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Infection and Immunity, September 2008, p. 4251-4258, Vol. 76, No. 9
0019-9567/08/$08.00+0     doi:10.1128/IAI.01570-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Galactose Residues on the Lipooligosaccharide of Moraxella catarrhalis 26404 Form the Epitope Recognized by the Bactericidal Antiserum from Conjugate Vaccination{triangledown}

Shengqing Yu,1 Hang Xie,1 Anup Datta,2 Natasha Naidu,2 and Xin-Xing Gu1*

Vaccine Research Facility, National Institute on Deafness and Other Communication Disorders, Rockville, Maryland 20850,1 Glycotechnology Core Resource, Glycobiology Research and Training Center, University of California, San Diego, La Jolla, California 920932

Received 28 November 2007/ Returned for modification 27 January 2008/ Accepted 17 April 2008

Lipooligosaccharide (LOS) from Moraxella catarrhalis has the potential to elicit bactericidal antibodies against the pathogen. We generated LOS-based conjugate vaccines that elicited bactericidal antibodies in animal models. However, epitopes on the LOS recognized by the functional anti-LOS antibodies remain unidentified. In this study, a mutant strain, D4, which lost the recognition by a bactericidal anti-LOS rabbit serum in Western blotting was generated from a serotype C strain 26404 by random transposon mutagenesis. Sequence analysis revealed there was an insertion of a kanamycin resistance gene in the lgt2 gene of D4, which encodes β(1-4)-galactosyltransferase. An isogenic lgt2 mutant, 26404lgt2, was constructed. Structural analysis indicated that the mutant strain produced a truncated LOS lacking terminal galactoses from 4- and 6-linked oligosaccharide chains of strain 26404. Further studies showed that the antiserum lost the recognition of both mutant cells and LOSs in Western blotting, an enzyme-linked immunosorbent assay (ELISA), or a flow cytometry assay. The antiserum also lost the ability to kill the mutant strain in a bactericidal assay, whereas it showed a bactericidal titer of 1:80 to strain 26404. In an inhibition ELISA, D-(+)-galactose or 26404lgt2 LOS showed no inhibition. However, the 26404 LOS and a serotype A O35E LOS with terminal galactoses on its 6-linked oligosaccharide chain showed >90% inhibition, while a serotype B 26397 LOS showed >60% inhibition. These studies suggest that the terminal {alpha}-Gal-(1->4)-β-Gal on the 6-linked oligosaccharide chain of 26404 LOS plays a critical role in forming the epitope recognized by the bactericidal antiserum induced by immunization with our conjugate vaccine.

* Corresponding author. Mailing address: 5 Research Court, Room 2A31, Rockville, MD 20850. Phone: (301) 402-2456. Fax: (301) 435-4040. E-mail: guxx{at}nidcd.nih.gov

{triangledown} Published ahead of print on 16 June 2008.

Editor: A. Camilli

Infection and Immunity, September 2008, p. 4251-4258, Vol. 76, No. 9
0019-9567/08/$08.00+0     doi:10.1128/IAI.01570-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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