Biomedical subjects

G Sandström

Publications and source records attributed to G Sandström.

At least 37 records · Page 2Linked to original sources

PubMed1996

Cytokine expression in the liver of mice infected with a highly virulent strain of Francisella tularensis.

Cytokine mRNA expression was determined in the liver of mice subcutaneously inoculated with a lethal dose of the highly virulent strain FSC 041 of Francisella tularensis subvar, tularensis or a sublethal dose of the live vaccine strain of F. tularensis subvar. palaearctica. Expression of mRNA for TNF-alpha, IL-12, IFN-gamma, and IL-10 was demonstrated within 48 h of inoculation, the kinetics being similar irrespective of bacterial strain used. Thus, the expression of a cytokine response believed to be important in the early host defence against live vaccine strain seemed insufficient to prevent the lethality of a more virulent strain.

Animals↗

PubMed1996

Construction of a shuttle vector for use in Francisella tularensis.

The characterisation of virulence factors of Francisella tularensis has been hampered by the lack of genetic system for the bacterium. In this study, a shuttle vector was constructed that can replicate autonomously in F. tularensis and Escherichia coli. To obtain this vector, the p15A replication origin of F. coli plasmid pACYC184 was introduced into a plasmid derivative of plasmid pFNL2OO, a plasmid which only can replicate in F. tularensis. The resulting shuttle vector, designated pKK2O2, harboured resistance genes for chloramphenicol and tetracycline. This vector might be used as a basis for the studies of virulence factors of F. tularensis.

Bacteriological Techniques↗

PubMed1996

Inability of the Francisella tularensis lipopolysaccharide to mimic or to antagonize the induction of cell activation by endotoxins.

We studied the ability of the lipopolysaccharide (LPS) extracted from a vaccine strain of Francisella tularensis (LPS-Ft) to mimic LPSs from other gram-negative bacteria for activation of various murine cell types or to antagonize the effects of other LPSs. We found that activation of macrophages for the production of tumor necrosis factor alpha and NO, of pre-B lymphocytes for the expression of surface immunoglobulins, and of bone marrow cells for the expression of LPS-binding sites was either undetectable with LPS-Ft or required concentrations 100 to 1,000 times higher than for standard LPSs. Preexposure of macrophages to LPS-Ft also failed to trigger down-regulation of tumor necrosis factor alpha (desensitization) or up-regulation of NO responses to an endotoxin challenge. In contrast to other atypical LPSs, LPS-Ft was also unable to antagonize any of the endotoxin-induced cellular responses mentioned above, suggesting that this LPS does not interact with LPS receptors.

Animals↗

PubMed1996

Characterization of two unusual clinically significant Francisella strains.

We have isolated two phenotypically distinct nonfastidious Francisella strains (Fx1 and Fx2) from the blood of compromised patients with pneumonia and compared them with eight other Francisella strains, including Francisella tularensis biovar tularensis, F. tularensis biovar novicida, and F. philomiragia. Our isolates grew well on sheep blood agar, chocolate agar, modified Thayer-Martin agar, and Trypticase soy agar. Fx1 and Fx2 were determined to be within the Francisella genus by cellular fatty acid analysis and by the utilization of glucose, production of H2S and catalase, and lack of motility, oxidase, nitrate reductase, and gelatinase. They were additionally shown to belong to the species F. tularensis by sequencing of two variable regions comprising approximately 500 nucleotides of the 16S rRNA gene. Also, RNA probe hybridization confirmed their belonging to the species F. tularensis. However, the new strains, which are not identical, are distinguished from other F. tularensis strains by growth characteristics, repetitive extragenic palindromic PCR fragment pattern, and some biochemical tests. Key biochemical differences included the findings that Fx1 was positive for beta-galactosidase and arabinose hydrolysis and that both strains were citrulline ureidase positive and glycerol negative. Commercial F. tularensis antiserum agglutinated stock F. tularensis strains but not Fx1, Fx2, F. tularensis biovar novicida, or F. philomiragia; serum from either patient failed to agglutinate or only weakly agglutinated commercial antigen but showed agglutination when tested against each patient's respective isolate. Fx1 and Fx2 produced beta-lactamase. Because of their good growth, negative serology, and biochemical profile, the organisms could be misidentified in the clinical laboratory if standard strategies or commercial identification systems are used.

