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Structural characterization of Haemophilus parainfluenzae lipooligosaccharide and elucidation of its role in adherence using an outer core mutant

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Abstract:

The opportunistic pathogen Haemophilus parainfluenzae is a gram-negative bacterium found in the oropharynx of humans. Haemophilus parainfluenzae is a member of the Pasteurellaceae family in which it is most closely related to Haemophilus sengis and Actinobacillus. Characterization of surface displayed lipooligosaccharide has identified components that are crucial in adherence. We examined the oligosaccharide structure of lipooligosaccharide from 2 clinical isolates of H. parainfluenzae. Core oligosaccharide was isolated by standard methods from purified lipooligosaccharide. Structural information was established by a combination of monosaccharide and methylation analyses, nuclear magnetic resonance spectroscopy, and mass spectrometry revealing the following structures: R-(1-6)--Glc-(1-4)-d,d-α-Hep-(1-6)--Glc-(1-4)- substituting a tri-heptose-Kdo inner core of L,d-α-Hep-(1-2)-l,d-α-Hep-(1-3)-l,d-α-Hep-(1-5)-α-Kdo at the 4-position of the proximal l,d-α-Hep residue to Kdo, and with a PEtn residue at the 6-position of the central l,d-α-Hep residue. In strain 4282, the R substituent is -galactose and in strain 4201 there is no substituent at the distal glucose. These analyses have revealed that multiple structural aspects of H. parainfluenzae lipooligosaccharide are comparable with nontypeable Haemophilus influenzae lipooligosaccharide. This study also identified a galactan in strain 4201 and a glucan in strain 4282. Haemophilus parainfluenzae was shown to adhere to a bronchial epithelial cell line to the same degree as nontypeable H. influenzae. However, an H. parainfluenzae mutant lacking the outer core of the lipooligosaccharide showed diminished adherence to the epithelial cells, suggesting that H. parainfluenzae lipooligosaccharide plays a role in tissue colonization.

Le pathogène opportuniste Haemophilus parainfluenzae est une bactérie négative à gram retrouvée dans l’oropharynx chez l’humain. Haemophilus parainfluenzae est membre de la famille des Pasteurellaceae plus étroitement apparenté à Haemophilus sengis et Actinobacillus. La caractérisation du lipooligosaccharide exposé à la surface a permis d’identifier les composantes cruciales à l’adhérence. Nous avons examiné la structure des oligosaccharides du lipooligosaccharide de 2 isolats cliniques de H. parainfluenzae. L’oligosaccharide central a été isolé par des méthodes standards à partir du lipooligosaccharide purifié. L’information structurale a été obtenue grâce à une combinaison d’analyses des monosaccharides et de la méthylation, de spectroscopie à résonnance magnétique nucléaire et de spectrométrie de masse qui ont mis en évidence les structures suivantes : R-(1-6)--Glc-(1-4)-d,d-α-Hep-(1-6)--Glc-(1-4)- substituant un tri-heptose-Kdo central du l,d-α-Hep-(1-2)-l,d-α-Hep-(1-3)-l,d-α-Hep-(1-5)-α-Kdo situé en position 4 du résidu l,d-α-Hep proximal au Kdo, et un résidu PEtn, situé en position 6 du résidu l,d-α-Hep central. Chez la souche 4282, le substituant R est le -galactose alors qu’il n’y a pas de substituant attaché au glucose distal chez la souche 4201. Ces analyses ont révélé que les aspects structuraux multiples du lipooligosaccharide de H. parainfluenzae sont comparables à ceux du lipooligosaccharide de H. influenzae non typable. Cette étude a aussi permis d’identifier un galactane chez la souche 4201 et un glucane chez la souche 4282. Haemophilus parainfluenzae peut adhérer à des cellules d’une lignée épithéliale bronchique au même degré que H. influenzae non typable. Cependant, un mutant de H. parainfluenzae dépourvu des composantes extérieures du lipooligosaccharide présente une adhérence réduite aux cellules épithéliales, suggérant que le lipooligosaccharide de H. parainfluenzae joue un rôle dans la colonisation du tissu.

Document Type: Research Article

Publication date: November 1, 2008

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  • Published since 1954, this monthly journal contains new research in the field of microbiology including applied microbiology and biotechnology; microbial structure and function; fungi and other eucaryotic protists; infection and immunity; microbial ecology; physiology, metabolism and enzymology; and virology, genetics, and molecular biology. It also publishes review articles and notes on an occasional basis, contributed by recognized scientists worldwide.
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