Chapter 14
In Vivo Imaging of Bioluminescent Leptospires
Fre ´ de ´ rique Vernel-Pauillac and Catherine Werts
Abstract
The study of pathological processes is often limited to in vitro or ex vivo assays, while understanding
pathogenesis of an infectious disease requires in vivo analysis. The use of pathogens, genetically modified to
express with luminescent enzymes, combined to charge-coupled device (CCD) cameras, constitutes a major
technological advance for assessing the course of infection in an intact, living host in real time and in a
noninvasive way. This technology, also called bioluminescence imaging, detects the photons emitted from
biological sources of light through animal tissues. Here, we describe the method we developed to monitor
leptospirosis in a mouse model, by following in a spatiotemporal scale, the dissemination and spread of
leptospires. These bacteria have been genetically modified to express the firefly luciferase, which produces
light in the presence of the substrate D-luciferin. This useful and accessible technology facilitates the study
of the kinetics of blood and tissue dissemination of live leptospires, and the pharmacological impact of
treatments and host directed therapeutics.
Key words Bioluminescence, Luciferase, Leptospires, In vivo live imaging, Bacterial dissemination,
Spatiotemporal kinetics, In vivo therapeutic monitoring
1 Introduction
In vivo assays are an essential step to advance our understanding of
pathological processes involved in an infectious disease. To assess
features leading to the spread of an infection and the preferential
localization of the pathogen, or tissue niche, conventional techniques have relied on end point assays or repeated sacrifices of animals
at designated time points.
Nowadays, each experiment conducted with live animals is
ethically evaluated and animal welfare must be ensured. Therefore,
in vivo live imaging constitutes an alternative and useful way to
reduce and refine experiments; it allows multi-animal, highthroughput studies at relatively low cost, useful to assess infection
evolution with a high signal sensitivity [1–5]. Bioluminescence
imaging is based on the activity of luminescent reporter genes
stably inserted into the genome of the microorganism of interest
[6–8]. The expression of the reporter and its bioluminescent
Nobuo Koizumi and Mathieu Picardeau (eds.), Leptospira spp.: Methods and Protocols, Methods in Molecular Biology, vol. 2134,
https://doi.org/10.1007/978-1-0716-0459-5_14, © Springer Science+Business Media, LLC, part of Springer Nature 2020
149
In Vivo Imaging of Bioluminescent Leptospires
Fre ´ de ´ rique Vernel-Pauillac and Catherine Werts
Abstract
The study of pathological processes is often limited to in vitro or ex vivo assays, while understanding
pathogenesis of an infectious disease requires in vivo analysis. The use of pathogens, genetically modified to
express with luminescent enzymes, combined to charge-coupled device (CCD) cameras, constitutes a major
technological advance for assessing the course of infection in an intact, living host in real time and in a
noninvasive way. This technology, also called bioluminescence imaging, detects the photons emitted from
biological sources of light through animal tissues. Here, we describe the method we developed to monitor
leptospirosis in a mouse model, by following in a spatiotemporal scale, the dissemination and spread of
leptospires. These bacteria have been genetically modified to express the firefly luciferase, which produces
light in the presence of the substrate D-luciferin. This useful and accessible technology facilitates the study
of the kinetics of blood and tissue dissemination of live leptospires, and the pharmacological impact of
treatments and host directed therapeutics.
Key words Bioluminescence, Luciferase, Leptospires, In vivo live imaging, Bacterial dissemination,
Spatiotemporal kinetics, In vivo therapeutic monitoring
1 Introduction
In vivo assays are an essential step to advance our understanding of
pathological processes involved in an infectious disease. To assess
features leading to the spread of an infection and the preferential
localization of the pathogen, or tissue niche, conventional techniques have relied on end point assays or repeated sacrifices of animals
at designated time points.
Nowadays, each experiment conducted with live animals is
ethically evaluated and animal welfare must be ensured. Therefore,
in vivo live imaging constitutes an alternative and useful way to
reduce and refine experiments; it allows multi-animal, highthroughput studies at relatively low cost, useful to assess infection
evolution with a high signal sensitivity [1–5]. Bioluminescence
imaging is based on the activity of luminescent reporter genes
stably inserted into the genome of the microorganism of interest
[6–8]. The expression of the reporter and its bioluminescent
Nobuo Koizumi and Mathieu Picardeau (eds.), Leptospira spp.: Methods and Protocols, Methods in Molecular Biology, vol. 2134,
https://doi.org/10.1007/978-1-0716-0459-5_14, © Springer Science+Business Media, LLC, part of Springer Nature 2020
149