ABSTRACT: These microarray studies are part of a larger study characterizing a deletion mutant of the putative transcriptional regulator IclR in Francisella tularensis LVS and SchuS4 strains. The microarrays were performed using RNA isolated from wild-type LVS and a LVS iclR deletion mutant after growing in Chamberlain?s defined media pH 6.3 to early mid-log phase. Results suggest that IclR affects expression of several genes after determining statistically significant differences by SAM. Overall design: Comparison of gene expression between LVS iclR deletion mutant vs. wild-type LVS.
INSTRUMENT(S): JCVI PFGRC Francisella tularensis 12K v2 array designed primarily based on strain Schu 4
Project description:These microarray studies are part of a larger study characterizing a deletion mutant of the putative transcriptional regulator IclR in Francisella tularensis LVS and SchuS4 strains. The microarrays were performed using RNA isolated from wild-type LVS and a LVS iclR deletion mutant after growing in Chamberlain?s defined media pH 6.3 to early mid-log phase. Results suggest that IclR affects expression of several genes after determining statistically significant differences by SAM. Comparison of gene expression between LVS iclR deletion mutant vs. wild-type LVS.
Project description:Prior aerosol exposure to F. tularensis subsp. tularensis, but not the live attenuated strain (LVS) of F. tularensis subsp. holarctica or F. novicida, significantly antagonized the transcriptional response in the lungs of infected mice exposed to aerosolized TLR4 ligand E. coli LPS. Overall design: The ability of a Toll-like receptor 4 (TLR4) agonist to induce a pulmonary inflammatory response in Francisella-infected animals was examined to distinguish between these two possible mechanisms, and also to investigate potential differences between three Francisella strains that exhibit varying levels of virulence in humans.
Project description:Francisella tularensis is a category A select agent based on its infectivity and virulence but disease mechanisms in F. tularensis infection remain poorly understood. A murine pulmonary model of infection was therefore employed to characterize the host immune response to F. tularensis infection and to discern differences in responses to infection with the highly virulent Type A F. tularensis strain Schu4 and to the less virulent Type B live vaccine strain (LVS). Mice were infected intranasally with F. tularensis Schu4 or LVS and organ lesions were assessed 48 and 120 hours after infection. Experiments were also conducted to assess and compare the host immune response to infection using global transcriptional analysis. Mice were infected via low dose aerosol with F. tularensis Schu4 or LVS strains, and transcriptional response in the lungs and spleen was monitored using full mouse genome microarrays at 12, 24, 48, and 120 hours after infection. We found differential and temporal differences in expression of cytokine and chemokine genes, CD molecules, apoptosis, leukocytes and adhesion receptors, and immunological signaling between infection with Schu4 and LVS strains. The transcriptional differences coincided with marked differences in the timing and extent of organ lesions in mice infected with the LVS and Schu4 strains. Thus, these studies revealed both temporal and qualitative differences in the host immune response to infection with virulent F. tularensis Schu4 compared to infection with LVS. Overall design: 18 mice total, 2 uninfected controls, 2 mice per time point, time points were 12 hour/24 hour/48 hour/ 120 hour post infection, 2 infection groups: LVS and Schu4 infection. Lung and spleen were harvested at each time point, poly A RNA was extracted, converted to cDNA, labeled and hybridized in triplicate
Project description:Francisella tularensis LVS was grown in MH broth in the presence or absence of 200uM spermine. RNA was harvasted from overnight (16 hour) cultures and processed for microarray hybridization.
Project description:We demonstrated recently that both constitutive and FAS-triggered apoptosis of human neutrophils are profoundly impaired by Francisella tularensis, but how this is achieved is largely unknown. To test the hypothesis that changes in neutrophil gene expression contribute to this phenotype, we used human oligonucleotide microarrays to identify differentially regulated genes in cells infected with F. tularensis strain LVS compared with uninfected controls. In order to examine the effect of F. tularensis on the neutrophil transcriptome, we performed microarray expression analysis on human neutrophils treated with F. tularensis subsp. holarctica live vaccine strain (LVS). Polymorphonuclear leukocytes (PMNs) were isolated from the blood of healthy donors. Control and F. tularensis-exposed PMNs were incubated at 37C for 0, 3, 6, 12, 24, and 48 hours.
Project description:Francisella tularensis LVS was grown to mid log phase and culture split into two. One culture served as control and other added H2O2 to final concentration of 0.03%. After 20 min at 37 C, RNA was isolated from both cultures