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| 2. |
- Desvars, Amélie, 1980-, et al.
(författare)
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Epidemiology and Ecology of Tularemia in Sweden, 1984-2012
- 2015
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Ingår i: Emerging Infectious Diseases. - : Centers for Disease Control and Prevention (CDC). - 1080-6040 .- 1080-6059. ; 21:1, s. 32-39
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Tidskriftsartikel (refereegranskat)abstract
- The zoonotic disease tularemia is endemic in large areas of the Northern Hemisphere, but research is lacking on patterns of spatial distribution and connections with ecologic factors. To describe the spatial epidemiology of and identify ecologic risk factors for tularemia incidence in Sweden, we analyzed surveillance data collected over 29 years (1984-2012). A total of 4,830 cases were notified, of which 3,524 met all study inclusion criteria. From the first to the second half of the study period, mean incidence increased 10-fold, from 0.26/100,000 persons during 1984-1998 to 2.47/100,000 persons during 1999 2012 (p<0.001). The incidence of tularemia was higher than expected in the boreal and alpine ecologic regions (p<0.001), and incidence was positively correlated with the presence of lakes and rivers (p<0.001). These results provide a comprehensive epidemiologic description of tularemia in Sweden and illustrate that incidence is higher in locations near lakes and rivers.
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| 3. |
- Edin, Alicia, et al.
(författare)
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Development and Laboratory Evaluation of a Real-Time PCR Assay for Detecting Viruses and Bacteria of Relevance for Community-Acquired Pneumonia
- 2015
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Ingår i: Journal of Molecular Diagnostics. - : Elsevier BV. - 1525-1578 .- 1943-7811. ; 17:3, s. 315-324
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Tidskriftsartikel (refereegranskat)abstract
- Community-acquired pneumonia may present with similar clinical symptoms, regardless of viral or bacterial cause. Diagnostic assays are needed to rapidly discriminate between causes, because this will guide decisions on appropriate treatment. Therefore, a quantitative real-time PCR (qPCR) assay with duplex reactions targeting eight bacteria and six viruses was developed. Technical performance was examined with linear plasmids. Upper and Lower respiratory tract specimens were used to compare the qPCR assay with standard microbiological methods. The limit of detection was 5 to 20 DNA template copies with approximately 1000-fold differences in concentrations of the two competing templates. SDs for positive controls were <5%. The use of the qPCR assay resulted in 113 positive identifications in 94 respiratory specimens compared with 38 by using standard diagnostics. Diagnostic accuracy of the qPCR assay varied between 60% positive agreement with standard tests for Streptococcus pneumoniae and 100% for Mycoplasma pneumoniae, Moraxella catarrhalis, and Staphylococcus aureus. Negative percentage of agreement was >95% for M. pneumoniae, Streptococcus pyogenes, respiratory syncytial virus, and influenza A virus; whereas it was only 56% for Haemophilus influenzae. Multiple microbial agents were identified in 19 of 44 sputum and 19 of 50 nasopharynx specimens. We conclude that in parallel qPCR detection of the targeted respiratory bacteria and viruses is feasible. The results indicate good technical performance of the assay in clinical specimens.
