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  • 1.
    Abbara, Aula
    et al.
    Imperial College, London, United Kingdom.
    Almalla, Mohamed
    American University of Beirut, Beirut, Lebanon.
    AlMasri, Ibrahim
    O'Brien Institute for Public Health, Cumming School of Medicine, University of Calgary, Calgary, Canada.
    AlKabbani, Hussam
    Department of Health and Nutrition Al-Ameen for Humanitarian Support, Gaziantep, Turkey.
    Karah, Nabil
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    El-Amin, Wael
    King's College Hospital London, United Arab Emirates.
    Rajan, Latha
    Tulane University School of Public Health and Tropical Medicine, Tulane University, LA, New Orleans, United States.
    Rahhal, Ibrahim
    Hand in Hand for Aid and Development, Gaziantep, Turkey.
    Alabbas, Mohammad
    Hand in Hand for Aid and Development, Gaziantep, Turkey.
    Sahloul, Zaher
    Department of Pulmonology and Critical Care, University of Illinois, IL, Chicago, United States.
    Tarakji, Ahmad
    Syrian American Medical Society, Washington DC, United States.
    Sparrow, Annie
    Department of Population Health Sciences and Policy, Icahn School of Medicine at Mount Sinai, New York, United States.
    The challenges of tuberculosis control in protracted conflict: The case of Syria2020Inngår i: International Journal of Infectious Diseases, ISSN 1201-9712, E-ISSN 1878-3511, International Journal of Infectious Diseases, ISSN 1201-9712, Vol. 90, s. 53-59Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Objectives: Syria's protracted conflict has resulted in ideal conditions for the transmission of tuberculosis (TB) and the cultivation of drug-resistant strains. This paper compares TB control in Syria before and after the conflict using available data, examines the barriers posed by protracted conflict and those specific to Syria, and discusses what measures can be taken to address the control of TB in Syria.

    Results: Forced mass displacement and systematic violations of humanitarian law have resulted in overcrowding and the destruction of key infrastructure, leading to an increased risk of both drug-sensitive and resistant TB, while restricting the ability to diagnose, trace contacts, treat, and follow-up. Pre-conflict, TB in Syria was officially reported at 22 per 100 000 population; the official figure for 2017 of 19 per 100 000 is likely a vast underestimate given the challenges and barriers to case detection. Limited diagnostics also affect the diagnosis of multidrug- and rifampicin-resistant TB, reported as comprising 8.8% of new diagnoses in 2017.

    Conclusions: The control of TB in Syria requires a multipronged, tailored, and pragmatic approach to improve timely diagnosis, increase detection, stop transmission, and mitigate the risk of drug resistance. Solutions must also consider vulnerable populations such as imprisoned and besieged communities where the risk of drug resistance is particularly high, and must recognize the limitations of national programming. Strengthening capacity to control TB in Syria with particular attention to these factors will positively impact other parallel conditions; this is key as attention turns to post-conflict reconstruction.

    Fulltekst (pdf)
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  • 2. Abbara, Aula
    et al.
    Rawson, Timothy M.
    Karah, Nabil
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    El-Amin, Wael
    Hatcher, James
    Tajaldin, Bachir
    Dar, Osman
    Dewachi, Omar
    Abu Sitta, Ghassan
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Sparrow, Annie
    A summary and appraisal of existing evidence of antimicrobial resistance in the Syrian conflict2018Inngår i: International Journal of Infectious Diseases, ISSN 1201-9712, E-ISSN 1878-3511, Vol. 75, s. 26-33Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Antimicrobial resistance (AMR) in populations experiencing war has yet to be addressed, despite the abundance of contemporary conflicts and the protracted nature of twenty-first century wars, in combination with growing global concern over conflict-associated bacterial pathogens. The example of the Syrian conflict is used to explore the feasibility of using existing global policies on AMR in conditions of extreme conflict. The available literature on AMR and prescribing behaviour in Syria before and since the onset of the conflict in March 2011 was identified. Overall, there is a paucity of rigorous data before and since the onset of conflict in Syria to contextualize the burden of AMR. However, post onset of the conflict, an increasing number of studies conducted in neighbouring countries and Europe have reported AMR in Syrian refugees. High rates of multidrug resistance, particularly Gram-negative organisms, have been noted amongst Syrian refugees when compared with local populations. Conflict impedes many of the safeguards against AMR, creates new drivers, and exacerbates existing ones. Given the apparently high rates of AMR in Syria, in neighbouring countries hosting refugees, and in European countries providing asylum, this requires the World Health Organization and other global health institutions to address the causes, costs, and future considerations of conflict-related AMR as an issue of global governance. (c) 2018 The Authors. Published by Elsevier Ltd on behalf of International Society for Infectious Diseases.

    Fulltekst (pdf)
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  • 3. Abbara, Aula
    et al.
    Rawson, Timothy M.
    Karah, Nabil
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    El-Amin, Wael
    Hatcher, James
    Tajaldin, Bachir
    Dar, Osman
    Dewachi, Omar
    Abu Sitta, Ghassan
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Sparrow, Annie
    Antimicrobial resistance in the context of the Syrian conflict: Drivers before and after the onset of conflict and key recommendations2018Inngår i: International Journal of Infectious Diseases, ISSN 1201-9712, E-ISSN 1878-3511, Vol. 73, s. 1-6Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Current evidence describing antimicrobial resistance (AMR) in the context of the Syrian conflict is of poor quality and sparse in nature. This paper explores and reports the major drivers of AMR that were present in Syria pre-conflict and those that have emerged since its onset in March 2011. Drivers that existed before the conflict included a lack of enforcement of existing legislation to regulate over-the-counter antibiotics and notification of communicable diseases. This contributed to a number of drivers of AMR after the onset of conflict, and these were also compounded by the exodus of trained staff, the increase in overcrowding and unsanitary conditions, the increase in injuries, and economic sanctions limiting the availability of required laboratory medical materials and equipment. Addressing AMR in this context requires pragmatic, multifaceted action at the local, regional, and international levels to detect and manage potentially high rates of multidrug-resistant infections. Priorities are (1) the development of a competent surveillance system for hospital-acquired infections, (2) antimicrobial stewardship, and (3) the creation of cost-effective and implementable infection control policies. However, it is only by addressing the conflict and immediate cessation of the targeting of health facilities that the rehabilitation of the health system, which is key to addressing AMR in this context, can progress. 

