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francisella tularensis deltapyrf mutants show that replication in nonmacrophages is sufficient for pathogenesis in vivo.the pathogenesis of francisella tularensis has been associated with this bacterium's ability to replicate within macrophages. f. tularensis can also invade and replicate in a variety of nonphagocytic host cells, including lung and kidney epithelial cells and hepatocytes. as uracil biosynthesis is a central metabolic pathway usually necessary for pathogens, we characterized deltapyrf mutants of both f. tularensis lvs and schu s4 to investigate the role of these mutants in intracellular growth. as ...201020385757
indoleamine 2,3-dioxygenase 1 is a lung-specific innate immune defense mechanism that inhibits growth of francisella tularensis tryptophan auxotrophs.upon microbial challenge, organs at various anatomic sites of the body employ different innate immune mechanisms to defend against potential infections. accordingly, microbial pathogens evolved to subvert these organ-specific host immune mechanisms to survive and grow in infected organs. francisella tularensis is a bacterium capable of infecting multiple organs and thus encounters a myriad of organ-specific defense mechanisms. this suggests that f. tularensis may possess specific factors that ai ...201020385761
lung cd4-cd8- double-negative t cells are prominent producers of il-17a and ifn-gamma during primary respiratory murine infection with francisella tularensis live vaccine strain.for several intracellular infections, pulmonary vaccination provides measurably better protection against pulmonary challenge. the unique factors that contribute to pulmonary immune responses are not well characterized. in this study, we show that cd4(-)cd8(-) double negative (dn) t cells are a major responding t cell subset in the lungs of mice during pulmonary francisella tularensis live vaccine strain (lvs) infection. dn t cells were a minor (<2%) subset in spleens and lungs of mice during su ...201020393138
antimicrobial and antibiofilm activity of cathelicidins and short, synthetic peptides against francisella.francisella infects the lungs causing pneumonic tularemia. focusing on the lung's host defense, we have examined antimicrobial peptides as part of the innate immune response to francisella infection. interest in antimicrobial peptides, such as the cathelicidins, has grown due their potential therapeutic applications and the increasing problem of bacterial resistance to commonly used antibiotics. only one human cathelicidin, ll-37, has been characterized. helical cathelicidins have also been disc ...201020399752
sensing cytoplasmic danger signals by the inflammasome.introduction: the innate immune system depends on molecules collectively known as pattern recognition receptors (prrs) to survey the extracellular space and the cytoplasm for the presence of dangerous pathogens, pathogen-derived molecules, or even self-derived molecular danger signals, which arise from tissue damage. absent in melanoma 2 (aim2) is a newly discovered prr involved in the sensing of dangerous cytosolic dna produced by infection with dna viruses. discussion: remarkably, recent studi ...201020401524
[tularaemia - an overview of the current knowledge].francisella tularensis belongs to the family francisellaceae. it is the aetiological agent of a zoonosis called tularaemia, spread throughout the northern hemisphere. currently, several subspecies of f. tularensis may be distinguished with various pathogenicity and geographical distribution. in human medicine, only sporadic infections or local epidemics are reported. given the fact that f. tularensis is highly pathogenic for humans and is easily spread by aerosol, water or food, it may be exploi ...201020401834
[detection of francisella tularensis by blood culture].francisella tularensis, from the family francisellaceae, is the aetiological agent of a zoonosis called tularaemia, spread throughout the northern hemisphere. the infectious dose is extremely low (10 cfu/ml) and the infection causes severe diseases or even death if untreated. the transmission to humans is always related to animals, either by a direct contact or by a contact with the environment contaminated by them. clinical symptoms of the disease can vary depending on the point of entry of the ...201020401835
mass mortality of giant abalone haliotis gigantea caused by a francisella sp. bacterium.in february 2005, a mass mortality of giant abalone haliotis (nordotis) gigantea gmelin, 1791 occurred on a private abalone farm in shimane prefecture, japan. the cumulative mortality rate reached about 84%. in histological observations, bacteria-like spherical particles were found in affected animals, suggesting a bacterial infection. many of the bacteria-like particles were found in the cells that were presumably host phagocytes. dna was extracted from the hemolymph of a diseased abalone and a ...201020402232
[identifying bacteria using a matrix-assisted laser desorption ionization time-of-flight (maldi-tof) mass spectrometer. comparison with routine methods used in clinical microbiology laboratories].the methods routinely used for bacterial identification in clinical microbiology laboratory, although miniaturized and automated, are still based on the same basic principles as classical identification methods. nevertheless, technological advances are emerging which could modify these routine methods. we report a comparative study between conventional identification methods and mass spectrometry maldi-tof (ms maldi-tof) for bacterial identification in the clinical microbiology laboratory.201020409613
azithromycin effectiveness against intracellular infections of francisella.macrolide antibiotics are commonly administered for bacterial respiratory illnesses. azithromycin (az) is especially noted for extremely high intracellular concentrations achieved within macrophages which is far greater than the serum concentration. clinical strains of type b francisella (f.) tularensis have been reported to be resistant to az, however our laboratory francisella strains were found to be sensitive. we hypothesized that different strains/species of francisella (including type a) m ...201020416090
virulence differences among francisella tularensis subsp. tularensis clades in mice.francisella tularensis subspecies tularensis (type a) and holarctica (type b) are of clinical importance in causing tularemia. molecular typing methods have further separated type a strains into three genetically distinct clades, a1a, a1b and a2. epidemiological analyses of human infections in the united states suggest that a1b infections are associated with a significantly higher mortality rate as compared to infections caused by a1a, a2 and type b. to determine if genetic differences as define ...201020419133
ecological niche modeling of francisella tularensis subspecies and clades in the united states.two subspecies of francisella tularensis are recognized: f. tularensis subsp. tularensis (type a) and f. tularensis subsp. holartica (type b). type a has been subdivided further into a1a, a1b, and a2, which differ geographically and clinically. the aim of this work was to determine whether or not differences among subspecies and clades translate into distinct ecological niches. we used 223 isolates from humans and wildlife representing all six genotypes (type a, b, a1, a2, a1a, or a1b). ecologic ...201020439975