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| 4. |
- Edin, Alicia, 1985-
(författare)
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Improved diagnosis and prediction of community-acquired pneumonia
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Community-acquired pneumonia (CAP) is a major cause of morbidity and mortality worldwide. Although there is wide variation in the microbial etiology, CAP may manifest with similar symptoms, making institution of proper treatment challenging. Therefore, etiological diagnosis is important to ensure that correct treatment and necessary infection control measures are instituted. This provides a challenge for conventional microbial diagnostic methods, typically based on culture and direct antigen tests. Moreover, existing molecular biomarkers have poor prognostic value. Few studies have investigated the global metabolic response during infection and virtually nothing is known about early responses after the start of antimicrobial treatment. The aim of this work was to improve diagnostic and predictive methods for CAP.In paper I, a qPCR panel targeting 15 pathogens known to cause CAP was developed and evaluated. It combined identification of bacterial pathogens and viruses in the same diagnostic platform. The method proved to be robust and the results consistent with those obtained by standard methods. The panel approach, compared to conventional, selective diagnostics, detected a larger number of pathogens. In Paper II, whole blood samples from 65 patients with bacteremic sepsis were analyzed for metabolite profiles. Forty-nine patients with symptoms of sepsis, but later attributed to other diagnoses, were matched according to age and sex and served as a control group. Six metabolites were identified, all of which predicted growth of bacteria in blood culture. One of the metabolites, myristic acid, alone predicted bacteremic sepsis with a sensitivity of 100% and a specificity of 95%. Paper III and IV were based on a clinical study enrolling 35 patients with suspected CAP in need of hospital care. The aim was to study the metabolic response during the early phase of acute infection. The qPCR panel developed in Paper I was used to obtain the microbial etiological diagnosis. Paper IV focused on the global metabolic response and highlighted the dynamics of changes in major metabolic pathways during early recovery. A specific metabolite pattern for M. pneumoniae etiology was found. Four metabolites accurately predicted all but one patient as either M. pneumoniae etiology or not. Paper III looked at phospholipid levels during the first 48 hours after hospital admission. It was found that all major phospholipid species, especially the lysophosphatidyl-cholines, were pronouncedly decreased during acute infection. Levels started to increase the day after admission, reaching statistical significance at 48 hours. Paper II-IV showed that metabolomics might be used to study a number of different aspects of infection, such as etiology, disease progress and recovery. Knowledge of the metabolic profiles of patients may not only be utilized for biomarker discovery, as proposed in this work, but also for the future development of targeted therapies and supportive treatment.
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| 5. |
- Kauppi, Anna M., et al.
(författare)
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Metabolites in Blood for Prediction of Bacteremic Sepsis in the Emergency Room
- 2016
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Ingår i: PLOS ONE. - : Public Library Science. - 1932-6203. ; 11:1
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Tidskriftsartikel (refereegranskat)abstract
- A metabolomics approach for prediction of bacteremic sepsis in patients in the emergency room (ER) was investigated. In a prospective study, whole blood samples from 65 patients with bacteremic sepsis and 49 ER controls were compared. The blood samples were analyzed using gas chromatography coupled to time-of-flight mass spectrometry. Multivariate and logistic regression modeling using metabolites identified by chromatography or using conventional laboratory parameters and clinical scores of infection were employed. A predictive model of bacteremic sepsis with 107 metabolites was developed and validated. The number of metabolites was reduced stepwise until identifying a set of 6 predictive metabolites. A 6-metabolite predictive logistic regression model showed a sensitivity of 0.91(95% CI 0.69-0.99) and a specificity 0.84 (95% CI 0.58-0.94) with an AUC of 0.93 (95% CI 0.89-1.01). Myristic acid was the single most predictive metabolite, with a sensitivity of 1.00 (95% CI 0.85-1.00) and specificity of 0.95 (95% CI 0.74-0.99), and performed better than various combinations of conventional laboratory and clinical parameters. We found that a metabolomics approach for analysis of acute blood samples was useful for identification of patients with bacteremic sepsis. Metabolomics should be further evaluated as a new tool for infection diagnostics.
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| 6. |
- Müller, Daniel C., et al.
(författare)
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Phospholipid Levels in Blood during Community-Acquired Pneumonia
- 2019
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Ingår i: PLOS ONE. - : Public Library of Science. - 1932-6203. ; 14:5
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Tidskriftsartikel (refereegranskat)abstract
- Phospholipids, major constituents of bilayer cell membranes, are present in large amounts in pulmonary surfactant and play key roles in cell signaling. Here, we aim at finding clinically useful disease markers in community-acquired pneumonia (CAP) using comprehensive phospholipid profiling in blood and modeling of changes between sampling time points. Serum samples from 33 patients hospitalized with CAP were collected at admission, three hours after the start of intravenous antibiotics, Day 1 (at 12–24 h), Day 2 (at 36–48 h), and several weeks after recovery. A profile of 75 phospholipid species including quantification of the bioactive lysophosphatidylcholines (LPCs) was determined using liquid chromatography coupled to time-of-flight mass spectrometry. To control for possible enzymatic degradation of LPCs, serum autotaxin levels were examined. Twenty-two of the 33 patients with a clinical diagnosis of CAP received a laboratory-verified CAP diagnosis by microbial culture or microbial DNA detection by qPCR. All major phospholipid species, especially the LPCs, were pronouncedly decreased in the acute stage of illness. Total and individual LPC concentrations increased shortly after the initiation of antibiotic treatment, concentrations were at their lowest 3h after the initiation, and increased after Day 1. The total LPC concentration increased by a change ratio of 1.6–1.7 between acute illness and Day 2, and by a ratio of 3.7 between acute illness and full disease resolution. Autotaxin levels were low in acute illness and showed little changes over time, contradicting a hypothesis of enzymatic degradation causing the low levels of LPCs. In this sample of patients with CAP, the results demonstrate that LPC concentration changes in serum of patients with CAP closely mirrored the early transition from acute illness to recovery after the initiation of antibiotics. LPCs should be further explored as potential disease stage biomarkers in CAP and for their potential physiological role during recovery.
