Construction of a MeOPN Transferase Knockout Mutant in C . jejuni 81 – 176

Campylobacter jejuni is a major cause of bacterial gastroenteritis worldwide, and the capsular polysaccharide (CPS) of this organism is required for persistence and disease. C. jejuni produces over 47 different capsular structures, including a unique O-methyl phosphoramidate (MeOPN) modification present on most C. jejuni isolates. Although the MeOPN structure is rare in nature it has structural similarity to some synthetic pesticides. In this study, we have demonstrated, by whole genome comparisons and high resolution magic angle spinning NMR, that MeOPN modifications are common to several Campylobacter species. Using MeOPN biosynthesis and transferase mutants generated in C. jejuni strain 81–176, we observed that loss of MeOPN from the cell surface correlated with increased invasion of Caco-2 epithelial cells and reduced resistance to killing by human serum. In C. jejuni, the observed serum mediated killing was determined to result primarily from activation of the classical complement pathway. The C. jejuni MeOPN transferase mutant showed similar levels of colonization relative to the wild-type in chickens, but showed a five-fold drop in colonization when co-infected with the wild-type in piglets. In Galleria mellonella waxmoth larvae, the MeOPN transferase mutant was able to kill the insects at wildtype levels. Furthermore, injection of the larvae with MeOPN-linked monosaccharides or CPS purified from the wild-type strain did not result in larval killing, indicating that MeOPN does not have inherent insecticidal activity. Citation: van Alphen LB, Wenzel CQ, Richards MR, Fodor C, Ashmus RA, et al. (2014) Biological Roles of the O-Methyl Phosphoramidate Capsule Modification in Campylobacter jejuni. PLoS ONE 9(1): e87051. doi:10.1371/journal.pone.0087051 Editor: Mikael Skurnik, University of Helsinki, Finland Received September 7, 2013; Accepted December 18, 2013; Published January 30, 2014 This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication. Funding: LVA held an Alberta Innovates Health Solutions Postdoctoral Fellowship. RAA is an Alberta Innovates Technology Futures Scholarship awardee. CMS is an Alberta Innovates Scholar. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: cszymans@ualberta.ca . These authors contributed equally to this work.