    Fulltekst (pdf)
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  • 4. Abraham, Nabil M.
    et al.
    Liu, Lei
    Jutras, Brandon Lyon
    Yadav, Akhilesh K.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Narasimhan, Sukanya
    Gopalakrishnan, Vissagan
    Ansari, Juliana M.
    Jefferson, Kimberly K.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Jacobs-Wagner, Christine
    Fikrig, Erol
    Pathogen-mediated manipulation of arthropod microbiota to promote infection2017Inngår i: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 114, nr 5, s. E781-E790Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Arthropods transmit diverse infectious agents; however, the ways microbes influence their vector to enhance colonization are poorly understood. Ixodes scapularis ticks harbor numerous human pathogens, including Anaplasma phagocytophilum, the agent of human granulocytic anaplasmosis. We now demonstrate that A. phagocytophilum modifies the I. scapularis microbiota to more efficiently infect the tick. A. phagocytophilum induces ticks to express Ixodes scapularis antifreeze glycoprotein (iafgp), which encodes a protein with several properties, including the ability to alter bacterial biofilm formation. IAFGP thereby perturbs the tick gut microbiota, which influences the integrity of the peritrophic matrix and gut barrier-critical obstacles for Anaplasma colonization. Mechanistically, IAFGP binds the terminal D-alanine residue of the pentapeptide chain of bacterial peptidoglycan, resulting in altered permeability and the capacity of bacteria to form biofilms. These data elucidate the molecular mechanisms by which a human pathogen appropriates an arthropod antibacterial protein to alter the gut microbiota and more effectively colonize the vector.

  • 5. Achouiti, A.
    et al.
    Vogl, T.
    Urban, Constantin
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Hommes, T. J.
    van Zoelen, M. A.
    Florquin, S.
    Roth, J.
    van 't Veer, C.
    de Vos, A. F.
    van der Poll, T.
    Myeloid related protein (mrp) 8/14 contributes to an antibacterial host response against klebsiella (k.) pneumoniae2012Inngår i: Shock, ISSN 1073-2322, E-ISSN 1540-0514, Vol. 37, nr S1, s. 56-56Artikkel i tidsskrift (Annet vitenskapelig)
  • 6. Achouiti, Ahmed
    et al.
    Vogl, Thomas
    Urban, Constantin F
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Röhm, Marc
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi.
    Hommes, Tijmen J
    van Zoelen, Marieke AD
    Florquin, Sandrine
    Roth, Johannes
    van't Veer, Cornelis
    de Vos, Alex F
    van der Poll, Tom
    Myeloid-related protein-14 contributes to protective immunity in gram-negative pneumonia derived sepsis2012Inngår i: PLoS Pathogens, ISSN 1553-7374, Vol. 8, nr 10, s. e1002987-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Klebsiella (K.) pneumoniae is a common cause of pneumonia-derived sepsis. Myeloid related protein 8 (MRP8, S100A8) and MRP14 (S100A9) are the most abundant cytoplasmic proteins in neutrophils. They can form MRP8/14 heterodimers that are released upon cell stress stimuli. MRP8/14 reportedly exerts antimicrobial activity, but in acute fulminant sepsis models MRP8/14 has been found to contribute to organ damage and death. We here determined the role of MRP8/14 in K. pneumoniae sepsis originating from the lungs, using an established model characterized by gradual growth of bacteria with subsequent dissemination. Infection resulted in gradually increasing MRP8/14 levels in lungs and plasma. Mrp14 deficient (mrp14(-/-)) mice, unable to form MRP8/14 heterodimers, showed enhanced bacterial dissemination accompanied by increased organ damage and a reduced survival. Mrp14(-/-) macrophages were reduced in their capacity to phagocytose Klebsiella. In addition, recombinant MRP8/14 heterodimers, but not MRP8 or MRP14 alone, prevented growth of Klebsiella in vitro through chelation of divalent cations. Neutrophil extracellular traps (NETs) prepared from wildtype but not from mrp14(-/-) neutrophils inhibited Klebsiella growth; in accordance, the capacity of human NETs to kill Klebsiella was strongly impaired by an anti-MRP14 antibody or the addition of zinc. These results identify MRP8/14 as key player in protective innate immunity during Klebsiella pneumonia.

    Fulltekst (pdf)
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  • 7. Adderley, Jack D.
    et al.
    von Freyend, Simona John
    Jackson, Sarah A.
    Bird, Megan J.
    Burns, Amy L.
    Anar, Burcu
    Metcalf, Tom
    Semblat, Jean-Philippe
    Billker, Oliver
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Wellcome Trust Sanger Institute, Hinxton, Cambridgeshire, UK.
    Wilson, Danny W.
    Doerig, Christian
    Analysis of erythrocyte signalling pathways during Plasmodium falciparum infection identifies targets for host-directed antimalarial intervention2020Inngår i: Nature Communications, E-ISSN 2041-1723, Vol. 11, nr 1, artikkel-id 4015Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Intracellular pathogens mobilize host signaling pathways of their host cell to promote their own survival. Evidence is emerging that signal transduction elements are activated in a-nucleated erythrocytes in response to infection with malaria parasites, but the extent of this phenomenon remains unknown. Here, we fill this knowledge gap through a comprehensive and dynamic assessment of host erythrocyte signaling during infection with Plasmodium falciparum. We used arrays of 878 antibodies directed against human signaling proteins to interrogate the activation status of host erythrocyte phospho-signaling pathways at three blood stages of parasite asexual development. This analysis reveals a dynamic modulation of many host signalling proteins across parasite development. Here we focus on the hepatocyte growth factor receptor (c-MET) and the MAP kinase pathway component B-Raf, providing a proof of concept that human signaling kinases identified as activated by malaria infection represent attractive targets for antimalarial intervention.

    Fulltekst (pdf)
    fulltext
  • 8.
    Adolfsson, Dan E.
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Tyagi, Mohit
    Singh, Pardeep
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Kaur, Amandeep
    Ådén, Jörgen
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Bharate, Jaideep B.
    Almqvist, Fredrik
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Enhanement of amyloid fibril binding by ring expansion of thiazolino fused 2-pyridone peptidomimeticsManuskript (preprint) (Annet vitenskapelig)
    Abstract [en]

    Thiazolino fused 2-pyridones undergo thiazoline ring opening by reaction with 2-nitrobenzyl bromide through thi- oether attack, and base promoted fragmentation of the resulting sulfonium ions. Subsequent deprotonation of the benzylic carbon and intramolecular 1,4-addition leads to ring closure, generating dihydrothiazine fused 2-pyridones by net ring expansion of the thiazoline ring. Application of the ring expansion procedure to the pyridine and pyrimidine fused thiazolino 2-pyridones provided compounds with enhanced fibril binding activity.

  • 9. Afset, J. E.
    et al.
    Larssen, K. W.
    Bergh, K.
    Larkeryd, A.
    Sjodin, A.
    Johansson, Anders
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi.
    Forsman, M.
    Phylogeographical pattern of Francisella tularensis in a nationwide outbreak of tularaemia in Norway, 20112015Inngår i: Eurosurveillance, ISSN 1025-496X, E-ISSN 1560-7917, Vol. 20, nr 19, s. 9-14, artikkel-id 21125Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    In 2011, a nationwide outbreak of tularaemia occurred in Norway with 180 recorded cases. It was associated with the largest peak in lemming density seen in 40 years. Francisella tularensis was isolated from 18 patients. To study the geographical distribution of F. tularensis genotypes in Norway and correlate genotype with epidemiology and clinical presentation, we performed whole genome sequencing of patient isolates. All 18 genomes from the outbreak carried genetic signatures of F. tularensis subsp. holarctica and were assigned to genetic clades using canonical single nucleotide polymorphisms. Ten isolates were assigned to major genetic clade B.6 (subclade B.7), seven to clade B.12, and one to clade B.4. The B.6 subclade B.7 was most common in southern and central Norway, while clade B.12 was evenly distributed between the southern, central and northern parts of the country. There was no association between genotype and clinical presentation of tularaemia, time of year or specimen type. We found extensive sequence similarity with F. tularensis subsp. holarctica genomes from high-endemic tularaemia areas in Sweden. Finding nearly identical genomes across large geographical distances in Norway and Sweden imply a life cycle of the bacterium without replication between the outbreaks and raise new questions about long-range migration mechanisms.