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| 7. |
- Alam, Athar, et al.
(författare)
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ClpB mutants of Francisella tularensis subspecies holarctica and tularensis are defective for type VI secretion and intracellular replication
- 2018
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Ingår i: Scientific Reports. - : Nature Publishing Group. - 2045-2322. ; 8
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Tidskriftsartikel (refereegranskat)abstract
- Francisella tularensis, a highly infectious, intracellular bacterium possesses an atypical type VI secretion system (T6SS), which is essential for the virulence of the bacterium. Recent data suggest that the HSP100 family member, ClpB, is involved in T6SS disassembly in the subspecies Francisella novicida. Here, we investigated the role of ClpB for the function of the T6SS and for phenotypic characteristics of the human pathogenic subspecies holarctica and tularensis. The Delta clpB mutants of the human live vaccine strain, LVS, belonging to subspecies holarctica, and the highly virulent SCHU S4 strain, belonging to subspecies tularensis, both showed extreme susceptibility to heat shock and low pH, severely impaired type VI secretion (T6S), and significant, but impaired intracellular replication compared to the wild-type strains. Moreover, they showed essentially intact phagosomal escape. Infection of mice demonstrated that both Delta clpB mutants were highly attenuated, but the SCHU S4 mutant showed more effective replication than the LVS strain. Collectively, our data demonstrate that ClpB performs multiple functions in the F. tularensis subspecies holarctica and tularensis and its function is important for T6S, intracellular replication, and virulence.
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| 8. |
- Binesse, Johan, et al.
(författare)
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Roles of Reactive Oxygen Species-Degrading Enzymes of Francisella tularensis SCHU S4
- 2015
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Ingår i: Infection and Immunity. - 0019-9567 .- 1098-5522. ; 83:6, s. 2255-2263
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Tidskriftsartikel (refereegranskat)abstract
- Francisella tularensis is a facultative intracellular bacterium utilizing macrophages as its primary intracellular habitat and is therefore highly capable of resisting the effects of reactive oxygen species (ROS), potent mediators of the bactericidal activity of macrophages. We investigated the roles of enzymes presumed to be important for protection against ROS. Four mutants of the highly virulent SCHU S4 strain with deletions of the genes encoding catalase (katG), glutathione peroxidase (gpx), a DyP-type peroxidase (FTT0086), or double deletion of FTT0086 and katG showed much increased susceptibility to hydrogen peroxide (H2O2) and slightly increased susceptibility to paraquat but not to peroxynitrite (ONOO-) and displayed intact intramacrophage replication. Nevertheless, mice infected with the double deletion mutant showed significantly longer survival than SCHU S4-infected mice. Unlike the aforementioned mutants, deletion of the gene coding for alkyl-hydroperoxide reductase subunit C (ahpC) generated a mutant much more susceptible to paraquat and ONOO- but not to H2O2. It showed intact replication in J774 cells but impaired replication in bone marrow-derived macrophages and in internal organs of mice. The live vaccine strain, LVS, is more susceptible than virulent strains to ROS-mediated killing and possesses a truncated form of FTT0086. Expression of the SCHU S4 FTT0086 gene rendered LVS more resistant to H2O2, which demonstrates that the SCHU S4 strain possesses additional detoxifying mechanisms. Collectively, the results demonstrate that SCHU S4 ROS-detoxifying enzymes have overlapping functions, and therefore, deletion of one or the other does not critically impair the intracellular replication or virulence, although AhpC appears to have a unique function.
