 of a β-lactam-susceptible H. parainfluenzae strain using site-directed mutagenesis. Recombinants were also generated using PCR-amplified ftsI from clinical strains encoding multiple amino acid substitutions. MICs of ampicillin, cefuroxime, cefotaxime and ceftriaxone were determined using Etest®.<bold>Results: </bold>Transformation of a susceptible strain with ftsI from clinical strains encoding four substitutions in the transpeptidase region of PBP3 conferred resistance to ampicillin, but not to cephalosporins. Introduction of ftsI from a clinical strain encoding eight substitutions conferred resistance to ampicillin, cefotaxime and ceftriaxone. MICs for recombinants were lower than those for the donor strains. Using site-directed mutagenesis, no single substitution conferred resistance to the tested β-lactams, although V511A increased the MIC of cefuroxime to the intermediate category for intravenous administration. Recombinants encoding N526K/H/S in combination with V511A were resistant to ampicillin. Substitution S385T increased the MICs of third-generation cephalosporins if V511A was also present.<bold>Conclusions: </bold>Substitutions in PBP3 are sufficient to confer resistance to both ampicillin and third-generation cephalosporins in H. parainfluenzae. A combination of substitutions at positions Val-511 and Asn-526 confers resistance to ampicillin. Resistance to third-generation cephalosporins probably requires more than four substitutions in PBP3.
- Subjects
HAEMOPHILUS parainfluenzae; AMPICILLIN; PENICILLIN-binding proteins; CEPHALOSPORINS; SITE-specific mutagenesis; GENETIC recombination; BETA lactam antibiotics; DRUG resistance in bacteria; THERAPEUTICS; BACTERIA; AMINO acids; ANTIBIOTICS; DRUG resistance in microorganisms; GENETICS; HAEMOPHILUS; HAEMOPHILUS diseases; MICROBIAL sensitivity tests; PHARMACODYNAMICS
- Publication
Journal of Antimicrobial Chemotherapy (JAC), 2017, Vol 72, Issue 9, p2544
- ISSN
0305-7453
- Publication type
journal article
- DOI
10.1093/jac/dkx157