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  • 301.
    Lindgren, Marie
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Bröms, Jeanette E.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Meyer, Lena
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Golovliov, Igor
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    The Francisella tularensis LVS ΔpdpC mutant exhibits a unique phenotype during intracellular infection2013In: BMC Microbiology, ISSN 1471-2180, E-ISSN 1471-2180, Vol. 13, article id 20Article in journal (Refereed)
    Abstract [en]

    Background: A prerequisite for the virulence of the facultative intracellular bacterium Francisella tularensis is effective intramacrophage proliferation, which is preceded by phagosomal escape into the cytosol, and ultimately leads to host cell death. Many components essential for the intracellular life cycle are encoded by a gene cluster, the Francisella pathogenicity island (FPI), constituting a type VI secretion system.

    Results: We characterized the FPI mutant ΔpdpC of the live vaccine strain (LVS) of F. tularensis and found that it exhibited lack of intracellular replication, incomplete phagosomal escape, and marked attenuation in the mouse model, however, unlike a phagosomally contained FPI mutant, it triggered secretion of IL-1β, albeit lower than LVS, and markedly induced LDH release.

    Conclusions: The phenotype of the ΔpdpC mutant appears to be unique compared to previously described F. tularensis FPI mutants.

  • 302.
    Lindgren, Marie
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Eneslätt, Kjell
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Bröms, Jeanette
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Importance of PdpC, IglC, IglI, and IglG for Modulation of a Host Cell Death Pathway Induced by Francisella tularensis2013In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 81, no 6, p. 2076-2084Article in journal (Refereed)
    Abstract [en]

    Modulation of host cell death pathways appears to be a prerequisite for the successful lifestyles of many intracellular pathogens. The facultative intracellular bacterium Francisella tularensis is highly pathogenic, and effective proliferation in the macrophage cytosol leading to host cell death is a requirement for its virulence. To better understand the prerequisites of this cell death, macrophages were infected with the F. tularensis live vaccine strain (LVS), and the effects were compared to those resulting from infections with deletion mutants lacking expression of either of the pdpC, iglC, iglG, or iglI genes, which encode components of the Francisella pathogenicity island (FPI), a type VI secretion system. Within 12 h, a majority of the J774 cells infected with the LVS strain showed production of mitochondrial superoxide and, after 24 h, marked signs of mitochondrial damage, caspase-9 and caspase-3 activation, phosphatidylserine expression, nucleosome formation, and membrane leakage. In contrast, neither of these events occurred after infection with the Delta iglI or Delta iglC mutants, although the former strain replicated. The Delta iglG mutant replicated effectively but induced only marginal cytopathogenic effects after 24 h and intermediate effects after 48 h. In contrast, the Delta pdpC mutant showed no replication but induced marked mitochondrial superoxide production and mitochondrial damage, caspase-3 activation, nucleosome formation, and phosphatidylserine expression, although the effects were delayed compared to those obtained with LVS. The unique phenotypes of the mutants provide insights regarding the roles of individual FPI components for the modulation of the cytopathogenic effects resulting from the F. tularensis infection.

  • 303.
    Lindgren, Marie
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Tancred, Linda
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Golovlev, Igor
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Twine, Susan
    National Research Council of Canada, Institute for Biological Sciences, Ottawa, Ontario, Canada.
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Characterization of the spontaneous mutant FSC043 of Francisella tularensis subspecies tularensisManuscript (preprint) (Other academic)
  • 304.
    Lindgren, Marie
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Tancred, Linda
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Golovliov, Igor
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Conlan, Wayne
    Twine, Susan M.
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Identification of Mechanisms for Attenuation of the FSC043 Mutant of Francisella tularensis SCHU S42014In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 82, no 9, p. 3622-3635Article in journal (Refereed)
    Abstract [en]

    Previously, we identified a spontaneous, essentially avirulent mutant, FSC043, of the highly virulent strain SCHU S4 of Francisella tularensis subsp. tularensis. We have now characterized the phenotype of the mutant and the mechanisms of its attenuation in more detail. Genetic and proteomic analyses revealed that the pdpE gene and most of the pdpC gene were very markedly downregulated and, as previously demonstrated, that the strain expressed partially deleted and fused fupA and fupB genes. FSC043 showed minimal intracellular replication and induced no cell cytotoxicity. The mutant showed delayed phagosomal escape; at 18 h, colocalization with LAMP-1 was 80%, indicating phagosomal localization, whereas the corresponding percentages for SCHU S4 and the Delta fupA mutant were < 10%. However, a small subset of the FSC043-infected cells contained up to 100 bacteria with LAMP-1 colocalization of around 30%. The unusual intracellular phenotype was similar to that of the Delta pdpC and Delta pdpC Delta pdpE mutants. Complementation of FSC043 with the intact fupA and fupB genes did not affect the phenotype, whereas complementation with the pdpC and pdpE genes restored intracellular replication and led to marked virulence. Even higher virulence was observed after complementation with both double-gene constructs. After immunization with the FSC043 strain, moderate protection against respiratory challenge with the SCHU S4 strain was observed. In summary, FSC043 showed a highly unusual intracellular phenotype, and based on our findings, we hypothesize that the mutation in the pdpC gene makes an essential contribution to the phenotype.

  • 305. Lindh, Markus V.
    et al.
    Pinhassi, Jarone
    Sensitivity of Bacterioplankton to Environmental Disturbance: A Review of Baltic Sea Field Studies and Experiments2018In: Frontiers in Marine Science, E-ISSN 2296-7745, Vol. 5, article id UNSP 361Article, review/survey (Refereed)
    Abstract [en]

    Bacterioplankton communities regulate energy and matter fluxes fundamental to all aquatic life. The Baltic Sea offers an outstanding ecosystem for interpreting causes and consequences of bacterioplankton community composition shifts resulting from environmental disturbance. Yet, a systematic synthesis of the composition of Baltic Sea bacterioplankton and their responses to natural or human-induced environmental perturbations is lacking. We review current research on Baltic Sea bacterioplankton dynamics in situ (48 articles) and in laboratory experiments (38 articles) carried out at a variety of spatiotemporal scales. In situ studies indicate that the salinity gradient sets the boundaries for bacterioplankton composition, whereas, regional environmental conditions at a within-basin scale, including the level of hypoxia and phytoplankton succession stages, may significantly tune the composition of bacterial communities. Also the experiments show that Baltic Sea bacteria are highly responsive to environmental conditions, with general influences of e.g. salinity, temperature and nutrients. Importantly, nine out of ten experiments that measured both bacterial community composition and some metabolic activities showed empirical support for the sensitivity scenario of bacteria - i.e., that environmental disturbance caused concomitant change in both community composition and community functioning. The lack of studies empirically testing the resilience scenario, i.e., experimental studies that incorporate the long-term temporal dimension, precludes conclusions about the potential prevalence of resilience of Baltic Sea bacterioplankton. We also outline outstanding questions emphasizing promising applications in incorporating bacterioplankton community dynamics into biogeochemical and food-web models and the lack of knowledge for deep-sea assemblages, particularly bacterioplankton structure-function relationships. This review emphasizes that bacterioplankton communities rapidly respond to natural and predicted human-induced environmental disturbance by altering their composition and metabolic activity. Unless bacterioplankton are resilient, such changes could have severe consequences for the regulation of microbial ecosystem services.

  • 306.
    Lindholm, Mark
    et al.
    Umeå University, Faculty of Medicine, Department of Odontology.
    Aung, Kyaw Min
    Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Wai, Sun Nyunt
    Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Oscarsson, Jan
    Umeå University, Faculty of Medicine, Department of Odontology.
    Role of OmpA1 and OmpA2 in Aggregatibacter actinomycetemcomitans and Aggregatibacter aphrophilus serum resistance2019In: Journal of Oral Microbiology, ISSN 2000-2297, E-ISSN 2000-2297, Vol. 11, no 1, article id 1536192Article in journal (Refereed)
    Abstract [en]

    Aggregatibacter actinomycetemcomitans and Aggregatibacter aphrophilus belong to the HACEK group of fastidious Gram-negative organisms, a recognized cause of infective endocarditis. A. actinomycetemcomitans is also implicated in aggressive forms of periodontitis. We demonstrated that A. aphrophilus strains, as A. actinomycetemcomitans are ubiquitously serum resistant. Both species encode two Outer membrane protein A paralogues, here denoted OmpA1 and OmpA2. As their respective pangenomes contain several OmpA1 and OmpA2 alleles, they represent potential genotypic markers. A naturally competent strain of A. actinomycetemcomitans and A. aphrophilus, respectively were used to elucidate if OmpA1 and OmpA2 contribute to serum resistance. Whereas OmpA1 was critical for survival of A. actinomycetemcomitans D7SS in 50% normal human serum (NHS), serum resistant ompA1 mutants were fortuitously obtained, expressing enhanced levels of OmpA2. Similarly, OmpA1 rather than OmpA2 was a major contributor to serum resistance of A. aphrophilus HK83. Far-Western blot revealed that OmpA1AA, OmpA2AA, and OmpA1AP can bind to C4-binding protein, an inhibitor of classical and mannose-binding lectin (MBL) complement activation. Indeed, ompA1 mutants were susceptible to these pathways, but also to alternative complement activation. This may at least partly reflect a compromised outer membrane integrity but is also consistent with alternative mechanisms involved in OmpA-mediated serum resistance.

  • 307.
    Lindkvist, Marie
    Umeå University, Faculty of Medicine, Clinical Microbiology, Infectious Diseases.
    Genetic and serologic characterization of a Swedish human hantavirus isolate2008Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Hantaviruses are found practically all over the world and cause hemorrhagic fevers in man. Each year about 150,000 people are hospitalized in these zoonotic infections which can be of two types: hemorrhagic fever with renal syndrome (HFRS) or hantavirus cardiopulmonary syndrome (HCPS), depending on the infecting virus. Hantavirus infections are emerging infectious diseases. That is, the number of reported cases of hantaviral disease is increasing, new hantaviruses are discovered continually, and already known hantaviruses are expected to spread to new areas. Therefore, knowledge and monitoring of these viruses are imperative from a public health perspective.

    In this thesis, the characterization of a local human Puumala (PUUV) virus isolate is described. Genetic and serological relationships to other hantaviruses are investigated and the viral protein interactions, critical for genome packaging and assembly, are studied. We found that the nucleotide and amino acid sequences of the local PUUV strains are significantly different from the PUUV prototype strain Sotkamo, a difference that indicates that there might be a risk of misdiagnosing PUUV infected patients when using reagents derived from the prototype strain. These data contributed to the introduction of locally derived diagnostic tools to the Laboratory of Clinical Virology at the Umeå University hospital, which is the reference centre for hantaviral diseases in Sweden. Furthermore, when studying the underlying mechanisms of genome packaging, we identified several regions and amino acids absolutely required for nucleocapsid protein interactions. Also, a region that appeared to regulate this interaction was discovered. Finally, the serological immune responses in DNA-vaccinated mice and PUUV infected patients were investigated. We found that the cross-reactive antibody response in vaccinated mice and in infected individuals was unique and independent of homologous titres. Furthermore, four immunodominant epitopes with specific cross-reactive characteristics were identified.

