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  • 1.
    Aili, Margareta
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Telepnev, Max
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Hallberg, Bengt
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Wolf-Watz, Hans
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Rosqvist, Roland
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    In vitro GAP activity towards RhoA, Rac1 and Cdc42 is not a prerequisite for YopE induced HeLa cell cytotoxicity2003In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 34, no 6, p. 297-308Article in journal (Refereed)
    Abstract [en]

    The YopE cytotoxin of Yersinia is an essential virulence determinant that is translocated into the eukaryotic target cell via a plasmid-encoded type III secretion system. YopE possess a GTPase activating protein activity that in vitro has been shown to down regulate RhoA, Rac1, and Cdc42. Translocated YopE induces de-polymerisation of the actin microfilament structure in the eukaryotic cell which results in a rounding up of infected cells described as a cytotoxic effect. Here, we have investigated the importance of different regions of YopE for induction of cytotoxicity and in vitro GAP activity. Sequential removal of the N- and C-terminus of YopE identified the region between amino acids 90 and 215 to be necessary for induction of cytotoxicity. Internal deletions containing the essential arginine at position 144 resulted in a total loss of cytotoxic response. In-frame deletions flanking the arginine finger defined a region important for the cytotoxic effect to amino acids 166–183. Four triple-alanine substitution mutants in this region, YopE166-8A, 169-71A, 175-7A and 178-80A were still able to induce cytotoxicity on HeLa cells although they did not show any in vitro GAP activity towards RhoA, Rac1 or Cdc42. A substitution mutant in position 206-8A showed the same phenotype, ability to induce cytotoxic response but no in vitro GAP activity. We speculate that YopE may have additional unidentified targets within the eukaryotic cell.

  • 2.
    Bang, Charlotte Sahlberg
    et al.
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden.
    Kruse, Robert
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. Örebro University Hospital.
    Demirel, Isak
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden.
    Önnberg, Anna
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. Dept Lab Med, Örebro University Hospital, Örebro, Sweden.
    Söderquist, Bo
    Örebro University, School of Medicine, Örebro University, Sweden. Örebro University Hospital. Dept Lab Med, Örebro University Hospital, Örebro, Sweden.
    Persson, Katarina
    Örebro University, School of Medicine, Örebro University, Sweden.
    Multiresistant uropathogenic extended-spectrum β-lactamase (ESBL)-producing Escherichia coli are susceptible to the carbon monoxide releasing molecule-2 (CORM-2).2014In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 66, p. 29-35Article in journal (Refereed)
    Abstract [en]

    Carbon monoxide (CO) releasing molecules (CO-RMs) have been shown to inhibit growth of commensal Escherichia coli (E. coli). In the present study we examined the effect of CORM-2 on uropathogenic E. coli (UPEC) that produces extended-spectrum β-lactamase (ESBL). Viability experiments showed that CORM-2 inhibited the growth of several different ESBL-producing UPEC isolates and that 500 μM CORM-2 had a bactericidal effect within 4 h. The bactericidal effect of CORM-2 was significantly more pronounced than the effect of the antibiotic nitrofurantoin. CORM-2 demonstrated a low level of cytotoxicity in eukaryotic cells (human bladder epithelial cell line 5637) at the concentrations and time-points where the antibacterial effect was obtained. Real-time RT-PCR studies of different virulence genes showed that the expression of capsule group II kpsMT II and serum resistance traT was reduced and that some genes encoding iron acquisition systems were altered by CORM-2. Our results demonstrate that CORM-2 has a fast bactericidal effect against multiresistant ESBL-producing UPEC isolates, and also identify some putative UPEC virulence factors as targets for CORM-2. CO-RMs may be candidate drugs for further studies in the field of finding new therapeutic approaches for treatment of uropathogenic ESBLproducing E. coli.

  • 3. Belibasakis, GN
    et al.
    Bostanci, N
    Hashim, A
    Johansson, Anders
    Umeå University, Faculty of Medicine, Odontology, Periodontology.
    Aduse-Opoku, J
    Curtis, MA
    Hughes, FJ
    Regulation of RANKL and OPG gene expression in human gingival fibroblasts and periodontal ligament cells by Porphyromonas gingivalis: a putative role of the Arg-gingipains2007In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 43, no 1, p. 46-53Article in journal (Refereed)
    Abstract [en]

    Porphyromonas gingivalis is highly implicated in the pathogenesis of periodontitis, which is characterized by the destruction of periodontal connective tissues and the supporting alveolar bone. Receptor Activator of NF-kappaB Ligand (RANKL) stimulates bone resorption, whereas osteoprotegerin (OPG) blocks its action, and this bi-molecular system is implicated in periodontitis. The aim of this work was (a) to investigate the regulation of RANKL and OPG gene expression in human periodontal ligament (PDL) cells and gingival fibroblasts (GF), in response to P. gingivalis culture supernatants, by quantitative real-time PCR and (b) to attempt to identify putative virulence factors involved in this process. The results indicated that P. gingivalis induced RANKL and reduced OPG mRNA expression by the studied cells, resulting in an increased RANKL/OPG expression ratio. Heat-inactivation of P. gingivalis resulted in significant reduction of RANKL mRNA expression. A Lys-gingipain mutant strain did not affect, whereas an Arg-gingipain mutant strain further enhanced RANKL mRNA expression, compared to their parental wild-type strain. In conclusion, P. gingivalis up-regulates the RANKL/OPG expression ratio in GF and PDL cells, denoting an enhanced osteoclastogenic potential by the cells. The component mainly responsible for RANKL induction appears to be proteinaceous, and it may be regulated by the Arg-gingipains.

  • 4.
    Demirel, Isak
    et al.
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden.
    Kruse, Robert
    Örebro University, School of Medicine, Örebro University, Sweden.
    Önnberg, Anna
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden.
    Persson, Katarina
    Örebro University, School of Medicine, Örebro University, Sweden.
    Ceftibuten-induced filamentation of extended spectrum beta lactamase (ESBL)-producing uropathogenic Escherichia coli alters host cell responses during an in vitro infection2015In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 78, p. 52-62Article in journal (Refereed)
    Abstract [en]

    Inadequate and delayed antibiotic treatment of extended spectrum beta-lactamase (ESBL)-producing isolates have been associated with increased mortality of affected patients. The purpose of this study was to compare the host response of human renal epithelial cells and polymorphonuclear leucocyte (PMN) cells when infected by ESBL-producing uropathogenic Escherichia coli (UPEC) isolates in the presence or absence of ineffective antibiotics.

