Characterization of the Origin of DNA Replication of the <i>Coxiella burnetii</i> Chromosome

Chen, Shuyin; Hoover, Timothy A.; Thompson, Herbert A.; Williams, Jim C.

doi:10.1111/j.1749-6632.1990.tb42259.x

Cited by 15 publications

(12 citation statements)

References 52 publications

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“…These values are about one-third the size of the DNA present in E. coli and are well within the sizes found in free living bacteria. Of considerable interest is the recent isolation of the origin of replication of the C. burnetii genome (7). Characterization of this region may provide insight into any unique aspects of rickettsial DNA replication.…”

Section: Genomementioning

confidence: 99%

Genetics of rickettsiae

1991

Eur J Epidemiol

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Classical genetic approaches useful with free-living bacteria are difficult to apply to the rickettsiae. Although rickettsial mutants have been isolated over the years, the genetic basis of these mutants is unknown, limiting their usefulness. The application of molecular biological techniques to rickettsial studies has provided the opportunity to isolate and study specific genes. Genes encoding metabolic enzymes from rickettsiae were cloned in Escherichia coli and shown to retain their regulatory properties, suggesting that recombinant DNA technology may be useful for studies of rickettsial enzymes and regulatory mechanisms. The potential use of rickettsial surface components, or virulence factors as possible antigens for protective subunit vaccines, has led to the cloning and expression in E. coli, of rickettsial chromosomal and plasmid genes encoding outer membrane proteins. The number of genes characterized in recent years has increased dramatically giving rise to an increasing source of information on rickettsial gene structure. Plasmids have only been identified in C. burnetii and possibly Rochalimaea quintana. The plasmid sequences present in C. burnetii are highly conserved suggesting that they are important to the growth and virulence of this organism. To understand the role of genes in the rickettsia-host relationship, it is critical that a genetic exchange system be developed. The recent description of transformation of R. quintana by electroporation is an important first step in this direction. The ability to introduce genetic material is necessary to address questions that cannot be resolved by studying rickettsial gene expression in E. coli.

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Section: Genomementioning

confidence: 99%

Genetics of rickettsiae

1991

Eur J Epidemiol

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“…The first successful introduction of exogenous plasmid DNA into Coxiella burnetii was reported more than 10 years ago and involved the transformation of eukaryotic host cell-propagated C. burnetii with a shuttle vector containing a 5.8 kb autonomous replication sequence (ARS) 127,128 . The generation of new genetic tools has benefited from the development of axenic media to propagate C. burnetii .…”

mentioning

confidence: 99%

“…The original C. burnetii Himar1 mutants were isolated after electroporation and infection of Vero cells, a technical and laborious process 131 . A second important genetic tool was a stably maintained shuttle vector plasmid for C. burnetii containing the plasmid RSF1010 origin of replication (adapted from use in Legionella pneumophila ) 127 . As an alternative approach to transformation with plasmids, a site-specific Tn 7 transposition system was recently developed 132 .…”

mentioning

confidence: 99%

Molecular pathogenesis of the obligate intracellular bacterium Coxiella burnetii

et al. 2013

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The agent of Q fever, Coxiella burnetii, is an obligate intracellular bacterium that causes acute and chronic infections. The study of C. burnetii pathogenesis has benefited from two recent fundamental advances: improved genetic tools and the ability to grow the bacterium in extracellular media. In this Review, we describe how these recent advances have improved our understanding of C. burnetii invasion and host cell modulation, including the formation of replication-permissive Coxiella-containing vacuoles. Furthermore, we describe the Dot/Icm (defect in organelle trafficking/intracellular multiplication) system, which is used by C. burnetii to secrete a range of effector proteins into the host cell, and we discuss the role of these effectors in remodelling the host cell.

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“…The organism is an obligate intracellular parasite that grows in acidic phagolysosomes of eukaryotic cells [2,3]. Genetic manipulation was possible after the isolation of an autonomous replication sequence ( ars ) from the C. burnetii genome [4,5]. The ars was cloned into pBluescript resulting in plasmid pSKO(+)1000 which was introduced into C. burnetii by electroporation.…”

Section: Introductionmentioning

confidence: 99%

Expression of Î²-lactamase inCoxiella burnetiitransformants

Suhan

Thompson

2000

FEMS Microbiology Letters

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Coxiella burnetii can be transformed to ampicillin resistance by electroporation with plasmids encoding beta-lactamase. However, non-plasmid emergence of resistance to ampicillin also develops. To validate the usefulness of the bla gene marker for selection and detection, transformed C. burnetii were examined for beta-lactamase expression by use of immunoblotting after SDS-PAGE. The 29-kDa mature form of the beta-lactamase protein was detected in C. burnetii lysates. Quantitation of these immunoblot signals showed that C. burnetii surprisingly expressed low levels of beta-lactamase. The results validate the use of plasmid-encoded ampicillin resistance as a means for selecting C. burnetii transformants; they also suggest that levels of ampicillin used for selection pressure should be empirically determined and that detection of beta-lactamase by antibody blotting done to confirm transformants.

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Characterization of the Origin of DNA Replication of the Coxiella burnetii Chromosome

Cited by 15 publications

References 52 publications

Genetics of rickettsiae

Genetics of rickettsiae

Molecular pathogenesis of the obligate intracellular bacterium Coxiella burnetii

Expression of Î²-lactamase inCoxiella burnetiitransformants

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