    Our findings have highlighted the complexity of the serological immune responses to hantavirus infections, and they emphasize the importance of customizing the diagnostic tools and performing clinical analyses on locally derived strains. In conclusion, we believe that these results are valuable in the development of new serological, genetic, and epidemiological tools.

  • 308.
    Lindmark, Barbro
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Rompikuntal, Pramod Kumar
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Vaitkevicius, Karolis
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Song, Tianyan
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Mizunoe, Yoshimitsu
    Uhlin, Bernt Eric
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Guerry, Patricia
    Wai, Sun Nyunt
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Outer membrane vesicle-mediated release of cytolethal distending toxin (CDT) from Campylobacter jejuni2009In: BMC Microbiology, ISSN 1471-2180, E-ISSN 1471-2180, Vol. 16, no 9, p. 220-Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Cytolethal distending toxin (CDT) is one of the well-characterized virulence factors of Campylobacter jejuni, but it is unknown how CDT becomes surface-exposed or is released from the bacterium to the surrounding environment.

    RESULTS: Our data suggest that CDT is secreted to the bacterial culture supernatant via outer membrane vesicles (OMVs) released from the bacteria. All three subunits (the CdtA, CdtB, and CdtC proteins) were detected by immunogold labeling and electron microscopy of OMVs. Subcellular fractionation of the bacteria indicated that, apart from the majority of CDT detected in the cytoplasmic compartment, appreciable amounts (20-50%) of the cellular pool of CDT proteins were present in the periplasmic compartment. In the bacterial culture supernatant, we found that a majority of the extracellular CDT was tightly associated with the OMVs. Isolated OMVs could exert the cell distending effects typical of CDT on a human intestinal cell line, indicating that CDT is present there in a biologically active form.

    CONCLUSION: Our results strongly suggest that the release of outer membrane vesicles is functioning as a route of C. jejuni to deliver all the subunits of CDT toxin (CdtA, CdtB, and CdtC) to the surrounding environment, including infected host tissue.

  • 309.
    Lindquist, Richard
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Överby, Anna K.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    The Role of Viperin in Antiflavivirus Responses2018In: DNA and Cell Biology, ISSN 1044-5498, E-ISSN 1557-7430, Vol. 37, no 9, p. 725-730Article in journal (Refereed)
    Abstract [en]

    Viperin is an interferon (IFN)-stimulated gene product, which is part of the first line of the intracellular response against viral infection. It is a potent antiviral protein, strongly upregulated after IFN-stimulation and virus infection. Viperin is antivirally active against many different viruses from different families and has been shown to inhibit several flaviviruses. Flaviviruses are an important group of arthropod-borne viruses that cause millions of infections annually. In this review, we focus on the recent advances of the antiviral mechanisms of viperin against these flaviviruses, both pointing to similarities and differences between viruses within the same genera.

  • 310. Lindqvist, Richard
    Brain region- and cell type-specific role of viperin in neurotropic flavivirus infectionManuscript (preprint) (Other academic)
  • 311.
    Lindqvist, Richard
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    The role of the type I interferons and viperin during neurotropic flavivirus infection2017Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Flaviviruses are globally distributed pathogens that cause millions of human infections annually. One of the most detrimental outcomes of flavivirus infection is encephalitis, which is caused by neurotropic flaviviruses such as West Nile virus (WNV), Japanese encephalitis virus (JEV), and Tick-borne encephalitis virus (TBEV). The type I interferons (IFNs) are powerful cytokines, and they are known as the first line of defense against viral infection. IFNs are expressed at low or undetectable levels at the basal state, but recognition of invading pathogens triggers a robust IFN response. After synthesis, IFN is secreted and acts in an autocrine or paracrine manner by binding to the interferon-α/β receptor (IFNAR) receptor, which is expressed on the surface of all nucleated cells. Binding to IFNAR mediates a downstream cascade that triggers expression of hundreds of interferon-stimulated genes (ISGs). Some ISGs express signaling molecules to amplify the response while others are potent antiviral proteins that can efficiently limit viral infection. The impact of the type I IFN response in tick-borne flavivirus infection was not previously known. We found that the type I IFN response was crucial for protection of mice against neurotropic infection with tick-borne flaviviruses such as TBEV and Langat virus (LGTV). The response was needed both in the periphery as well as in the central nervous system (CNS), as transgenic mice lacking either peripherally or CNS-located IFNAR both succumbed to LGTV infection. Although we found that the local IFN response within the CNS is essential for protection against lethal LGTV infection, the cells responsible for the local IFN production were not known.

    Astrocytes are one of the most abundant cell types within the CNS, but their role in neurotropic flavivirus infection was not fully characterized. In other viral infections, astrocytes are potent IFN producers, thus we were interested in characterizing the role of the type I IFN response in astrocytes during neurotropic flavivirus infection and its contribution to flavivirus pathogenesis. We found that upon flavivirus infection, astrocytes mount a strong type I IFN response that protects neighboring astrocytes from TBEV, JEV, WNV, and ZIKV infection. Furthermore, IFN signaling was found to protect astrocytes from TBEV-induced cytopathic effects. However, the ISGs that mediated these effects were not known.

    In vitro studies of viperin, which was discovered in 2001 as an ISG with broad antiviral activity, has shown strong antiviral activity against TBEV, but its role in vivo and mode of action in flavivirus infection was not known. Using mice deficient in viperin, we wanted to determine the role of viperin in flavivirus infection. We found that viperin plays a region-specific role in the brain by controlling LGTV replication in the olfactory bulb and cerebrum. Remarkably, viperin was able to inhibit TBEV replication in primary cortical neurons isolated from the cerebrum but not in granule cell neurons isolated from the cerebellum. Furthermore, IFN treatment failed to compensate for loss of viperin in cortical neurons, indicating that viperin might be the most important ISG against TBEV in cortical neurons. Interestingly, we also found that viperin is needed for the IFN-mediated antiviral response against WNV and ZIKV in cortical neurons. Thus, viperin showed broad but region-specific antiviral mechanisms against different flaviviruses.

    Although viperin has been shown to inhibit many viruses, the molecular antiviral mechanism is not clear and appears to differ between viruses. We performed a co-immunoprecipitation (CoIP) screen to identify TBEV proteins that could interact with viperin, and prM, E, NS2A, NS2B, and NS3 were identified. Interaction of viperin with NS3 resulted in degradation of the viral protein. We screened NS3 of JEV, yellow fever virus (YFV), ZIKV, and TBEV. Interestingly, although all NS3 proteins tested interacted with viperin, only those of ZIKV, and TBEV were significantly degraded by viperin. The degradation of NS3 correlated well with the antiviral activity of viperin, as only TBEV and ZIKV were inhibited.

    In summary, this work revealed the importance of the local type I IFN response in the brain during neurotropic infections by flaviviruses. We identified astrocytes to be an important IFN producer within the CNS during neurotropic flavivirus infection. Astrocytes release type I IFN quickly after viral infection, and this interferon protects neighboring neurons and astrocytes from infection. Furthermore, viperin, a very potent antiviral ISG, is highly expressed in astrocytes and it is essential for controlling viral replication and mediating viral clearance in both neurons and astrocytes of the cerebrum. We also found that viperin specifically targeted the NS3 proteins of TBEV and ZIKV for degradation.

  • 312.
    Lindqvist, Richard
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience, Clinical Neuroscience.
    Kurhade, Chaitanya
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Gilthorpe, Jonathan D.
    Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience.
    Överby, Anna K.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Cell-type- and region-specific restriction of neurotropic flavivirus infection by viperin2018In: Journal of Neuroinflammation, ISSN 1742-2094, E-ISSN 1742-2094, Vol. 15, article id 80Article in journal (Refereed)
    Abstract [en]

    Background: Flaviviruses are a group of diverse and emerging arboviruses and an immense global health problem. A number of flaviviruses are neurotropic, causing severe encephalitis and even death. Type I interferons (IFNs) are the first line of defense of the innate immune system against flavivirus infection. IFNs elicit the concerted action of numerous interferon-stimulated genes (ISGs) to restrict both virus infection and replication. Viperin (virus-inhibitory protein, endoplasmic reticulum-associated, IFN-inducible) is an ISG with broad-spectrum antiviral activity against multiple flaviviruses in vitro. Its activity in vivo restricts neurotropic infections to specific regions of the central nervous system (CNS). However, the cell types in which viperin activity is required are unknown. Here we have examined both the regional and cell-type specificity of viperin in the defense against infection by several model neurotropic flaviviruses.

    Methods: Viral burden and IFN induction were analyzed in vivo in wild-type and viperin(-/-) mice infected with Langat virus (LGTV). The effects of IFN pretreatment were tested in vitro in primary neural cultures from different brain regions in response to infection with tick-borne encephalitis virus (TBEV), West Nile virus (WNV), and Zika virus (ZIKV).

    Results: Viperin activity restricted nonlethal LGTV infection in the spleen and the olfactory bulb following infection via a peripheral route. Viperin activity was also necessary to restrict LGTV replication in the olfactory bulb and the cerebrum following CNS infection, but not in the cerebellum. In vitro, viperin could restrict TBEV replication in primary cortical neurons, but not in the cerebellar granule cell neurons. Interferon-induced viperin was also very important in primary cortical neurons to control TBEV, WNV, and ZIKV.

    Conclusions: Our findings show that viperin restricts replication of neurotropic flaviviruses in the CNS in a region- and cell-type-specific manner. The most important sites of activity are the olfactory bulb and cerebrum. Activity within the cerebrum is required in the cortical neurons in order to restrict spread. This study exemplifies cell type and regional diversity of the IFN response within the CNS and shows the importance of a potent broad-spectrum antiviral ISG.

  • 313.
    Lindqvist, Richard
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Upadhyay, Arunkumar S.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Överby, Anna K.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Tick-Borne Flaviviruses and the Type I Interferon Response2018In: Viruses, ISSN 1999-4915, E-ISSN 1999-4915, Vol. 10, no 7, article id 340Article, review/survey (Refereed)
    Abstract [en]

    Flaviviruses are globally distributed pathogens causing millions of human infections every year. Flaviviruses are arthropod-borne viruses and are mainly transmitted by either ticks or mosquitoes. Mosquito-borne flaviviruses and their interactions with the innate immune response have been well-studied and reviewed extensively, thus this review will discuss tick-borne flaviviruses and their interactions with the host innate immune response.