cloning, expression, purification, crystallization and preliminary x-ray diffraction analysis of macrophage growth locus a (mgla) protein from francisella tularensis.francisella tularensis, a potential bioweapon, causes a rare infectious disease called tularemia in humans and animals. the macrophage growth locus a (mgla) protein from f. tularensis associates with rna polymerase to positively regulate the expression of multiple virulence factors that are required for its survival and replication within macrophages. the mgla protein was overproduced in escherichia coli, purified and crystallized. the crystals diffracted to 7.5 a resolution at the advanced phot ...201020445258
absent in melanoma 2 is required for innate immune recognition of francisella tularensis.macrophages respond to cytosolic nucleic acids by activating cysteine protease caspase-1 within a complex called the inflammasome. subsequent cleavage and secretion of proinflammatory cytokines il-1beta and il-18 are critical for innate immunity. here, we show that macrophages from mice lacking absent in melanoma 2 (aim2) cannot sense cytosolic double-stranded dna and fail to trigger inflammasome assembly. caspase-1 activation in response to intracellular pathogen francisella tularensis also req ...201020457908
tularemia of european brown hare (lepus europaeus): a pathological, histopathological, and immunohistochemical study.the european brown hare (lepus europaeus) plays an important role in the ecology of tularemia, and it may serve as a significant source of human infection. the aim of the present study was to examine the lesions induced by francisella tularensis in 50 cases of naturally infected seropositive european brown hares. gross pathological examination revealed scant to numerous grayish-white foci with diameters of 0.1 to 1.0 cm in single organs (24 cases) or multiple organs (20 cases) in 44 of 50 cases ...201020466857
differentiation of bacteria using fatty acid profiles from gas chromatography-tandem mass spectrometry.fatty acids are essential components in cell membranes of bacteria, and they determine the different structures of lipids and lipid a. therefore, fatty acids are good targets for development of rapid and reliable methods for differentiation of bacteria.201020474059
directed screen of francisella novicida virulence determinants using drosophila melanogaster.francisella tularensis is a highly virulent, facultative intracellular human pathogen whose virulence mechanisms are not well understood. occasional outbreaks of tularemia and the potential use of f. tularensis as a bioterrorist agent warrant better knowledge about the pathogenicity of this bacterium. thus far, genome-wide in vivo screens for virulence factors have been performed in mice, all however restricted by the necessity to apply competition-based, negative-selection assays. we wanted to ...201020479082
development of a real-time pcr assay for identification and quantification of the fish pathogen francisella noatunensis subsp. orientalis.members of the genus francisella are small gram-negative facultative intracellular bacteria that cause francisellosis in a wide variety of fish species worldwide. f. noatunensis subsp. orientalis has been recently described as a warm-water pathogen of tilapia oreochromis spp. in this study, a quantitative real-time polymerase chain reaction (qpcr) taqman probe assay was developed to rapidly and accurately detect and quantify f. noatunensis subsp. orientalis from fish tissue. the target region of ...201020481087
molecular bases of proliferation of francisella tularensis in arthropod vectors.arthropod vectors are important vehicles for transmission of francisella tularensis between mammals, but very little is known about the f. tularensis-arthropod vector interaction. drosophila melanogaster has been recently developed as an arthropod vector model for f. tularensis. we have shown that intracellular trafficking of f. tularensis within human monocytes-derived macrophages and d. melanogaster-derived s2 cells is very similar. within both evolutionarily distant host cells, the francisell ...201020482589
molecular complexity orchestrates modulation of phagosome biogenesis and escape to the cytosol of macrophages by francisella tularensis.upon entry of francisella tularensis to macrophages, the francisella-containing phagosome (fcp) is trafficked into an acidified late endosome-like phagosome with limited fusion to the lysosomes followed by rapid escape into the cytosol where the organism replicates. although the francisella pathogenicity island (fpi), which encodes a type vi-like secretion apparatus, is required for modulation of phagosome biogenesis and escape into the cytosol, the mechanisms involved are not known. to decipher ...201020482590
involvement of the aim2, nlrc4, and nlrp3 inflammasomes in caspase-1 activation by listeria monocytogenes.infection with listeria monocytogenes can cause meningitis and septicemia in newborn, elderly, or immunocompromised individuals. pregnant women are particularly susceptible to listeria, leading to a potentially fatal infection. cytosolic listeria activates the proinflammatory caspase-1 and induces the processing and secretion of interleukins il-1beta and il-18 as well as caspase-1-dependent pyroptosis. this study elucidates the role of various inflammasome components of host macrophages in the p ...201020490635
j774 macrophage-like cell line cytokine and chemokine patterns are modulated by francisella tularensis lvs strain infection.mutual interactions were investigated between intracellular parasitic bacterium francisella tularensis (f.t.; highly virulent bacterium responsible for tularemia, replicating within the host macrophages) and murine macrophage-like cell line j774. recombinant murine lymphokine inf-gamma and/or lps derived from e. coli were determined to stimulate in vitro antimicrobial activity of macrophage-like j774 cell line against the live vaccine strain (lvs) of f.t. through their ability to produce proinfl ...201020490763
modulation of hepatic ppar expression during ft lvs lps-induced protection from francisella tularensis lvs infection.it has been shown previously that administration of francisella tularensis (ft) live vaccine strain (lvs) lipopolysaccharide (lps) protects mice against subsequent challenge with ft lvs and blunts the pro-inflammatory cytokine response.201020082697
identification of ciprofloxacin resistance by simpleprobe, high resolution melt and pyrosequencing nucleic acid analysis in biothreat agents: bacillus anthracis, yersinia pestis and francisella tularensis.the potential for genetic modification of biological warfare agents makes rapid identification of antibiotic resistant strains critical for the implementation of suitable infection control measures. the fluorinated quinolone, ciprofloxacin, is an antibiotic effective for treating bacterial infections by inhibiting the enzyme activity of the dna type ii topoisomerases dna gyrase and topoisomerase iv. the genes that encode the subunits of dna gyrase (gyra and gyrb) and topo iv (par c and pare) con ...201020100564