  • 10.
    Ahmad, Irfan
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Institute of Biomedical and Allied Health Sciences, University of Health Sciences, Lahore, Pakistan.
    Karah, Nabil
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Nadeem, Aftab
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Wai, Sun Nyunt
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Analysis of colony phase variation switch in Acinetobacter baumannii clinical isolates2019Inngår i: PLOS ONE, E-ISSN 1932-6203, Vol. 14, nr 1, artikkel-id e0210082Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Reversible switching between opaque and translucent colony formation is a novel feature of Acinetobacter baumannii that has been associated with variations in the cell morphology, surface motility, biofilm formation, antibiotic resistance and virulence. Here, we assessed a number of phenotypic alterations related to colony switching in A. baumannii clinical isolates belonging to different multi-locus sequence types. Our findings demonstrated that these phenotypic alterations were mostly strain-specific. In general, the translucent subpopulations of A. baumannii produced more dense biofilms, were more piliated, and released larger amounts of outer membrane vesicles (OMVs). In addition, the translucent subpopulations caused reduced fertility of Caenorhabditis elegans. When assessed for effects on the immune response in RAW 264.7 macrophages, the OMVs isolated from opaque subpopulations of A. baumannii appeared to be more immunogenic than the OMVs from the translucent form. However, also the OMVs from the translucent subpopulations had the potential to evoke an immune response. Therefore, we suggest that OMVs may be considered for development of new immunotherapeutic treatments against A. baumannii infections.

    Fulltekst (pdf)
    fulltext
  • 11.
    Ahmad, Irfan
    et al.
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Institute of Biomedical and Allied Health Sciences, University of Health Sciences, Lahore, Pakistan.
    Nadeem, Aftab
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Mushtaq, Fizza
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Institute of Biomedical and Allied Health Sciences, University of Health Sciences, Lahore, Pakistan.
    Zlatkov, Nikola
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Shahzad, Muhammad
    Department of Pharmacology, University of Health Sciences, Lahore, Pakistan.
    Zavialov, Anton V.
    Department of Biochemistry, University of Turku, Tykistökatu 6A, Turku, Finland.
    Wai, Sun Nyunt
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Csu pili dependent biofilm formation and virulence of Acinetobacter baumannii2023Inngår i: npj Biofilms and Microbiomes, E-ISSN 2055-5008, Vol. 9, nr 1, artikkel-id 101Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Acinetobacter baumannii has emerged as one of the most common extensive drug-resistant nosocomial bacterial pathogens. Not only can the bacteria survive in hospital settings for long periods, but they are also able to resist adverse conditions. However, underlying regulatory mechanisms that allow A. baumannii to cope with these conditions and mediate its virulence are poorly understood. Here, we show that bi-stable expression of the Csu pili, along with the production of poly-N-acetyl glucosamine, regulates the formation of Mountain-like biofilm-patches on glass surfaces to protect bacteria from the bactericidal effect of colistin. Csu pilus assembly is found to be an essential component of mature biofilms formed on glass surfaces and of pellicles. By using several microscopic techniques, we show that clinical isolates of A. baumannii carrying abundant Csu pili mediate adherence to epithelial cells. In addition, Csu pili suppressed surface-associated motility but enhanced colonization of bacteria into the lungs, spleen, and liver in a mouse model of systemic infection. The screening of c-di-GMP metabolizing protein mutants of A. baumannii 17978 for the capability to adhere to epithelial cells led us to identify GGDEF/EAL protein AIS_2337, here denoted PdeB, as a major regulator of Csu pili-mediated virulence and biofilm formation. Moreover, PdeB was found to be involved in the type IV pili-regulated robustness of surface-associated motility. Our findings suggest that the Csu pilus is not only a functional component of mature A. baumannii biofilms but also a major virulence factor promoting the initiation of disease progression by mediating bacterial adherence to epithelial cells.

    Fulltekst (pdf)
    fulltext
  • 12.
    Ahmad, Irfan
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Institute of Biomedical and Allied Health Sciences, University of Health Sciences, Lahore, Pakistan.
    Nygren, Evelina
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Khalid, Fizza
    Myint, Si Lhyam
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    A Cyclic-di-GMP signalling network regulates biofilm formation and surface associated motility of Acinetobacter baumannii 179782020Inngår i: Scientific Reports, E-ISSN 2045-2322, Vol. 10, nr 1, artikkel-id 1991Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Acinetobacter baumannii has emerged as an increasing multidrug-resistant threat in hospitals and a common opportunistic nosocomial pathogen worldwide. However, molecular details of the pathogenesis and physiology of this bacterium largely remain to be elucidated. Here we identify and characterize the c-di-GMP signalling network and assess its role in biofilm formation and surface associated motility. Bioinformatic analysis revealed eleven candidate genes for c-di-GMP metabolizing proteins (GGDEF/EAL domain proteins) in the genome of A. baumannii strain 17978. Enzymatic activity of the encoded proteins was assessed by molecular cloning and expression in the model organisms Salmonella typhimurium and Vibrio cholerae. Ten of the eleven GGDEF/EAL proteins altered the rdar morphotype of S. typhimurium and the rugose morphotype of V. cholerae. The over expression of three GGDEF proteins exerted a pronounced effect on colony formation of A. baumannii on Congo Red agar plates. Distinct panels of GGDEF/EAL proteins were found to alter biofilm formation and surface associated motility of A. baumannii upon over expression. The GGDEF protein A1S_3296 appeared as a major diguanylate cyclase regulating macro-colony formation, biofilm formation and the surface associated motility. AIS_3296 promotes Csu pili mediated biofilm formation. We conclude that a functional c-di-GMP signalling network in A. baumannii regulates biofilm formation and surface associated motility of this increasingly important opportunistic bacterial pathogen.

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  • 13.
    Aili, Margareta
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Isaksson, Elin L
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Carlsson, Sara E
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Wolf-Watz, Hans
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Rosqvist, Roland
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Francis, Matthew S
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Regulation of Yersinia Yop-effector delivery by translocated YopE2008Inngår i: International Journal of Medical Microbiology, ISSN 1438-4221, E-ISSN 1618-0607, Vol. 298, nr 3-4, s. 183-192Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The bacterial pathogen Yersinia pseudotuberculosis uses a type III secretion (T3S) system to translocate Yop effectors into eukaryotic cells. Effectors are thought to gain access to the cytosol via pores formed in the host cell plasma membrane. Translocated YopE can modulate this pore formation through its GTPase-activating protein (GAP) activity. In this study, we analysed the role of translocated YopE and all the other known Yop effectors in the regulation of effector translocation. Elevated levels of Yop effector translocation into HeLa cells occurred by YopE-defective strains, but not those defective for other Yop effectors. Only Yersinia devoid of YopK exhibits a similar hyper-translocation phenotype. Since both yopK and yopE mutants also failed to down-regulate Yop synthesis in the presence of eukaryotic cells, these data imply that translocated YopE specifically regulates subsequent effector translocation by Yersinia through at least one mechanism that involves YopK. We suggest that the GAP activity of YopE might be working as an intra-cellular probe measuring the amount of protein translocated by Yersinia during infection. This may be a general feature of T3S-associated GAP proteins, since two homologues from Pseudomonas aeruginosa, exoenzyme S (ExoS) and exoenzyme T (ExoT), can complement the hyper-translocation phenotypes of the yopE GAP mutant.

