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Using Enhanced Green Fluorescence Protein-expressing Escherichia Coli to Assess Mouse Peritoneal Macrophage Phagocytosis
Published on: January 4, 2019
The co-transcriptome of uropathogenic Escherichia coli-infected mouse macrophages reveals new insights into
Charalampos Harris Mavromatis1, Nilesh J Bokil, Makrina Totsika
1Division of Biological and Environmental Sciences and Engineering, Division of Computer, Electrical and Mathematical Sciences and Engineering, King Abdullah University of Science and Technology, Thuwal, Kingdom of Saudi Arabia; Division of Medical Genetics, Department of Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA, USA.
Abstract:
Urinary tract infections (UTI) are among the most common infections in humans. Uropathogenic Escherichia coli (UPEC) can invade and replicate within bladder epithelial cells, and some UPEC strains can also survive within macrophages. To understand the UPEC transcriptional programme associated with intramacrophage survival, we performed host-pathogen co-transcriptome analyses using RNA sequencing. Mouse bone marrow-derived macrophages (BMMs) were challenged over a 24 h time course with two UPEC reference strains that possess contrasting intramacrophage phenotypes: UTI89, which survives in BMMs, and 83972, which is killed by BMMs. Neither of these strains caused significant BMM cell death at the low multiplicity of infection that was used in this study. We developed an effective computational framework that simultaneously separated, annotated and quantified the mammalian and bacterial transcriptomes. Bone marrow-derived macrophages responded to the two UPEC strains with a broadly similar gene expression programme. In contrast, the transcriptional responses of the UPEC strains diverged markedly from each other. We identified UTI89 genes up-regulated at 24 h post-infection, and hypothesized that some may contribute to intramacrophage survival. Indeed, we showed that deletion of one such gene (pspA) significantly reduced UTI89 survival within BMMs. Our study provides a technological framework for simultaneously capturing global changes at the transcriptional level in co-cultures, and has generated new insights into the mechanisms that UPEC use to persist within the intramacrophage environment.
Insights
Uropathogenic Escherichia coli (UPEC) survival within macrophages was studied using co-transcriptome analysis. A key gene, pspA, was identified as crucial for UPEC persistence inside host immune cells.
Area of Science:
- Microbiology
- Immunology
- Genomics
Background:
- Urinary tract infections (UTIs) are common, often caused by Uropathogenic Escherichia coli (UPEC).
- Some UPEC strains can survive within macrophages, a critical factor in persistent infections.
- Understanding the genetic basis of UPEC intramacrophage survival is essential for developing new treatments.
Purpose of the Study:
- To investigate the transcriptional changes in UPEC during intramacrophage survival.
- To identify specific UPEC genes contributing to survival within macrophages.
- To develop a computational framework for analyzing host-pathogen co-transcriptomes.
Main Methods:
- Host-pathogen co-transcriptome analysis using RNA sequencing.
- Challenging mouse bone marrow-derived macrophages (BMMs) with two UPEC strains (UTI89 and 83972) with differing survival phenotypes.
- Developing a computational framework for simultaneous transcriptome analysis of host and pathogen.
Main Results:
- UPEC transcriptional responses diverged significantly between the two strains within macrophages.
- Several UTI89 genes were upregulated at 24 hours post-infection, suggesting a role in survival.
- Deletion of the pspA gene in UPEC significantly reduced its survival within BMMs.
Conclusions:
- The study provides a novel technological framework for co-culture transcriptome analysis.
- Identified specific UPEC genes, like pspA, are critical for intramacrophage survival.
- These findings offer new insights into UPEC persistence mechanisms within the host.
