Sequencing of the<i>Francisella tularensis</i>Strain Schu 4 Genome Reveals the Shikimate and Purine Metabolic Pathways, Targets for the Construction of a Rationally Attenuated Auxotrophic Vaccine

Karlsson, J. L.; Prior, Richard G.; Williams, Katrina; Lindler, Luther E.; Brown, Katherine A.; Chatwell, Nicola; Hjalmarsson, Karin; Loman, Nicholas J.; Mack, Kerri; Pallen, Mark J.; Popek, Michael; Sandström, Gunnar; Sjöstedt, Anders; Svensson, Thomas; Tamaš, Ivica; Andersson, Siv G. E.; Wren, Brendan W.; Oyston, Petra C. F.; Titball, Richard W.

doi:10.1089/10906590050145249

Cited by 50 publications

(51 citation statements)

References 51 publications

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“…The sequencing of the F. tularensis genome (22,28) and the development of simple genetic tools with F. tularensis subsp. novicida have facilitated analysis of virulence factors of Francisella.…”

Section: Resultsmentioning

confidence: 99%

A Francisella tularensis Pathogenicity Island Required for Intramacrophage Growth

et al. 2004

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Francisella tularensis is a gram-negative, facultative intracellular pathogen that causes the highly infectious zoonotic disease tularemia. We have discovered a ca. 30-kb pathogenicity island of F. tularensis (FPI) that includes four large open reading frames (ORFs) of 2.5 to 3.9 kb and 13 ORFs of 1.5 kb or smaller. Previously, two small genes located near the center of the FPI were shown to be needed for intramacrophage growth. In this work we show that two of the large ORFs, located toward the ends of the FPI, are needed for virulence. Although most genes in the FPI encode proteins with amino acid sequences that are highly conserved between high-and low-virulence strains, one of the FPI genes is present in highly virulent type A F. tularensis, absent in moderately virulent type B F. tularensis, and altered in F. tularensis subsp. novicida, which is highly virulent for mice but avirulent for humans. The G؉C content of a 17.7-kb stretch of the FPI is 26.6%, which is 6.6% below the average G؉C content of the F. tularensis genome. This extremely low G؉C content suggests that the DNA was imported from a microbe with a very low G؉C-containing chromosome.

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Section: Resultsmentioning

confidence: 99%

A Francisella tularensis Pathogenicity Island Required for Intramacrophage Growth

et al. 2004

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“…In L. monocytogenes, the required proteins have been characterized and comprise the pore-forming lysteriolysin (LLO) and two phospholipases C, PlcA and PlcB. An ongoing sequencing analysis of the genome of the highly virulent isolate Schu S4 has not revealed the presence of any hemolysins or phospholipases (26,36). Thus, it is likely that F. tularensis utilizes mechanisms distinct from those of other intracellular bacterial pathogens, like L. monocytogenes, to escape from the phagosome.…”

Section: Discussionmentioning

confidence: 99%

An Attenuated Strain of the Facultative Intracellular BacteriumFrancisella tularensisCan Escape the Phagosome of Monocytic Cells

Golovliov¹,

Baranov

Kročová³

et al. 2003

Infect Immun

273

335

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The facultative intracellular bacterium Francisella tularensis is a highly virulent and contagious organism, and little is known about its intracellular survival mechanisms. We studied the intracellular localization of the attenuated human vaccine strain, F. tularensis LVS, in adherent mouse peritoneal cells, in mouse macrophagelike cell line J774A.1, and in human macrophage cell line THP-1. Confocal microscopy of infected J774A.1 cells indicated that during the first hour of infection the bacteria colocalized with the late endosomal-lysosomal glycoprotein LAMP-1, but within 3 h this colocalization decreased significantly from approximately 60% to 30%. Transmission electron microscopy revealed that >90% of bacteria were not enclosed by a phagosomal membrane after 2 h of infection, and some bacteria were in vacuoles that were only partially surrounded by a limiting membrane. Similar findings were obtained with all three host cell types. Immunoelectron microscopy performed with an F. tularensis LVS-specific polyclonal rabbit antiserum showed that the antiserum stained a thick, evenly distributed capsule-like material in bacteria grown in broth. In contrast, intracellular F. tularensis LVS cells were only marginally stained with this antiserum. Instead, most of the immunoreactive material was diffusely localized in the phagosomes or was associated with the phagosomal membrane. Our findings indicate that F. tularensis LVS is able to escape from the phagosomes of macrophages via a mechanism that may involve degradation of the phagosomal membrane.

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“…It has been postulated previously that mutations affecting the F. tularensis purine synthesis pathway could be used to generate a live attenuated tularemia vaccine (25). In fact, defined allelic replacement mutants with mutations that disrupt this pathway have been used to produce vaccine strains attenuated for replication in host cells in a variety of other bacterial species (36).…”

Section: Discussionmentioning

confidence: 99%

AttenuatedFrancisella novicidaTransposon Mutants Protect Mice against Wild-Type Challenge

et al. 2006

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Francisella tularensis is the bacterial pathogen that causes tularemia in humans and a number of animals. To date, there is no approved vaccine for this widespread and life-threatening disease. The goal of this study was to identify F. tularensis mutants that can be used in the development of a live attenuated vaccine. We screened F. novicida transposon mutants to identify mutants that exhibited reduced growth in mouse macrophages, as these cells are the preferred host cells of Francisella and an essential component of the innate immune system. This approach yielded 16 F. novicida mutants that were 100-fold more attenuated for virulence in a mouse model than the wild-type parental strain. These mutants were then tested to determine their abilities to protect mice against challenge with high doses of wild-type bacteria. Five of the 16 attenuated mutants (with mutations corresponding to dsbB, FTT0742, pdpB, fumA, and carB in the F. tularensis SCHU S4 strain) provided mice with protection against challenge with high doses (>8 ؋ 10 5 CFU) of wild-type F. novicida. We believe that these findings will be of use in the design of a vaccine against tularemia.

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Sequencing of theFrancisella tularensisStrain Schu 4 Genome Reveals the Shikimate and Purine Metabolic Pathways, Targets for the Construction of a Rationally Attenuated Auxotrophic Vaccine

Cited by 50 publications

References 51 publications

A Francisella tularensis Pathogenicity Island Required for Intramacrophage Growth

A Francisella tularensis Pathogenicity Island Required for Intramacrophage Growth

An Attenuated Strain of the Facultative Intracellular BacteriumFrancisella tularensisCan Escape the Phagosome of Monocytic Cells

AttenuatedFrancisella novicidaTransposon Mutants Protect Mice against Wild-Type Challenge

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