  • 14. Akkaya, Munir
    et al.
    Bansal, Abhisheka
    Sheehan, Patrick W.
    Pena, Mirna
    Cimperman, Clare K.
    Qi, Chen Feng
    Yazew, Takele
    Otto, Thomas D.
    Billker, Oliver
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Miller, Louis H.
    Pierce, Susan K.
    Testing the impact of a single nucleotide polymorphism in a Plasmodium berghei ApiAP2 transcription factor on experimental cerebral malaria in mice2020Inngår i: Scientific Reports, E-ISSN 2045-2322, Vol. 10, nr 1, artikkel-id 13630Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Cerebral malaria (CM) is the deadliest form of severe Plasmodium infections. Currently, we have limited understanding of the mechanisms by which Plasmodium parasites induce CM. The mouse model of CM, experimental CM (ECM), induced by infection with the rodent parasite, Plasmodium berghei ANKA (PbANKA) has been extensively used to study the pathophysiology of CM. Recent genomic analyses revealed that the coding regions of PbANKA and the closely related Plasmodium berghei NK65 (PbNK65), that does not cause ECM, differ in only 21 single nucleotide polymorphysims (SNPs). Thus, the SNP-containing genes might contribute to the pathogenesis of ECM. Although the majority of these SNPs are located in genes of unknown function, one SNP is located in the DNA binding site of a member of the Plasmodium ApiAP2 transcription factor family, that we recently showed functions as a virulence factor alternating the host's immune response to the parasite. Here, we investigated the impact of this SNP on the development of ECM. Our results using CRISPR-Cas9 engineered parasites indicate that despite its immune modulatory function, the SNP is neither necessary nor sufficient to induce ECM and thus cannot account for parasite strain-specific differences in ECM phenotypes.

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  • 15. Akkaya, Munir
    et al.
    Bansal, Abhisheka
    Sheehan, Patrick W.
    Pena, Mirna
    Molina-Cruz, Alvaro
    Orchard, Lindsey M.
    Cimperman, Clare K.
    Qi, Chen-Feng
    Ross, Philipp
    Yazew, Takele
    Sturdevant, Daniel
    Anzick, Sarah L.
    Thiruvengadam, Girija
    Otto, Thomas Dan
    Billker, Oliver
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Llinas, Manuel
    Miller, Louis H.
    Pierce, Susan K.
    A single-nucleotide polymorphism in a Plasmodium berghei ApiAP2 transcription factor alters the development of host immunity2020Inngår i: Science Advances, E-ISSN 2375-2548, Vol. 6, nr 6, artikkel-id eaaw6957Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The acquisition of malaria immunity is both remarkably slow and unpredictable. At present, we know little about the malaria parasite genes that influence the host's ability to mount a protective immune response. Here, we show that a single-nucleotide polymorphism (SNP) resulting in a single amino acid change (S to F) in an ApiAP2 transcription factor in the rodent malaria parasite Plasmodium berghei (Pb) NK65 allowed infected mice to mount a T helper cell 1 (T(H)1)-type immune response that controlled subsequent infections. As compared to PbNK65(S), PbNK65(F) parasites differentially expressed 46 genes, most of which are predicted to play roles in immune evasion. PbNK65(F) infections resulted in an early interferon-gamma response and a later expansion of germinal centers, resulting in high levels of infected red blood cell-specific T(H)1-type immunoglobulin G2b (IgG2b) and IgG2c antibodies. Thus, the Pb ApiAP2 transcription factor functions as a critical parasite virulence factor in malaria infections.

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  • 16.
    Akopyan, Karen
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Edgren, Tomas
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Wang-Edgren, Helen
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Rosqvist, Roland
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Fahlgren, Anna
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Wolf-Watz, Hans
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Fällman, Maria
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Translocation of surface-localized effectors in type III secretion2011Inngår i: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 108, nr 4, s. 1639-1644Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Pathogenic Yersinia species suppress the host immune response by using a plasmid-encoded type III secretion system (T3SS) to translocate virulence proteins into the cytosol of the target cells. T3SS-dependent protein translocation is believed to occur in one step from the bacterial cytosol to the target-cell cytoplasm through a conduit created by the T3SS upon target cell contact. Here, we report that T3SS substrates on the surface of Yersinia pseudotuberculosis are translocated into target cells. Upon host cell contact, purified YopH coated on Y. pseudotuberculosis was specifically and rapidly translocated across the target-cell membrane, which led to a physiological response in the infected cell. In addition, translocation of externally added YopH required a functional T3SS and a specific translocation domain in the effector protein. Efficient, T3SS-dependent translocation of purified YopH added in vitro was also observed when using coated Salmonella typhimurium strains, which implies that T3SS-mediated translocation of extracellular effector proteins is conserved among T3SS-dependent pathogens. Our results demonstrate that polarized T3SS-dependent translocation of proteins can be achieved through an intermediate extracellular step that can be reconstituted in vitro. These results indicate that translocation can occur by a different mechanism from the assumed single-step conduit model.

  • 17.
    Alam, Athar
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Bröms, Jeanette E
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Kumar, Rajender
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Sjöstedt, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    The Role of ClpB in Bacterial Stress Responses and Virulence2021Inngår i: Frontiers in Molecular Biosciences, E-ISSN 2296-889X, Vol. 8, artikkel-id 668910Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Bacterial survival within a mammalian host is contingent upon sensing environmental perturbations and initiating an appropriate counter-response. To achieve this, sophisticated molecular machineries are used, where bacterial chaperone systems play key roles. The chaperones are a prerequisite for bacterial survival during normal physiological conditions as well as under stressful situations, e.g., infection or inflammation. Specific stress factors include, but are not limited to, high temperature, osmolarity, pH, reactive oxidative species, or bactericidal molecules. ClpB, a member of class 1 AAA+ proteins, is a key chaperone that via its disaggregase activity plays a crucial role for bacterial survival under various forms of stress, in particular heat shock. Recently, it has been reported that ClpB also regulates secretion of bacterial effector molecules related to type VI secretion systems. In this review, the roles of ClpB in stress responses and the mechanisms by which it promotes survival of pathogenic bacteria are discussed.