insect infection model for campylobacter jejuni reveals that o-methyl phosphoramidate has insecticidal activity.galleria mellonella (wax moth) larvae have elsewhere been shown to be susceptible to pathogens such as francisella tularensis, burkholderia mallei, and pseudomonas aeruginosa. we report that the larvae are rapidly killed by campylobacter jejuni at 37c. three strains of c. jejuni tested, 11168h (human diarrheal isolate), g1 (human guillain-barré syndrome isolate), and 81-176 (human diarrheal isolate), were equally effective at killing g. mellonella larvae. a panel of defined mutants of c. jejuni ...201020113177
investigating an airborne tularemia outbreak, germany.in november 2005, an outbreak of tularemia occurred among 39 participants in a hare hunt in hesse, germany. previously reported tularemia outbreaks in germany dated back to the 1950s. we conducted a retrospective cohort study among participants and investigated the environment to identify risk factors for infection. ten participants had serologic evidence of acute francisella tularensis infection; 1 other participant died before laboratory confirmation was obtained. presence within 5 meters of t ...201020113553
[rabbit fever--a neck tumor].twenty-two-year-old previously healthy male is referred to the ent clinic with a neck tumour. the patient was an animal caretaker. ultrasonic examination showed a lymph gland conglomerate on the left side of the neck. primary serologic examination was negative and the lymph tumour was removed. the biopsy showed necrotic lymphadenitis. three weeks after the first consultation the patient tests positive for f. tularensis. the patient was treated with ciprofloxacin 750 mg x 2 by the oral route for ...201020122334
[a case of tularemia in a danish hunter]."rabbit fever" (francisella tularensis) is a rare infection in denmark. it was first described in denmark in 1987. it is most likely to affect people who come into close contact with infected animals or ticks, such as hunters, butchers and veterinarians. the diagnosis should be suspected in such persons presenting with fever, headache, lethargy, lymphadenitis and bite wounds. we present a danish case describing the diagnosis and treatment of a hunter infected with t. tularensis.201020122335
toll-like receptor 3 agonist protection against experimental francisella tularensis respiratory tract infection.we investigated whether toll-like receptor 3 (tlr3) stimulation would protect the host from inhaled francisella tularensis. tlr3 is expressed by respiratory epithelial cells and macrophages and can be activated by a synthetic double-stranded rna ligand called polyinosine-polycytosine [poly(i:c)]. thus, we evaluated poly(i:c) as a novel treatment against inhaled f. tularensis. in vivo, balb/c mice intranasally (i.n.) treated with poly(i:c) (100 microg/mouse) 1 h before or after schu 4 or lvs (100 ...201020123717
groel and lipopolysaccharide from francisella tularensis live vaccine strain synergistically activate human macrophages.francisella tularensis, the causative agent of tularemia, interacts with host cells of innate immunity in an atypical manner. for most gram-negative bacteria, the release of lipopolysaccharide (lps) from their outer membranes stimulates an inflammatory response. when lps from the attenuated live vaccine strain (lvs) or the highly virulent schu s4 strain of f. tularensis was incubated with human umbilical vein endothelial cells, neither species of lps induced expression of the adhesion molecule e ...201020123721
identification of trkh, encoding a potassium uptake protein required for francisella tularensis systemic dissemination in mice.francisella tularensis is a highly infectious bacterium causing the zoonotic disease tularaemia. during its infectious cycle, f. tularensis is not only exposed to the intracellular environment of macrophages but also resides transiently in extracellular compartments, in particular during its systemic dissemination. the screening of a bank of f. tularensis lvs transposon insertion mutants on chemically defined medium (cdm) led us to identify a gene, designated trkh, encoding a homolog of the pota ...201020126460
a tolc mutant of francisella tularensis is hypercytotoxic compared to the wild type and elicits increased proinflammatory responses from host cells.the highly infectious bacterium francisella tularensis is a facultative intracellular pathogen and the causative agent of tularemia. tolc, which is an outer membrane protein involved in drug efflux and type i protein secretion, is required for the virulence of the f. tularensis live vaccine strain (lvs) in mice. here, we show that an lvs deltatolc mutant colonizes livers, spleens, and lungs of mice infected intradermally or intranasally, but it is present at lower numbers in these organs than in ...201020028804
inhibition of acpa phosphatase activity with ascorbate attenuates francisella tularensis intramacrophage survival.acid phosphatase activity in the highly infectious intracellular pathogen francisella tularensis is directly related with the ability of these bacteria to survive inside host cells. pharmacological inactivation of acid phosphatases could potentially help in the treatment of tularemia or even be utilized to neutralize the infection. in the present work, we report inhibitory compounds for three of the four major acid phosphatases produced by f. tularensis schu4: acpa, acpb, and acpc. the inhibitor ...201020028980
real-time pcr for diagnosis of oculoglandular tularemia. 201020031067
the unraveling panoply of francisella tularensis virulence attributes.francisella tularensis is a highly infectious gram-negative bacterium causing the zoonotic disease tularemia. this facultative intracellular pathogen multiplies in vivo mainly inside macrophages, but has the capacity to infect and survive in many other cell types, including other phagocytic and nonphagocytic cells. in vitro, f. tularensis escapes rapidly from the phagosomal compartment and replicates in the cytoplasm of infected cells. an impressive number of novel genes related to f. tularensis ...201020034843
francisella tularensis t-cell antigen identification using humanized hla-dr4 transgenic mice.there is no licensed vaccine against the intracellular pathogen francisella tularensis. the use of conventional mouse strains to screen protective vaccine antigens may be problematic, given the differences in the major histocompatibility complex (mhc) binding properties between murine and human antigen-presenting cells. we used engineered humanized mice that lack endogenous mhc class ii alleles but that express a human hla allele (hla-dr4 transgenic [tg] mice) to identify potential subunit vacci ...201020016043
differential ability of novel attenuated targeted deletion mutants of francisella tularensis subspecies tularensis strain schu s4 to protect mice against aerosol challenge with virulent bacteria: effects of host background and route of immunization.francisella tularensis subspecies tularensis is a highly virulent facultative intracellular pathogen of humans and a potential biological weapon. a live vaccine strain, f. tularensis lvs, was developed more than 50 years ago by pragmatic attenuation of a strain of the less virulent holarctica subspecies. lvs was demonstrated to be highly effective in human volunteers who were exposed to intradermal challenge with fully virulent subsp. tularensis, but was less effective against aerosol exposure. ...201020018266
