Investigating Salmonella CsgD - Mediated Bet-Hedging Using a Dual Reporter Strain
Date
2023-10-04
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
ORCID
Type
Thesis
Degree Level
Masters
Abstract
Non-typhoidal Salmonella are a leading cause of foodborne illness. These bacteria cycle
between infecting hosts and persisting in nature as biofilms. The host to environment transition
of Salmonella is a critical stage in the transmission cycle that is not fully understood.
In non-typhoidal Salmonella bistable synthesis of the master biofilm regulator CsgD
results in a population of CsgD⁺ biofilm aggregates CsgD⁻ single cells. The single cells
synthesize the Salmonella pathogenicity island 1 type III secretion system (SPI-1 T3SS) which
is a major virulence factor for host cell invasion. We believe this population splitting is a bet hedging strategy to improve transmission, providing a way to cause disease immediately (single
cells) or after long time periods (aggregates). We hypothesize that Salmonella enterica serovar
Typhimurium (S. Typhimurium) differentiates into biofilm⁺ cells and SPI-1 T3SS⁺ cells in vivo.
To study bet-hedging, I built a fluorescent reporter strain of S. Typhimurium that tracks
biofilm⁺ cells (green fluorescent protein) and SPI-1 T3SS⁺ cells (mCherry) simultaneously. I
quantified the number of cells that form in a biofilm flask model using confocal microscopy. S.
Typhimurium formed four cell populations: biofilm⁺ cells (49%), SPI-1 T3SS⁺ cells (26%),
biofilm and SPI-1 T3SS⁺ cells (2%), and non-fluorescent cells (23%). As SPI-1 is bistably
regulated, we think non-fluorescent cells are a SPI-1 T3SS⁻ population.
To determine if population splitting happens in vivo, I developed a model using
Caenorhabditis elegans. I evaluated survival of worms exposed to S. Typhimurium and
quantified Salmonella inside worms. Exposure to Salmonella shortened worm lifespan, and the
intestinal load of Salmonella increased over time. Worms infected with S. Typhimurium were
imaged by confocal microscopy. Both SPI-1 T3SS⁺ cells and biofilm⁺ cells were observed in the
intestine.
My MSc research has laid the groundwork for future studies on Salmonella bet-hedging
and host-pathogen interactions. The S. Typhimurium dual reporter strain provides a unique way
to study different Salmonella cell types in real-time and in more detail than before. Studying how
biofilm and virulent cells interact with the host immune system will aid in development of
Salmonella vaccines and will help us better understand the importance of bet-hedging for
Salmonella transmission.
Description
Keywords
Salmonella enterica, bet-hedging, bistability, biofilms, SPI-1 type III secretion, persistence, transmission
Citation
Degree
Master of Science (M.Sc.)
Department
Microbiology and Immunology
Program
Microbiology and Immunology