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Updated: Feb 26, 2026
An Electroporation Method to Transform Rickettsia spp. with a Fluorescent Protein-Expressing Shuttle Vector in Tick Cell Lines
Published on: October 11, 2022
Transcriptional profiling of Rickettsia prowazekii coding and non-coding transcripts during in vitro host-pathogen
Casey L C Schroeder1, Hema P Narra1, Abha Sahni1
1Department of Pathology, University of Texas Medical Branch, 301 University Boulevard Galveston, TX 77555, USA.
Abstract:
Natural pathogen transmission of Rickettsia prowazekii, the etiologic agent of epidemic typhus, to humans is associated with arthropods, including human body lice, ticks, and ectoparasites of eastern flying squirrel. Recently, we have documented the presence of small RNAs in Rickettsia species and expression of R. prowazekii sRNAs during infection of cultured human microvascular endothelial cells (HMECs), which represent the primary target cells during human infections. Bacterial noncoding transcripts are now well established as critical post-transcriptional regulators of virulence and adaptation mechanisms in varying host environments. Despite their importance, little is known about the expression profile and regulatory activities of R. prowazekii sRNAs (Rp_sRs) in different host cells encountered as part of the natural life-cycle. To investigate the sRNA expression profile of R. prowazekii during infection of arthropod host cells, we employed an approach combining in vitro infection, bioinformatics, RNA sequencing, and PCR-based quantitation. Global analysis of R. prowazekii transcriptome by strand-specific RNA sequencing enabled us to identify 67 cis-acting (antisense) and 26 trans-acting (intergenic) Rp_sRs expressed during the infection of Amblyomma americanum (AAE2) cells. Comparative evaluation of expression during R. prowazekii infection of HMECs and AAE2 cells by quantitative RT-PCR demonstrated significantly higher expression of four selected Rp_sRs in tick AAE2 cells. Examination of the coding transcriptome revealed differential up-regulation of >150 rickettsial genes in either HMECs or AAE2 cells and yielded evidence for host cell-dependent utilization of alternative transcription start sites by 18 rickettsial genes. Our results thus suggest noticeable differences in the expression of both Rp_sRs as well as the coding transcriptome and the exploitation of multiple transcription initiation sites for select genes during the infection of human endothelium and tick vector cells as the host and yield new insights into rickettsial virulence and transmission mechanisms.
Insights
This study reveals that Rickettsia prowazekii small RNAs (Rp_sRs) and coding genes show different expression patterns in human versus tick cells. This finding offers new insights into bacterial virulence and transmission dynamics.
Area of Science:
- Microbiology
- Molecular Biology
- Genomics
Background:
- Rickettsia prowazekii causes epidemic typhus and transmits via arthropods.
- Small RNAs (sRNAs) regulate bacterial virulence and adaptation.
- The sRNA expression profile of R. prowazekii in different host cells is largely unknown.
Purpose of the Study:
- To investigate the sRNA expression profile of R. prowazekii during infection of arthropod host cells.
- To compare sRNA and coding gene expression in human and tick host cells.
- To gain insights into rickettsial virulence and transmission.
Main Methods:
- In vitro infection models using human microvascular endothelial cells (HMECs) and Amblyomma americanum (AAE2) cells.
- Strand-specific RNA sequencing for global transcriptome analysis.
- Quantitative RT-PCR for expression profiling of selected sRNAs and genes.
Main Results:
- Identified 67 cis-acting and 26 trans-acting Rp_sRs during AAE2 cell infection.
- Four selected Rp_sRs showed significantly higher expression in tick AAE2 cells compared to HMECs.
- >150 rickettsial genes were differentially expressed between HMECs and AAE2 cells.
- Evidence for host cell-dependent alternative transcription start sites in 18 rickettsial genes.
Conclusions:
- R. prowazekii exhibits distinct sRNA and coding gene expression profiles in human and tick host cells.
- The bacterium utilizes alternative transcription start sites in a host-dependent manner.
- These findings enhance understanding of rickettsial virulence and transmission mechanisms.
