Construction of a reporter plasmid for screening in vivo promoter activity in Francisella tularensis

Kuoppa, K

doi:10.1016/s0378-1097(01)00446-3

Cited by 20 publications

(28 citation statements)

References 9 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Of these, approximately 1 % fluoresced on agar when examined under UV light and were removed from the library. This is similar to the proportion of in vitroexpressed promoters identified by Kuoppa et al (2001) when screening for cat activity. The non-fluorescent clones were assembled into pools of eight for screening in macrophages.…”

Section: Resultssupporting

confidence: 75%

“…As electroporation is poorly efficient in Francisella compared with conjugation, the oriT locus of pPV2 was inserted into pKK202 to produce a mobilizable vector. A promoter trapping vector based on induction of chloramphenicol acetyltransferase has been shown to be able to identify intracellularly induced genes (Kuoppa et al, 2001). The drawback of the use of antibiotic resistance as an inducible marker is that it requires the introduction of multiple resistance genes into a human pathogen.…”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

An intracellularly inducible gene involved in virulence and polyphosphate production in Francisella

Richards

Michell

Oyston

2008

Journal of Medical Microbiology

View full text Add to dashboard Cite

Section: Resultssupporting

confidence: 75%

Section: Resultsmentioning

confidence: 99%

An intracellularly inducible gene involved in virulence and polyphosphate production in Francisella

Richards

Michell

Oyston

2008

Journal of Medical Microbiology

View full text Add to dashboard Cite

“…PCR primers are listed in Supplementary Table S1. Plasmid pKK214 (Kuoppa et al, 2001) was PCR-amplified with primers pKK214-EcoRI and pKK214-EcoNotI, followed by digestion with EcoRI and religation, which resulted in pKEK648, which lacks Cm R . Then the FTN1451 promoter (Gallagher et al, 2007) was digested from pKEK886 (Liu et al, 2007) with EcoRI and NotI, and ligated into pKEK648 digested similarly, resulting in pKEK894.…”

Section: Methodsmentioning

confidence: 99%

Characterization of the Francisella tularensis subsp. novicida type IV pilus

et al. 2008

View full text Add to dashboard Cite

Francisella tularensis causes the disease tularaemia. Type IV pili (Tfp) genes are present in the genomes of all F. tularensis subspecies. We show that the wild-type F. tularensis subsp. novicida expresses pilus fibres on its surface, and mutations in the Tfp genes pilF and pilT disrupt pilus biogenesis. Mutations in other Tfp genes (pilQ and pilG) do not eliminate pilus expression. A mutation in pilE4 eliminates pilus expression, whereas mutations in the other pilin subunits pilE1-3 and pilE5 do not, suggesting that pilE4 is the major pilus structural subunit. The virulence regulator MglA is required for pilus expression, and it regulates the transcription of a putative Tfp glycosylation gene (FTN0431). However, MglA does not regulate transcription of pilF, pilT or pilE4, and a strain lacking FTN0431 still expresses pili; thus, it is unclear how MglA regulates pilus expression. Only pilF was also required for protein secretion, while pilE4 and pilT were not, indicating that there is very little overlap of the protein secretion/Tfp functions of the pil genes. The protein secretion component pilE1 was more important for in vitro intramacrophage growth and mouse virulence than the Tfp component pilE4. Our results provide the first genetic characterization of the novel Tfp system of F. tularensis. INTRODUCTIONFrancisella tularensis is a highly infectious bacterium that causes tularaemia. F. tularensis is found in many different animal hosts and can be transmitted by arthropod vectors (Ellis et al., 2002). Humans can acquire the bacteria by a number of different routes, but inhalation of low doses of the organism can lead to a serious pneumonic form of disease that has a high mortality rate, and this has led to the classification of this bacterium as a category A biothreat agent (Dennis et al., 2001). F. tularensis is further divided into different subspecies (Oyston et al., 2004). F. tularensis subsp. tularensis has historically been associated with bioweapons development, and is reported to have the highest virulence for humans (McLendon et al., 2006). F. tularensis subsp. holarctica is also infectious in humans but typically with a less severe outcome (Sjostedt, 2003). An attenuated 'live vaccine strain' (LVS) was derived by repeated passage of subsp. holarctica in the laboratory; despite the unclear nature of the attenuating mutation(s) in this strain, it is frequently used as a laboratory model for tularaemia (Anthony & Kongshavn, 1987;Eigelsbach et al., 1951). F. tularensis subsp. novicida has low virulence for humans, but maintains high virulence in mice (Kieffer et al., 2003;Lauriano et al., 2004). Whole-genome sequencing has revealed that all three subspecies are closely related, with the less virulent (for humans) subsp. novicida showing less genomic decay than the more virulent subsp. holarctica and subsp. tularensis (Larsson et al., 2005;Petrosino et al., 2006;Rohmer et al., 2006).The ability of F. tularensis to survive and replicate within macrophages has been linked to its virulence (Anthony et al., 19...

show abstract

“…The origin of Francisella replication in this plasmid was determined by deletion and sequence analysis (96,100). Using pFNL10, E. coli-F. tularensis shuttle vectors were created, but these plasmids are not stably maintained within F. tularensis (73,86,94). A stable Francisella plasmid, pFNLTP1, was created by spontaneous deletion during passage through LVS (86).…”

Section: Genetic Tools For Manipulation Of Francisella Tularensismentioning

confidence: 99%

Francisella tularensis:Taxonomy, Genetics, and Immunopathogenesis of a Potential Agent of Biowarfare

McLendon

Apicella

Allen

2006

Annu. Rev. Microbiol.

208

292

View full text Add to dashboard Cite

Tularemia is a zoonosis of humans caused by infection with the facultative intracellular bacterium Francisella tularensis. Interest in F. tularensis has increased markedly in the past few years because of its potential use as an agent of bioterrorism. Five subspecies of this organism are found in the Northern hemisphere, but only F. tularensis subsp. tularensis and subsp. holarctica cause disease in humans. This review summarizes what is known about the pathogenesis of tularemia with a focus on bacterial surface components such as lipopolysaccharide and capsule as well as information obtained from the F. tularensis subsp. tularensis SCHU S4 genome. In particular, the mechanisms of action of recently identified virulence factors are discussed in the context of bacterial replication in macrophages and manipulation of the host inflammatory response. Throughout this report shared and unique features of F. tularensis subsp. tularensis, subsp. holarctica, and subsp. novicida are discussed.

show abstract

Construction of a reporter plasmid for screening in vivo promoter activity in Francisella tularensis

Cited by 20 publications

References 9 publications

An intracellularly inducible gene involved in virulence and polyphosphate production in Francisella

An intracellularly inducible gene involved in virulence and polyphosphate production in Francisella

Characterization of the Francisella tularensis subsp. novicida type IV pilus

Francisella tularensis:Taxonomy, Genetics, and Immunopathogenesis of a Potential Agent of Biowarfare

Contact Info

Product

Resources

About