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  • 18.
    Alam, Athar
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Golovliov, Igor
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Javed, Eram
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi.
    Kumar, Rajender
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Ådén, Jörgen
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sjöstedt, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Dissociation between the critical role of ClpB of Francisella tularensis for the heat shock response and the DnaK interaction and its important role for efficient type VI secretion and bacterial virulence2020Inngår i: PLoS Pathogens, ISSN 1553-7366, E-ISSN 1553-7374, Vol. 16, nr 4, s. 1-27, artikkel-id e1008466Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Author summary Type VI secretion systems (T6SSs) are essential virulence determinants of many Gram-negative pathogens, including Francisella tularensis. This highly virulent bacterium encodes an atypical T6SS lacking ClpV, the ATPase crucial for prototypic T6SS sheath disassembly. It, however, possesses ClpB, a protein critical for heat shock survival via its interaction with DnaK. Since ClpB possesses ATPase activity, it has been hypothesized to provide a compensatory function for the absence of ClpV, a hypothesis supported by the recent findings from us and others. Here, we investigated how F. tularensis ClpB controls T6S. In silico modelling of the ClpB-DnaK complex identified key interactions that were experimentally verified. For example, mutating one of the DnaK-interacting residues rendered the bacterium exquisitely susceptible to heat shock, but had no effect on T6S and virulence. In contrast, removing the N-terminal of ClpB only had a slight effect on the heat shock response, but strongly compromised both T6S and virulence. Intriguingly, the Escherichia coli ClpB could fully complement the function of F. tularensis ClpB. The data demonstrate that the two critical roles of ClpB, mediating heat shock survival and effective T6S, are dissociated and that the N-terminal is crucial for T6S and virulence. Francisella tularensis, a highly infectious, intracellular bacterium possesses an atypical type VI secretion system (T6SS), which is essential for its virulence. The chaperone ClpB, a member of the Hsp100/Clp family, is involved in Francisella T6SS disassembly and type VI secretion (T6S) is impaired in its absence. We asked if the role of ClpB for T6S was related to its prototypical role for the disaggregation activity. The latter is dependent on its interaction with the DnaK/Hsp70 chaperone system. Key residues of the ClpB-DnaK interaction were identified by molecular dynamic simulation and verified by targeted mutagenesis. Using such targeted mutants, it was found that the F. novicida ClpB-DnaK interaction was dispensable for T6S, intracellular replication, and virulence in a mouse model, although essential for handling of heat shock. Moreover, by mutagenesis of key amino acids of the Walker A, Walker B, and Arginine finger motifs of each of the two Nucleotide-Binding Domains, their critical roles for heat shock, T6S, intracellular replication, and virulence were identified. In contrast, the N-terminus was dispensable for heat shock, but required for T6S, intracellular replication, and virulence. Complementation of the Delta clpB mutant with a chimeric F. novicida ClpB expressing the N-terminal of Escherichia coli, led to reconstitution of the wild-type phenotype. Collectively, the data demonstrate that the ClpB-DnaK interaction does not contribute to T6S, whereas the N-terminal and NBD domains displayed critical roles for T6S and virulence.

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  • 19. Aldick, Thomas
    et al.
    Bielaszewska, Martina
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Humpf, Hans-Ulrich
    Wai, Sun Nyunt
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Karch, Helge
    Vesicular stabilization and activity augmentation of enterohaemorrhagic Escherichia coli haemolysin2009Inngår i: Molecular Microbiology, ISSN 0950-382X, E-ISSN 1365-2958, Vol. 71, nr 6, s. 1496-508Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Haemolysin from enterohaemorrhagic Escherichia coli (EHEC-Hly), a putative EHEC virulence factor, belongs to the RTX (repeat-in-toxin) family whose members rapidly inactivate themselves by self-aggregation. By investigating the status of EHEC-Hly secreted extracellularly, we found the toxin both in a free, soluble form and associated, with high tendency and independently of its acylation status, to outer membrane vesicles (OMVs) extruded by EHEC. We compared the interaction of both toxin forms with erythrocytes using scanning electron microscopy and binding assays. The OMV-associated toxin was substantially (80 times) more stable under physiological conditions than the free EHEC-Hly as demonstrated by prolonged haemolytic activity (half-life time 20 h versus 15 min). The haemolysis was preceded by calcium-dependent binding of OMVs carrying EHEC-Hly to erythrocytes; this binding was mediated by EHEC-Hly. We demonstrate that EHEC-Hly is a biologically active cargo in OMVs with dual roles: a cell-binding protein and a haemolysin. These paired functions produce a biologically potent form of the OMV-associated RTX toxin and augment its potential towards target cells. Our findings provide a general concept for stabilization of RTX toxins and open new insights into the biology of these important virulence factors.

  • 20.
    Aliashkevich, Alena
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Molecular mechanisms and biological consequences of the production of non-canonical D-amino acids in bacteria2021Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Most bacteria possess a vital net-like macromolecule – peptidoglycan (PG). PG encases bacteria around the cytoplasmic membrane to withstand the high internal turgor pressure and thereby protect the cell from bursting. In addition, PG is a major morphological determinant of bacteria being both required and sufficient to maintain cell shape. During cell growth PG hydrolysis and synthesis are tightly controlled to keep proper cell shape and integrity at all times. Given the essentiality of PG for bacterial growth and survival, the synthesis of this polymer is a major target of many natural and synthetic antibiotics (e.g. penicillins, glycopeptides).

    For a long time, PG composition was considered to be conserved and static, however it’s now being recognized as a dynamic and plastic macromolecule. The structure and chemistry of PG is influenced by a myriad of environmental cues that include interkingdom/interspecies interactions. Recently, it was found that a wide set of non-canonical D-amino acids (D-amino acids different from D-Ala and D-Glu, NCDAAs) are produced and released to the extracellular milieu by diverse bacteria. In Vibrio cholerae these NCDAAs are produced by broad-spectrum racemase enzyme (BsrV) and negatively regulate PG synthesis through their incorporation into PG. We have shown that in addition to D-Met and D-Leu, which were reported previously, V. cholerae also releases high amounts of D-Arg, which inhibits a broader range of phylogenetically diverse bacteria. Thus, NCDAAs affect not only the producer, but might target other species within the same environmental niche. However, in contrast to D-Met, D-Arg targets cell wall independent pathways. 

    We have shown that non-proteinogenic amino acids also can be racemized by Bsr. A plant amino acid L-canavanine (L-CAN) is converted into D-CAN by a broad-spectrum amino acid racemase (BSAR) of the soil bacterium Pseudomonas putida and subsequently released to the environment. D-CAN gets highly incorporated into the PG of Rhizobiales (such as Agrobacterium tumefaciens, Sinorhizobium meliloti) thereby affecting the overall PG structure, bacterial morphogenesis and growth fitness. We found that detrimental effect of D-CAN in A. tumefaciens can be suppressed by a single amino acid substitution in the cell division PG transpeptidase penicillin-binding protein 3a (PBP3a). 