the genome of the amoeba symbiont "candidatus amoebophilus asiaticus" reveals common mechanisms for host cell interaction among amoeba-associated bacteria.protozoa play host for many intracellular bacteria and are important for the adaptation of pathogenic bacteria to eukaryotic cells. we analyzed the genome sequence of "candidatus amoebophilus asiaticus," an obligate intracellular amoeba symbiont belonging to the bacteroidetes. the genome has a size of 1.89 mbp, encodes 1,557 proteins, and shows massive proliferation of is elements (24% of all genes), although the genome seems to be evolutionarily relatively stable. the genome does not encode pat ...201020023027
a novel brain heart infusion broth supports the study of common francisella tularensis serotypes.francisella tularensis schu s4, lvs and u112 have become model organisms for the study of francisella pathogenesis, and represent a cross section of the different f. tularensis subspecies. both schu s4 and lvs are fastidious organisms, requiring medium fortified with supplements and nutrients for enhanced growth. chamberlains defined medium, tryptone soy broth supplemented with cysteine (tsbc), and cation-adjusted mueller-hinton broth (camhb) supplemented with 2% isovitalex are typically used in ...201020005265
identification, cloning, expression, and purification of francisella lpp3: an immunogenic lipoprotein.the severe and fatal human disease, tularemia, results from infection with the gram-negative pathogen francisella tularensis. identification of surface outer membrane proteins, specifically lipoproteins, has been of interest for vaccine development and understanding the initiation of disease. we sought to identify francisella live vaccine strain lipoproteins that could be a component of a subunit vaccine and have adjuvant properties as tlr2 agonists. we have identified a membrane lipoprotein of ...201020006480
a rapid multiplex assay for nucleic acid-based diagnostics.we have developed a rapid (under 4 hours), multiplex, nucleic acid assay, adapted to a microsphere array detection platform. we call this assay multiplex oligonucleotide ligation-pcr (mol-pcr). unlike other ligation-based assays that require multiple steps, our protocol consists of a single tube reaction, followed by hybridization to a luminex microsphere array for detection. we demonstrate the ability of this assay to simultaneously detect diverse nucleic acid signatures (e.g., unique sequences ...201020006656
contributions of francisella tularensis subsp. novicida chitinases and sec secretion system to biofilm formation on chitin.francisella tularensis, the zoonotic cause of tularemia, can infect numerous mammals and other eukaryotes. although studying f. tularensis pathogenesis is essential to comprehending disease, mammalian infection is just one step in the ecology of francisella species. f. tularensis has been isolated from aquatic environments and arthropod vectors, environments in which chitin could serve as a potential carbon source and as a surface for attachment and growth. we show that f. tularensis subsp. novi ...201019948864
mechanisms of bacterial virulence in pulmonary infections.to consider the relevance to severe human lung infections of recently discovered virulence mechanisms of staphylococcus aureus and francisella tularensis.201019956071
gallium disrupts iron uptake by intracellular and extracellular francisella strains and exhibits therapeutic efficacy in a murine pulmonary infection model.francisella tularensis requires iron (fe) for growth, but the biologic sources of fe for this organism are largely unknown. we found that francisella sp. growing in broth culture or within human macrophages can acquire fe from the two major host fe-binding proteins, lactoferrin (lf) and transferrin (tf). fe acquisition is a potential target for novel therapies. gallium (ga) is a transition metal that interferes with cellular fe metabolism by competing with fe for uptake/utilization. growth of ei ...201019917753
an outbreak of holarctica-type tularemia in pediatric patients.holarctica-type tularemia is endemic in the northern hemisphere. despite recurrent epidemics tularemia is not well known in children and the pediatric cases are often misdiagnosed.201019918211
cell biology and molecular ecology of francisella tularensis.francisella tularensis is a highly infectious intracellular bacterium that causes the fulminating disease tularemia, which can be transmitted between mammals by arthropod vectors. genomic studies have shown that the f. tularensis has been undergoing genomic decay with the most virulent strains having the lowest number of functional genes. entry of f. tularensis into macrophages is mediated by looping phagocytosis and is associated with signalling through syk tyrosine kinase. within macrophages a ...201019863554
utilization of an unstable plasmid and the i-scei endonuclease to generate routine markerless deletion mutants in francisella tularensis.we engineered an efficient system to make francisella tularensis deletion mutations using an unstable, poorly maintained plasmid to enhance the likelihood of homologous recombination. for counterselection, we adapted a strategy using i-scei, which causes a double-stranded break in the integrated suicide vector, forcing a second recombination to mediate allelic replacement.201019879904
phagosomal retention of francisella tularensis results in tirap/mal-independent tlr2 signaling.tlr2 plays a central role in the activation of innate immunity in response to ft, the causative agent of tularemia. we reported previously that ft lvs elicited strong, dose-dependent nf-kappab reporter activity in tlr2-expressing human embryo kidney 293 t cells and that ft lvs-induced murine macrophage proinflammatory cytokine gene and protein expression is tlr2-dependent. we demonstrated further that ft can signal through tlr2 from within the phagosome and that phagosomal retention of ft leads ...201019889726
acid phosphatases do not contribute to the pathogenesis of type a francisella tularensis.the intracellular pathogen francisella tularensis is the causative agent of tularemia, a zoonosis that can affect humans with potentially lethal consequences. essential to francisella virulence is its ability to survive and proliferate within phagocytes through phagosomal escape and cytosolic replication. francisella spp. encode a variety of acid phosphatases, whose roles in phagosomal escape and virulence have been documented yet remain controversial. here we have examined in the highly virulen ...201019858304
description of francisella hispaniensis sp. nov., isolated from human blood, reclassification of francisella novicida (larson et al. 1955) olsufiev et al. 1959 as francisella tularensis subsp. novicida comb. nov. and emended description of the genus francisella.strain fhsp1t, isolated from human blood in spain in 2003, was studied for its taxonomic allocation. by 16s rrna and reca gene sequencing, the strain was shown to belong to the genus francisella. in the 16s rrna gene sequence, francisella sp. fhsp1t shared similarity of more than 99% with strains of francisella tularensis subspecies and francisella novicida u112t, 98% with francisella piscicida gm2212t and 98.4% with francisella philomiragia atcc 25015t. in the reca gene sequence, francisella sp ...201019783615
