ArticleApplied and environmental microbiology2012
Two systems for targeted gene deletion in Coxiella burnetii.
Article in Applied and environmental microbiology, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 78 papers.
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Who cites it
78 citing papers in PubMed, 106 citations in OpenAlex.
- TGF-β/IFN-γ Antagonism in Subversion and Self-Defense of Phase II Coxiella burnetiiInfection and immunity · 2023Trial
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- Stepwise assembly of virulence-associated traits in the intracellular pathogenbioRxiv : the preprint server for biology · 2026Article
- Bacterial Puppeteering: How the Stealth BacteriumPathogens (Basel, Switzerland) · 2025Review
- Coxiella burnetii as a model system for understanding host immune response against obligate intracellular, vacuolar pathogens.PLoS pathogens · 2025Article
- CRISPR-Cas9-based approaches for genetic analysis and epistatic interaction studies inmSphere · 2024Article
- Caspase-8 activity mediates TNFα production and restrictsInfection and immunity · 2024Article
- Caspase-8 activity mediates TNFα production and restrictsbioRxiv : the preprint server for biology · 2024Article
- Metabolism and physiology of pathogenic bacterial obligate intracellular parasites.Frontiers in cellular and infection microbiology · 2024Review
- Psittacosis: An Underappreciated and Often Undiagnosed Disease.Pathogens (Basel, Switzerland) · 2023Review
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- A Novel Marine MammalPathogens (Basel, Switzerland) · 2023Article
- Complex Signaling Networks Control Coxiella burnetii.Journal of bacteriology · 2023Article
- Article
- Comparative Transcriptomics and Genomics from Continuous Axenic Media Growth IdentifiesbioRxiv : the preprint server for biology · 2023Article
- Advances in genetic manipulation ofFrontiers in immunology · 2023Review
- Recent advances in genetic systems in obligate intracellular human-pathogenic bacteria.Frontiers in cellular and infection microbiology · 2023Review
- SURE editing: combining oligo-recombineering and programmable insertion/deletion of selection markers to efficiently edit the Mycoplasma pneumoniae genome.Nucleic acids research · 2022Article
- The endogenous Coxiella burnetii plasmid encodes a functional toxin-antitoxin system.Molecular microbiology · 2022Article
- Primary Murine Macrophages as a Tool for Virulence Factor Discovery in Coxiella burnetii.Microbiology spectrum · 2022Article
18 more citing papers are in PubMed but not listed here.
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Authors and funding
4 authors at 2 institutions in 1 country.
Funding
Abstract
Coxiella burnetii is a ubiquitous zoonotic bacterial pathogen and the cause of human acute Q fever, a disabling influenza-like illness. C. burnetii's former obligate intracellular nature significantly impeded the genetic characterization of putative virulence factors. However, recent host cell-free (axenic) growth of the organism has enabled development of shuttle vector, transposon, and inducible gene expression technologies, with targeted gene inactivation remaining an important challenge. In the present study, we describe two methods for generating targeted gene deletions in C. burnetii that exploit pUC/ColE1 ori-based suicide plasmids encoding sacB for positive selection of mutants. As proof of concept, C. burnetii dotA and dotB, encoding structural components of the type IVB secretion system (T4BSS), were selected for deletion. The first method exploited Cre-lox-mediated recombination. Two suicide plasmids carrying different antibiotic resistance markers and a loxP site were integrated into 5' and 3' flanking regions of dotA. Transformation of this strain with a third suicide plasmid encoding Cre recombinase resulted in the deletion of dotA under sucrose counterselection. The second method utilized a loop-in/loop-out strategy to delete dotA and dotB. A single suicide plasmid was first integrated into 5' or 3' target gene flanking regions. Resolution of the plasmid cointegrant by a second crossover event under sucrose counterselection resulted in gene deletion that was confirmed by PCR and Southern blot. ΔdotA and ΔdotB mutants failed to secrete T4BSS substrates and to productively infect host cells. The repertoire of C. burnetii genetic tools now allows ready fulfillment of molecular Koch's postulates for suspected virulence genes.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.