    Rhizobiales are a polar-growing species that encode multiple LD-transpeptidases (LDTs), enzymes that normally perform PG crosslinking, but that can also incorporate NCDAAs into termini of the PG peptides. As these species incorporate high amounts of D-CAN in their PG, we hypothesized that LDTs might represent the main path used by NCDAAs to edit A. tumefaciens’ PG and cause their detrimental effects. Therefore, we decided to further explore the significance of LDT proteins for growth and morphogenesis in A. tumefaciens. While in the Gram-negative model organism E. coli LDT proteins are non-essential under standard laboratory conditions, we found that A. tumefaciens needs at least one LDT for growth out of the 14 putative LDTs encoded in its genome. Moreover, clustering the LDT proteins based on their sequence similarity revealed that A. tumefaciens has 7 LDTs that are exclusively present among Rhizobiales. Interestingly, the loss of this group of LDTs (but not the rest) leads to reduced growth, lower PG crosslinkage and rounded cell phenotype, which suggests that this group of Rhizobiales- specific LDTs have a major role in maintaining LD-crosslinking homeostasis, which in turn is important for cell elongation and proper shape maintenance in A. tumefaciens.

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  • 21.
    Aliashkevich, Alena
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Alvarez, Laura
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    New Insights Into the Mechanisms and Biological Roles of D-Amino Acids in Complex Eco-Systems2018Inngår i: Frontiers in Microbiology, E-ISSN 1664-302X, Vol. 9, artikkel-id 683Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    In the environment bacteria share their habitat with a great diversity of organisms, from microbes to humans, animals and plants. In these complex communities, the production of extracellular effectors is a common strategy to control the biodiversity by interfering with the growth and/or viability of nearby microbes. One of such effectors relies on the production and release of extracellular D-amino acids which regulate diverse cellular processes such as cell wall biogenesis, biofilm integrity, and spore germination. Non-canonical D-amino acids are mainly produced by broad spectrum racemases (Bsr). Bsr's promiscuity allows it to generate high concentrations of D-amino acids in environments with variable compositions of L-amino acids. However, it was not clear until recent whether these molecules exhibit divergent functions. Here we review the distinctive biological roles of D-amino acids, their mechanisms of action and their modulatory properties of the biodiversity of complex eco-systems.

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  • 22.
    Aliashkevich, Alena
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    LD-transpeptidases: the great unknown among the peptidoglycan cross-linkers2022Inngår i: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 289, nr 16, s. 4718-4730Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    The peptidoglycan (PG) cell wall is an essential polymer for the shape and viability of bacteria. Its protective role is in great part provided by its mesh-like character. Therefore, PG-cross-linking enzymes like the penicillin-binding proteins (PBPs) are among the best targets for antibiotics. However, while PBPs have been in the spotlight for more than 50 years, another class of PG-cross-linking enzymes called LD-transpeptidases (LDTs) seemed to contribute less to PG synthesis and, thus, has kept an aura of mystery. In the last years, a number of studies have associated LDTs with cell wall adaptation to stress including β-lactam antibiotics, outer membrane stability, and toxin delivery, which has shed light onto the biological meaning of these proteins. Furthermore, as some species display a great abundance of LD-cross-links in their cell wall, it has been hypothesized that LDTs could also be the main synthetic PG-transpeptidases in some bacteria. In this review, we introduce these enzymes and their role in PG biosynthesis and we highlight the most recent advances in understanding their biological role in diverse species.

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  • 23.
    Aliashkevich, Alena
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Howell, Matthew
    Division of Biological Sciences, University of Missouri, Columbia, MO, USA; Department of Biology and Environmental Science, Westminster College, Fulton, MO, USA.
    Brown, Pamela J. B.
    Division of Biological Sciences, University of Missouri, Columbia, MO, USA.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    D-canavanine affects peptidoglycan structure, morphogenesis and fitness in Rhizobiales2021Inngår i: Environmental Microbiology, ISSN 1462-2912, E-ISSN 1462-2920, Vol. 23, nr 10, s. 5823-5836Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The bacterial cell wall is made of peptidoglycan (PG), a polymer that is essential for maintenance of cell shape and survival. Many bacteria alter their PG chemistry as a strategy to adapt their cell wall to external challenges. Therefore, identifying these environmental cues is important to better understand the interplay between microbes and their habitat. Here we used the soil bacterium Pseudomonas putida to uncover cell wall modulators from plant extracts and found canavanine (CAN), a non-proteinogenic amino acid. We demonstrated that cell wall chemical editing by CAN is licensed by P. putida BSAR, a broad-spectrum racemase which catalyzes production of DL-CAN from L-CAN, which is produced by many legumes. Importantly, D-CAN diffuses to the extracellular milieu thereby having a potential impact on other organisms inhabiting the same niche. Our results show that D-CAN alters dramatically the PG structure of Rhizobiales (e.g. Agrobacterium tumefaciens, Sinorhizobium meliloti), impairing PG crosslinkage and cell division. Using A. tumefaciens we demonstrated that the detrimental effect of D-CAN is suppressed by a single amino acid substitution in the cell division PG transpeptidase penicillin binding protein 3a. Collectively, this work highlights the role of amino acid racemization in cell wall chemical editing and fitness.

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  • 24.
    Aliashkevich, Alena
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Schiffthaler, Bastian
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Umeå Plant Science Centre (UPSC). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysiologisk botanik.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Genetic dissection of LD-transpeptidation in Agrobacterium tumefaciensManuskript (preprint) (Annet vitenskapelig)
  • 25. Allas, Ular
    et al.
    Toom, Lauri
    Selyutina, Anastasia
    Maeorg, Uno
    Medina, Ricardo
    Merits, Andres
    Rinken, Ago
    Hauryliuk, Vasili
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). University of Tartu, Institute of Technology, Nooruse 1, Tartu 50411, Estonia.
    Kaldalu, Niilo
    Tenson, Tanel
    Antibacterial activity of the nitrovinylfuran G1 (Furvina) and its conversion products2016Inngår i: Scientific Reports, E-ISSN 2045-2322, Vol. 6, artikkel-id 36844Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    2-Bromo-5-(2-bromo-2-nitrovinyl) furan (G1 or Furvina) is an antimicrobial with a direct reactivity against thiol groups. It is active against Gram-positive and Gram-negative bacteria, yeasts and filamentous fungi. By reacting with thiol groups it causes direct damage to proteins but, as a result, is very short-living and interconverts into an array of reaction products. Our aim was to characterize thiol reactivity of G1 and its conversion products and establish how much of antimicrobial and cytotoxic effects are due to the primary activity of G1 and how much can be attributed to its reaction products. Stability of G1 in growth media as well as its conversion in the presence of thiols was characterized. The structures of G1 decomposition products were determined using NMR and mass-spectroscopy. Concentration-and time-dependent killing curves showed that G1 is bacteriostatic for Escherichia coli at the concentration of 16 mu g/ml and bactericidal at 32 mu g/ml. However, G1 is inefficient against non-growing E. coli. Addition of cysteine to medium reduces the antimicrobial potency of G1. Nevertheless, the reaction products of G1 and cysteine enabled prolonged antimicrobial action of the drug. Therefore, the activity of 2-bromo-5-(2-bromo-2-nitrovinyl) furan is a sum of its immediate reactivity and the antibacterial effects of the conversion products.

