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Updated: Jul 26, 2025
High-throughput Assay to Phenotype Salmonella enterica Typhimurium Association, Invasion, and Replication in Macrophages
Published on: August 11, 2014
Salmonella enterica serovar Typhi uses two type 3 secretion systems to replicate in human macrophages and colonize
Meagan Hamblin1, Ruth Schade1, Ramya Narasimhan1
1Department of Microbiology and Immunology, Stanford University School of Medicine , Stanford, California, USA.
Abstract:
Salmonella enterica serovar Typhi (S. Typhi) is a human-restricted pathogen that replicates in macrophages. In this study, we investigated the roles of the S. Typhi type 3 secretion systems (T3SSs) encoded on Salmonella pathogenicity islands (SPI)-1 (T3SS-1) and SPI-2 (T3SS-2) during human macrophage infection. We found that mutants of S. Typhi deficient for both T3SSs were defective for intramacrophage replication as measured by flow cytometry, viable bacterial counts, and live time-lapse microscopy. T3SS-secreted proteins PipB2 and SifA contributed to S. Typhi replication and were translocated into the cytosol of human macrophages through both T3SS-1 and T3SS-2, demonstrating functional redundancy for these secretion systems. Importantly, an S. Typhi mutant strain that is deficient for both T3SS-1 and T3SS-2 was severely attenuated in the ability to colonize systemic tissues in a humanized mouse model of typhoid fever. Overall, this study establishes a critical role for S. Typhi T3SSs during its replication within human macrophages and during systemic infection of humanized mice. IMPORTANCE Salmonella enterica serovar Typhi is a human-restricted pathogen that causes typhoid fever. Understanding the key virulence mechanisms that facilitate S. Typhi replication in human phagocytes will enable rational vaccine and antibiotic development to limit the spread of this pathogen. While S. Typhimurium replication in murine models has been studied extensively, there is limited information available about S. Typhi replication in human macrophages, some of which directly conflict with findings from S. Typhimurium murine models. This study establishes that both of S. Typhi's two type 3 secretion systems (T3SS-1 and T3SS-2) contribute to intramacrophage replication and virulence.
Insights
Salmonella Typhi uses two type 3 secretion systems (T3SS-1 and T3SS-2) to replicate inside human macrophages. Both T3SSs are crucial for bacterial virulence and colonization during typhoid fever infections.
Area of Science:
- Microbiology
- Immunology
- Infectious Diseases
Background:
- Salmonella enterica serovar Typhi (S. Typhi) is a human-specific pathogen causing typhoid fever.
- S. Typhi replicates within human macrophages, a key aspect of its pathogenesis.
- Understanding S. Typhi's virulence factors in human cells is vital for developing new treatments.
Purpose of the Study:
- To investigate the roles of S. Typhi's type 3 secretion systems (T3SS-1 and T3SS-2) in human macrophage infection.
- To determine the contribution of T3SSs to intramacrophage replication and systemic virulence.
Main Methods:
- Generating S. Typhi mutants deficient in T3SS-1 and/or T3SS-2.
- Assessing bacterial replication within human macrophages using flow cytometry, viable counts, and live microscopy.
- Evaluating bacterial colonization in a humanized mouse model of typhoid fever.
Main Results:
- Mutants lacking both T3SS-1 and T3SS-2 showed significantly reduced intramacrophage replication.
- T3SS-secreted proteins PipB2 and SifA were essential for S. Typhi replication and translocated via both T3SSs.
- A double T3SS mutant was severely attenuated in colonizing systemic tissues in a humanized mouse model.
Conclusions:
- Both T3SS-1 and T3SS-2 are critical for S. Typhi replication within human macrophages.
- Functional redundancy exists between T3SS-1 and T3SS-2 in mediating bacterial virulence.
- Targeting these T3SSs could be a promising strategy for typhoid fever therapeutics.