  • 314.
    Lindström, Mona
    et al.
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB).
    Tjust, Anton E.
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB). Umeå University, Faculty of Medicine, Department of Clinical Sciences, Ophthalmology.
    Domellöf, Fatima Pedrosa
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB). Umeå University, Faculty of Medicine, Department of Clinical Sciences, Ophthalmology.
    Pax7-Positive Cells/Satellite Cells in Human Extraocular Muscles2015In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, E-ISSN 1552-5783, Vol. 56, no 10, p. 6132-6143Article in journal (Refereed)
    Abstract [en]

    PURPOSE. We quantified and investigated the distribution of Pax7-positive cells/satellite cells (SCs) in the human extraocular muscles (EOMs). METHODS. An immunofluorescence multiple-marker method simultaneously combining two SC markers (Pax7, NCAM), detection of the basement membrane (laminin) and cell nuclei (4',6-diamidino-2-phenylindole [DAPI]), was used on the anterior, middle, and posterior portions of EOMs from five healthy donors. Pax7-positive cell and SC content, myonuclear content, myofiber cross-sectional area, and myonuclear domain were analyzed in single cross-sections. Between 3915 and 13,536 myofibers per muscle cross-section and myofibers from the entire EOM cross-section were analyzed for quantification of Pax7-positive cells per myofiber (Pax7/F).

    RESULTS. The number of Pax7/F in the human EOMs varies along the length of the muscle with twice as high Pax7/F in the anterior part of the EOMs, but within the range of what has been previously reported for normal adult limb muscles. Furthermore, there are Pax7-positive cells in positions other than the classical SC position and the myonuclear domain size of adult EOMs is noticeably smaller than that previously reported for other adult skeletal muscles.

    CONCLUSIONS. Previous data on differences in Pax7-positive cell/SC abundance between EOMs and limb muscles must be reconsidered and the characteristics of different Pax7-positive cell populations further investigated. Higher numbers of Pax7-positive cells in the anterior portion of the EOMs may have a bearing for strabismus surgery involving sectioning of the muscle fibers.

  • 315. Liu, Hui
    et al.
    Fero, Jutta B.
    Mendez, Melissa
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Carpenter, Beth M.
    Servetas, Stephanie L.
    Rahman, Arifur
    Goldman, Matthew D.
    Boren, Thomas
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Salama, Nina R.
    Merrell, D. Scott
    Dubois, Andre
    Analysis of a single Helicobacter pylori strain over a 10-year period in a primate model2015In: International Journal of Medical Microbiology, ISSN 1438-4221, E-ISSN 1618-0607, Vol. 305, no 3, p. 392-403Article in journal (Refereed)
    Abstract [en]

    Helicobacter pylori from different individuals exhibits substantial genetic diversity. However, the kinetics of bacterial diversification after infection with a single strain is poorly understood. We investigated evolution of H. pylori following long-term infection in the primate stomach; Rhesus macaques were infected with H. pylori strain USU101 and then followed for 10 years. H. pylori was regularly cultured from biopsies, and single colony isolates were analyzed. At 1-year, DNA fingerprinting showed that all output isolates were identical to the input strain; however, at 5-years, different H. pylori fingerprints were observed. Microarray-based comparative genomic hybridization revealed that long term persistence of USU101 in the macaque stomach was associated with specific whole gene changes. Further detailed investigation showed that levels of the BabA protein were dramatically reduced within weeks of infection. The molecular mechanisms behind this reduction were shown to include phase variation and gene loss via intragenomic rearrangement, suggesting strong selective pressure against BabA expression in the macaque model. Notably, although there is apparently strong selective pressure against babA, babA is required for establishment of infection in this model as a strain in which babA was deleted was unable to colonize experimentally infected macaques.

  • 316.
    Liu, Junfa
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Thanikkal, Edvin
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Obi, Ikenna
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Francis, Matthew
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Elevated CpxR~P levels repress the Ysc-Yop type III secretion system of Yersinia pseudotuberculosis2012In: Research in Microbiology, ISSN 0923-2508, E-ISSN 1769-7123, Vol. 163, no 8, p. 518-530Article in journal (Refereed)
    Abstract [en]

    One way that Gram-negative bacteria respond to extracytoplasmic stress is through the CpxA-CpxR system. An activated CpxA sensor kinase phosphorylates the CpxR response regulator to instigate positive auto-amplification of Cpx pathway activation, as well as synthesis of various bacterial survival factors. In the absence of CpxA, human enteropathogenic Yersinia pseudotuberculosis accumulates high CpxR~P levels aided by the action of low molecular weight phosphodonors such as acetyl~P. Critically, these bacteria are also defective for plasmid encoded Ysc-Yop-dependent type III synthesis and secretion, an essential determinant of virulence. Herein, we investigated whether elevated CpxR~P levels account for lost Ysc-Yop function. Decisively, reducing CpxR~P in Yersinia defective for CpxA phosphatase activity - through incorporating second-site suppressor mutations in ackA-pta or cpxR - dramatically restored Ysc-Yop T3S function. Moreover, the repressive effect of accumulated CpxR~P is a direct consequence of binding to the promoter regions of the T3S genes. Thus, Cpx pathway activation has two consequences in Yersinia; one, to maintain quality control in the bacterial envelope, and the second, to restrict ysc-yop gene expression to those occasions where it will have maximal effect.

  • 317.
    Login, Frederic H.
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Wolf-Watz, Hans
    Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    YscU/FlhB of Yersinia pseudotuberculosis Harbors a C-terminal Type III Secretion Signal2015In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 290, no 43, p. 26282-26291Article in journal (Refereed)
    Abstract [en]

    All type III secretion systems (T3SS) harbor a member of the YscU/FlhB family of proteins that is characterized by an auto-proteolytic process that occurs at a conserved cytoplasmic NPTH motif. We have previously demonstrated that YscU(CC), the C-terminal peptide generated by auto-proteolysis of Yersinia pseudotuberculosis YscU, is secreted by the T3SS when bacteria are grown in Ca2+ -depleted medium at 37 degrees C. Here, we investigated the secretion of this early T3S-substrate and showed that YscU(CC) encompasses a specific C-terminal T3S signal within the 15 last residues (U-15). U-15 promoted C-terminal secretion of reporter proteins like GST and YopE lacking its native secretion signal. Similar to the "classical" N-terminal secretion signal, U-15 interacted with the ATPase YscN. Although U-15 is critical for YscU(CC) secretion, deletion of the C-terminal secretion signal of YscU(CC) did neither affect Yop secretion nor Yop translocation. However, these deletions resulted in increased secretion of YscF, the needle subunit. Thus, these results suggest that YscU via its C-terminal secretion signal is involved in regulation of the YscF secretion.

  • 318.
    Lopes, Jose Pedro
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Stylianou, Marios
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Nilsson, Gunnar
    Stockholm, Sweden.
    Urban, Constantin F.
    Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Opportunistic pathogen Candida albicans elicits a temporal response in primary human mast cells2015In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 5, article id 12287Article in journal (Refereed)
    Abstract [en]

    Immunosuppressed patients are frequently afflicted with severe mycoses caused by opportunistic fungal pathogens. Besides being a commensal, colonizing predominantly skin and mucosal surfaces, Candida albicans is the most common human fungal pathogen. Mast cells are present in tissues prone to fungal colonization being expectedly among the first immune cells to get into contact with C. albicans. However, mast cell-fungus interaction remains a neglected area of study. Here we show that human mast cells mounted specific responses towards C. albicans. Collectively, mast cell responses included the launch of initial, intermediate and late phase components determined by the secretion of granular proteins and cytokines. Initially mast cells reduced fungal viability and occasionally internalized yeasts. C. albicans could evade ingestion by intracellular growth leading to cellular death. Furthermore, secreted factors in the supernatants of infected cells recruited neutrophils, but not monocytes. Late stages were marked by the release of cytokines that are known to be anti-inflammatory suggesting a modulation of initial responses. C. albicans-infected mast cells formed extracellular DNA traps, which ensnared but did not kill the fungus. Our results suggest that mast cells serve as tissue sentinels modulating antifungal immune responses during C. albicans infection. Consequently, these findings open new doors for understanding fungal pathogenicity.

  • 319.
    Lopes, José Pedro
    et al.
    Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Stylianou, Marios
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology. Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Backman, Emelie
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology. Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Holmberg, Sandra
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology. Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Ekoff, Maria
    Nilsson, Gunnar
    Urban, Constantin F.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology. Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Cryptococcus neoformans Induces MCP-1 Release and Delays the Death of Human Mast Cells2019In: Frontiers in Cellular and Infection Microbiology, E-ISSN 2235-2988, Vol. 9, article id 289Article in journal (Refereed)
    Abstract [en]

    Cryptococcosis, caused by the basidiomycete Cryptococcus neoformans, is a life-threatening disease affecting approximately one million people per year worldwide. Infection can occur when C. neoformans cells are inhaled by immunocompromised people. In order to establish infection, the yeast must bypass recognition and clearance by immune cells guarding the tissue. Using in vitro infections, we characterized the role of mast cells (MCs) in cryptococcosis. We found that MCs recognize C. neoformans and release inflammatory mediators such as tryptase and cytokines. From the latter group MCs released mainly CCL-2/MCP-1, a strong chemoattractant for monocytic cells. We demonstrated that supernatants of infected MCs recruit monocytes but not neutrophils. During infection with C. neoformans, MCs have a limited ability to kill the yeast depending on the serotype. C. neoformans, in turn, modulates the lifespan of MCs both, by presence of its polysaccharide capsule and by secreting soluble modulators. Taken together, MCs might have important contributions to fungal clearance during early stages of cryptocococis where these cells regulate recruitment of monocytes to mucosal tissues.

  • 320. Loures, Flávio V
    et al.
    Röhm, Marc
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Lee, Chrono K
    Santos, Evelyn
    Wang, Jennifer P
    Specht, Charles A
    Calich, Vera L G
    Urban, Constantin F
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Levitz, Stuart M
    Recognition of Aspergillus fumigatus Hyphae by Human Plasmacytoid Dendritic Cells Is Mediated by Dectin-2 and Results in Formation of Extracellular Traps2015In: PLoS Pathogens, ISSN 1553-7366, E-ISSN 1553-7374, Vol. 11, no 2, article id e1004643Article in journal (Refereed)
    Abstract [en]

    Plasmacytoid dendritic cells (pDCs) were initially considered as critical for innate immunity to viruses. However, our group has shown that pDCs bind to and inhibit the growth of Aspergillus fumigatus hyphae and that depletion of pDCs renders mice hypersusceptible to experimental aspergillosis. In this study, we examined pDC receptors contributing to hyphal recognition and downstream events in pDCs stimulated by A. fumigatus hyphae. Our data show that Dectin-2, but not Dectin-1, participates in A. fumigatus hyphal recognition, TNF-α and IFN-α release, and antifungal activity. Moreover, Dectin-2 acts in cooperation with the FcRγ chain to trigger signaling responses. In addition, using confocal and electron microscopy we demonstrated that the interaction between pDCs and A. fumigatus induced the formation of pDC extracellular traps (pETs) containing DNA and citrullinated histone H3. These structures closely resembled those of neutrophil extracellular traps (NETs). The microarray analysis of the pDC transcriptome upon A. fumigatus infection also demonstrated up-regulated expression of genes associated with apoptosis as well as type I interferon-induced genes. Thus, human pDCs directly recognize A. fumigatus hyphae via Dectin-2; this interaction results in cytokine release and antifungal activity. Moreover, hyphal stimulation of pDCs triggers a distinct pattern of pDC gene expression and leads to pET formation.

