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-␣, IL1-␤, 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.

Interactions ofHaemophilus parasuisand its LOS with porcine brain microvascular endothelial cells

“…We now demonstrated that LOS significantly impaired the adhesion of H. parasuis to PBMEC, suggesting a role for this bacterial component as an adhesin. Likewise, LPS from other bacterial species, including swine pathogens of the same family, such as Actinobacillus pleuropneumoniae have been shown to be important adhesins [30,32]. Nevertheless, other adhesins may also contribute to bacteria-cell interactions based on our finding that the addition of LOS to a competitive assay did not completely abolish the adhesion of H. parasuis to PBMEC.…”

A Complete Lipopolysaccharide Inner Core Oligosaccharide Is Required for Resistance of Burkholderia cenocepacia to Antimicrobial Peptides and Bacterial Survival In Vivo

“…Additional mutant strains must be constructed in order to test the importance of LPS outer core sugars for antimicrobial peptide resistance. In other organisms, such as Y. enterocolitica and Actinobacillus pleuropneumoniae, the outer core residues contribute to antimicrobial peptide resistance (45,50). SAL1 does not survive in the rat agar bead model of chronic lung infection.…”

Identification of Novel Glycosyltransferases Required for Assembly of the Pasteurella multocida A:1 Lipopolysaccharide and Their Involvement in Virulence

“…Antimicrobial peptides are a critical component of the innate immune system of higher organisms (18). A (21,25). We have shown recently that the terminal PCho residues on the P. multocida LPS contribute to resistance to fowlicidin 1 (14).…”