Adult↗

PubMed1995

Integration in trigeminal premotor interneurones in the cat. 3. Input characteristics and synaptic actions of neurones in subnucleus-gamma of the oral nucleus of the spinal trigeminal tract with a projection to the masseteric motoneurone subnucleus.

A population of last-order interneurones within the rostrodorsal part of the oral nucleus of the spinal trigeminal tract (NVspo-gamma) has been investigated in 21 chloralose anaesthetised cats. The neurones were identified by their antidromic (AD) response to microstimulation (median current 9 microA, range 3-39 microA) of the ipsior the contralateral masseteric subnucleus of the trigeminal motor nucleus. Fifty-one of 113 interneurones tested were discharged from the ipsilateral and eight from the contralateral motor nucleus. The average conduction time was 0.50 ms from the ipsilateral and 0.74 ms from the contralateral motoneurone pool. Conduction velocities of the axons ranged from 2.0 to 14.0 ms. The pattern of primary afferent input onto the selected neurones was analysed by graded electrical stimulation of dissected trigeminal nerves. Low-threshold afferents innervating the intraoral mucosa including the tongue and the perioral skin of the lower lip were the most effective inputs, as judged from both the frequency of occurrence and from the latencies of the evoked spike discharges. Ninety-six percent of the neurones responded to stimulation of the inferior alveolar nerve (Alv inf) and 83% responded to stimulation of the lingual nerve (Ling). The median threshold strength required to evoke the Alv inf and the Ling responses was 1.7 T (range 1.0-3.6 T) and 1.3 T (range 1.0-5.0 T), respectively. The median latency to spike discharges evoked by the Alv inf was 2.0 ms (range 1.3-4.8 ms) and to the Ling it was 2.5 ms (range 1.4-7.0 ms). Action potentials elicited by stimulation of the masseteric and digastric nerves were observed in 40% and 10% of the neurones, respectively. These responses, which had median latencies of more than 8 ms (range 4.7-16.0 ms), were only seen at stimulation intensities above 2 T (range 2.5-25 T). An input from the maxillary whisker nerve was seen in only one case. Postspike averages of the extracellular field potentials within the trigeminal motoneurone subnuclei evoked by interneuronal spikes were made in a subsample of 51 NVspo-gamma neurones activated by iontophoresis of L-glutamic acid. Excitatory synaptic effects within the masseteric subnucleus were observed in eight cases. An inhibitory effect was seen in one case. One specific neurone gave an excitatory extracellular field potential within the digastric motoneurone subnucleus. This interneurone was AD activated from the digastric, but not from the masseteric subnucleus.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗

PubMed1995

Adjuvanticity of ISCOMs incorporating a T cell-reactive lipoprotein of the facultative intracellular pathogen Francisella tularensis.

Immunostimulating complexes (ISCOMs) are known to be highly effective adjuvants for envelope antigens of viral agents, but have not been evaluated for use with antigens of intracellular bacteria. Balb/c mice were subcutaneously immunized with ISCOMs into which the T cell-reactive membrane protein TUL4 of Francisella tularensis had been incorporated. Spleen cells from the immunized mice responded in vitro to TUL4 and to heat-killed F. tularensis live vaccine strain (LVS) with proliferation and production of gamma-interferon, whereas spleen cells from control mice immunized with TUL4 only did not respond to the antigens. When mice immunized with TUL4 ISCOMs were challenged with F. tularensis LVS, bacterial counts in spleen and liver were lower than in non-immunized mice. Again, TUL4 had no effect when used without ISCOMs. When proteins of a total membrane preparation of F. tularensis LVS were incorporated in ISCOMs and used for immunization, a decrease in bacterial counts was obtained which was similar in magnitude to that of TUL4 ISCOMs. Generally, the adjuvant effects demonstrated did not compare with the excellent protective effect of live tularaemia vaccine. Nonetheless, ISCOMs provide a means whereby protective antigens of F. tularensis can be tested.

