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Infection and Immunity, March 2006, p. 1828-1836, Vol. 74, No. 3
0019-9567/06/$08.00+0     doi:10.1128/IAI.74.3.1828-1836.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Biofilm Growth Increases Phosphorylcholine Content and Decreases Potency of Nontypeable Haemophilus influenzae Endotoxins

Shayla West-Barnette, Andrea Rockel, and W. Edward Swords*

Department of Microbiology and Immunology, Wake Forest University Health Sciences, Winston-Salem, North Carolina 27157

Received 24 June 2005/ Returned for modification 1 August 2005/ Accepted 2 December 2005

Nontypeable Haemophilus influenzae (NTHI) is a common respiratory commensal and opportunistic pathogen. NTHI is normally contained within the airways by host innate defenses that include recognition of bacterial endotoxins by Toll-like receptor 4 (TLR4). NTHI produces lipooligosaccharide (LOS) endotoxins which lack polymeric O side chains and which may contain host glycolipids. We recently showed that NTHI biofilms contain variants with sialylated LOS glycoforms that are essential to biofilm formation. In this study, we show that NTHI forms biofilms on epithelial cell layers. Confocal analysis revealed that sialylated variants were distributed throughout the biofilm, while variants expressing phosphorylcholine (PCho) were found within the biofilm. Consistent with this observation, PCho content of LOS purified from NTHI biofilms was increased compared to LOS from planktonic cultures. Hypothesizing that the observed changes in endotoxin composition could affect bioactivity, we compared inflammatory responses to NTHI LOS purified from biofilm and planktonic cultures. Our results show that endotoxins from biofilms induced weaker host innate responses. While we observed a minimal effect of sialylation on LOS bioactivity, there was a significant decrease in bioactivity associated with PCho substitutions. We thus conclude that biofilm growth increases the proportion of PCho+ variants in an NTHI population, resulting in a net decrease in LOS bioactivity. Thus, in addition to their well-documented resistance phenotypes, our data show that biofilm communities of NTHI bacteria contain variants that evoke less potent host responses.

* Corresponding author. Mailing address: Department of Microbiology and Immunology, Wake Forest University Health Sciences, 5101A Gray Building, Medical Center Blvd., Winston-Salem, NC 27157. Phone: (336) 713-5049. Fax: (336) 716-9928. E-mail: wswords{at}wfubmc.edu.

Editor: J. N. Weiser

Infection and Immunity, March 2006, p. 1828-1836, Vol. 74, No. 3
0019-9567/06/$08.00+0     doi:10.1128/IAI.74.3.1828-1836.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

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