  • 321.
    Luan, Shi-Lu
    Umeå University, Faculty of Medicine, Clinical Microbiology.
    Molecular epidemiology of streptococcus agalactiae: mobile elements as genetic markers.2006Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Streptococcus agalactiae, also designated group B streptococcus (GBS), is a Gram-positive coccus, and it is an important pathogen that causes invasive disease in neonates, pregnant adults, and non-pregnant adults with predisposing conditions. The group II intron GBSi1 is one of the major mobile genetic elements identified in S. agalactiae. The aim of this thesis was to characterize the GBSi1 distribution pattern, the population structure, and the influence of serotype- and clone-specific properties on the invasive capacity among clinical invasive and non-invasive isolates of S. agalactiae.

    Two additional copies of GBSi1 were identified at sites different from the primarily identified scpB-lmb locus. The distribution of GBSi1 was uneven among different serotypes. Three intron copies were only found in isolates of serotype III, and these targeted all the three identified gene loci. In contrast, a single copy of GBSi1 was found in isolates of serotype II and V and only located at the scpB-lmb locus. Furthermore, at the 5′ flanking region of the scpB-lmb gene locus, a novel 2.1 kb DNA fragment with plasmid features was identified only in intron carrying isolates. This may suggest that GBSi1 once was brought into the S. agalactiae genome by an integrated plasmid.

    Multilocus sequence typing was used to characterize totally 314 invasive and non-invasive S. agalactiae isolates collected in Northern and Western Sweden from the years 1988 to 2004. Five major genetic lineages (clonal complexes) were identified among both invasive and non-invasive isolates, including serotype Ia, Ib, and II to V, indicating a clonal population structure of S. agalactiae isolates. A number of genetically highly related isolates were found to express different capsular types, suggesting that capsule switching occurs rather frequently between isolates. Furthermore, non-invasive isolates belonging to the same clonal complexes displayed more heterogeneity in capsule expression as well as in the distribution patterns of mobile genetic elements than invasive isolates. This indicates that less variability is allowed in a highly selective environment such as the blood. All major clonal complexes and serotypes caused invasive disease, although their ability to do so varied greatly. CC17 was significantly associated with neonatal invasive disease; whereas CC19 was equally common among isolates from adult and neonatal disease, despite that both CC17 and CC19 expressed capsular type III. This striking difference seen between CC17 and CC19 suggests that clonal complex associated properties, in addition to capsular type, play important roles in the virulence of S. agalactiae. CC1, a new emerging clone since early 1990s, has caused substantial amount of disease among adults. In addition, mutually exclusive distribution of mobile elements GBSi1 and IS1548 was seen, and they were shown to constitute genetic markers for serotype III CC17 and CC19 isolates, respectively.

  • 322. Luna, Brian
    et al.
    Kubler, André
    Larsson, Christer
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Foster, Brent
    Bagci, Ulas
    Mollura, Daniel J
    Jain, Sanjay K
    Bishai, William R
    In Vivo Prediction of Tuberculosis-Associated Cavity Formation in Rabbits2015In: The Journal of infectious diseases, ISSN 1537-6613, Vol. 211, no 3, p. 481-485Article in journal (Refereed)
    Abstract [en]

    The presence of cavitary lesions in patients with tuberculosis poses a significant clinical concern due to the risk of infectivity and the risk of antibiotic treatment failure. We describe 2 algorithms that use noninvasive positron emission tomography (PET) and computed tomography (CT) to predict the development of cavitary lesions in rabbits. Analysis of the PET region of interest predicted cavitary disease with 100% sensitivity and 76% specificity, and analysis of the CT region of interest predicted cavitary disease with 83.3% sensitivity and 76.9% specificity. Our results show that restricting our analysis to regions with high [(18)F]-fluorodeoxyglucose uptake provided the best combination of sensitivity and specificity.

  • 323.
    Lundqvist, Jenny
    Umeå University, Faculty of Medicine, Molecular Biology (Faculty of Medicine).
    Malaria and relapsing fever Borrelia: interactions and potential therapy2009Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Infectious diseases such as malaria and relapsing fever borreliosis (RF), cause severe human mortality and morbidity in developing countries. Malaria, caused by Plasmodium spp. parasites, is estimated by the World Health Organization to cause 1.5-2.7 million deaths annually. RF, caused by Borrelia spirochetes, has the highest prevalence described for any bacterial disease in Africa, with infection outcomes ranging from asymptomatic to fatal. RF borreliosis manifests in humans as a recurring fever and with other symptoms very similar to those of malaria.

    RF borreliosis has been regarded as a transient infection of the blood. However, B. duttonii exploits the brain as an immunoprivileged site escaping the host immune response while spirochetes in the blood are cleared. To investigate whether residual bacteria are dormant or actively dividing, mice with residual brain infection were administered ceftriaxone, a β-lactam antibiotic interfering with cell wall synthesis. Hence, it only affects actively dividing bacteria. Ceftriaxone eradicated brain RF infection in all treated mice, demonstrating that the bacteria are actively multiplying rather than in a dormant state. The findings support the therapeutic use of ceftriaxone for RF neuroborreliosis since penetration into cerebrospinal fluid is greater for ceftriaxone than for the often recommended doxycycline.

    The clinical features of malaria and RF are similar and diagnosis is further complicated by the frequently occurring concomitant malaria-RF infections. Therefore, we established a mouse model to study the pathogenesis and immunological response to Plasmodium/Borrelia mixed infection. Interestingly, malaria was suppressed in the co-infected animals whereas spirochete numbers were elevated 21-fold. The immune response in the concomitantly infected mice was polarized towards malaria leaving the spirochetes unharmed. Mice with co-infections also exhibited severe anemia and internal damages, probably attributed to escalating spirochete numbers. A secondary malaria infection reactivated the residual brain RF infection in 60% of the mice. This highlights the importance of co-infections as diagnostic pitfalls as well as the need for novel treatment strategies.

    Currently there is no commercial malaria vaccine and increasing drug resistance presents an urgent need for new malaria chemotherapeutics. Blood-stage malaria parasites are rapidly growing with high metabolic and biosynthetic activity, making them highly sensitive to limitations in polyamine supply. Disrupting polyamine synthesis in vivo with trans-4-methylcyclohexylamine (4MCHA) eradicated the malaria infection gradually, resulting in protective immunity. This leads the way for further biochemical and pharmacological development of the polyamine inhibitor 4MCHA and similar compounds as antimalarial drugs

  • 324.
    Löfgren, Niklas
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Physiological chemistry.
    Role of ANGPTL-4 in the regulation of the functional pool of LPL in mouse heart2017Independent thesis Basic level (professional degree), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    The heart is constantly pumping blood through the body to provide tissues with oxygen and nutrients and retrieve waste products. The cardiomyocytes therefore have a high energy demand. The mostly used energy substrate is fatty acids, covering approximately 70 % of the energy need under normal conditions. Fatty acids reach the heart either bound to albumin or delivered from circulating lipoproteins after liberation from triglycerides through the action of the enzyme lipoprotein lipase (LPL). LPL is produced in parenchymal cells like cardiomyocytes and is then transported to the luminal side of capillary endothelium where it hydrolyses lipoproteins. The LPL activity is rapidly modulated according to the needs of the tissue and the regulation occurs mostly by post-translational mechanisms. Derangements in the normal function of the LPL system of the heart have implications for ischemia, diabetic myopathy and other forms of heart failure.

    One candidate protein that regulates LPL activity in other tissues is angiopoietin-like protein 4 (ANGPTL-4). Binding of this protein to the active LPL dimer causes dissociation to inactive LPL monomers. In mice, the expression of Angptl4 is seen in liver and adipose tissue, but also in skeletal muscle and heart.

    The aim of this study was to identify the role of ANGPTL-4 for control of LPL on the cardiac capillary endothelium in mice. This pool of functional LPL in the heart can only be studied by extracorporal retrograde perfusion of the coronary arteries by heparin in order to release the LPL that is bound to the endothelium. Hearts from mice with knock-out or transgenic overexpression of Angptl4 was perfused, and the LPL activity was determined in perfusates from fasted and fed mice. In the Angptl4 knock-out mice, the nutritional regulation of the amount of LPL activity on the endothelium was intact, but acted from a higher baseline level. In the transgenic mice the LPL activity was pressed down to a low level in both fed and fasted mice, and the typical nutritional regulation of LPL activity was wiped out. The large pressure on the LPL system by the elevated amounts of ANGPTL-4 had disabled the regulatory system.

    In conclusion, ANGPTL-4 seems to be responsible for control of the baseline level of LPL activity on the endothelium of mouse hearts. Other factors than ANGPTL-4 must, however, be responsible for the normal, rapid modulation of LPL activity according to the nutritional requirement of the tissue.

  • 325. Löfling, Jonas
    et al.
    Diswall, Mette
    Eriksson, Sara
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Borén, Thomas
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Breimer, Michael E
    Holgersson, Jan
    Studies of Lewis antigens and H. pylori adhesion in CHO cell lines engineered to express Lewis b determinants2008In: Glycobiology, ISSN 0959-6658, E-ISSN 1460-2423, Vol. 18, no 7, p. 494-501Article in journal (Refereed)
    Abstract [en]

    Many microbes bind and adhere via adhesins to host cell carbohydrates as an initial step for infection. Therefore, cell lines expressing Lewis b (Le(b)) determinants were generated as a potential model system for Helicobacter pylori colonization and infection, and their expression of blood group Lewis determinants was characterized. CHO-K1 cells were stably transfected with selected glycosyltransferase cDNAs, and two Le(b) positive clones, 1C5 and 2C2, were identified. Expression of Lewis (Le(a), Le(b), Le(x), and Le(y)) determinants was analyzed by flow cytometry of intact cells, SDS-PAGE/Western blot of solubilized glycoproteins, and thin layer chromatography immunostaining of isolated glycolipids (GL). Binding of H. pylori to cells was examined by microscopy and quantified. Flow cytometry showed that 1C5 and 2C2 were Le(a) and Le(b) positive. 1C5 expressed Le(b) on O-linked, but not N-linked, glycans and only weakly on GLs. In contrast, 2C2 expressed Le(b) on N-, O-glycans, and GLs. Furthermore, both clones expressed Le(a) on N- and O-glycans but not on GLs. 2C2, but not 1C5, stained positively for Le(y) on N-linked glycans and GLs. Both clones, as well as the parental CHO-K1 cells, expressed Le(x) on GLs. A Le(b)-binding H. pylori strain bound to the 1C5 and 2C2 cells. In summary, two glycosyltransferase transfected CHO-K1 cell clones differed regarding Lewis antigen expression on N- and O-linked glycans as well as on GLs. Both clones examined supported adhesion of a Le(b)-binding H. pylori strain and may thus be a useful in vitro model system for H. pylori colonization/infection studies.