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  • 26.
    Almyroudis, Nikolaos G.
    et al.
    Roswell Park Cancer Institute, Buffalo, New York, United States of America.
    Grimm, Melissa J.
    Roswell Park Cancer Institute, Buffalo, New York, United States of America.
    Davidson, Bruce A.
    Roswell Park Cancer Institute, Buffalo, New York, United States of America.
    Röhm, Marc
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Urban, Constantin F.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Segal, Brahm H.
    Roswell Park Cancer Institute, Buffalo, New York, United States of America.
    NETosis and NADPH oxidase: at the intersection of host defense, inflammation, and injury2013Inngår i: Frontiers in Immunology, E-ISSN 1664-3224, Vol. 4, artikkel-id 45Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Neutrophils are armed with both oxidant-dependent and -independent pathways for killing pathogens. Activation of the phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase constitutes an emergency response to infectious threat and results in the generation of antimicrobial reactive oxidants. In addition, NADPH oxidase activation in neutrophils is linked to activation of granular proteases and generation of neutrophil extracellular traps (NETs). NETosis involves the release of nuclear and granular components that can target extracellular pathogens. NETosis is activated during microbial threat and in certain conditions mimicking sepsis, and can result in both augmented host defense and inflammatory injury. In contrast, apoptosis, the physiological form of neutrophil death, not only leads to non-inflammatory cell death but also contributes to alleviate inflammation. Although there are significant gaps in knowledge regarding the specific contribution of NETs to host defense, we speculate that the coordinated activation of NADPH oxidase and NETosis maximizes microbial killing. Work in engineered mice and limited patient experience point to varying susceptibility of bacterial and fungal pathogens to NADPH oxidase versus NET constituents. Since reactive oxidants and NET constituents can injure host tissue, it is important that these pathways be tightly regulated. Recent work supports a role for NETosis in both acute lung injury and in autoimmunity. Knowledge gained about mechanisms that modulate NETosis may lead to novel therapeutic approaches to limit inflammation-associated injury.

    Fulltekst (pdf)
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  • 27.
    Alvarez, Laura
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Aliashkevich, Alena
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    de Pedro, Miguel A.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Bacterial secretion of D-arginine controls environmental microbial biodiversity2018Inngår i: The ISME Journal, ISSN 1751-7362, E-ISSN 1751-7370, Vol. 12, nr 2, s. 438-450Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Bacteria face tough competition in polymicrobial communities. To persist in a specific niche, many species produce toxic extracellular effectors to interfere with the growth of nearby microbes. These effectors include the recently reported non-canonical D-amino acids (NCDAAs). In Vibrio cholerae, the causative agent of cholera, NCDAAs control cell wall integrity in stationary phase. Here, an analysis of the composition of the extracellular medium of V. cholerae revealed the unprecedented presence of D-Arg. Compared with other D-amino acids, D-Arg displayed higher potency and broader toxicity in terms of the number of bacterial species affected. Tolerance to D-Arg was associated with mutations in the phosphate transport and chaperone systems, whereas D-Met lethality was suppressed by mutations in cell wall determinants. These observations suggest that NCDAAs target different cellular processes. Finally, even though virtually all Vibrio species are tolerant to D-Arg, only a few can produce this D-amino acid. Indeed, we demonstrate that D-Arg may function as part of a cooperative strategy in vibrio communities to protect non-producing members from competing bacteria. Because NCDAA production is widespread in bacteria, we anticipate that D-Arg is a relevant modulator of microbial subpopulations in diverse ecosystems.

  • 28.
    Alvarez, Laura
    et al.
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Bacterial Competition Assay Based on Extracellular D-amino Acid Production2018Inngår i: Bio-protocol, E-ISSN 2331-8325, Vol. 8, nr 7, artikkel-id e2787Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Bacteria live in polymicrobial communities under tough competition. To persist in a specific niche many species produce toxic extracellular effectors as a strategy to interfere with the growth of nearby microbes. One of such effectors are the non-canonical D-amino acids. Here we describe a method to test the effect of D-amino acid production in fitness/survival of bacterial subpopulations within a community. Co-cultivation methods usually involve the growth of the competing bacteria in the same container. Therefore, within such mixed cultures the effect on growth caused by extracellular metabolites cannot be distinguished from direct physical interactions between species (e.g., T6SS effectors). However, this problem can be easily solved by using a filtration unit that allows free diffusion of small metabolites, like L- and D-amino acids, while keeping the different subpopulations in independent compartments. With this method, we have demonstrated that D-arginine is a bactericide effector produced by Vibrio cholerae, which strongly influences survival of diverse microbial subpopulations. Moreover, D-arginine can be used as a cooperative instrument in mixed Vibrio communities to protect non-producing members from competing bacteria.

  • 29.
    Alvarez, Laura
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Espaillat, Akbar
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Hermoso, Juan A.
    de Pedro, Miguel A.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Peptidoglycan Remodeling by the Coordinated Action of Multispecific Enzymes2014Inngår i: Microbial Drug Resistance, ISSN 1076-6294, E-ISSN 1931-8448, Vol. 20, nr 3, s. 190-198Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The peptidoglycan (PG) cell wall constitutes the main defense barrier of bacteria against environmental insults and acts as communication interface. The biochemistry of this macromolecule has been well characterized throughout the years but recent discoveries have unveiled its chemical plasticity under environmental stresses. Non-canonical D-amino acids (NCDAA) are produced and released to the extracellular media by diverse bacteria. Such molecules govern cell wall adaptation to challenging environments through their incorporation into the polymer, a widespread capability among bacteria that reveals the inherent catalytic plasticity of the enzymes involved in the cell wall metabolism. Here, we analyze the recent structural and biochemical characterization of Bsr, a new family of broad spectrum racemases able to generate a wide range of NCDAA. We also discuss the necessity of a coordinated action of PG multispecific enzymes to generate adequate levels of modification in the murein sacculus. Finally, we also highlight how this catalytic plasticity of NCDAA-incorporating enzymes has allowed the development of new revolutionary methodologies for the study of PG modes of growth and in vivo dynamics.

  • 30.
    Alvarez, Laura
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Hernandez, Sara B.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Cell Wall Biology of Vibrio cholerae2021Inngår i: Annual Review of Microbiology, ISSN 0066-4227, E-ISSN 1545-3251, Vol. 75, s. 151-174Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Most bacteria are protected from environmental offenses by a cell wall consisting of strong yet elastic peptidoglycan. The cell wall is essential for preserving bacterial morphology and viability, and thus the enzymes involved in the production and turnover of peptidoglycan have become preferred targets for many of our most successful antibiotics. In the past decades, Vibrio cholerae, the gram-negative pathogen causing the diarrheal disease cholera, has become a major model for understanding cell wall genetics, biochemistry, and physiology. More than 100 articles have shed light on novel cell wall genetic determinants, regulatory links, and adaptive mechanisms. Here we provide the first comprehensive review of V. cholerae's cell wall biology and genetics. Special emphasis is placed on the similarities and differences with Escherichia coli, the paradigm for understanding cell wall metabolism and chemical structure in gram-negative bacteria.