the presence of cd14 overcomes evasion of innate immune responses by virulent francisella tularensis in human dendritic cells in vitro and pulmonary cells in vivo.francisella tularensis is a gram-negative bacterium that causes acute, lethal disease following inhalation. we have previously shown that viable f. tularensis fails to stimulate secretion of proinflammatory cytokines following infection of human dendritic cells (hdc) in vitro and pulmonary cells in vivo. here we demonstrate that the presence of the cd14 receptor is critical for detection of virulent f. tularensis strain schus4 by dendritic cells, monocytes, and pulmonary cells. addition of solub ...201019841074
development of real-time pcr assays for the specific detection of francisella tularensis ssp. tularensis, holarctica and mediaasiatica.real-time polymerase chain reaction (pcr) assays were developed to detect francisella tularensis (ft), the causative agent of tularaemia in humans. two real-time pcrs (ftt0376 and ftt0523) were designed in genetic sequences identified by the insignia genome comparison tool (http://insignia.cbcb.umd.edu/) as being unique to pathogenic subspecies of f. tularensis. both pcrs identified all pathogenic f. tularensis subspecies but did not cross react with avirulent francisella philomiragia or f. tula ...201019833196
aged mice display an altered pulmonary host response to francisella tularensis live vaccine strain (lvs) infections.aging is a complex phenomenon that has been shown to affect many organ systems including the innate and adaptive immune systems. the current study was designed to examine the potential effect of immunosenescence on the pulmonary immune response using a francisella tularensis live vaccine strain (lvs) inhalation infection model. f. tularensis is a gram-negative intracellular pathogen that can cause a severe pneumonia. in this study both young (8-12 week old) and aged (20-24 month old) mice were i ...201019825409
francisella novicida forms in vitro biofilms mediated by an orphan response regulator.francisella tularensis is associated with water and waterways and infects many species of animals, insects, and protists. the mechanism francisella utilizes to persist in the environment and in tick vectors is currently unknown. we have demonstrated for the first time that francisella novicida, a model organism of f. tularensis, forms a biofilm in vitro. selected f. novicida transposon mutants were tested for their ability to form biofilm compared to the wildtype f. novicida strain. mutation of ...201019763680
survival of secondary lethal systemic francisella lvs challenge depends largely on interferon gamma.although survival of primary infection with the live vaccine strain (lvs) of francisella tularensis depends on interferon gamma (ifn-gamma), the relative importance of ifn-gamma to secondary protective immunity in vivo has not been clearly established. here we examine the role of ifn-gamma in t cell priming and expression of vaccine-induced protection against lethal intraperitoneal challenge of mice. large amounts of ifn-gamma were detected between days 3 and 7 in the sera of lvs-immunized mice, ...201019781659
characterization of francisella tularensis strains, comparing their carbon source utilization.thirteen francisella tularensis strains were isolated from 22 seropositive brown hares (lepus europaeus) originating from different parts of hungary, and further two from a patas monkey (erythrocebus patas) and vervet monkey (chlorocebus aethiops). the isolates were identified as f. tularensis ssp. holarctica on the basis of culture, morphological and biochemical characteristics. the identification was verified by polymerase chain reaction and the sequencing of the partial 16s rrna gene. utiliza ...201019538455
detection of francisella piscicida in atlantic cod (gadus morhua l) by the loop-mediated isothermal amplification (lamp) reaction.the loop-mediated isothermal amplification (lamp) reaction was evaluated for its speed and sensitivity in detecting the presence of francisella piscicida, the causative agent of francisellosis in atlantic cod (gadus morhua). four primer sets, consisting of two outer and two inner, were designed from the groel gene of the pathogen. the lamp reaction was optimised at 63 degrees c for 1h using bacterial genomic dna as the template and the products were visualised under ultra-violet light and analys ...201019398357
the role of birds in dissemination of francisella tularensis: first direct molecular evidence for bird-to-human transmission.during a recent large tularemia outbreak in bulgaria we found several cases that were remote from the main focus. one case had an unusual mode of transmission. a hunter acquired tularemia through a nail scratch from a buzzard (buteo buteo) and consequently developed a typical ulceroglandular form of the disease. the diagnosis was confirmed by serological methods and successful cultivation. comparative strain typing was performed by high-resolution multi-locus variable-number tandem repeat analys ...201019664305
diversity of francisella species in environmental samples from martha's vineyard, massachusetts.we determined whether francisella spp. are present in water, sediment, and soil from an active tularemia natural focus on martha's vineyard, massachusetts, during a multiyear outbreak of pneumonic tularemia. environmental samples were tested by polymerase chain reaction (pcr) targeting francisella species 16s rrna gene and succinate dehydrogenase a (sdha) sequences; evidence of the agent of tularemia was sought by amplification of francisella tularensis-specific sequences for the insertion eleme ...201019669828
[tularemia: a decade in the province of soria (spain)].tularemia is a zoonotic disease that has been regularly reported in spain since 1997. this study analyzes suspected, probable, and confirmed cases of tularemia in the province of soria, and compares them with tularemia cases recorded in the autonomous community of castilla y léon, which, with the exception of 1 sporadic case, occurred in 2 epidemic outbreaks in 1997/1998 and 2007/2008.201019419798
a possible novel francisella genomic species isolated from blood and urine of a patient with severe illness.two identical isolates were recovered in pure culture from the blood and urine of a patient suffering from severe septicaemia associated with obstructive pyelonephritis secondary to lithotripsy. preliminary phenotypic and genotypic characterizations based on serological, biochemical and sequence analyses following pcr amplification of selected gene regions indicate that this organism represents a potential new francisella genomic species.201019709068
