Live and Dead GFP-Tagged Bacteria Showed Indistinguishable Fluorescence in Caenorhabditis elegans Gut

Ju-Ya Hsiao; Chun-Yao Chen; Mei-Jun Yang; Han-Chen Ho

doi:10.1007/s12275-013-2589-8

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Research Support, Non-U.S. Gov't
Live and Dead GFP-Tagged Bacteria Showed Indistinguishable Fluorescence in Caenorhabditis elegans Gut: Ju-Ya Hsiao ¹, Chun-Yao Chen ², Mei-Jun Yang ¹, Han-Chen Ho ¹; Journal of Microbiology 2013;51(3):367-372.
DOI: https://doi.org/10.1007/s12275-013-2589-8
Published online: June 28, 2013

¹Department of Anatomy, Tzu Chi University, Hualien 97004, Taiwan, ²Department of Life Science, Tzu Chi University, Hualien 97004, Taiwan

Corresponding author: Han-Chen Ho , Tel: +886-3-856-5301,

Received: 29 October 2012 • Accepted: 28 January 2013

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Abstract

Caenorhabditis elegans has been used for studying hostpathogen interactions since long, and many virulence genes of pathogens have been successfully identified. In several studies, fluorescent pathogens were fed to C. elegans and fluorescence observed in the gut was considered an indicator for bacterial colonization. However, the grinder in the pharynx of these nematodes supposedly crushes the bacterial cells, and the ground material is delivered to the intestine for nutrient absorption. Therefore, it remains unclear whether intact bacteria pass through the grinder and colonize in the intestine. Here we investigated whether the appearance of fluorescence is indicative of intact bacteria in the gut using both fluorescence microscopy and transmission electron microscopy. In wild-type N2 C. elegans, Escherichia coli DH5α, and Vibrio vulnificus 93U204, both of which express the green fluorescence protein, were found intact only proximal to the grinder, while crushed bacterial debris was found in the post-pharyngeal lumen. Nevertheless, the fluorescence was evident throughout the lumen of worm intestines irrespective of whether the bacteria were intact or not. We further investigated the interaction of the bacteria with C. elegans phm-2 mutant, which has a dysfunctional grinder. Both strains of bacteria were found to be intact and accumulated in the pharynx and intestine owing to the defective grinder. The fluorescence intensity of intact bacteria in phm-2 worms was indistinguishable from that of crushed bacterial debris in N2 worms. Therefore, appearance of fluorescence in the C. elegans intestine should not be directly interpreted as successful bacterial colonization in the intestine.

Keywords: Vibrio vulnificus;phm-2;electron microscopy

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Live and Dead GFP-Tagged Bacteria Showed Indistinguishable Fluorescence in Caenorhabditis elegans Gut

J. Microbiol. 2013;51(3):367-372. Published online June 28, 2013

DOI: https://doi.org/10.1007/s12275-013-2589-8

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