ArticleScientific reports2017
A genetic system for targeted mutations to disrupt and restore genes in the obligate bacterium, Ehrlichia chaffeensis.
Article in Scientific reports, 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers.
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Who cites it
13 citing papers in PubMed, 16 citations in OpenAlex.
- A targeted mutational strategy aiding in generating antisense RNA to knockdown theJournal of bacteriology · 2026Article
- Article
- RipE expression correlates with high ATP levels inFrontiers in cellular and infection microbiology · 2024Article
- Recent advances in genetic systems in obligate intracellular human-pathogenic bacteria.Frontiers in cellular and infection microbiology · 2023Review
- Type 1 secretion system and effectors in Rickettsiales.Frontiers in cellular and infection microbiology · 2023Review
- Targeted mutagenesis in Anaplasma marginale to define virulence and vaccine development against bovine anaplasmosis.PLoS pathogens · 2022Article
- Analysis of Orientia tsutsugamushi promoter activity.Pathogens and disease · 2021Article
- Functional Characterization of MultipleInternational journal of molecular sciences · 2021Article
- Multiple Ehrlichia chaffeensis Genes Critical for Its Persistent Infection in a Vertebrate Host Are Identified by Random Mutagenesis Coupled withInfection and immunity · 2020Article
- Mutational analysis of gene function in the Anaplasmataceae: Challenges and perspectives.Ticks and tick-borne diseases · 2019Article
- Sequence Determinants Spanning -10 Motif and Spacer Region Implicated in UniqueFrontiers in microbiology · 2019Article
- Protein and DNA synthesis demonstrated in cell-free Ehrlichia chaffeensis organisms in axenic medium.Scientific reports · 2018Article
- Antigen-Specific CD4Frontiers in immunology · 2018Article
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Authors and funding
5 authors at 2 institutions in 1 country.
Funding
Abstract
Obligate intracellular bacteria (obligates) belonging to Rickettsiales and Chlamydiales cause diseases in hundreds of millions of people worldwide and in many animal species. Lack of an efficient system for targeted mutagenesis in obligates remains a major impediment in understanding microbial pathogenesis. Challenges in creating targeted mutations may be attributed to essential nature of majority of the genes and intracellular replication dependence. Despite success in generating random mutations, a method that works well in creating mutations in specific genes of interest followed by complementation remains problematic for obligates and is a highly sought-after goal. We describe protocols to generate stable targeted mutations by allelic exchange in Ehrlichia chaffeensis, an obligate intracellular tick-borne bacterium responsible for human monocytic ehrlichiosis. Targeted mutations in E. chaffeensis were created to disrupt two genes, and also to restore one gene by another allelic exchange mutation leading to the restoration of transcription and protein expression from the inactivated gene and the recovered organisms also express mCherry, which distinguishes from the wild type. We expect that the methods developed are broadly applicable to other obligates, particularly to rickettsial pathogens, to routinely perform targeted mutations to enable studies focused on protein structure-function analyses, host-pathogen interactions and in developing vaccines.
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