Visualisation of bioluminescent Vibrio midae SY9 K811 in juvenile Haliotis midae abalone using non-invasive bioluminescence imagining
Master Thesis
2014
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University of Cape Town
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Abstract
Bioluminescence imaging (BLI) is a powerful technique that enables real time monitoring of various processes such as gene regulation, host-pathogen interaction and distribution of bacterial cells in vivo. Most of these processes however have only been monitored in murine models or mammalian tissue culture systems. In order to determine whether BLI could be used to monitor bacterial-host interactions in an invertebrate system, the probiotic bacterium Vibrio midae SY9 was genetically labelled with a bioluminescent plasmid (pKluxCat) containing the lux operon from Photorhabdus luminescens. The plasmid was modified by the addition of the Tn9 chloramphenicol acetyltransferase (cat) gene to enable selection of Vibrio transconjugants. The bacterial cells were labelled using a conjugal mating technique and the resulting transconjugant cells were highly bioluminescent with a minimum of 5.264 x104 colony forming units detectable in liquid culture. The bioluminescent cells emitted light stably and consistently without compromising the growth of the strain, and bioluminescence emission was at its highest during the logarithmic growth phase. A strong linear correlation between bioluminescence emitted and bacterial numbers (r2=0.99) was found, indicating that the amount of bacteria present in vivo could be accurately quantitated using bioluminescence. Administration of the bioluminescent bacterial cells by oral gavage to juvenile Haliotis midae abalone allowed non-invasive visualisation of bioluminescence within abalone for up to 10 hours post-gavage. This study therefore demonstrates the potential for using bioluminescence imaging to investigate the interaction of bacteria with an invertebrate host.
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Includes bibliographical references.
Reference:
Moonsamy, K. 2014. Visualisation of bioluminescent Vibrio midae SY9 K811 in juvenile Haliotis midae abalone using non-invasive bioluminescence imagining. University of Cape Town.