  • 326.
    Lövgren, J. Mattias
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Wikström, P. Mikael
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    The rlmB Gene Is Essential for Formation of Gm2251 in 23S rRNA but Not for Ribosome Maturation in Escherichia coli2001In: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 183, no 23, p. 6957-6960Article in journal (Refereed)
    Abstract [en]

    In Saccharomyces cerevisiae, the rRNA Gm2270 methyltransferase, Pet56p, has an essential role in the maturation of the mitochondrial large ribosomal subunit that is independent of its methyltransferase activity. Here we show that the proposed Escherichia coli ortholog, RlmB (formerly YjfH), indeed is essential for the formation of Gm in position 2251 of 23S rRNA. However, a DeltarlmB mutant did not show any ribosome assembly defects and was not outgrown by a wild-type strain even after 120 cell mass doublings. Thus, RlmB has no important role in ribosome assembly or function in E. coli.

  • 327.
    Lövheim, Hugo
    et al.
    Umeå University, Faculty of Medicine, Department of Community Medicine and Rehabilitation, Geriatric Medicine.
    Gilthorpe, Jonathan
    Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience, Clinical Neuroscience.
    Adolfsson, Rolf
    Umeå University, Faculty of Medicine, Department of Clinical Sciences, Psychiatry.
    Nilsson, Lars-Göran
    Umeå University, Faculty of Medicine, Umeå Centre for Functional Brain Imaging (UFBI). Aging Research Center, Karolinska Institutet and Stockholm University, Stockholm, Sweden.
    Elgh, Fredrik
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Reactivated herpes simplex infection increases the risk of Alzheimer's disease2015In: Alzheimer's & Dementia, ISSN 1552-5260, E-ISSN 1552-5279, Vol. 11, no 6, p. 593-599Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Previous studies have suggested a link between herpes simplex virus (HSV) type 1 and the development of Alzheimer's disease (AD).

    METHODS: The present analysis included 3432 persons (53.9% women, mean age at inclusion 62.7 ± 14.4 years) with a mean follow-up time of 11.3 years. The number of incident AD cases was 245. Serum samples were analyzed for anti-HSV antibodies (immunoglobulin (Ig)G and IgM) by enzyme-linked immunosorbent assays.

    RESULTS: The presence of anti-HSV IgG antibodies was not associated with an increased risk for AD, controlled for age and sex (hazard ratio, HR, 0.993, P = .979). However, the presence of anti-HSV IgM at baseline was associated with an increased risk of developing AD (HR 1.959, P = .012).

    CONCLUSION: Positivity for anti-HSV IgM, a sign of reactivated infection, was found to almost double the risk for AD, whereas the presence of anti-HSV IgG antibodies did not affect the risk.

  • 328. Maddocks, Sarah E.
    et al.
    Wright, Christopher J.
    Nobbs, Angela H.
    Brittan, Jane L.
    Franklin, Linda
    Strömberg, Nicklas
    Umeå University, Faculty of Medicine, Department of Odontology, Cariology.
    Kadioglu, Aras
    Jepson, Mark A.
    Jenkinson, Howard F.
    Streptococcus pyogenes antigen I/II-family polypeptide AspA shows differential ligand-binding properties and mediates biofilm formation2011In: Molecular Microbiology, ISSN 0950-382X, E-ISSN 1365-2958, Vol. 81, no 4, p. 1034-1049Article in journal (Refereed)
    Abstract [en]

    The streptococcal antigen I/II (AgI/II)-family polypeptides are cell wall-anchored adhesins expressed by most indigenous oral streptococci. Proteins sharing 30-40% overall amino acid sequence similarities with AgI/II-family proteins are also expressed by Streptococcus pyogenes. The S. pyogenes M28_Spy1325 polypeptide (designated AspA) displays an AgI/II primary structure, with alanine-rich (A) and prolinerich (P) repeats flanking a V region that is projected distal from the cell. In this study it is shown that AspA from serotype M28 S. pyogenes, when expressed on surrogate host Lactococcus lactis, confers binding to immobilized salivary agglutinin gp-340. This binding was blocked by antibodies to the AspA-VP region. In contrast, the N-terminal region of AspA was deficient in binding fluid-phase gp-340, and L. lactis cells expressing AspA were not agglutinated by gp-340. Deletion of the aspA gene from two different M28 strains of S. pyogenes abrogated their abilities to form biofilms on saliva-coated surfaces. In each mutant strain, biofilm formation was restored by trans complementation of the aspA deletion. In addition, expression of AspA protein on the surface of L. lactis conferred biofilm-forming ability. Taken collectively, the results provide evidence that AspA is a biofilm-associated adhesin that may function in host colonization by S. pyogenes.

  • 329. Magalhaes, Ana
    et al.
    Marcos-Pinto, Ricardo
    Gomes, Joana
    Nairn, Alison V.
    Rossez, Yannick
    Robbe-Masselot, Catherine
    Maes, Emmanuel
    Bugaytsova, Zhanna
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Figueiredo, Ceu
    Borén, Thomas
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Moremen, Kelley W.
    Reis, Celso A.
    The glycan receptors of Helicobacter pylori: decoding the pathways underlying gastric glycophenotype modulation2016In: Glycobiology, ISSN 0959-6658, E-ISSN 1460-2423, Vol. 26, no 12, p. 1401-1402Article in journal (Refereed)
    Abstract [en]

    The gastrointestinal tract is covered by a complex extracellular mucus layer that protects the gastric epithelium fromexternal aggressions such as chemical agents, microorganismsand shear stress. Although this mucus barrier confers protec-tion against certain pathogens, it may also provide a niche formicrobial binding.Helicobacter pyloriexploits the host glycoconjugates present in the gastric mucus layer and lining thesurface epithelium of the gastric mucosa to colonize the stomach. Infection can persist for decades promoting chronicinflammation, and in a subset of individuals lesions cansilently progress to cancer. The secreted MUC5AC mucin isthe main component of the gastric mucus layer, andH. pyloriBabA-mediated binding to MUC5AC confers increased riskfor overt disease. We have shown that FUT2 determines theO-glycosylation pattern of Muc5ac, with Fut2 knock-outleading to a marked decrease inα1,2-fucosylated structuresand increased expression of the terminal type 1 glycan structure Lewisa. Importantly, for thefirst time, we structurallyvalidated the expression of Lewisain murine gastric mucosa(1). We further demonstrated that loss of mucin FUT2-mediated fucosylation impairs gastric mucosal binding ofH.pyloriBabA adhesin, which is a recognized feature of patho-genicity. UponH. pyloriinfection,concomitantly with tissueinflammation, there is a remodeling of the gastric glycopheno-type. We showed that increased expression of sialyl-Lewisa/xantigens is due to transcriptional up-regulation of theB3GNT5,B3GALT5,andFUT3genes. In addition, weobserved thatH. pyloriinfected individuals present a markedgastric local pro-inflammatory signature with significantlyhigher TNF-αlevels and demonstrated that TNF-induced activation of the NF-kappaB pathway results in B3GNT5 up-regulation (2). Furthermore, we showed that this gastric glycosylation shift, characterized by increased sialylation pat-terns, favors SabA-mediatedH. pyloriattachment to humaninflamed gastric mucosa. Our data provides clinically relevantinsights into the regulatory mechanisms underlyingH. pylorimodulation of host glycosylation machinery, and phenotypicalterations crucial for life-long infection and gastric disease.

  • 330. Magalhaes, Ana
    et al.
    Rossez, Yannick
    Robbe-Masselot, Catherine
    Maes, Emmanuel
    Gomes, Joana
    Shevtsova, Anna
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Bugaytsova, Jeanna
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Boren, Thomas
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Reis, Celso A.
    Muc5ac gastric mucin glycosylation is shaped by FUT2 activity and functionally impacts Helicobacter pylori binding2016In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, article id 25575Article in journal (Refereed)
    Abstract [en]

    The gastrointestinal tract is lined by a thick and complex layer of mucus that protects the mucosal epithelium from biochemical and mechanical aggressions. This mucus barrier confers protection against pathogens but also serves as a binding site that supports a sheltered niche of microbial adherence. The carcinogenic bacteria Helicobacter pylori colonize the stomach through binding to host glycans present in the glycocalyx of epithelial cells and extracellular mucus. The secreted MUC5AC mucin is the main component of the gastric mucus layer, and BabA-mediated binding of H. pylori to MUC5AC confers increased risk for overt disease. In this study we unraveled the O-glycosylation profile of Muc5ac from glycoengineered mice models lacking the FUT2 enzyme and therefore mimicking a non-secretor human phenotype. Our results demonstrated that the FUT2 determines the O-glycosylation pattern of Muc5ac, with Fut2 knock-out leading to a marked decrease in alpha 1,2-fucosylated structures and increased expression of the terminal type 1 glycan structure Lewis-a. Importantly, for the first time, we structurally validated the expression of Lewis-a in murine gastric mucosa. Finally, we demonstrated that loss of mucin FUT2-mediated fucosylation impairs gastric mucosal binding of H. pylori BabA adhesin, which is a recognized feature of pathogenicity.