Adjuvants, Immunologic↗

PubMed1995

Cytokine expression in the liver during the early phase of murine tularemia.

Cytokine expression was determined in the livers of mice inoculated subcutaneously with Francisella tularensis LVS. During the first 48 h of infection, there was a logarithmic increase of bacteria in the liver, with a doubling time of 2.5 h. Within 48 h, tumor necrosis factor alpha (TNF-alpha), interleukin 10 (IL-10), IL-12, and gamma interferon (IFN-gamma) mRNAs were expressed, and production of TNF-alpha and IFN-gamma was demonstrated. There was no expression within 96 h of mRNA from IL-2, IL-3, or IL-4. After subcutaneous inoculation of heat-killed LVS, no expression of any of the cytokine mRNAs and no increase in the levels of TNF-alpha or IFN-gamma occurred. The expression of TNF-alpha, IL-12, and IFN-gamma is held to be important to evoke an early T-cell-independent host defense against F. tularensis as well as to drive the expansion of a protective Th1 cell response.

Animals↗

PubMed1995

Perceived sound quality of three bone-anchored hearing aid models.

Eighteen patients judged the sound quality of the new Classic 300 and the existing HC 100 or HC 200 on 4 five-stage sound quality scales. Two of the scales relate to the perceptual dimensions softness and clarity, one relates to interference or noise and one relates to the overall impression. The evaluations were made, based on a questionnaire, in the patients' own homes and comprised the listening situations: TV news, music, conversation with two to three people in a noisy environment and one optional choice. The patients also stated which hearing aid was best in each situation and made a confidence rating for their selection. The patients were accustomed to wearing the HC 100 or HC 200. In these comparisons of the different bone-anchored hearing aid models, the Classic 300 received the best ratings overall. Seventeen patients changed their previously fitted hearing aid to a Classic 300 and satisfactory sound quality was undoubtedly one of the principal reasons. When it came to the confidence rating for their selections, some two-thirds of the patients stated that it was without hesitation or with little hesitation. Our overall conclusion is that the sound quality of bone-anchored hearing aids is satisfactory when it comes to clarity and overall impression. However, it may be expected that these patients with conductive losses get a better sound quality than other patients with sensorineural losses if the same type of aid is used.

Adolescent↗

PubMed1994

The tularaemia vaccine.

Tularaemia is a disease caused by the facultative intracellular bacterium Francisella tularensis. Vaccination resulting in protective immunity is induced by live vaccine only. Such vaccination can be performed by scarification utilizing the live vaccine strain of F. tularensis (F. tularensis LVS), which results in good but not complete protection. Humoral as well as cell-mediated immunity are induced by vaccination and it has been shown that cell-mediated immunity is a prerequisite for protection. Since the live vaccine strain is attenuated and the genetic background of attenuation is unknown it is important to consider process parameters so that the immunogenicity of the vaccine is preserved.

Animals↗

PubMed1994

Persistence of cell-mediated immunity and decline of humoral immunity to the intracellular bacterium Francisella tularensis 25 years after natural infection.

The humoral and cell-mediated immune responses to the facultative intracellular pathogen Francisella tularensis were studied in subjects who had tularemia 25 years earlier. Serum agglutinin titers were low. Of 53 subjects, only 2 had a titer > 40. The F. tularensis-induced T cell response, on the other hand, was vigorous and in fact similar in magnitude to the response to purified protein derivative (PPD) of Mycobacterium tuberculosis. T cells from 44 of 52 subjects showed a significant (P < .05) proliferative response in vitro to each of four membrane proteins of F. tularensis. Only 2 subjects did not respond to any of the proteins. In most cultures from 18 subjects tested, interferon-gamma was produced in response to the proteins. During the last 25 years, tularemia has been reported only rarely in the region where the subjects lived, suggesting that an antigen-specific T cell response may persist after tularemia independently of reexposure to F. tularensis.