    The renal epithelial cell line A498 and PMN cells were stimulated with ESBL-producing UPEC isolates in the presence or absence of three different antibiotics (trimetoprim, ceftibuten and ciprofloxacin). Host cell responses were evaluated as release of cytokines (IL-6, IL-8), reactive oxygen species (ROS), ATP and endotoxins. Bacterial morphology and PMN phagocytosis were evaluated by microscopy.

    In the presence of ceftibuten, 2 out of 3 examined ESBL-isolates changed their morphology into a filamentous form. The presence of ceftibuten enhanced IL-6, IL-8 and ROS-production from host cells, but only from cells stimulated by the filamentous isolates. The bacterial supernatant and not the filamentous bacteria per se was responsible for the increased release of IL-6, IL-8 and ROS. Increased endotoxin and ATP levels were found in the bacterial supernatants from filamentous isolates. Apyrase decreased IL-6 secretion from A498 cells and polymyxin B abolished the increased ROS-production from PMN cells. PMN were able to inhibit the bacterial growth of some ESBL-isolates in the presence of ceftibuten.

    In conclusion, antibiotic-induced filamentation of ESBL-producing UPEC isolates and the associated release of ATP and endotoxins can alter the host cell response in the urinary tract.

  • 5.
    Fröman, Gunnar
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Amin, Kawa
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Yacoub, Alia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Mård, Per-Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Binding of type-I collagen to Chlamydia trachomatis1994In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 17, no 4, p. 253-259Article in journal (Refereed)
    Abstract [en]

    Denatured type-I collagen was found to bind to Chlamydia trachomatis elementary bodies (EBs) in a time-dependent and specific manner. Specificity was tested by having a large excess of other proteins present in the binding mixture. Only denatured type-I collagen efficiently competed for binding. Radiolabelled fibronectin did not bind under the test conditions used. The binding was temperature-dependent and the interaction increased at the melting temperature of the collagen. Evidence was found for two binding sites: one with high affinity (Kd 3.3 x 10(-9)) and one with low affinity (Kd 1.7 x 10(-7)), with an estimated number of binding sites per EB of 590 and 2900, respectively. The interaction between C. trachomatis and collagen may also be relevant in vivo, since 50% binding occurred at 37 degrees C. The binding to denatured collagen may be of importance for the development of sexually acquired reactive arthritis.

  • 6. Hamid, N
    et al.
    Gustavsson, A
    Andersson, K
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology.
    McGee, K
    Persson, C
    Rudd, CE
    Fällman, M
    YopH dephosphorylates Cas and Fyn-binding protein in macrophages. 1999In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 27, p. 231-242Article in journal (Refereed)
  • 7.
    Hamid, Nivia
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Gustavsson, Anna
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Andersson, Kerstin
    McGee, Karen
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Persson, Cathrine
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Rudd, Christopher E.
    Fällman, Maria
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    YopH dephosphorylates Cas and Fyn-binding protein in macrophages1999In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 27, no 4, p. 231-242Article in journal (Refereed)
    Abstract [en]

    The tyrosine phosphatase YopH is an essential virulence effector of pathogenic Yersinia spp. YopH, which is translocated from extracellularly located bacteria into interacting target cells, blocks phagocytosis by professional phagocytes. We show here that immunoprecipitation of YopH from lysates of J774 cells infected with Y. pseudotuberculosis expressing an inactive form of YopH resulted in co-precipitation of certain phosphotyrosine proteins. The association between the inactive YopH and phosphotyrosine proteins in the 120 kDa range was rapid and could be detected after 2 min of infection. The proteins were identified as the docking proteins Cas and Fyn-binding protein (FYB). Upon infection of J774 cells with Y. pseudotuberculosis lacking YopH expression both of these proteins became tyrosine phosphorylated. Moreover, this infection caused recruitment of Cas to peripheral focal complexes, and FYB was relocalized to areas surrounding these structures. Both Cas and FYB became dephosphorylated upon infection with Y. pseudotuberculosis expressing active YopH, and this was associated with disruption of focal complexes. With regard to the previous identification of Cas and focal complexes as targets of YopH in HeLa cells, the present study supports an important role for these targets in a general mechanism of bacterial uptake. 

  • 8.
    Jonsson, Maria
    et al.
    Umeå University, Faculty of Medicine, Microbiology.
    Elmros, Teodor
    Umeå University, Faculty of Medicine, Microbiology.
    Bergström, Sven
    Umeå University, Faculty of Medicine, Microbiology.
    Subcutaneous implanted chambers in different mouse strains as an animal model to study genetic stability during infection with Lyme disease Borrelia1995In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 18, no 2, p. 109-14Article in journal (Refereed)
    Abstract [en]

    Tissue metal net cages were implanted subcutaneously in BALB/cJ and C3H/Tif mice as an experimental model of Borrelia burgdorferi infection. B. burgdorferi sensu stricto strain Sh2-82 could be isolated up to 14 weeks after the inoculation. However, a significant difference in infectivity between the two mice strains was observed. C3H/Tif mice were more susceptible to developing chronic B. burgdorferi s.s. infections than BALB/cJ mice. Although a B. burgdorferi infection was established, no rearrangements in the ospA and ospB genes were observed in any of the infected mice.

