title: Truncation of the lipopolysaccharide outer core affects susceptibility to antimicrobial peptides and virulence of Actinobacillus pleuropneumoniae serotype 1.
abstract: We reported previously that the core oligosaccharide region of the lipopolysaccharide (LPS) is essential for optimal adhesion of Actinobacillus pleuropneumoniae, an important swine pathogen, to respiratory tract cells. Rough LPS and core LPS mutants of A. pleuropneumoniae serotype 1 were generated by using a mini-Tn10 transposon mutagenesis system. Here we performed a structural analysis of the oligosaccharide region of three core LPS mutants that still produce the same O-antigen by using methylation analyses and mass spectrometry. We also performed a kinetic study of proinflammatory cytokines production such as interleukin (IL)-6, tumor necrosis factor-alpha, IL1-beta, MCP-1, and IL8 by LPS-stimulated porcine alveolar macrophages, which showed that purified LPS of the parent strain, the rough LPS and core LPS mutants, had the same ability to stimulate the production of cytokines. Most interestingly, an in vitro susceptibility test of these LPS mutants to antimicrobial peptides showed that the three core LPS mutants were more susceptible to cationic peptides than both the rough LPS mutant and the wild type parent strain. Furthermore, experimental pig infections with these mutants revealed that the galactose (Gal I) and d,d-heptose (Hep IV) residues present in the outer core of A. pleuropneumoniae serotype 1 LPS are important for adhesion and overall virulence in the natural host, whereas deletion of the terminal GalNAc-Gal II disaccharide had no effect. Our data suggest that an intact core-lipid A region is required for optimal protection of A. pleuropneumoniae against cationic peptides and that deletion of specific residues in the outer LPS core results in the attenuation of the virulence of A. pleuropneumoniae serotype 1.
authors.affiliation: Groupe de Recherche sur les Maladies Infectieuses du Porc and the Département de Pathologie et Microbiologie, Faculté de Médecine Vétérinaire, Université de Montréal, St-Hyacinthe, Quebec J2S 7C6, Canada.
authors.forename: Mahendrasingh
authors.initials: M
authors.lastname: Ramjeet
authors.forename: Vincent
authors.initials: V
authors.lastname: Deslandes
authors.forename: Frank
authors.initials: F
authors.lastname: St Michael
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Cox
authors.forename: Marylène
authors.initials: M
authors.lastname: Kobisch
authors.forename: Marcelo
authors.initials: M
authors.lastname: Gottschalk
authors.forename: Mario
authors.initials: M
authors.lastname: Jacques
mesh_terms: D016977:Actinobacillus pleuropneumoniae / Q000737:chemistry* / Q000145:classification / Q000235:genetics / Q000472:pathogenicity*; D000595:Amino Acid Sequence; D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry / Q000494:pharmacology; D001483:Base Sequence; D002240:Carbohydrate Sequence; D016207:Cytokines / Q000096:biosynthesis / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D066298:In Vitro Techniques; D018836:Inflammation Mediators / Q000378:metabolism; D008070:Lipopolysaccharides / Q000737:chemistry* / Q000633:toxicity*; D016676:Macrophages, Alveolar / Q000187:drug effects / Q000276:immunology; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016296:Mutagenesis; D019081:O Antigens / Q000737:chemistry / Q000235:genetics / Q000633:toxicity; D012703:Serotyping; D034421:Sus scrofa; D014774:Virulence
chemical_list: D023181:Antimicrobial Cationic Peptides; D016207:Cytokines; D018076:DNA, Complementary; D018836:Inflammation Mediators; D008070:Lipopolysaccharides; D019081:O Antigens