title: Low-level resistance of Staphylococcus aureus to thrombin-induced platelet microbicidal protein 1 in vitro associated with qacA gene carriage is independent of multidrug efflux pump activity.
abstract: Thrombin-induced platelet microbial protein 1 (tPMP-1), a cationic antimicrobial polypeptide released from thrombin-stimulated rabbit platelets, targets the Staphylococcus aureus cytoplasmic membrane to initiate its microbicidal effects. In vitro resistance to tPMP-1 correlates with survival advantages in vivo. In S. aureus, the plasmid-carried qacA gene encodes a multidrug transporter, conferring resistance to organic cations (e.g., ethidium [Et]) via proton motive force (PMF)-energized export. We previously showed that qacA also confers a tPMP-1-resistant (tPMP-1r) phenotype in vitro. The current study evaluated whether (i) transporters encoded by the qacB and qacC multidrug resistance genes also confer tPMP-1r and (ii) tPMP-1r mediated by qacA is dependent on efflux pump activity. In contrast to tPMP-1r qacA-bearing strains, the parental strain and its isogenic qacB- and qacC-containing strains were tPMP-1 susceptible (tPMP-1s). Efflux pump inhibition by cyanide m-chlorophenylhydrazone abrogated Etr, but not tPMP-1r, in the qacA-bearing strain. In synergy assays, exposure of the qacA-bearing strain to tPMP-1 did not affect the susceptibility of Et (ruling out Et-tPMP-1 cotransport). The following cytoplasmic membrane parameters did not differ significantly between the qacA-bearing and parental strains: contents of the major phospholipids; asymmetric distributions of the positively charged species, lysyl-phosphotidylglycerol; fatty acid composition; and relative surface charge. Of note, the qacA-bearing strain exhibited greater membrane fluidity than that of the parental, qacB-, or qacC-bearing strain. In conclusion, among these families of efflux pumps, only the multidrug transporter encoded by qacA conferred a tPMP-1r phenotype. These data suggest that qacA-encoded tPMP-1r results from the impact of a specific transporter upon membrane structure or function unrelated to PMF-dependent peptide efflux.
authors.affiliation: Division of Infectious Diseases, Habor-UCLA Medical Center, Torrance, CA, USA. bayer@humc.edu
authors.forename: A S
authors.initials: AS
authors.lastname: Bayer
authors.forename: L I
authors.initials: LI
authors.lastname: Kupferwasser
authors.forename: M H
authors.initials: MH
authors.lastname: Brown
authors.forename: R A
authors.initials: RA
authors.lastname: Skurray
authors.forename: S
authors.initials: S
authors.lastname: Grkovic
authors.forename: T
authors.initials: T
authors.lastname: Jones
authors.forename: K
authors.initials: K
authors.lastname: Mukhopadhay
authors.forename: M R
authors.initials: MR
authors.lastname: Yeaman
mesh_terms: D000818:Animals; D017920:Antiporters / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001798:Blood Proteins / Q000494:pharmacology*; D024881:Drug Resistance, Bacterial; D029968:Escherichia coli Proteins; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D011817:Rabbits; D013211:Staphylococcus aureus / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development; D013917:Thrombin / Q000378:metabolism*
chemical_list: D017920:Antiporters; D001426:Bacterial Proteins; D001798:Blood Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; C099272:QacB protein, Staphylococcus aureus; C077150:platelet microbicidal protein, Oryctolagus cuniculus; C068498:qacA protein, Staphylococcus aureus; C075436:EmrE protein, E coli; D013917:Thrombin