  • 9.
    Kumar, Anjani
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Manisha, --
    Sangha, Gurkamaljit Kaur
    Shrivastava, Anju
    Kaur, Jagdeep
    The immunosuppressive effects of a novel recombinant LipQ (Rv2485c) protein of Mycobacterium tuberculosis on human macrophage cell lines2017In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 107, p. 361-367Article in journal (Refereed)
    Abstract [en]

    Mycobacterium tuberculosis (MTB), an intracellular pathogen, still represents a major global health challenge. A number of mycobacterial macromolecules have been shown to target biological processes within host macrophages; however, the exact mechanism for the majority of these host pathogen interactions is still poorly understood. Moreover, the lipid metabolic pathway is one of the most important physiologic pathways that plays a vital role in the survival and infection of Mycobacterium tuberculosis. In present study, we investigated the effect of rLipQ from Mycobacterium tuberculosis H37Rv on macrophage functions invitro.Our results demonstrate that rLipQ significantly lowers the expression level of pro-inflammatory cytokines (TNF-alpha& IFN-gamma) and augments the level of anti inflammatory cytokines such as IL-4 & IL-10as compared to LPS stimulated macrophages. An up-regulation of anti-inflammatory and down-regulation of pro-inflammatory cytokines levels in rLipQ pretreated macrophages implies immuno-modulatory functions in TB patients. Interestingly, rLipQ also inhibited the expression of iNOS, TLR-2 and transcription factor NF-kB in LPS stimulated macrophages whereas the expression of TLR-4 remains unchanged. The inhibition in the expression of these signaling molecules has been correlated to the inhibition of NO production in macrophages. Taken together, these studies demonstrate that rLipQ is a novel lipase that is highly immunogenic and may play an important role in the virulence and pathogenesis of M.tuberculosis infection, by altering the balance of cytokines, which might help to assess prognosis and contribute to a better understanding against host-pathogen interactions.

  • 10.
    Lai, Xin-He
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Golovliov, Igor
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Expression of iglC is mecessary for intracellular growth and induction of apoptosis in murine macrophages by Francisella tularensis2004In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 37, no 5, p. 225-230Article in journal (Refereed)
    Abstract [en]

    Francisella tularensis is a facultative intracellular bacterium capable of inducing apoptosis in murine macrophages. In a previous study, an iglC null mutant of F. tularensis live vaccine strain LVS was generated by allelic replacement and in the current study this iglC mutant was successfully complemented in trans. We characterized the capacity of this iglC mutant and the complemented strain to induce macrophage apoptosis. The iglC mutant did not induce apoptosis in the infected cells. In contrast, the complemented iglC strain was able to multiply in the murine macrophage-like cell line J774A.1 and induced apoptosis similar to that of the wild-type strain. It is the first successful example of complementation in trans of a F. tularensis mutant strain and more importantly this work provides direct evidence that the intracellular growth ability is essential for F. tularensis to induce macrophage apoptosis.

  • 11.
    Larsson, Christer
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Lundqvist, Jenny
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Bergström, Sven
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Residual brain infection in murine relapsing fever borreliosis can be successfully treated with ceftriaxone2008In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 44, no 3, p. 262-264Article in journal (Refereed)
    Abstract [en]

    Like several other spirochetes, relapsing fever Borrelia can cause persistent infection of the central nervous system (CNS). By treating mice harboring residual Borrelia duttonii brain infection with the bacteriocidal, cell wall inhibiting antibiotic ceftriaxone, bacteria were cleared from the brain. This shows that the residual infection is not latent but actively growing.

  • 12.
    Leary, Sophie E. C.
    et al.
    Biomedical Sciences Department, Defence Evaluation and Research Agency, Porton Down, Salisbury, Wiltshire, SP4 0JQ, U.K..
    Griffin, Kate F.
    Biomedical Sciences Department, Defence Evaluation and Research Agency, Porton Down, Salisbury, Wiltshire, SP4 0JQ, U.K..
    Galyov, Edouard E.
    Institute for Animal Health, Compton, Nr Newbury, Berkshire RG20 7NN, U.K..
    Hewer, Jason
    Biomedical Sciences Department, Defence Evaluation and Research Agency, Porton Down, Salisbury, Wiltshire, SP4 0JQ, U.K..
    Williamson, E. Diane
    Biomedical Sciences Department, Defence Evaluation and Research Agency, Porton Down, Salisbury, Wiltshire, SP4 0JQ, U.K..
    Holmström, Anna
    Department of Microbiology, Defence Research Establishment, S-901 87 Umeå, Sweden.
    Forsberg, Åke Forsberg
    Department of Microbiology, Defence Research Establishment, S-901 87 Umeå, Sweden.
    Titball, Richard W.
    Biomedical Sciences Department, Defence Evaluation and Research Agency, Porton Down, Salisbury, Wiltshire, SP4 0JQ, U.K..
    Yersinia outer proteins (YOPS) E, K and N are antigenic but non-protective compared to V antigen, in a murine model of bubonic plague1999In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 26, no 3, p. 159-169Article in journal (Refereed)
    Abstract [en]

    The pathogenic Yersiniae produce a range of virulence proteins, encoded by a 70 kb plasmid, which are essential for infection, and also form part of a contact-dependent virulence mechanism. One of these proteins, V antigen, has been shown to confer a high level of protection against parenteral infection with Y. pestis in murine models, and is considered to be a protective antigen. In this study, the protective efficacy of V antigen has been compared in the same model with that of other proteins (YopE, YopK and YopN), which are part of the contact-dependent virulence mechanism. Mice immunised with two intraperitoneal doses of V antigen or each of the Yops, administered with either Alhydrogel or interleukin-12, produced high antigen-specific serum IgG titres. As shown in previous studies, V+Alhydrogel was fully protective, and 5/5 mice survived a subcutaneous challenge with 90 or 9x10(3) LD50's of Y. pestis GB. In addition, these preliminary studies also showed that V+IL-12 was partially protective: 4/5 or 3/5 mice survived a challenge with 90 or 9x10(3) LD50's, respectively. In contrast, none of the mice immunised with the Yops survived the challenges, and there was no significant delay in the mean time to death compared to mice receiving a control protein. These results show that using two different vaccine regimens, Yops E, K and N, failed to elicit protective immune responses in a murine model of plague, whereas under the same conditions, V antigen was fully or partially protective.