Adult↗

PubMed1994

Analysis of 16S ribosomal DNA sequences of Francisella strains and utilization for determination of the phylogeny of the genus and for identification of strains by PCR.

The 16S ribosomal DNAs (rDNAs) of two strains of Francisella tularensis and one strain of Francisella philomiragia were sequenced. On the basis of phylogenetic analysis data, the genus Francisella was placed in the gamma subclass of the Proteobacteria. The most closely related organism was the intracellular bacterium Wolbachia persica. The sequenced 16S rDNA molecules of the Francisella species exhibited very high levels of similarity (98.5 to 99.9%). Two variable regions, comprising 390 to 450 nucleotides of the 16S rDNA molecules of 17 additional Francisella strains, including members of the species F. tularensis and F. philomiragia, were also sequenced. At most, six nucleotide differences were observed among the sequences of the F. tularensis strains. The sequence of Francisella novicida was virtually identical to the sequences of the F. tularensis strains, thereby supporting the hypothesis that these organisms are members of the same species. On the basis of the observed differences, primer pairs were designed to distinguish strains by using the PCR at the genus, species, and subspecies levels. This permitted sensitive identification of strains belonging to the genus Francisella and discrimination of the species F. tularensis and F. philomiragia.

Base Sequence↗

PubMed1994

Increased synthesis of DnaK, GroEL, and GroES homologs by Francisella tularensis LVS in response to heat and hydrogen peroxide.

The response of the facultative intracellular bacterium Francisella tularensis LVS to stress was assayed by pulse-labeling with [35S]methionine followed by two-dimensional gel electrophoresis and autoradiography. A temperature increase from 37 to 42 degrees C or exposure to 5 mM hydrogen peroxide induced increased syntheses of at least 15 proteins. Among these proteins were a 75-, a 60-, and a 10-kDa protein. By N-terminal sequence analysis, these three proteins were found to be extensively homologous to the highly conserved chaperone proteins DnaK, GroEL, and GroES of Escherichia coli. Antibodies specific to the DnaK homolog of E. coli reacted with the 75-kDa protein, and antibodies to the GroEL homolog of Legionella micdadei reacted with the 60-kDa protein. A readiness to respond to hydrogen peroxide with synthesis of the chaperone components may be fundamental to the intracellular survival of pathogens such as F. tularensis, which are exposed to oxidative stress while invading the host macrophages.

Amino Acid Sequence↗

PubMed1993

Identification and classification of different isolates of Francisella tularensis.

The causative agent of tularemia, Francisella tularensis, occurs in two main biovars, the highly virulent F. t. biovar tularensis, found in North America; and the less virulent biovar palaearctica, found all over the northern hemisphere. Two other biovars have been proposed, F. t. biovar mediaasiatica and F. t. biovar palaearctica var. japonica. In Sweden tularemia is most frequently observed in man and varying hares (Lepus timidus), and occasionally in other species. Tularemia in hares is normally an acute fatal disease, although less fatal infections have been reported. The diagnosis of tularemia is routinely based on immunological reactions. We studied 10 different isolates of F. tularensis from varying hares, one isolate from an Ural owl (Strix uralensis), one vaccine strain, one strain of F. t. biovar japonica, and six isolates from a virulence study of F. tularensis, by biochemical tests and by hybridization experiments with probes complementary to 16S rRNA. All isolates, except the isolate F. t. biovar japonica, were characterized as F. t. biovar palaearctica by biochemical tests. In the 16S rRNA analysis all isolates were positive to the probe for Francisella tularensis and the probe for F. t. biovar palaearctica with the exception that F. t. biovar japonica reacted with the probe specific to F. t. biovar tularensis. To further confirm that the strains used belonged to F. t. biovar palaearctica virulence tests in rabbits were performed which disclosed this phenotype. The results presented in this work show that the isolated strains from the western part of Europe were F. t. biovar palaearctica, irrespective of animal origin or virulence.

