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Applied and Environmental Microbiology, May 2003, p. 2864-2874, Vol. 69, No. 5
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.5.2864-2874.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

In Vivo Tracking of Campylobacter jejuni by Using a Novel Recombinant Expressing Green Fluorescent Protein

Philip F. Mixter, John D. Klena, Gary A. Flom, Amy M. Siegesmund, and Michael E. Konkel*

School of Molecular Biosciences, Washington State University, Pullman, Washington 99164-4234

Received 4 October 2002/ Accepted 7 February 2003

Campylobacter jejuni is a leading cause of food-borne disease in developed countries. The goal of this study was to develop a plasmid-based reporter system with green fluorescent protein (GFP) to facilitate the study of C. jejuni in a variety of niches. C. jejuni transformants harboring the pMEK91 GFP gene (gfp)-containing vector were readily detectable by both fluorescence microscopy and flow cytometry. Given the ease of detecting these organisms, additional experiments were performed in which BALB/c mice were injected intraperitoneally with C. jejuni harboring the gfp-containing vector. Four hours after injection of the mice, flow cytometry analyses determined that C. jejuni synthesizing GFP were predominantly associated with granulocytes. More specifically, the proportion of CD11b+ Gr-1+ lavage neutrophils with green fluorescence ranged from 99.7 to 100%, while the proportion of CD11b+ Gr-1- lavage macrophages ranged from 77.0 to 80.0%. In contrast, few CD11b- CD45R+ B lymphocytes from the lavage of the C. jejuni-injected mice were associated with green-fluorescent C. jejuni (proportions ranged from 0.75 to 0.77%). Cell-free C. jejuni was recovered from tissue homogenates after intraperitoneal injection. Macrorestriction profiling with pulsed-field gel electrophoresis identified a genotypic variant of the C. jejuni F38011 wild-type isolate. In vivo this variant displayed a phenotype identical to that of the wild-type isolate. In summary, we demonstrate that C. jejuni associates with marker-defined cellular subsets in vivo with a novel gfp reporter system and that C. jejuni genotypic variants can be isolated from both in vitro and in vivo systems.

* Corresponding author. Mailing address: School of Molecular Biosciences, Washington State University, Pullman, WA 99164-4234. Phone: (509) 335-5039. Fax: (509) 335-1907. E-mail: konkel{at}mail.wsu.edu.

Applied and Environmental Microbiology, May 2003, p. 2864-2874, Vol. 69, No. 5
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.5.2864-2874.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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