  • 13.
    Majeed, Meytham
    et al.
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Perskvist, Nasrin
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Ernst, Joel D.
    Division of Infectious Diseases and Rosalind Russell Research Laboratory, University of California, San Francisco and San Francisco General Hospital, San Francisco, CA, U.S.A..
    Orselius, Kristina
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Stendahl, Olle
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Roles of calcium and annexins in phagocytosis and elimination of an attenuated strain of Mycobacterium tuberculosisin human neutrophils1998In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 24, no 5, p. 309-320Article in journal (Refereed)
    Abstract [en]

    The phagocytic function of neutrophils is a crucial element in the host defence against invading microorganisms. We investigated phagocytosis and intracellular killing of an attenuated strain of Mycobacterium tuberculosis(H37Ra) by human neutrophils focusing on the role of the cytosolic free calcium concentration [Ca2+]iand certain cytosolic calcium-dependent membrane-binding proteins annexins. Phagocytic uptake did not trigger a calcium rise and occurred independently of different calcium conditions, and in a serum-dependent manner. Changes in the viability of H37Ra were determined by agar plate colony count and a radiometric assay. Neutrophils showed a capacity to kill ingested mycobacteria and this occurred without a rise in [Ca2+]i. The ability to kill H37Ra decreased in the absence of extracellular calcium and when intra-extracellular calcium was reduced. Immunofluorescence staining revealed that during phagocytosis of H37Ra, annexins III, IV and VI translocated from cytoplasm to the proximity of the H37Ra-containing phagosomes, whereas the localization of annexin I and V remained unchanged. The translocation of annexin IV occurred even when Ca2+-depleted neutrophils ingested H37Ra in the absence of extracellular calcium. We concluded that neutrophil-mediated killing of mycobacteria is a Ca2+-dependent process. The fact that the association of certain annexins to the membrane vesicle containing H37Ra differ from other phagosomes suggests a selective regulatory mechanism during phagocytosis of mycobacteria by neutrophils.

  • 14.
    Melhus, Åsa
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Ryan, Allen F.
    Expression of molecular markers for bone formation increases during experimental acute otitis media2001In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 30, no 3, p. 111-120Article in journal (Refereed)
    Abstract [en]

    Bony tissues are integral parts of the function of the middle ear and the protection of adjacent vital structures. To explore the reaction of middle ear bone to acute otitis media, rats were challenged with Streptococcus pneumoniae and Haemophilus influenzae. Local changes were monitored for up to 1 month. After reverse transcription, competitive polymerase chain reaction was used to determine the expression levels of two molecular markers of bone formation, osteocalcin and procollagen I, and the two cytokines interleukin (IL)-6 and tumor necrosis factor (TNF)-alpha, in the bone. Middle ear bone responded rapidly to bacterial challenge, and the reaction depended upon the causative agent. On day 1, IL-6 and TNF-alpha transcripts were detected in the bone from all middle ears. After a short period of decreased expression of osteocalcin, during which the otitis diagnosis could not be made clinically, the levels of bone formation markers increased dramatically. The maximum levels of these markers were reached on days 6 and 14 for animals challenged with H. influenzae and pneumococci, respectively. Infections induced by pneumococci had a longer duration, and after the initial phase the production of osteocalcin and procollagen transcript were significantly higher in the pneumococcus-infected animals. The results indicate that even in an uncomplicated infection, the bone of the bulla reacts to an acute otitis media with a short period of inhibited osteoblast activity followed by a longer period of new bone formation.

  • 15. O'Toole, Ronan
    et al.
    von Hofsten, Jonas
    Rosqvist, Roland
    Olsson, Per-Erik
    Örebro University, School of Science and Technology.
    Wolf-Watz, Hans
    Visualisation of zebrafish infection by GFP-labelled Vibrio anguillarum2004In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 37, no 1, p. 41-46Article in journal (Refereed)
    Abstract [en]

    Vibrio anguillarum is an invasive pathogen of fish causing a septicaemia called vibriosis. In this work, transparent zebrafish were immersed in water containing green fluorescent protein labelled V. anguillarum. The infection was visualised at the whole fish and single bacterium levels using microscopy. The gastrointestinal tract was the first site where the pathogen was detected. This enteric localisation occurred independently of the flagellum or motility. On the other hand, chemotactic motility was essential for association of the pathogen with the fish surface. In conclusion, the zebrafish infection model provides evidence that the intestine and skin represent sites of infection by V. anguillarum and suggests a host site where chemotaxis may function in virulence.

  • 16.
    Persson, Alexander
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Blomgran, Robert
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Eklund, Daniel
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences.
    Lundstrom, Charlotte
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Stendahl, Olle
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Induction of apoptosis in human neutrophils by Mycobacterium tuberculosis is dependent on mature bacterial lipoproteins2009In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 47, no 3, p. 143-150Article in journal (Refereed)
    Abstract [en]

    Modulation of immune cell apoptosis is a key evasion strategy utilized by Mycobacterium tuberculosis (Mtb). To be able to multiply within macrophages, the bacterium delays apoptosis and down-regulates pro-inflammatory activation in these cells, whereas apoptosis is rapidly induced in the potently bactericidal neutrophils. Initial host-pathogen interactions between neutrophils and Mtb, subsequently leading to apoptosis, need to be investigated to understand the early features during Mtb infections. Opsonized Mtb were readily phagocytosed, and the immuno-mediated phagocytosis triggered early activation of anti-apoptotic Akt in the neutrophils but the bacteria still induced apoptosis to the same extent as non-phagocytosed Mtb. Mtb-induced apoptosis was strictly dependent on NADPH oxidase-generated reactive oxygen species, compounds shown to damage lysosomal granules. Despite this, we found no involvement of damaged azurophilic granules in Mtb-induced apoptosis in human neutrophils. Instead, the Mtb-induced apoptosis was p38 MAPK dependent and induced through the mitochondrial pathway. Moreover, Mtb deficient of mature lipoproteins lacked the determinants required for induction of neutrophil apoptosis. These results show that Mtb exert a strong intrinsic capacity to induce apoptosis in neutrophils that is capable of overcoming the anti-apoptotic signaling in the cell.