Animals↗

PubMed1992

The 17 kDa lipoprotein and encoding gene of Francisella tularensis LVS are conserved in strains of Francisella tularensis.

A T-cell-stimulating 17 kDa protein of the vaccine strain Francisella tularensis LVS has previously been cloned, sequenced and shown to be a lipoprotein. In the present study, it was investigated whether the protein, denoted TUL4, and its gene are present in various strains of the genus Francisella. By Western blot analysis, it was demonstrated that a TUL4-specific monoclonal antibody bound to a protein present in each of the Francisella strains. The immunoreactive proteins had an M(r) of 17 kDa in all F. tularensis strains and in the strain Francisella novicida, whereas the M(r) in strains of Francisella philomiragia was 20 kDa. When genomic preparations were probed with a radioactive DNA fragment of F. tularensis LVS encoding TUL4, hybridization was demonstrated in all strains of Francisella, although the F. philomiragia strains did not hybridize under conditions of high stringency. The hybridizing chromosomal DNA fragment of the F. philomiragia strains was larger than that of the other Francisella strains. No hybridization or Western blot reactivity was seen when various other Gram-negative and Gram-positive bacteria were probed. In summary, the 17 kDa lipoprotein of F. tularensis LVS appears to be Francisella-specific and present in the species F. tularensis and F. novicida, whereas an immunologically related protein is present in F. philomiragia.

Bacterial Proteins↗

PubMed1992

Immunogenicity and toxicity of lipopolysaccharide from Francisella tularensis LVS.

Lipopolysaccharide (LPS) from the live vaccine strain of Francisella tularensis (F. tularensis LVS) was isolated and purified. The LPS did not stimulate lymphocytes from previously tularaemia-vaccinated individuals or lymphocytes from non-primed individuals. However, serum antibodies from tularaemia vaccinees reacted with the LPS whereas virtually no reactivity was found with antibodies from individuals not exposed to F. tularensis LVS. Antibodies of immunoglobulin class M displayed the antibody reactivity predominantly. The LPS failed to induce the mononuclear cell-derived cytokine interleukin-1 and only low levels of tumour necrosis factor were detected. Furthermore, no LPS endotoxin properties were found in galactosamine-treated mice or in the Limulus amoebocyte lysate assay. From these results it can be concluded that F. tularensis LVS possesses a lipopolysaccharide-like molecule, which does not exhibit properties of a classical endotoxin.

Animals↗

PubMed1992

Humoral and cell-mediated immunity in mice to a 17-kilodalton lipoprotein of Francisella tularensis expressed by Salmonella typhimurium.

A 17-kDa lipoprotein, TUL4, of the facultative intracellular bacterium Francisella tularensis is one of several membrane proteins that induce an in vitro response in T cells from F. tularensis-primed humans. A DNA fragment of the live vaccine strain F. tularensis LVS encoding TUL4 was cloned into Salmonella typhimurium chi 4072, an attenuated delta cya delta crp mutant. Expression of the protein by the recombinant S. typhimurium chi 4072 (pTUL4-15) was maintained after passage in BALB/cJ mice. When mice were immunized with S. typhimurium chi 4072(pTUL4-15), some animals showed an antibody response and a T-cell response to TUL4. When the immunized mice were challenged with the live vaccine strain F. tularensis LVS, bacterial counts in the liver and spleen were lower than in animals immunized with S. typhimurium chi 4072. Immunization with F. tularensis LVS caused a much stronger protection against the challenge than did immunization with S. typhimurium chi 4072(pTUL4-15). The present study demonstrated that the 17-kDa lipoprotein TUL4 of F. tularensis is involved in a protective immunity to tularemia. Possibly, several T-cell-reactive proteins of the organism have to contribute for optimal protection to be achieved.