  • 331.
    Mahdavi, Jafar
    et al.
    Umeå University, Faculty of Medicine, Department of Odontology, Oral Microbiology.
    Sondén, B
    Umeå University, Faculty of Medicine, Department of Odontology, Oral Microbiology.
    Hurtig, Marina
    Umeå University, Faculty of Medicine, Department of Odontology, Oral Microbiology.
    Olfat, Farzad O
    Umeå University, Faculty of Medicine, Department of Odontology, Oral Microbiology.
    Forsberg, Lina
    Umeå University, Faculty of Medicine, Department of Odontology, Oral Microbiology.
    Roche, Niamh
    Ångström, Jonas
    Larsson, Thomas
    Teneberg, Susann
    Karlsson, Karl-Anders
    Altraja, Siiri
    Wadström, Torkel
    Kersulyte, Dangeruta
    Berg, Douglas E
    Dubois, Andre
    Petersson, Christoffer
    Magnusson, Karl-Eric
    Norberg, Thomas
    Lindh, Frank
    Lundskog, Bertil B
    Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Arnqvist, Anna
    Umeå University, Faculty of Medicine, Department of Odontology, Oral Microbiology. Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Hammarström, Lennart
    Borén, Thomas
    Umeå University, Faculty of Medicine, Department of Odontology, Oral Microbiology.
    Helicobacter pylori SabA adhesin in persistent infection and chronic inflammation2002In: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 297, no 5581, p. 573-578Article in journal (Refereed)
    Abstract [en]

    Helicobacter pylori adherence in the human gastric mucosa involves specific bacterial adhesins and cognate host receptors. Here, we identify sialyl-dimeric-Lewis x glycosphingolipid as a receptor for H. pylori and show that H. pylori infection induced formation of sialyl-Lewis x antigens in gastric epithelium in humans and in a Rhesus monkey. The corresponding sialic acid-binding adhesin (SabA) was isolated with the "retagging" method, and the underlying sabA gene (JHP662/HP0725) was identified. The ability of many H. pylori strains to adhere to sialylated glycoconjugates expressed during chronic inflammation might thus contribute to virulence and the extraordinary chronicity of H. pylori infection.

  • 332. Manger, Mari S.
    et al.
    Strand, Tor A.
    Taneja, Sunita
    Refsum, Helga
    Ueland, Per M.
    Nygard, Ottar
    Schneede, Jörn
    Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience.
    Sommerfelt, Halvor
    Bhandari, Nita
    Cobalamin Status Modifies the Effect of Zinc Supplementation on the Incidence of Prolonged Diarrhea in 6-to 30-Month-Old North Indian Children2011In: Journal of Nutrition, ISSN 0022-3166, E-ISSN 1541-6100, Vol. 141, no 6, p. 1108-1113Article in journal (Refereed)
    Abstract [en]

    The observed effect of zinc supplementation on diarrheal morbidity varies between trials and there is a need to identify subgroups most likely to benefit from improved zinc nutriture. In a randomized, double-blind trial in 2296 children in New Delhi, India, we assessed whether baseline cobalamin or folate status modified the effect of zinc supplementation on the incidence of prolonged (>= 7 d duration) and acute diarrhea. Children aged 6-30 mo received zinc or placebo daily for 4 mo. We measured plasma concentrations of folate, cobalamin, total homocysteine (tHcy), and methylmalonic acid (MMA) at enrollment and assessed the efficacy of zinc supplementation in subgroups based on these variables. The efficacy of zinc on reducing the risk of prolonged diarrhea was higher in those with plasma cobalamin concentrations below the 25th percentile and in those with tHcy and MMA concentrations above the 75th percentile. The OR (95% Cl) for children below and above the 25th percentile for cobalamin were 0.53 (0.35-0.78) and 0.90 (0.73-1.11), respectively (P-interaction = 0.015). There were similar differences for the OR when comparing efficacy in those above and below the 75th percentile for tHcy and MMA (P-interaction = 0.045 and 0.188, respectively). Baseline folate status did not modify the effect of zinc on prolonged diarrhea. Neither cobalamin nor folate status influenced the effect of zinc on acute diarrhea. Children with poor cobalamin status benefited more from zinc supplementation for the prevention of prolonged diarrhea J. Nutr. 141: 1108-1113, 2011.

  • 333. Marinho, Catarina M.
    et al.
    Dos Santos, Patricia T.
    Kallipolitis, Birgitte H.
    Johansson, Jörgen
    Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Ignatov, Dmitriy
    Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Guerreiro, Duarte N.
    Piveteau, Pascal
    O'Byrne, Conor P.
    The σB-dependent regulatory sRNA Rli47 represses isoleucine biosynthesis in Listeria monocytogenes through a direct interaction with the ilvA transcript2019In: RNA Biology, ISSN 1547-6286, E-ISSN 1555-8584Article in journal (Refereed)
    Abstract [en]

    The facultative intracellular pathogen Listeria monocytogenes can persist and grow in a diverse range of environmental conditions, both outside and within its mammalian host. The alternative sigma factor Sigma B (sigma(B)) plays an important role in this adaptability and is critical for the transition into the host. While some of the functions of the sigma(B) regulon in facilitating this transition are understood the role of sigma(B)-dependent small regulatory RNAs (sRNAs) remain poorly characterized. In this study, we focused on elucidating the function of Rli47, a sigma(B)-dependent sRNA that is highly induced in the intestine and in macrophages. Using a combination of in silico and in vivo approaches, a binding interaction was predicted with the Shine-Dalgarno region of the ilvA mRNA, which encodes threonine deaminase, an enzyme required for branched-chain amino acid biosynthesis. Both ilvA transcript levels and threonine deaminase activity were increased in a deletion mutant lacking the rli47 gene. The Delta rli47 mutant displayed a shorter growth lag in isoleucine-depleted growth media relative to the wild-type, and a similar phenotype was also observed in a mutant lacking sigma(B). The impact of the Delta rli47 on the global transcription profile of the cell was investigated using RNA-seq, and a significant role for Rli47 in modulating amino acid metabolism was uncovered. Taken together, the data point to a model where Rli47 is responsible for specifically repressing isoleucine biosynthesis as a way to restrict growth under harsh conditions, potentially contributing to the survival of L. monocytogenes in niches both outside and within the mammalian host.

  • 334.
    Maripuu, Linda
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Biomedical Laboratory Science.
    Superantigens in group A streptococcus: gene diversity and humoral immune response2011Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Group A streptococcus (GAS) is a strictly human pathogen that causes infections ranging from asymptomatic carriage to the highly lethal streptococcal toxic shock syndrome (STSS). GAS are classified according to the sequence of the variable 5’ end of the emm-gene that encodes the surface associated M-protein. In the late 1980s, outbreaks of GAS infections with high rates of STSS were reported in several parts of the world, including Sweden. Superantigens (SAgs), a group of exotoxins, have been described as key mediators of STSS due to their capacity to polyclonally activate T-cells and induce a massive release of inflammatory cytokines. Previous reports have revealed that sera from STSS patients have lower capacity to neutralize this SAg-mediated immune stimulation and a higher prevalence of GAS isolates with specific emm-genotypes during disease outbreaks. The aims of this thesis were to analyse the protective antibody response mounted by the host against SAgs produced by the infecting GAS isolate and to characterise the isolates emm-genotypes and SAg gene profiles. The clinical material examined was collected from patients with STSS, sepsis, erysipelas, or tonsillitis in Sweden between 1986 and 2001. Both acute- and convalescence-phase sera were analyzed, along with the infecting GAS isolates. The 92 clinical GAS isolates examined were found to exhibit a high degree of genetic diversity in terms of the number and identity of their SAg genes. Isolates with a given emm-genotype could be divided into subgroups on the basis of their SAg gene profiles. Ten different SAg gene profiles were identified in the 45 emm1 isolates examined; one of these ten was highly persistent, being observed in 22 isolates collected over 14 years. Two of the 11 known SAg genes in GAS, smeZ-1 and speA, were more prevalent in the emm1 associated profiles than in the SAg gene profiles of isolates with other emm-genotypes. Patients infected by GAS with the emm1-genotype were less likely to produce acute-phase sera that could effectively neutralize the T-cell mitogenicity induced by the infecting isolate’s extracellular products (EP). Sepsis patients whose sera exhibited this lack of neutralizing ability were more prone to developing STSS. Most patients whose acute-phase sera did not effectively neutralize the EP from the infecting isolate lacked protective antibodies in their convalescent-phase sera despite having elevated ELISA titers. The results reported herein show that combining SAg gene profiling with emm-genotyping may be useful for tracking the spread of GAS clones in the community. It was also shown that a lack of neutralizing activity in convalescence-phase sera might be due to an inability of those patients to mount a protective immune response against SAgs produced by the infecting GAS isolate.

  • 335.
    Maripuu, Linda
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Ekström, M
    Norgren, M
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Characterization of mitogenic and DNase activity of recombinantly produced mitogenic factor from group A streptococcusManuscript (preprint) (Other academic)
  • 336.
    Marttila, Marko
    Umeå University, Faculty of Medicine, Clinical Microbiology.
    Cellular receptors for species B adenoviruses2007Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Adenoviruses belong to the most common human pathogens. The severity of infection varies greatly, from subclinical to lethal, depending on the virus type and immune status of the infected host. The 51 known human adenovirus serotypes are divided into six species (A-F) based on characteristics such as tropism. Species B adenoviruses, which are the subjects of this thesis, are further divided into subspecies B:1 that contains Ad3, Ad7, Ad16, Ad21 and Ad50 and subspecies B:2 that contains Ad11, Ad14, Ad34 and Ad35. Species B adenoviruses primarily cause ocular and respiratory tract infections, but certain serotypes (Ad11, Ad34 and Ad35) are also associated with renal disease.

    The main aim of this thesis was to identify and characterize cellular receptors for species B adenoviruses. This will ultimately help to understand the diverse tropism shown by different adenoviruses and perhaps contribute to development of antivirals. Also, since adenoviruses are among the most commonly used vector for gene therapy it is of importance to characterize the initial steps of adenovirus life cycle.

    Members of species B adenoviruses have been shown to utilize both the complement regulating membrane cofactor protein (MCP), i.e. CD46, and a still unknown receptor. CD80 and CD86, usually found on antigen-presenting cells, have also been suggested as receptors

    We found first that Ad11 used CD46 as a cellular receptor on respiratory A549 cells, and subsequently that CD46 is a cellular receptor for all species B adenovirus serotypes, except for adenovirus types 3 and 7, using cells that represent the tropism of species B adenoviruses, i.e. respiratory, conjunctival and renal epithelial cells.

    We further compared the relative roles of CD46 with CD80 and CD86 using cells that represent species B adenovirus tropism. Using soluble candidate receptors and antibodies against corresponding receptors to challenge virus binding to and infection of cells, we found that on these cells, CD46 is a cellular receptor for all species B adenoviruses except Ad3 and Ad7, and that CD80 and CD86 do not play an important role.

    We have further pinpointed the interaction site for Ad11 on CD46 by X-ray crystallography. The extracellular region of CD46 contains four short consensus repeats (SCR1-4) of which the outermost N-terminal SCR1 and SCR2 mediate binding to Ad11. This interaction was confirmed by inhibiting infection and binding of Ad11 to A549 cells using soluble SCR1-2 fragments. Surprisingly the conformation of bound CD46 differs profoundly from its unbound state, with the bent surface structure straightened into an elongated rod. Viral proteins can sometimes undergo large conformational changes upon receptor binding, but this is, to the best of our knowledge, the first example of a virus protein dramatically changing the overall structure of its receptor. CD46 serves as a receptor for a large number of viral and bacterial pathogens and it is structurally and functionally related to other viral receptors such as CD21 and CD55. The mode of interaction presented here may serve as a conceptual framework for studies of many other receptors that are constructed from SCR domains.