  • 17.
    Persson, Alexander
    et al.
    Department of Molecular and Clinical Medicine, Linköping University, Sweden.
    Blomgran-Julinder, Robert
    Division of Medical Microbiology, Department of Molecular and Clinical Medicine, Faculty of Health Sciences, Linköping University, Linköping, Sweden.
    Eklund, Daniel
    Division of Medical Microbiology, Department of Molecular and Clinical Medicine, Faculty of Health Sciences, Linköping University, Linköping, Sweden.
    Lundström, Charlotte
    Division of Medical Microbiology, Department of Molecular and Clinical Medicine, Faculty of Health Sciences, Linköping University, Linköping, Sweden.
    Stendahl, Olle
    Division of Medical Microbiology, Department of Molecular and Clinical Medicine, Faculty of Health Sciences, Linköping University, Linköping, Sweden.
    Induction of apoptosis in human neutrophils by Mycobacterium tuberculosis is dependent on mature bacterial lipoproteins2009In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 47, no 3, p. 143-150Article in journal (Refereed)
    Abstract [en]

    Modulation of immune cell apoptosis is a key evasion strategy utilized by Mycobacterium tuberculosis (Mtb). To be able to multiply within macrophages, the bacterium delays apoptosis and down-regulates pro-inflammatory activation in these cells, whereas apoptosis is rapidly induced in the potently bactericidal neutrophils. Initial host-pathogen interactions between neutrophils and Mtb, subsequently leading to apoptosis, need to be investigated to understand the early features during Mtb infections. Opsonized Mtb were readily phagocytosed, and the immuno-mediated phagocytosis triggered early activation of anti-apoptotic Akt in the neutrophils but the bacteria still induced apoptosis to the same extent as non-phagocytosed Mtb. Mtb-induced apoptosis was strictly dependent on NADPH oxidase-generated reactive oxygen species, compounds shown to damage lysosomal granules. Despite this, we found no involvement of damaged azurophilic granules in Mtb-induced apoptosis in human neutrophils. Instead, the Mtb-induced apoptosis was p38 MAPK dependent and induced through the mitochondrial pathway. Moreover, Mtb deficient of mature lipoproteins lacked the determinants required for induction of neutrophil apoptosis. These results show that Mtb exert a strong intrinsic capacity to induce apoptosis in neutrophils that is capable of overcoming the anti-apoptotic signaling in the cell.

  • 18.
    Schöier, Johan
    et al.
    Linköping University, Department of Molecular and Clinical Medicine, Clinical Microbiology. Linköping University, Faculty of Health Sciences.
    Öllinger, Karin
    Linköping University, Department of Neuroscience and Locomotion, Pathology. Linköping University, Faculty of Health Sciences.
    Kvarnström, Maria
    Linköping University, Department of Molecular and Clinical Medicine, Clinical Immunology. Linköping University, Faculty of Health Sciences.
    Söderlund, Gustaf
    Linköping University, Department of Biomedicine and Surgery, Oncology. Linköping University, Faculty of Health Sciences.
    Kihlström, Erik
    Linköping University, Department of Molecular and Clinical Medicine, Clinical Microbiology. Linköping University, Faculty of Health Sciences.
    Chlamydia trachomatis -induced apoptosis occurs in uninfected McCoy cells late in the developmental cycle and is regulated by the intracellular redox state2001In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 31, no 4, p. 173-184Article in journal (Refereed)
    Abstract [en]

    Infections with the obligate intracellular bacterium Chlamydia trachomatis are characterized by avoidance of fusion between chlamydia-containing endosomes and lysosomes, bacterial persistence and development of post-infectious sequelae. In this report we show that C. trachomatis induces apoptosis in McCoy and HeLa cells. Apoptosis was monitored by three different techniques; enzyme-linked immunoassay (EIA) of fragmented nucleosomes, terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) and flow cytometry of propidium iodide-stained cells. Apoptosis occurred in uninfected cells, was induced late in the chlamydial developmental cycle, beyond 24 h post-infection and was dependent on bacterial protein synthesis. Apoptosis was not significantly increased in infected, inclusion-containing cells. Treatment of cells with the antioxidants ascorbic acid (10 μM) and α-tocopherol (10 μM) reduced the degree of apoptosis. These results suggest that host cells infected with C. trachomatis generate proapoptotic stimuli that induce apoptosis in uninfected, neighbouring cells and that the redox state of the cell is a regulator in chlamydia-induced apoptosis.

  • 19. Sjöström, Annika
    Analysis of the sfaXIIlocus in the Escherichia coli meningitis isolate IHE3034 reveals two novel regulatory genes within the promoter-distal region of the main S fimbrial operon2009In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208Article in journal (Refereed)
  • 20.
    Sjöström, Annika E.
    et al.
    Umeå University, Faculty of Medicine, Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Balsalobre, Carlos
    Umeå University, Faculty of Medicine, Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Emödy, Levente
    Department of Medical Microbiology and Immunology, University medical School, Pecs, Hungary.
    Westerlund-Wikström, Benita
    General Microbiology, Faculty of Biosciences, University of Helsinki, Finland.
    Hacker, Jörg
    Robert-Koch-Institut, Nordufer, Berlin, Germany.
    Uhlin, Bernt Eric
    Umeå University, Faculty of Medicine, Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    The SfaXII protein from newborn meningitis E. coli is involved in regulation of motility and type 1 fimbriae expression2009In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 46, no 5, p. 243-252Article in journal (Refereed)
    Abstract [en]

    The genomes of pathogenic E. coli may contain several different fimbrial operons. How bacteria regulate and coordinate the choice of fimbrial expression under different circumstances remains largely unanswered. In this report we have investigated the role of the sfaXII gene associated to the SfaII fimbrial determinant in the E. coli isolate IHE3034. sfaXII belongs to a subfamily of genes, the 17kDa genes, located near different fimbrial operons in uropathogenic and newborn meningitis E. coli (NMEC) strains. Using the NMEC isolate IHE3034 and non-pathogenic E. coli strains we found that the sfaXII gene had an inhibitory effect on type 1 fimbriae expression. Down regulation of type 1 fimbriae was exerted at transcriptional level both by inhibiting expression from the fimA promoter and by reducing the frequency of OFF-to-ON switching. The effect of sfaXII on expression of the recombinase FimB that catalyzes OFF to ON switching might explain the described reduction in percentage of ON cells. Moreover, expression of the sfaXII gene strongly influenced motility and flagella production of the NMEC isolate IHE3034. We propose that the sfaXII gene, and presumably other members in the 17kDa gene family, may play a role in the control of virulence related gene expression in pathogenic E. coli.