Amino Acid Sequence↗

PubMed1992

Characterization and classification of strains of Francisella tularensis isolated in the central Asian focus of the Soviet Union and in Japan.

The two subspecies of Francisella tularensis, F. tularensis subsp. tularensis (type A) and F. tularensis subsp. palaearctica (type B), differ from each other in biochemistry and virulence. Strains of F. tularensis subsp. tularensis are believed to be confined to North America, whereas strains of F. tularensis subsp. palaearctica occur in Europe, in Asia, and in North America. Moreover, the existence of two other subspecies, designated F. tularensis subsp. mediaasiatica and F. tularensis subsp. palaearcitica japonica, has been suggested for strains of F. tularensis isolated in the central Asian focus of the Soviet Union and in Japan, respectively. In the present study, strains biochemically classified as F. tularensis subsp. mediaasiatica or F. tularensis subsp. palaearctica japonica have been investigated by hybridization with probes specific to 16S rRNAs of the two main subspecies. Furthermore, the virulence and biochemical characteristics of the strains were compared with those of strains belonging to F. tularensis subsp. palaearctica and F. tularensis subsp. tularensis. It was found that 16S rRNAs of F. tularensis subsp. mediaasiatica and F. tularensis subsp. palaearctica japonica hybridize with the probe specific to a genotype proposed herein, genotype A (F. tularensis subsp. tularensis), which shows that strains genetically related to this subspecies are found outside North America. However, the central Asian strains differed from F. tularensis subsp. palaearctica and F. tularensis subsp. tularensis strains when investigated by fermentation of glucose. The results of the biochemical tests could not be unambiguously used for differentiation of strains into F. tularensis subsp. palaearctica or F. tularensis subsp. tularensis. These drawbacks suggest that classification of strains of Francisella on the basis of 16S rRNA analysis may be preferable to classification on the basis of biochemical analysis.

Animals↗

PubMed1992

Various membrane proteins of Francisella tularensis induce interferon-gamma production in both CD4+ and CD8+ T cells of primed humans.

Tularaemia is an intracellular infection, which is controlled by the host as a result of an immunospecific T-cell response. A crucial product of the responding T cells is interferon-gamma (IFN-gamma), which acts by enhancing the microbicidal activity of macrophages. T cells of tularaemia-vaccinated individuals respond in vitro to a multitude of protein antigens of the vaccine strain Francisella tularensis LVS. In the present study, the responses to four of these antigens were shown to be confined mostly to the CD45RO+ memory T-cell subset. To characterize further the phenotype of the responding cells, purified CD4+ and CD8+ T cells were stimulated with the antigens. CD4+ T cells, but not CD8+ T cells, proliferated and produced IFN-gamma. However, when CD8+ T cells were isolated from bulk cultures of lymphocytes, which had been stimulated with antigen for 3 days, they responded to an extent similar to that of CD4+ T cells. Purified CD8+ T cells also responded when they were supplemented with interleukin-2 (IL-2). There was a direct quantitative correlation between the proliferative response of CD4+ and CD8+ T cells and their production of IFN-gamma. IL-2 was produced in the cultures, the amounts being higher in the cultures of CD4+ than in those of CD8+ cells. IL-4 was not detected in the culture medium of any of the T-cell subsets. Seventeen human alpha beta + CD4+ CD8- CD3+ T-cell clones, specific to antigens of F. tularensis, were raised. When proliferating, these clones did invariably produce IL-2 and IFN-gamma but no IL-4. In conclusion, both CD4+ and CD8+ T cells of tularaemia-vaccinated individuals respond with proliferation to various protein antigens of F. tularensis, and the proliferative response is strictly associated with IFN-gamma production. The CD8+ T-cell response seems to depend on cytokines supplied by proliferating CD4+ T cells.

Antigens, Bacterial↗