tularemia.tularemia is a potentially fatal multi-systemic disease of humans and other animals caused by the bacterial pathogen francisella tularensis. the disease can be transmitted by ticks, biting flies, water exposure, food, and aerosols and occurs around the northern hemisphere including north america, europe, and asia. there are several defined species and subspecies, including f. tularensis subsp. tularensis (jellison type a) which is pathogenic for rabbits and occurs in north america, f. tularensis ...201019713053
the francisella intracellular life cycle: toward molecular mechanisms of intracellular survival and proliferation.the tularemia-causing bacterium francisella tularensis is a facultative intracellular organism with a complex intracellular lifecycle that ensures its survival and proliferation in a variety of mammalian cell types, including professional phagocytes. because this cycle is essential to francisella pathogenesis and virulence, much research has focused on deciphering the mechanisms of its intracellular survival and replication and characterizing both bacterial and host determinants of the bacterium ...201021687806
francisella tularensis metabolism and its relation to virulence.francisella tularensis is a gram-negative bacterium capable of causing the zoonotic disease tularaemia in a large number of mammalian species and in arthropods. f. tularensis is a facultative intracellular bacterium that infects and replicates in vivo mainly inside macrophages. during its systemic dissemination, f. tularensis must cope with very different life conditions (such as survival in different target organs or tissues and/or survival in the blood streamôçª) and may thus encounter a broad ...201021687763
francisella tularensis blue-gray phase variation involves structural modifications of lipopolysaccharide o-antigen, core and lipid a and affects intramacrophage survival and vaccine efficacy.francisella tularensis is a cdc category a biological agent and a potential bioterrorist threat. there is no licensed vaccine against tularemia in the united states. a long-standing issue with potential francisella vaccines is strain phase variation to a gray form that lacks protective capability in animal models. comparisons of the parental strain (lvs) and a gray variant (lvsg) have identified lipopolysaccharide (lps) alterations as a primary change. the lps of the f. tularensis variant strain ...201021687776
the role of the francisella tularensis pathogenicity island in type vi secretion, intracellular survival, and modulation of host cell signaling.francisella tularensis is a highly virulent gram-negative intracellular bacterium that causes the zoonotic disease tularemia. essential for its virulence is the ability to multiply within host cells, in particular monocytic cells. the bacterium has developed intricate means to subvert host immune mechanisms and thereby facilitate its intracellular survival by preventing phagolysosomal fusion followed by escape into the cytosol, where it multiplies. moreover, it targets and manipulates numerous h ...201021687753
enediol mimics as inhibitors of the d-arabinose 5-phosphate isomerase (kdsd) from francisella tularensis.we explored the d-arabinose 5-phosphate isomerase (kdsd, e.c. 5.3.1.13) from francisella tularensis, a highly infectious gram-negative pathogen that has raised concern as a potential bioweapon, as a target for the development of novel chemotherapeutics. f. tularensis kdsd was expressed in escherichia coli from a synthetic gene, purified, and characterized. a group of hydroxamates designed to be mimics of the putative enediol intermediate in the enzyme's catalytic mechanism were prepared and test ...201021236668
genotyping of francisella tularensis, the causative agent of tularemia.francisella tularensis is a facultative, intracellular, zoonotic pathogen and the causative agent of tularemia. historically, f. tularensis has been subdivided into subspecies on the basis of phenotypic traits, including biochemical reactivity and virulence. more recently, a number of genotypic methods, ranging from relatively insensitive methods to full genome sequencing, have been used to investigate genetic diversity within f. tularensis. these analyses indicate that f. tularensis is a pathog ...201021313823
receptor mimicry as novel therapeutic treatment for biothreat agents.the specter of intentional release of pathogenic microbes and their toxins is a real threat. this article reviews the literature on adhesins of biothreat agents, their interactions with oligosaccharides and the potential for anti-adhesion compounds as an alternative to conventional therapeutics. the minimal binding structure of ricin has been well characterised and offers the best candidate for successful anti-adhesion therapy based on the galβ1-4glcnac structure. the botulinum toxin serotypes a ...201021327124
an outbreak of disease caused by francisella sp. in nile tilapia oreochromis niloticus at a recirculation fish farm in the uk.this study details the first diagnosis of francisella sp. in tilapia in the united kingdom. losses of tilapia fry at a recirculation fish farm in england were investigated, giving a presumptive positive diagnosis of infection with francisella sp. by histopathological examination. most fish sampled showed moderate to marked pathology of the major organs, with lesions being present in most tissues. the most obvious host response was granuloma formulation. a subsequent follow-up visit provided furt ...201021387995
[real time pcr hybridization for the rapid and specific identification of francisella tularensis].tularemia is highly infectious and fatal zoonotic disease caused by gram negative bacteria francisella tularensis. the necessity to undergo medical treatment in early phase of illness in humans and possibility of making use of bacterial aerosol by terrorists in an attack create an urgent need to implement a rapid and effective method which enables to identify the agent. in our study two primers fopa f/r and hybridization probes fopa s1/s2 designed from fopa gene sequence, were tested for their p ...201021473100
rapid onset of icam-1 expression is a marker of effective macrophages activation during infection of francisella tularensis lvs in vitro.francisella tularensis is capable to modulate immunobiological activities of the host cells. we focused on the expression of icam-1 (cd54) on j774.2 mouse macrophage cell line infected by f. tularensis live vaccine strain (lvs) in vitro as a putative marker of subsequent elimination of infection. j774.2 cell line cells were infected by f. tularensis lvs strain (multiplicity of infection, 1:100). cell cultures were stimulated either 3 h before infection or 3 h after infection by either lipopolysa ...201121476047
evasion of ifn-γ signaling by francisella novicida is dependent upon francisella outer membrane protein c.francisella tularensis is a gram-negative facultative intracellular bacterium and the causative agent of the lethal disease tularemia. an outer membrane protein (ftt0918) of f. tularensis subsp. tularensis has been identified as a virulence factor. we generated a f. novicida (f. tularensis subsp. novicida) ftn_0444 (homolog of ftt0918) fopc mutant to study the virulence-associated mechanism(s) of ftt0918.201121483828