  • 21.
    Sjöström, Annika E.
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Sondén, Berit
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Müller, Claudia
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Rydström, Anna
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Dobrindt, Ulrich
    University of Würzburg, Institute for Molecular Infection Biology, D-97070 Würzburg, Germany.
    Uhlin, Bernt Eric
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Analysis of the sfaXII locus in the Escherichia coli meningitis isolate IHE3034 reveals two novel regulatory genes within the promoter-distal region ofthe main S fimbrial operon2009In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 46, no 3, p. 150-158Article in journal (Refereed)
    Abstract [en]

    We describe the expression and regulation of the gene sfaXII located near the SfaII fimbrial determinant inthe newborn meningitis Escherichia coli (NMEC) isolate IHE3034. sfaXII belongs to a gene family, the 17-kDa genes, typically located downstream (300–3000 bp) of different fimbrial operons found in E. coli isolates of uropathogenic and newborn meningitis origin. Using transcriptional sfaXII reporter genefusions we found that different environmental conditions commonly affecting expression of fimbrialgenes also affected sfaXII expression. Analysis of the sfaXII transcripts showed that the gene is part of themain fimbrial operon as it is transcribed together with the rest of the fimbrial genes. In addition, the sfaXII gene can be expressed from a more proximal promoter and is found to be subject to strong downregulationby the nucleoid protein H-NS. Studies with an sfaXII mutant derivative of IHE3034 did notreveal effects on SfaII fimbrial biogenesis as monitored by e.g. immunofluorescence microscopy. Nevertheless,a mutation in sfaXII resulted in altered expression of other surface components. Moreover, we define a new gene, sfaYII, coding for a putative phosphodiesterase that is located in between the sfaXII gene and the fimbrial biogenesis genes. Our studies by ectopic expression of sfaYII in Vibrio cholerae showed that the gene product caused reduced biofilm formation and it is proposed that sfaYII caninfluence cyclic-di-GMP turnover in the bacteria. Our findings demonstrate that the operons typical for S-fimbriae of extraintestinal pathogenic E. coli include previously unrecognized novel regulatory genes.

  • 22.
    Sjöström, Annika E.
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Sondén, Berit
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Müller, Claudia
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Rydström, Anna
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Dobrindt, Ulrich
    Institute for Molecular Infection Biology, University of Würzburg, Germany.
    Wai, Sun Nyunt
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Uhlin, Bernt Eric
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Analysis of the sfaXII locus in the Escherichia coli meningitis isolate IHE3034 reveals two novel regulatory genes within the promoter-distal region of the main S fimbrial operon2009In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 46, no 3, p. 150-158Article in journal (Refereed)
    Abstract [en]

    We describe the expression and regulation of the gene sfaXII located near the SfaII fimbrial determinant in the newborn meningitis Escherichia coli (NMEC) isolate IHE3034. sfaXII belongs to a gene family, the 17kDa genes, typically located downstream (300 – 3000 bp) of different fimbrial operons found in E. coli isolates of uropathogenic and newborn meningitis origin. Using transcriptional sfaXII reporter gene fusions we found that different environmental conditions commonly affecting expression of fimbrial genes also affected sfaXII expression. Analysis of the sfaXII transcripts showed that the gene is part of the main fimbrial operon as it is transcribed together with the rest of the fimbrial genes. In addition, the sfaXII gene can be expressed from a more proximal promoter and is found to be subject to strong down-regulation by the nucleoid protein H-NS. Studies with an sfaXII mutant derivative of IHE3034 did not reveal effects on SfaII fimbrial biogenesis as monitored by e.g. immunofluorescence microscopy. Nevertheless, a mutation in sfaXII resulted in altered expression of other surface components. Moreover, we define a new gene, sfaYII, coding for a putative phosphodiesterase that is located in between the sfaXII gene and the fimbrial biogenesis genes. Our studies by ectopic expression of sfaYII in Vibrio cholerae showed that the gene product caused reduced biofilm formation and it is proposed that sfaYII can influence cyclic-di-GMP turnover in the bacteria. Our findings demonstrate that the operons typical for S-fimbriae of extraintestinal pathogenic E. coli include previously unrecognized novel regulatory genes.

  • 23.
    Sundin, Charlotta
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Thelaus, Johanna
    Bröms, Jeanette E
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Forsberg, Åke
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Polarisation of type III translocation by Pseudomonas aeruginosa requires PcrG, PcrV and PopN2004In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 37, no 6, p. 313-322Article in journal (Refereed)
    Abstract [en]

    Type III secretion (TTS) mediated translocation of exoenzymes is a key virulence strategy utilised by the opportunistic pathogen Pseudomonas aeruginosa to deliver exoenzyme effectors into the eukaryotic cell. We have previously shown that type III mediated translocation is a contact dependent process, which requires the secreted translocator proteins PerV, PopB and PopD. To further analyse this mechanism, HeLa cells were infected with the wild-type strain PAK as well as isogenic pcrV, popB, popD, pcrG and popN mutants. In the presence of eukaryotic cells, expression of exoenzyme S (ExoS) increased. When cells were infected with the wild-type strain PAK no ExoS was detected in the tissue culture medium. This confirms that ExoS translocation by P. aeruginosa occurs by a polarised mechanism. In contrast, high levels of ExoS were recovered in the tissue Culture medium when cells were infected with pcrG, pcrV and popN mutants. Additionally, ExoS expression levels were higher for these mutants regardless of inducing conditions. This suggests that PcrG, PcrV and PopN are involved in negative regulation of ExoS expression and secretion, and are required to ensure polarised delivery of effectors into target cells.

  • 24.
    Svensson, Lovisa
    et al.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Poljakovic, Mirjana
    Karolinska Institutet.
    Säve, Susanne
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Gilberthorpe, Nicola
    University of Sheffield, UK.
    Schön, Thomas
    Kalmar County Hospital.
    Strid, Sigge
    Stensö Health Center, Kalmar County Council.
    Corker, Hazel
    University of Sheffield, UK.
    Poole, Robert K.
    University of Sheffield, UK.
    Persson, Katarina
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences. Örebro University.
    Role of flavohemoglobin in combating nitrosative stress in uropathogenic Escherichia coli – implications for urinary tract infection2010In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 49, no 3, p. 59-66Article in journal (Refereed)
    Abstract [en]

    During the course of urinary tract infection (UTI) nitric oxide (NO) is generated as part of the host response. This study investigates the significance of the NO-detoxifying enzyme flavohemoglobin (Hmp) in protection of uropathogenic Escherichia coli (UPEC) against nitrosative stress. An hmp (J96Δhmp) knockout mutant of UPEC strain J96 was constructed using single-gene deletion. The viability of J96Δhmp was significantly reduced (P < 0.001) compared to the wild-type strain after exposure to the NO-donor DETA/NO. The NO consumption in J96Δhmp was significantly (P < 0.001) impaired compared to J96wt. Screening UPEC isolates from patients with UTI revealed increased hmp expression in all patients. In a competition-based mouse model of UTI, the hmp mutant strain was significantly (P < 0.05) out-competed by the wild-type strain. This study demonstrates, for the first time, that Hmp contributes to the protection of UPEC against NO-mediated toxicity in vitro. In addition, hmp gene expression occurs in UPEC isolates from the infected human urinary tract and UPEC that were hmp-deficient had a reduced ability to colonize the mouse urinary tract. Taken together the results suggest that NO detoxification by Hmp may be a fitness advantage factor in UPEC, and a potentially interesting target for development of novel treatment concepts for UTI.