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| 9. |
- Bröms, Jeanette E., et al.
(författare)
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A mutagenesis-based approach identifies amino acids in the N-terminal part of Francisella tularensis IglE that critically control type VI system-mediated secretion
- 2017
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Ingår i: Virulence. - : TAYLOR & FRANCIS INC. - 2150-5594 .- 2150-5608. ; 8:6, s. 821-847
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Tidskriftsartikel (refereegranskat)abstract
- The Gram-negative bacterium Francisella tularensis is the etiological agent of the zoonotic disease tularemia. Its life cycle is characterized by an ability to survive within phagocytic cells through phagosomal escape and replication in the cytosol, ultimately causing inflammasome activation and host cell death. Required for these processes is the Francisella Pathogenicity Island (FPI), which encodes a Type VI secretion system (T6SS) that is active during intracellular infection. In this study, we analyzed the role of the FPI-component IglE, a lipoprotein which we previously have shown to be secreted in a T6SS-dependent manner. We demonstrate that in F. tularensis LVS, IglE is an outer membrane protein. Upon infection of J774 cells, an Delta iglE mutant failed to escape from phagosomes, and subsequently, to multiply and cause cytopathogenicity. Moreover, Delta iglE was unable to activate the inflammasome, to inhibit LPS-stimulated secretion of TNF-alpha, and showed marked attenuation in the mouse model. In F. novicida, IglE was required for in vitro secretion of IglC and VgrG. A mutagenesis-based approach involving frameshift mutations and alanine substitution mutations within the first similar to 38 residues of IglE revealed that drastic changes in the sequence of the extreme N-terminus (residues 2-6) were well tolerated and, intriguingly, caused hyper-secretion of IglE during intracellular infection, while even subtle mutations further downstream lead to impaired protein function. Taken together, this study highlights the importance of IglE in F. tularensis pathogenicity, and the contribution of the N-terminus for all of the above mentioned processes.
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| 10. |
- Eneslätt, Kjell, et al.
(författare)
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Vaccine-mediated mechanisms controlling replication of Francisella tularensis in human peripheral blood mononuclear cells using a co-culture system
- 2018
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Ingår i: Frontiers in Cellular and Infection Microbiology. - : Frontiers Media S.A.. - 2235-2988. ; 8
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Tidskriftsartikel (refereegranskat)abstract
- Cell-mediated immunity (CMI) is normally required for efficient protection against intracellular infections, however, identification of correlates is challenging and they are generally lacking. Francisella tularensis is a highly virulent, facultative intracellular bacterium and CMI is critically required for protection against the pathogen, but how this is effectuated in humans is poorly understood. To understand the protective mechanisms, we established an in vitro co-culture assay to identify how control of infection of F. tularensis is accomplished by human cells and hypothesized that the model will mimic in vivo immune mechanisms. Non-adherent peripheral blood mononuclear cells (PBMCs) were expanded with antigen and added to cultures with adherent PBMC infected with the human vaccine strain, LVS, or the highly virulent SCHU S4 strain. Intracellular numbers of F. tularensis was followed for 72 h and secreted and intracellular cytokines were analyzed. Addition of PBMC expanded from naïve individuals, i.e., those with no record of immunization to F. tularensis, generally resulted in little or no control of intracellular bacterial growth, whereas addition of PBMC from a majority of F. tularensis-immune individuals executed static and sometimes cidal effects on intracellular bacteria. Regardless of infecting strain, statistical differences between the two groups were significant, P < 0.05. Secretion of 11 cytokines was analyzed after 72 h of infection and significant differences with regard to secretion of IFN-γ, TNF, and MIP-1β was observed between immune and naïve individuals for LVS-infected cultures. Also, in LVS-infected cultures, CD4 T cells from vaccinees, but not CD8 T cells, showed significantly higher expression of IFN-γ, MIP-1β, TNF, and CD107a than cells from naïve individuals. The co-culture system appears to identify correlates of immunity that are relevant for the understanding of mechanisms of the protective host immunity to F. tularensis.
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