title: Characterization of a Carbapenem-Hydrolyzing Enzyme, PoxB, in Pseudomonas aeruginosa PAO1.
abstract: Pseudomonas aeruginosa is an opportunistic pathogen often associated with severe and life-threatening infections that are highly impervious to treatment. This microbe readily exhibits intrinsic and acquired resistance to varied antimicrobial drugs. Resistance to penicillin-like compounds is commonplace and provided by the chromosomal AmpC β-lactamase. A second, chromosomally encoded β-lactamase, PoxB, has previously been reported in P. aeruginosa. In the present work, the contribution of this class D enzyme was investigated using a series of clean in-frame ampC, poxB, and oprD deletions, as well as complementation by expression under the control of an inducible promoter. While poxB deletions failed to alter β-lactam sensitivities, expression of poxB in ampC-deficient backgrounds decreased susceptibility to both meropenem and doripenem but had no effect on imipenem, penicillin, and cephalosporin MICs. However, when expressed in an ampCpoxB-deficient background, that additionally lacked the outer membrane porin-encoding gene oprD, PoxB significantly increased the imipenem as well as the meropenem and doripenem MICs. Like other class D carbapenem-hydrolyzing β-lactamases, PoxB was only poorly inhibited by class A enzyme inhibitors, but a novel non-β-lactam compound, avibactam, was a slightly better inhibitor of PoxB activity. In vitro susceptibility testing with a clinical concentration of avibactam, however, failed to reduce PoxB activity against the carbapenems. In addition, poxB was found to be cotranscribed with an upstream open reading frame, poxA, which itself was shown to encode a 32-kDa protein of yet unknown function.
authors.affiliation: Department of Biological Sciences, College of Arts and Sciences, Florida International University, Miami, Florida, USA.
authors.forename: Diansy
authors.initials: D
authors.lastname: Zincke
authors.affiliation: Department of Biological Sciences, College of Arts and Sciences, Florida International University, Miami, Florida, USA.
authors.forename: Deepak
authors.initials: D
authors.lastname: Balasubramanian
authors.affiliation: LL Silver Consulting, Springfield, New Jersey, USA.
authors.forename: Lynn L
authors.initials: LL
authors.lastname: Silver
authors.affiliation: Department of Human and Molecular Genetics, Herbert Wertheim College of Medicine, Florida International University, Miami, Florida, USA matheelabfor65roses@gmail.com.
authors.forename: Kalai
authors.initials: K
authors.lastname: Mathee
mesh_terms: D053961:Azabicyclo Compounds / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D015780:Carbapenems / Q000494:pharmacology*; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D015378:Imipenem / Q000494:pharmacology; D000077731:Meropenem; D008826:Microbial Sensitivity Tests; D009876:Operon; D018272:Porins / Q000235:genetics / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D013845:Thienamycins / Q000494:pharmacology; D018440:beta-Lactam Resistance / Q000187:drug effects / Q000235:genetics*; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism*
chemical_list: D053961:Azabicyclo Compounds; D001426:Bacterial Proteins; D015780:Carbapenems; D018272:Porins; D013845:Thienamycins; C077157:OprD protein, Pseudomonas aeruginosa; D015378:Imipenem; C543519:avibactam; C048795:AmpC beta-lactamases; D001618:beta-Lactamases; C000611877:beta-lactamase PoxB, Pseudomonas aeruginosa; D000077731:Meropenem