  • 25. Svensson, Lovisa
    et al.
    Poljakovic, Mirjana
    Säve, Susanne
    Gilberthorpe, Nicola
    Schön, Thomas
    Strid, Sigge
    Corker, Hazel
    Poole, Robert K.
    Persson, Katarina
    Örebro University, School of Health and Medical Sciences.
    Role of flavohemoglobin in combating nitrosative stress in uropathogenic Escherichia coli: implications for urinary tract infection2010In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 49, no 3, p. 59-66Article in journal (Refereed)
    Abstract [en]

    During the course of urinary tract infection (UTI) nitric oxide (NO) is generated as part of the host response. This study investigates the significance of the NO-detoxifying enzyme flavohemoglobin (Hmp) in protection of uropathogenic Escherichia coli (UPEC) against nitrosative stress. An hmp (J96Deltahmp) knockout mutant of UPEC strain J96 was constructed using single-gene deletion. The viability of J96Deltahmp was significantly reduced (P<0.001) compared to the wild-type strain after exposure to the NO-donor DETA/NO. The NO consumption in J96Deltahmp was significantly (P<0.001) impaired compared to J96wt. Screening UPEC isolates from patients with UTI revealed increased hmp expression in all patients. In a competition-based mouse model of UTI, the hmp mutant strain was significantly (P<0.05) out-competed by the wild-type strain. This study demonstrates, for the first time, that Hmp contributes to the protection of UPEC against NO-mediated toxicity in vitro. In addition, hmp gene expression occurs in UPEC isolates from the infected human urinary tract and UPEC that were hmp-deficient had a reduced ability to colonize the mouse urinary tract. Taken together the results suggest that NO detoxification by Hmp may be a fitness advantage factor in UPEC, and a potentially interesting target for development of novel treatment concepts for UTI.

  • 26.
    Tavares, Raquel
    et al.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Pathak, Sushil Kumar
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Induction of TNF, CXCL8 and IL-1 beta in macrophages by Helicobacter pylori secreted protein HP1173 occurs via MAP-kinases, NF-kappa B and AP-1 signaling pathways2018In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 125, p. 295-305Article in journal (Refereed)
    Abstract [en]

    Macrophages play an important role in the generation of host immune responses against H. pylori. In this study, we investigated the effect of a functionally uncharacterized H. pylori secreted protein HP1173 on macrophage responses. In a screen of eight H. pylori strains, similar expression levels of the HP1173 protein were observed in the whole-cell extracts, however; the amount of protein released into the culture medium varied significantly among strains. Recombinant purified HP1173 (rHP1173) was found to bind to THP-1 cells that were differentiated into macrophages via phorbol-12-myristate-13-acetate (PMA) treatment. The exposure of macrophages to rHP1173 led to the production of the proinflammatory cytokines TNF and IL-1 beta and the chemokine CXCL8 in a dose- and time-dependent manner. Under similar conditions, rHP1173 failed to induce apoptosis in macrophages. Furthermore, rHP1173-induced expression of TNF, IL-1 beta and CXCL8 was observed at the level of gene transcription. Incubation of macrophages in the conditioned medium from a mutant H. pylori strain 26695 lacking HP1173 protein expression resulted in the reduced induction of TNF, CXCL8 and IL-1 beta, confirming the role of the endogenous protein. Intracellular signaling involving MAPKs, NF-kappa B and the AP-1 family of transcription factors was required for rHP1173-induced TNF, CXCL8 and IL-1 beta release from macrophages. The blocking of MyD88, which is an adaptor for multiple toll-like receptors (TLRs), had no effect on rHP1173-induced TNF, CXCL8 and IL-1 beta release from macrophages, suggesting that Myd88-dependent TLR signaling was not involved in the recognition of and responses to rHP1173. These findings provide novel insights into the potential role of HP1173 in H. pylori infection-associated disease development.

  • 27.
    Telepnev, Max
    et al.
    Umeå University, Faculty of Medicine, Clinical Microbiology, Clinical Bacteriology.
    Golovliov, Igor
    Umeå University, Faculty of Medicine, Clinical Microbiology, Clinical Bacteriology.
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Clinical Microbiology, Clinical Bacteriology.
    Francisella tularensis LVS initially activates but subsequently down-regulates intracellular signaling and cytokine secretion in mouse monocytic and human peripheral blood mononuclear cells.2005In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 38, no 5-6, p. 239-247Article in journal (Refereed)
    Abstract [en]

    Monocytic cells constitute an important defense mechanism against invading pathogens by recognizing conserved pathogens components. The recognition leads to activation of intracellular pathways involving nuclear factor kappa B (NF-kappaB) and mitogen-activated protein kinases (MAPK), such as the c-Jun NH2-terminal kinase (JNK), and p38. We show that in vitro infection with Francisella tularensis results in activation of NF-kappaB, phosphorylation of p38 and c-Jun, and secretion of TNF-alpha in adherent mouse peritoneal cells, in the mouse macrophage-like cell line J774A.1, in the human macrophage cell line THP-1, and in human peripheral blood monocytic cells. This occurred after infection with the human live vaccine strain, F. tularensis LVS or a mutant strain denoted deltaiglC, which lacks expression of a 23-kDa protein, or after addition of killed F. tularensis LVS. Addition of purified F. tularensis LPS resulted in no discernible effects on the cells. When the effects were followed up to 5 h, activation persisted in cultures with killed bacteria or infected with the deltaiglC strain. In contrast, the signal transduction activation and secretion of TNF-alpha were down-regulated within the 5h period in mouse peritoneal cells, J774 cells or human peripheral blood mononuclear cells infected with F. tularensis LVS. Together, the results suggest that infection with live F. tularensis LVS bacteria leads to a rapid induction of a proinflammatory response in mouse and human cells but after internalization of bacteria, this response is completely or partly down-regulated in most cell types. This down-regulation does not occur when cells are infected with the mutant deltaiglC.

