title: Membrane-active mechanism of LFchimera against Burkholderia pseudomallei and Burkholderia thailandensis.
abstract: LFchimera, a construct combining two antimicrobial domains of bovine lactoferrin, lactoferrampin265-284 and lactoferricin17-30, possesses strong bactericidal activity. As yet, no experimental evidence was presented to evaluate the mechanisms of LFchimera against Burkholderia isolates. In this study we analyzed the killing activity of LFchimera on the category B pathogen Burkholderia pseudomallei in comparison to the lesser virulent Burkholderia thailandensis often used as a model for the highly virulent B. pseudomallei. Killing kinetics showed that B. thailandensis E264 was more susceptible for LFchimera than B. pseudomallei 1026b. Interestingly the bactericidal activity of LFchimera appeared highly pH dependent; B. thailandensis killing was completely abolished at and below pH 6.4. FITC-labeled LFchimera caused a rapid accumulation within 15 min in the cytoplasm of both bacterial species. Moreover, freeze-fracture electron microscopy demonstrated extreme effects on the membrane morphology of both bacterial species within 1 h of incubation, accompanied by altered membrane permeability monitored as leakage of nucleotides. These data indicate that the mechanism of action of LFchimera is similar for both species and encompasses disruption of the plasma membrane and subsequently leakage of intracellular nucleotides leading to cell dead.
authors: Department of Microbiology, Faculty of Medicine, Khon Kaen University, Khon Kaen, 40002, Thailand.
authors: Sakawrat
authors: S
authors: Kanthawong
authors: Aekkalak
authors: A
authors: Puknun
authors: Jan G M
authors: JG
authors: Bolscher
authors: Kamran
authors: K
authors: Nazmi
authors: Jan
authors: J
authors: van Marle
authors: Johannes J
authors: JJ
authors: de Soet
authors: Enno C I
authors: EC
authors: Veerman
authors: Surasakdi
authors: S
authors: Wongratanacheewin
authors: Suwimol
authors: S
authors: Taweechaisupapong
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D019117:Burkholderia / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D016957:Burkholderia pseudomallei / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects; D005614:Freeze Fracturing; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D007781:Lactoferrin / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D013045:Species Specificity
chemical_list: D023181:Antimicrobial Cationic Peptides; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; C502040:lactoferrampin; C557490:lactoferricin (17-30); D007781:Lactoferrin