  • 337.
    Marttila, Marko
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Ackelind, Pär
    Persson, David
    Stehle, Thilo
    Arnberg, Niklas
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    CD46 but not CD80 or CD86 serves as a cellular receptor for selected species B adenoviruses on human epithelial cellsManuscript (Other academic)
  • 338.
    Marttila, Marko
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Persson, David
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Gustafsson, Dan
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Liszewski, M Kathryn
    Atkinson, John P
    Wadell, Göran
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Arnberg, Niklas
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    CD46 is a cellular receptor for all species B adenoviruses except types 3 and 72005In: Journal of Virology, ISSN 0022-538X, E-ISSN 1098-5514, Vol. 79, no 22, p. 14429-14436Article in journal (Refereed)
    Abstract [en]

    The 51 human adenovirus serotypes are divided into six species (A to F). Adenovirus serotypes from all species except species B utilize the coxsackie-adenovirus receptor for attachment to host cells in vitro. Species B adenoviruses primarily cause ocular and respiratory tract infections, but certain serotypes are also associated with renal disease. We have previously demonstrated that adenovirus type 11 (species B) uses CD46 (membrane cofactor protein) as a cellular receptor instead of the coxsackie-adenovirus receptor (A. Segerman et al., J. Virol. 77:9183-9191, 2003). In the present study, we found that transfection with human CD46 cDNA rendered poorly permissive Chinese hamster ovary cells more permissive to infection by all species B adenovirus serotypes except adenovirus types 3 and 7. Moreover, rabbit antiserum against human CD46 blocked or efficiently inhibited all species B serotypes except adenovirus types 3 and 7 from infecting human A549 cells. We also sequenced the gene encoding the fiber protein of adenovirus type 50 (species B) and compared it with the corresponding amino acid sequences from selected serotypes, including all other serotypes of species B. From the results obtained, we conclude that CD46 is a major cellular receptor on A549 cells for all species B adenoviruses except types 3 and 7.

  • 339.
    Marwaha, Sania
    et al.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Uvell, Hanna
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Salin, Olli
    Umeå University, Faculty of Science and Technology, Department of Chemistry. Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Lindgren, Anders E. G.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Silver, Jim
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Elofsson, Mikael
    Umeå University, Faculty of Science and Technology, Department of Chemistry. Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Gylfe, Åsa
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    N-acylated derivatives of sulfamethoxazole and sulfafurazole inhibit intracellular growth of Chlamydia trachomatis2014In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 58, no 5, p. 2968-2971Article in journal (Refereed)
    Abstract [en]

    Antibacterial compounds with novel modes of action are needed for management of bacterial infections. Here we describe a high-content screen of 9,800 compounds identifying acylated sulfonamides as novel growth inhibitors of the sexually transmitted pathogen Chlamydia trachomatis. The effect was bactericidal and distinct from that of sulfonamide antibiotics, as para-aminobenzoic acid did not reduce efficacy. Chemical inhibitors play an important role in Chlamydia research as probes of potential targets and as drug development starting points.

  • 340. Massad, E.
    et al.
    Nascimento Burattini, M.
    Khan, K.
    Struchiner, C. J.
    Coutinho, F. A. B.
    Wilder-Smith, Annelies
    Umeå University, Faculty of Medicine, Department of Public Health and Clinical Medicine, Epidemiology and Global Health. Institute of Public Health, University of Heidelberg, Germany; Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore.
    On the origin and timing of Zika virus introduction in Brazil2017In: Epidemiology and Infection, ISSN 0950-2688, E-ISSN 1469-4409, Vol. 145, no 11, p. 2303-2312Article in journal (Refereed)
    Abstract [en]

    The timing and origin of Zika virus (ZIKV) introduction in Brazil has been the subject of controversy. Initially, it was assumed that the virus was introduced during the FIFA World Cup in June-July 2014. Then, it was speculated that ZIKV may have been introduced by athletes from French Polynesia (FP) who competed in a canoe race in Rio de Janeiro in August 2014. We attempted to apply mathematical models to determine the most likely time window of ZIKV introduction in Brazil. Given that the timing and origin of ZIKV introduction in Brazil may be a politically sensitive issue, its determination (or the provision of a plausible hypothesis) may help to prevent undeserved blame. We used a simple mathematical model to estimate the force of infection and the corresponding individual probability of being infected with ZIKV in FP. Taking into account the air travel volume from FP to Brazil between October 2013 and March 2014, we estimated the expected number of infected travellers arriving at Brazilian airports during that period. During the period between December 2013 and February 2014, 51 individuals travelled from FP airports to 11 Brazilian cities. Basing on the calculated force of ZIKV infection (the per capita rate of new infections per time unit) and risk of infection (probability of at least one new infection), we estimated that 18 (95% CI 12-22) individuals who arrived in seven of the evaluated cities were infected. When basic ZIKV reproduction numbers greater than one were assumed in the seven evaluated cities, ZIKV could have been introduced in any one of the cities. Based on the force of infection in FP, basic reproduction ZIKV number in selected Brazilian cities, and estimated travel volume, we concluded that ZIKV was most likely introduced and established in Brazil by infected travellers arriving from FP in the period between October 2013 and March 2014, which was prior to the two aforementioned sporting events.

  • 341.
    Mei, Ya-fang
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Harrach, Balzs
    Wadell, Göran
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Mastadenovirus: adenoviridae2012In: The Springer Index of Viruses / [ed] Christian Tidona, Gholamreza Darai, New York: Springer Science+Business Media B.V., 2012, p. 33-48Chapter in book (Refereed)
  • 342.
    Meyer, Lena
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    The Francisella pathogenicity island: its role in type VI secretion and intracellular infection2015Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Intracellular bacteria have developed various mechanisms to enter and persist in host cells and, at the same time, to evade the host immune response. One such pathogen is Francisella tularensis, the etiological agent of tularemia. After phagocytosis, this Gram-negative bacterium quickly escapes from the phagocytic compartment and replicates in the host cell cytosol. For this mode of infection, several components of the Francisella pathogenicity island (FPI) are critical. Interestingly, some FPI proteins share homology to components of Type VI Secretion Systems (T6SSs), but their assembly and functionality remains to be shown in Francisella.The thesis focused on the characterization of several of these FPI components; more specifically, how they contribute to the infection cycle as well as their possible role in the putative T6SS. We identified three unique mutants, ΔiglG, ΔiglI and ΔpdpE, which to various degrees were able to escape the phagosomal compartment, replicate in the host cytosol and cause host cell cytotoxicity. In contrast, ΔiglE as well as mutants within the conserved core components of T6SSs, VgrG and DotU, were defective for all of these processes. In the case of IglE, which is a lipoprotein and localized to the outer membrane of the bacterial cell wall, residues within its N-terminus were identified to be important for IglE function. Consistent with a suggested role as a trimeric membrane puncturing device, VgrG was found to form multimers. DotU stabilized the inner membrane protein IcmF, in agreement with its function as a core T6SS component. The functionality of the secretion system was shown by the translocation of several FPI proteins into the cytosol of infected macrophages, among them IglE, IglC and VgrG, of which IglE was the most prominently secreted protein. At the same time, the secretion was dependent on the core components VgrG, DotU but also on IglG. Although we and others have shown the importance of FPI proteins for the escape of F. tularensis, it has been difficult to assess their role in the subsequent replication, since mutants that fail to escape never reach the growth-permissive cytosol. For this reason, selected FPI mutants were microinjected into the cytosol of different cell types and their growth compared to their replication upon normal uptake. Our data suggest that not only the metabolic adaptation to the cytosolic compartment is important for the replication of intracytosolic bacteria, but also the mechanism of their uptake as well as the permissiveness of the cytosolic compartment per se.

  • 343.
    Meyer, Lena
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Bröms, Jeanette
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Alam, A.
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    A mutagenesisbased approach to map functional domain(s) within the N- terminus of Francisella tularensis IglEManuscript (preprint) (Other academic)
  • 344.
    Meyer, Lena
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Bröms, Jeanette
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Liu, Xijia
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Rottenberg, M.E.
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Microinjection of Francisella tularensis and Listeria monocytogenes reveals the importance of bacterial and host factors for successful replication2015In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 83, no 8, p. 3233-3242Article in journal (Other academic)
    Abstract [en]

    Certain intracellular bacteria use the host cell cytosol as the replicative niche. Although it has been hypothesized that the successful exploitation of this compartment requires a unique metabolic adaptation, supportive evidence is lacking. For Francisella tularensis, many genes of the Francisella pathogenicity island (FPI) are essential for intracellular growth, and therefore, FPI mutants are useful tools for understanding the prerequisites of intracytosolic replication. We compared the growth of bacteria taken up by phagocytic or nonphagocytic cells with that of bacteria microinjected directly into the host cytosol, using the live vaccine strain (LVS) of F. tularensis; five selected FPI mutants thereof, i.e., Delta iglA, Delta iglC, Delta iglG, Delta iglI, and Delta pdpE strains; and Listeria monocytogenes. After uptake in bone marrow-derived macrophages (BMDM), ASC(-/-) BMDM, MyD88(-/-) BMDM, J774 cells, or HeLa cells, LVS, Delta pdpE and Delta iglG mutants, and L. monocytogenes replicated efficiently in all five cell types, whereas the Delta iglA and Delta iglC mutants showed no replication. After microinjection, all 7 strains showed effective replication in J774 macrophages, ASC(-/-) BMDM, and HeLa cells. In contrast to the rapid replication in other cell types, L. monocytogenes showed no replication in MyD88(-/-) BMDM and LVS showed no replication in either BMDM or MyD88(-/-) BMDM after microinjection. Our data suggest that the mechanisms of bacterial uptake as well as the permissiveness of the cytosolic compartment per se are important factors for the intracytosolic replication. Notably, none of the investigated FPI proteins was found to be essential for intracytosolic replication after microinjection.

  • 345.
    Mistry, Nitesh
    Umeå University, Faculty of Medicine, Clinical Microbiology.
    Human papillomavirus tropism: determinants of viral tissue specificity2007Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Cervical cancer is the second most common cancer among women worldwide and human papillomavirus (HPV) is a prerequisit for the development of this cancer. HPV belongs to the Papillomaviridae family and infects the basal layer of epithelial cells where it generally progresses into warts or condylomas. HPV can only reproduce in differentiating epithelia and it is therefore difficult to study the natural infection of HPV. More than 100 HPV types exist and they are divided into different genera based on their L1 open reading frame sequence. Most of the HPV types in the alpha-papillomavirus genus infect the mucosal epithelium while HPVs from the beta-papillomavirus genus usually infect cutaneous epithelial cells. Presently, it is not known what decides the anatomical tropism and our aim was to study determinants of this tropism.