  • 28. Tobias, Joshua
    et al.
    Lebens, Michael
    Wai, Sun Nyunt
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Holmgren, Jan
    Svennerholm, Ann-Mari
    Surface expression of Helicobacter pylori HpaA adhesion antigen on Vibrio cholerae, enhanced by co-expressed enterotoxigenic Escherichia coli fimbrial antigens2017In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 105, p. 177-184Article in journal (Refereed)
    Abstract [en]

    Helicobacter pylori infection can cause peptic ulceration and is associated with gastric adenocarcinoma. This study aimed to construct and characterize a non-virulent Vibrio cholerae 01 strain, which grows more rapidly than H. pylori, as vector for H. pylori antigens for possible use as a vaccine strain against H. pylori. This was done by recombinant expression of the H. pylori adhesion antigen HpaA alone or, as a proof of principle, together with different colonization factor (CF) antigens of enterotoxigenic Escherichia coli (ETEC) which may enhance immune responses against HpaA. A recombinant V cholerae strain co expressing HpaA and a fimbrial CF antigens CFA/I or CS5, but not the non-fimbrial CF protein CS6, was shown to express larger amounts of HpaA on the surface when compared with the same V cholerae strain expressing HpaA alone. Mutations in the CFA/I operon showed that the chaperon, possibly together with the usher, was involved in enhancing the surface expression of HpaA. Oral immunization of mice with formaldehyde-inactivated recombinant V cholerae expressing HpaA alone or together with CFA/I induced significantly higher serum antibody responses against HpaA than mice similarly immunized with inactivated HpaA-expressing H. pylori bacteria. Our results demonstrate that a non-virulent V cholerae strain can be engineered to allow strong surface expression of HpaA, and that the expression can be further increased by co-expressing it with ETEC fimbrial antigens. Such recombinant V cholerae strains expressing HpaA, and possibly also other H. pylori antigens, may have the potential as oral inactivated vaccine candidates against H. pylori.

  • 29. Twine, Susan M
    et al.
    Shen, Hua
    Kelly, John F
    Chen, Wangxue
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Clinical Microbiology. Umeå University, Faculty of Medicine, Clinical Microbiology, Clinical Bacteriology.
    Conlan, J Wayne
    NRC, Kanada.
    Virulence comparison in mice of distinct isolates of type A Francisella tularensis.2006In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 40, no 3, p. 133-8Article in journal (Refereed)
    Abstract [en]

    Francisella tularensis subspecies tularensis (type A F. tularensis) is considered to be one of the most virulent of all bacterial pathogens. Mice are extremely susceptible to infection with this subspecies (LD100 via various inoculation routes is <10 cfu). However, it has not been established whether overt virulence differences exist amongst type A strains of F. tularensis. To this end, the present study compared the virulence of two distinct type A strains, FSC033 and SCHU S4, for naïve mice and mice immunized with the live vaccine strain of the pathogen, F. tularensis LVS. One nominal isolate of SCHU S4 was found to be completely avirulent. Another isolate was highly virulent, but all examined cases appeared somewhat less virulent than FSC033. The implication of these findings for future infection and immunity studies is discussed.

  • 30.
    Westermark, Linda
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Fahlgren, Anna
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Fällman, Maria
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Immune response to diphtheria toxin-mediated depletion complicates the use of the CD11c-DTR(tg) model for studies of bacterial gastrointestinal infections2012In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 53, no 3-4, p. 154-161Article in journal (Refereed)
    Abstract [en]

    Dendritic cells play an important role in the immune response against pathogens, as they are responsible for the activation and control of both innate and adaptive immune responses. The CD11c-DTR(tg) model, which allows transient elimination of dendritic cells by diphtheria toxin-treatment (DTx), has been extensively used to study the importance of this immune cell during steady-state and infection conditions in mice. Infecting dendritic cell-depleted mice orally with Yersinia pseudotuberculosis results in a markedly reduced level of infection compared with infection of non-depleted mice. We show here that it is not the lack of dendritic cells per se that is responsible for the reduced infection efficiency, instead it is an immune response induced by the DTx-treatment that prevents the bacteria from establishing colonization in Peyer's patches. The DTx-induced depletion initiates an immune response, with elevated serum levels of keratinocyte-derived cytokine (KC) and recruitment of polymorphonuclear neutrophils to dendritic cell-containing organs, such as Peyer's patches. Since the window for having an animal depleted of dendritic cells is limited in time for this model, the DTx-mediated effect on the immune system complicates the use of this model in studies of early events during bacterial infections.

  • 31. Wu, Z.
    et al.
    Milton, Debra L..
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Nybom, P.
    Sjö, A.
    Magnusson, K. E.
    Vibrio cholerae hemagglutinin/protease (HA/protease) causes morphological changes in cultured epithelial cells and perturbs their paracellular barrier function1996In: Microbial Pathogenesis, ISSN 0882-4010, E-ISSN 1096-1208, Vol. 21, no 2, p. 111-123Article in journal (Refereed)
    Abstract [en]

    In this report, we describe the cytotoxic activity of the cholera hemagglutinin/protease (HA/protease). A concentrated protein sample from the 37 degrees C overnight culture supernatant of CVD110, a delta ctxA, delta zot, delta Ace and hlyA::(ctxB mer) mutant of El Tor biotype Ogawa serotype strain E7946 caused morphological changes in cultured MDCK-I epithelial cells and altered their arrangement of filamentous actin (F-actin) and Zonula occludens-associated protein ZO-1. The drastic morphological changes can be inhibited by Zincov, a specific bacterial metalloprotease inhibitor. The cytotoxic fractions of the sample after FPLC gelfiltration fractionation showed two visible protein bands with molecular weights of approximately 34- and 32 kDa. Microsequencing of these two proteins revealed that they were the cholera HA/protease.

1 - 31 of 31
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