outbreak of tularaemia in central norway, january to march 2011.from january to march 2011, 39 cases of tularaemia were diagnosed in three counties in central norway: 21 cases of oropharyngeal type, 10 cases of glandular/ulceroglandular type, two of respiratory and two of typhoid type. three cases were asymptomatic and clinical information was unavailable for one case. the mean age was 40.3 years (range 2-89 years). thirty-four reported use of drinking water from private wells. an increased rodent (lemming) population and snow melting may have led to contami ...201121489376
formulation and stabilization of francisella tularensis live vaccine strain.francisella tularensis live vaccine strain (f. tularensis lvs), a promising vaccine candidate for protection against f. tularensis exposure, is a particularly thermolabile vaccine and difficult to stabilize sufficiently for storage under refrigerated conditions. our preliminary data show that f. tularensis lvs can be stabilized in the dried state using foam drying, a modified freeze drying method, with sugar-based formulations. the process was conducted under mild drying conditions, which result ...201121491457
tlr4-dependent activation of inflammatory cytokine response in macrophages by francisella elongation factor tu.the bacterial determinants of pulmonary francisella induced inflammatory responses and their interaction with host components are not clearly defined. in this study, proteomic and immunoblot analyses showed presence of a cytoplasmic protein elongation factor tu (ef-tu) in the membrane fractions of virulent francisella novicida, lvs and schus4, but not in an attenuated f. novicida mutant. ef-tu was immunodominant in mice vaccinated and protected from virulent f. novicida. moreover, recombinant ef ...201121497800
ulceroglandular tularemia.an increasing number of patients with the zoonosis tularemia have been reported in the last few years in europe. tularemia can be divided into different forms depending on its appearance. tularemia must be considered in the differential diagnosis of diseases that present with an ulcer and regional lymphadenopathy. the diagnosis can be confirmed by culturing francisella tularensis. with effective antibiotic intervention, the prognosis is favorable. typically tularemia develops after outdoor activ ...201121501382
structure of francisella tularensis peptidyl-trna hydrolase.the rational design of novel antibiotics for bacteria involves the identification of inhibitors for enzymes involved in essential biochemical pathways in cells. in this study, the cloning, expression, purification, crystallization and structure of the enzyme peptidyl-trna hydrolase from francisella tularensis, the causative agent of tularemia, was performed. the structure of f. tularensis peptidyl-trna hydrolase is comparable to those of other bacterial peptidyl-trna hydrolases, with most residu ...201121505237
comparative study of immune status to infectious agents in elderly patients with multiple myeloma, waldenstrom's macroglobulinemia, and monoclonal gammopathy of undetermined significance.whereas patients with multiple myeloma (mm) have a well-documented susceptibility to infections, this has been less studied in other b-cell disorders, such as waldenstrom's macroglobulinemia (wm) and monoclonal gammopathy of undetermined significance (mgus). we investigated the humoral immunity to 24 different pathogens in elderly patients with mm (n = 25), wm (n = 16), and mgus (n = 18) and in age-matched controls (n = 20). antibody titers against pneumococci, staphylococcal alpha-toxin, tetanu ...201121508164
brucellosis of the european brown hare (lepus europaeus).the european brown hare (lepus europaeus) is an important reservoir of brucella suis biovar 2 and also of the life-threatening zoonotic agent francisella tularensis. since both bacteria can produce similar gross pathological lesions in this species, laboratory tests are necessary for the final diagnosis. the aim of the present study was to develop an immunohistochemical method for the detection of b. suis infection and to describe the pathological and histological lesions caused by b. suis in eu ...201121511271
tularemia vaccines: recent developments and remaining hurdles.francisella tularensis subsp. tularensis is a facultative intracellular bacterial pathogen of humans and other mammals. its inhaled infectious dose is very low and can result in very high mortality. historically, subsp. tularensis was developed as a biological weapon and there are now concerns about its abuse as such by terrorists. a live attenuated vaccine developed pragmatically more than half a century ago from the less virulent holarctica subsp. is the sole prophylactic available, but it rem ...201121526941
transstadial transmission of francisella tularensis holarctica in mosquitoes, sweden.in sweden, human cases of tularemia caused by francisella tularensis holarctica are assumed to be transmitted by mosquitoes, but how mosquito vectors acquire and transmit the bacterium is not clear. to determine how transmission of this bacterium occurs, mosquito larvae were collected in an area where tularemia is endemic, brought to the laboratory, and reared to adults in their original pond water. screening of adult mosquitoes by real-time pcr demonstrated f. tularensis lpna sequences in 14 of ...201121529386
differential chitinase activity and production within francisella species, subspecies and subpopulations.genotyping of francisella tularensis (a1a, a1b, a2 and type b) and francisella novicida has identified multiple differences between species, and among f. tularensis subspecies and subpopulations. variations in virulence, geographic distribution and ecology are also known to exist among this group of bacteria, despite the >95% nucleotide identity in their genomes. this study expands the description of phenotypic differences by evaluating the ability of f. tularensis and f. novicida to degrade chi ...201121531796
a central metabolic circuit controlled by qsec in pathogenic escherichia coli.the qsec sensor kinase regulates virulence in multiple gram-negative pathogens, by controlling the activity of the qseb response regulator. we have previously shown that qsec deletion interferes with dephosphorylation of qseb thus unleashing what appears to be an uncontrolled positive feedback loop stimulating increased qseb levels. deletion of qsec downregulates virulence gene expression and attenuates enterohaemorrhagic and uropathogenic escherichia coli (ehec and upec), salmonella typhimurium ...201121542868
isolation and mutagenesis of a capsule-like complex (clc) from francisella tularensis, and contribution of the clc to f. tularensis virulence in mice.francisella tularensis is a category-a select agent and is responsible for tularemia in humans and animals. the surface components of f. tularensis that contribute to virulence are not well characterized. an electron-dense capsule has been postulated to be present around f. tularensis based primarily on electron microscopy, but this specific antigen has not been isolated or characterized.201121544194