    By using HPV virus like particles (VLP) and pseudovirus we found that VLPs from the two alpha-papillomaviruses HPV-6 and HPV-16 interacted with cell-surface heparan sulfate (HS) for initial attachment. When we labelled HPV VLPs with a fluorescent dye to study internalization HPV-6 was more strongly inhibited than HPV-16. Furthermore, a pseudovirus infection assay demonstrated that the beta-papillomavirus HPV-5 was less dependent on HS for infection than HPV-16. By analyzing the isoelectric point (p1) of the HPV L1 capsid protein we found that alpha HPV types were more positively charged than beta HPV types. Also, HPV-6 had a higher positive charge than HPV-16. Thus, the inhibition of the negatively charged heparin against HPV infection was clearly related to the charge of the HPV L1 capsid. This suggested that the initial interaction could be one of the determinants of tropism although not the sole factor.

    Lactoferrin is a protein found in milk, saliva, semen, tear fluid and endocervical secretions that has antiviral activities. Both human and bovine lactoferrin inhibited HPV infection but we found no significant differences in inhibition of alpha- and beta-papillomavirus infection. We could however demonstrate that different lactoferricins, small peptide derivates from the N-terminal part of lactoferrin, were able to inhibit HPV infection. This antiviral activity depended on lactoferricin peptide, HPV type and cell origin.

    The regulation of HPV gene expression in the host cell could also determine HPV tropism. The HPV long control region (LCR) contains cis-responsive elements that regulate HPV transcription and the epithelial tropism of HPV is determined by epithelial specific constitutive enhancers in the LCR. It has been hypothesized that the combination of transcription factors in the host cell determines the cell-type-specific expression. In cells with a skin origin the HPV-5 LCR was twice as efficient in transcriptional activation compared to HPV-16 LCR, while in cervical cells the HPV-16 LCR was almost twice as effective in activating transcription compared to HPV-5 LCR.

    To conclude, alpha- and beta-papillomaviruses differed regarding their ability to infect cells and regulate viral gene expression. These abilities corresponded with their natural host cells and suggested that HPV anatomical tropism could be determined at several steps in the HPV life cycle.

  • 346.
    Mistry, Nitesh
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Inoue, Hirotoshi
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Jamshidi, Fariba
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Storm, Rickard J.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Oberste, M. Steven
    Arnberg, Niklas
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Coxsackievirus A24 variant uses aialic acid-containing O-Linked glycoconjugates as cellular receptors on human ocular cells2011In: Journal of Virology, ISSN 0022-538X, E-ISSN 1098-5514, Vol. 85, no 21, p. 11283-11290Article in journal (Refereed)
    Abstract [en]

    Coxsackievirus A24 variant (CVA24v) is a main causative agent of acute hemorrhagic conjunctivitis (AHC), which is a highly contagious eye infection. Previously it has been suggested that CVA24v uses sialic acid-containing glycoconjugates as attachment receptors on corneal cells, but the nature of these receptors is poorly described. Here, we set out to characterize and identify the cellular components serving as receptors for CVA24v. Binding and infection experiments using corneal cells treated with deglycosylating enzymes or metabolic inhibitors of de novo glycosylation suggested that the receptor(s) used by CVA24v are constituted by sialylated O-linked glycans that are linked to one or more cell surface proteins but not to lipids. CVA24v bound better to mouse L929 cells overexpressing human P-selectin glycoprotein ligand-1 (PSGL-1) than to mock-transfected cells, suggesting that PSGL-1 is a candidate receptor for CVA24v. Finally, binding competition experiments using a library of mono- and oligosaccharides mimicking known PSGL-1 glycans suggested that CVA24v binds to Neu5Ac alpha 2,3Gal disaccharides (Neu5Ac is N-acetylneuraminic acid). These results provide further insights into the early steps of the CVA24v life cycle.

  • 347.
    Mohamed, Nahla
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Infectious Diseases.
    Nilsson, Elin
    Johansson, Patrik
    Klingström, Jonas
    Evander, Magnus
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Ahlm, Clas
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Infectious Diseases.
    Bucht, Göran
    Development and evaluation of a broad reacting SYBR-green based quantitative real-time PCR for the detection of different hantaviruses2013In: Journal of Clinical Virology, ISSN 1386-6532, E-ISSN 1873-5967, Vol. 56, no 4, p. 280-285Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Hantaviruses are endemic in most parts of the world and cause hundreds of thousand human cases of hemorrhagic fever with renal syndrome (HFRS) and hantavirus cardiopulmonary syndrome (HCPS) annually throughout Eurasia and the Americas. They are zoonotic viruses, most commonly transmitted to humans by aerosolized rodent excreta. New hantaviruses are frequently discovered in previously unknown reservoir species and geographic areas. Consequently, there is a need to improve hantavirus diagnostics. OBJECTIVES: This paper describes the design and evaluation of a rapid and robust quantitative real-time PCR (QRT-PCR) assay able to detect a wide range of hantaviruses. STUDY DESIGN: Primers with the potential to detect different hantaviruses were designed from conserved regions of different hantavirus L segments, as identified from multiple sequence alignments. RESULTS: By using SYBR-green-based QRT-PCR 100-1000 target molecules of in vitro produced RNA and less than 100 copies of hantavirus RNA from different hantavirus clades and regions of the world were detected. When using the assay on clinical samples from patients with acute HFRS, Puumala hantavirus (PUUV) RNA was confirmed in all previously positive samples. Notably, the broad reacting L-segment QRT-PCR also detected viral RNA in HFRS patient samples, previously negative by a QRT-PCR targeting the S segment of PUUV. CONCLUSIONS: This novel assay provides a powerful tool for diagnosis of hantaviruses from different clades and regions and may also be useful in surveys with the purpose of finding new hantaviruses in rodent or insectivore species.

  • 348.
    Monsen, Tor J.
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Holm, Stig E.
    Ferry, Björn Magnus
    Umeå University, Faculty of Social Sciences, Department of Education.
    Ferry, Sven A.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Department of Public Health and Clinical Medicine, Family Medicine.
    Mecillinam resistance and outcome of pivmecillinam treatment in uncomplicated lower urinary tract infection in women2014In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 122, no 4, p. 317-323Article in journal (Refereed)
    Abstract [en]

    Pivmecillinam (PIV) is a first-line antimicrobial for treatment of lower urinary tract infection in women (LUTIW). Mecillinam, the active substance of PIV, is bactericidal mainly against gram-negative uropathogens, whereas gram-positive species are considered intrinsically resistant. However, successful treatment of LUTIW caused by Staphylococcus saprophyticus has been reported, but more rarely for other gram-positive species. The aim of this study was to compare clinical and bacteriological outcome of PIV vs placebo treatment among uropathogens with special focus on mecillinam-resistant isolates. We analysed data from a prospective, multicentre, placebo-controlled, primary health care, therapy study performed in Sweden in 1995-1998 that included 1143 women with symptoms suggestive of LUTIW. Urine cultures were collected and symptoms registered at inclusion and at follow-up visits. Overall, the efficacy of PIV was superior to that of placebo. Clinical and bacteriological outcomes of PIV treatment were similar for S. saprophyticus, Escherichia coli as for most other uropathogens irrespective of their susceptibility to mecillinam. However, the occurrence of enterococci increased nearly fivefold shortly post PIV treatment, although with mild symptoms and a high spontaneous eradication. As susceptibility to mecillinam in vitro did not predict bacteriological and clinical outcome of PIV treatment, we suggest that the present breakpoints for mecillinam should be revised.

  • 349.
    Monsen, Tor
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Lövgren, Elisabeth
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Widerström, Micael
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Infectious Diseases.
    Wallinder, Lars
    Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences, Orthopaedics.
    In vitro effect of ultrasound on bacteria and suggested protocol for sonication and diagnosis of prosthetic infections2009In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 47, no 8, p. 2496-2501Article in journal (Refereed)
    Abstract [en]

    Sonication of implants has been shown to be a promising method for diagnosis of prosthetic infections due to its improved sensitivity, simplicity, and low cost. The aim of the present study was to evaluate the effects of ultrasound performed under different conditions regarding temperature, duration, and composition of sonication tubes on bacterial species often associated with prosthetic infections. We found that ultrasound had an inhibitory effect on bacteria, of which gram-negative bacteria, in particular Escherichia coli, were almost eradicated after 5 min of sonication at 35 degrees C. Gram-positive bacteria were found to be resistant to the effect of ultrasound. Four factors were important for the inhibitory effect of sonication: the type of microorganism, the temperature of the sonication buffer, the duration of exposure to ultrasound (minutes), and the material and composition of the sonication tube in which sonication is performed. On the basis of the results from the present study, we propose a protocol for sonication and recovery of bacteria associated with biofilm on infected implants prior to conventional culture. From the present protocol, we recommend sonication for 7 min at 22 degrees C at the maximum effect which permits survival of gram-negative bacteria.

  • 350.
    Monsen, Tor
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology. Umeå university hospital.
    Ryden, Patrik
    Umeå University, Faculty of Science and Technology, Department of Mathematics and Mathematical Statistics.
    A new concept and a comprehensive evaluation of SYSMEX UF-1000i flow cytometer to identify culture-negative urine specimens in patients with UTI2017In: European Journal of Clinical Microbiology and Infectious Diseases, ISSN 0934-9723, E-ISSN 1435-4373, Vol. 36, no 9, p. 1691-1703Article in journal (Refereed)
    Abstract [en]

    Urinary tract infections (UTIs) are among the most common bacterial infections in men and urine culture is gold standard for diagnosis. Considering the high prevalence of culture-negative specimens, any method that identifies such specimens is of interest. The aim was to evaluate a new screening concept for flow cytometry analysis (FCA). The outcomes were evaluated against urine culture, uropathogen species and three conventional screening methods. A prospective, consecutive study examined 1,312 urine specimens, collected during January and February 2012. The specimens were analyzed using the Sysmex UF1000i FCA. Based on the FCA data culture negative specimens were identified in a new model by use of linear discriminant analysis (FCA-LDA). In total 1,312 patients were included. In- and outpatients represented 19.6% and 79.4%, respectively; 68.3% of the specimens originated from women. Of the 610 culture-positive specimens, Escherichia coli represented 64%, enterococci 8% and Klebsiella spp. 7%. Screening with FCA-LDA at 95% sensitivity identified 42% (552/1312) as culture negative specimens when UTI was defined according to European guidelines. The proposed screening method was either superior or similar in comparison to the three conventional screening methods. In conclusion, the proposed/suggested and new FCA-LDA screening method was superior or similar to three conventional screening methods. We recommend the proposed screening method to be used in clinic to exclude culture negative specimens, to reduce workload, costs and the turnaround time. In addition, the FCA data may add information that enhance handling and support diagnosis of patients with suspected UTI pending urine culture.

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