147Stephen H. Gillespie (ed.), Antibiotic Resistance Protocols, Methods in Molecular Biology, vol. 1736,
https://doi.org/10.1007/978-1-4939-7638-6_14, © Springer Science+Business Media, LLC 2018
Chapter 14
Use of Larval Zebrafish Model to Study Within-Host
Infection Dynamics
Tomasz K. Prajsnar, Gareth McVicker, Alexander Williams,
Stephen A. Renshaw, and Simon J. Foster
Abstract
Investigating bacterial dynamics within the infected host has proved very useful for understanding mecha-
nisms of pathogenesis. Here we present the protocols we use to study bacterial dynamics within infected
embryonic zebrafish. This chapter encompasses basic techniques used to study bacterial infection within
larval zebrafish, including embryonic zebrafish maintenance, injections of morpholino oligonucleotides,
intravenous injections of bacterial suspensions, and fluorescence imaging of infected zebrafish. Specific
methods for studying bacterial within-host population dynamics are also described.
Key words Zebrafish, Infection, Bacterial population dynamics, Fluorescence microscopy1 Introduction
Animal models of human infection have proven an effective way to
elucidate the mechanisms of bacterial pathogenesis. Zebrafish are
commonly used in bacterial pathogenesis studies [ 1 ] mainly due to
their small size, ease of breeding, and similarities of their immune
system components to those of humans [ 2 ]. Macrophages and
neutrophils are already present in developing zebrafish at 30 h
postfertilization (hpf). In addition, the optical transparency of
embryonic and larval zebrafish allows visualization of immune cell
types interacting with invading pathogens in real time. Both host
and pathogen cells can be fluorescently labeled either genetically or
with fluorescent dyes [ 3 , 4 ] to visualize pathogen subcellular local-
ization as well as the pH in their intraphagocyte milieu [ 4 ].
Additionally, morpholino-modified antisense oligonucleotides
(morpholinos or MOs) have been extensively used in zebrafish to
knock down genes of interest by either blocking the translation or
interfering with the RNA splicing process. MO-mediated knock-
down can be easily used as a measure to study the influence of host
factors on the bacterial infection process. We would stress the