comparative evaluation of automated and manual commercial dna extraction methods for detection of francisella tularensis dna from suspensions and spiked swabs by real-time polymerase chain reaction.this study evaluated commercial automated and manual dna extraction methods for the isolation of francisella tularensis dna suitable for real-time polymerase chain reaction (pcr) analysis from cell suspensions and spiked cotton, foam, and polyester swabs. two automated methods, the magna pure compact and the qiacube, were compared to 4 manual methods, the it 1-2-3 dna sample purification kit, the masterpure complete dna and rna purification kit, the qiaamp dna blood mini kit, and the ultraclean ...201121546201
non-fcεr bearing mast cells secrete sufficient interleukin-4 to control francisella tularensis replication within macrophages.mast cells have classically been implicated in the triggering of allergic and anaphylactic reactions. however, recent findings have elucidated the ability of these cells to selectively release a variety of cytokines leading to bacterial clearance through neutrophil and dendritic cell mobilization, and suggest an important role in innate host defenses. our laboratory has established a primary bone marrow derived mast cell-macrophage co-culture system and found that mast cells mediated a significa ...201121565523
pathology in practice. francisella tularensis. 201121568771
inhibition of francisella tularensis lvs infection of macrophages results in a reduced inflammatory response: evaluation of a therapeutic strategy for intracellular bacteria.f. tularensis is an intracellular pathogen and is able to invade several different cell types, in particular macrophages, most commonly through phagocytosis. a flow cytometric assay was developed to measure bacterial uptake, using a fitc-labelled anti-f. tularensis lps antibody in conjunction with antibodies to cell surface markers, in order to determine specific cell phenotypes that were positive for the bacteria. several phagocytic inhibitors, were evaluated in macrophage cell lines and a lung ...201121569124
the rna chaperone hfq is important for growth and stress tolerance in francisella novicida.the rna-binding protein hfq is recognized as an important regulatory factor in a variety of cellular processes, including stress resistance and pathogenesis. hfq has been shown in several bacteria to interact with small regulatory rnas and act as a post-transcriptional regulator of mrna stability and translation. here we examined the impact of hfq on growth, stress tolerance, and gene expression in the intracellular pathogen francisella novicida. we present evidence of hfq involvement in the abi ...201121573133
tularaemia in berlin - two independent cases in travellers returning from central anatolia, turkey, february 2011. 201121586267
francisella tularensis membrane complexome by blue native/sds-page.the study of membrane proteins and membrane protein complexes (mpc) provides crucial information in the field of bacterial physiology and pathogenesis research. the method of blue native polyacrylamide gel electrophoresis and its combination with sds-page (bn/sds-page) were here employed to study the membrane complexome of an intracellular bacterium francisella tularensis, the causative agent of a severe disease tularemia. in the presented study we describe the subunit composition and stoichiome ...201121601022
impact of francisella tularensis pilin homologs on pilus formation and virulence.francisella tularensis is a facultative intracellular bacterium and the causative agent of tularemia. virulence factors for this bacterium, particularly those that facilitate host cell interaction, remain largely uncharacterized. however, genes homologous to those involved in type iv pilus structure and assembly, including six genes encoding putative major pilin subunit proteins, are present in the genome of the highly virulent schu s4 strain. to analyze the roles of three putative pilin genes i ...201121605655
discovery of a role for hsp82 in histoplasma virulence through a quantitative screen for macrophage lethality.the application of forward genetics can reveal new factors required for the virulence of intracellular pathogens. to facilitate such virulence screens, we developed macrophage cell lines with which the number of intact host cells following infection with intracellular pathogens can be rapidly and easily ascertained through the expression of a constitutive lacz transgene. using known virulence mutants of francisella novicida and histoplasma capsulatum, we confirmed the applicability of these host ...201121606189
evidence of circulation of an epidemic strain of francisella tularensis in france by multispacer typing.multispacer typing (mst) was used to type ten francisella tularensis strains detected in french patients. incorporating 79 swedish f. tularensis strains, phylogenetic analysis demonstrated that although tularemia appears as a sporadic disease in france, it is caused by an epidemic cluster of strains.201121611870
arthropod-borne tularemia in poland: a case report.abstract tularemia is a rare zoonosis. the most common way is ingestion of contaminated meat or water, but the infection may also be acquired by insect bite. the clinical picture of the disease may be nonspecific. due to polymorphisms of clinical picture, specific treatment is often delayed. in the last 50 years, in poland, the most infections were acquired by handling hares. in our article, we present the case of a patient who was infected with francisella tularensis due to arthropod bite. in t ...201121612529
differential mortality of dog tick vectors due to infection by diverse francisella tularensis tularensis genotypes.abstract the factors involved in the long-term perpetuation of francisella tularensis tularensis in nature are poorly understood. martha's vineyard, massachusetts, has become a site of sustained transmission of type a tularemia, with nearly 100 human cases reported from 2000 to 2010. we have identified a stable focus of f. tularensis transmission there, where the annual prevalence in host-seeking dermacentor variabilis is about 3%, suggesting that this tick perpetuates the agent. however, labora ...201121612530
francisella tularensis molecular typing using differential insertion sequence amplification (disa).tularemia is a potentially fatal disease that is caused by the highly infectious and zoonotic pathogen francisella tularensis. despite the monomorphic nature of sequenced f. tularensis genomes, there is a significant degree of plasticity in the organization of genetic elements. the observed variability in these genomes is primarily due to the transposition of direct repeats and insertion sequence (is) elements. since current methods used to genotype f. tularensis are time-consuming and require e ...201121613430
cellulitis, headache, and fever following tick bites. 201121617001
seroprevalence of 10 zoonotic infections in 2 canadian cree communities.we evaluated the seroprevalence of 10 zoonotic agents among the general population (15 years old and over) of eastmain and wemindji, james bay, quebec, in 2007. overall seroprevalence rates were similar between the 2 communities. nearly half the individuals tested (n = 251; 146 women, 105 men) were seropositive (n = 115) for at least one zoonosis. the highest seroprevalence rates were for leptospira sp. (23%), francisella tularensis (17%), and the california serogroup viruses (jc and ssh viruses ...201121398073
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