  • 31.
    Alvarez, Laura
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Hernandez, Sara B
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    de Pedro, Miguel A
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Ultra-sensitive, high-resolution liquid chromatography methods for the high-throughput quantitative analysis of bacterial cell wall chemistry and structure2016Inngår i: Bacterial cell wall homeostasis: methods and protocols /edited by Hee-Jeon Hong / [ed] Hee-Jeon Hong, New York: Humana Press, 2016, Vol. 1440, s. 11-27Kapittel i bok, del av antologi (Fagfellevurdert)
    Abstract [en]

    High-performance liquid chromatography (HPLC) analysis has been critical for determining the structural and chemical complexity of the cell wall. However this method is very time consuming in terms of sample preparation and chromatographic separation. Here we describe (1) optimized methods for peptidoglycan isolation from both Gram-negative and Gram-positive bacteria that dramatically reduce the sample preparation time, and (2) the application of the fast and highly efficient ultra-performance liquid chromatography (UPLC) technology to muropeptide separation and quantification. The advances in both analytical instrumentation and stationary-phase chemistry have allowed for evolved protocols which cut run time from hours (2-3 h) to minutes (10-20 min), and sample demands by at least one order of magnitude. Furthermore, development of methods based on organic solvents permits in-line mass spectrometry (MS) of the UPLC-resolved muropeptides. Application of these technologies to high-throughput analysis will expedite the better understanding of the cell wall biology.

  • 32.
    Alvarez, Laura
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Centro de Biología Molecular Severo Ochoa, Universidad Autónoma de Madrid- Consejo Superior de Investigaciones Científicas, Madrid, Spain.
    Sanchez-Hevia, Dione
    Sanchez, Mercedes
    Berenguer, Jose
    A new family of nitrate/nitrite transporters involved in denitrification2019Inngår i: International Microbiology, ISSN 1139-6709, E-ISSN 1618-1905, Vol. 22, nr 1, s. 19-28Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Denitrifying bacteria carry out nitrate and nitrite respiration inside and outside the cell, respectively. In Thermus thermophilus, nitrate and nitrite transport processes are carried out by major facilitator superfamily (MFS) transporters. The sequence of the nar operon of nitrate-only respiring strains of T. thermophilus includes two tandemly organized MFS transporter genes (narK and narT) of the NarK1 and NarK2 families. Both can function as nitrate/nitrite antiporters, but NarK has been proposed as more specific for nitrate whereas NarT more specific for nitrite. In some nitrate- and nitrite-respiring strains of the same species, a single MFS transporter (NarO) belonging to a different MFS subfamily appears. To analyze the role of this single MFS in the same genetic context, we transferred the two types of nar operon to the aerobic strain HB27, and further included in both of them the ability to respire nitrite. The new denitrifying strains HB27dn, with two MFS, and HB27dp, with a single one, were used to isolate mutants devoid of transporters. Through in trans complementation experiments, we demonstrate that the NarO single MFS works efficiently in the transport of both nitrate and nitrite.

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  • 33.
    Amer, Ayad
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Costa, Tiago
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Farag, Salah
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Avican, Ummehan
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Forsberg, Åke
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Francis, Matthew
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Genetically engineered frameshifted YopN-TyeA chimeras influence type III secretion system function in Yersinia pseudotuberculosis2013Inngår i: PLOS ONE, E-ISSN 1932-6203, Vol. 8, nr 10, artikkel-id e77767Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Type III secretion is a tightly controlled virulence mechanism utilized by many gram negative bacteria to colonize their eukaryotic hosts. To infect their host, human pathogenic Yersinia spp. translocate protein toxins into the host cell cytosol through a preassembled Ysc-Yop type III secretion device. Several of the Ysc-Yop components are known for their roles in controlling substrate secretion and translocation. Particularly important in this role is the YopN and TyeA heterodimer. In this study, we confirm that Y. pseudotuberculosis naturally produce a 42 kDa YopN-TyeA hybrid protein as a result of a +1 frame shift near the 3 prime of yopN mRNA, as has been previously reported for the closely related Y. pestis. To assess the biological role of this YopN-TyeA hybrid in T3SS by Y. pseudotuberculosis, we used in cis site-directed mutagenesis to engineer bacteria to either produce predominately the YopN-TyeA hybrid by introducing +1 frame shifts to yopN after codon 278 or 287, or to produce only singular YopN and TyeA polypeptides by introducing yopN sequence from Y. enterocolitica, which is known not to produce the hybrid. Significantly, the engineered 42 kDa YopN-TyeA fusions were abundantly produced, stable, and were efficiently secreted by bacteria in vitro. Moreover, these bacteria could all maintain functionally competent needle structures and controlled Yops secretion in vitro. In the presence of host cells however, bacteria producing the most genetically altered hybrids (+1 frameshift after 278 codon) had diminished control of polarized Yop translocation. This corresponded to significant attenuation in competitive survival assays in orally infected mice, although not at all to the same extent as Yersinia lacking both YopN and TyeA proteins. Based on these studies with engineered polypeptides, most likely a naturally occurring YopN-TyeA hybrid protein has the potential to influence T3S control and activity when produced during Yersinia-host cell contact.

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  • 34.
    Amer, Ayad
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Gurung, Jyoti
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Costa, Tiago
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Ruuth, Kristina
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Zavialov, Anton
    Joint Biotechnology Laboratory, Department of Chemistry, University of Turku, Turku, Finland.
    Forsberg, Åke
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Francis, Matthew S
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    YopN and TyeA Hydrophobic Contacts Required for Regulating Ysc-Yop Type III Secretion Activity by Yersinia pseudotuberculosis2016Inngår i: Frontiers in Cellular and Infection Microbiology, E-ISSN 2235-2988, Vol. 6, artikkel-id 66Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Yersinia bacteria target Yop effector toxins to the interior of host immune cells by the Ysc-Yop type III secretion system. A YopN-TyeA heterodimer is central to controlling Ysc-Yop targeting activity. A + 1 frameshift event in the 3-prime end of yopN can also produce a singular secreted YopN-TyeA polypeptide that retains some regulatory function even though the C-terminal coding sequence of this YopN differs greatly from wild type. Thus, this YopN C-terminal segment was analyzed for its role in type III secretion control. Bacteria producing YopN truncated after residue 278, or with altered sequence between residues 279 and 287, had lost type III secretion control and function. In contrast, YopN variants with manipulated sequence beyond residue 287 maintained full control and function. Scrutiny of the YopN-TyeA complex structure revealed that residue W279 functioned as a likely hydrophobic contact site with TyeA. Indeed, a YopNW279G mutant lost all ability to bind TyeA. The TyeA residue F8 was also critical for reciprocal YopN binding. Thus, we conclude that specific hydrophobic contacts between opposing YopN and TyeA termini establishes a complex needed for regulating Ysc-Yop activity.

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  • 35.
    Amer, Ayad
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Åhlund, Monika
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Bröms, Jeanette
    Department of Medical Countermeasures, Swedish Defense Research Agency, Division of NBC12 Defense, Umeå, Sweden.
    Forsberg, Åke
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Francis, Matthew
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Impact of the N-terminal secretor domain on YopD translocator function in Yersinia pseudotuberculosis type III secretion2011Inngår i: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 193, nr 23, s. 6683-6700Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Type III secretion systems (T3SSs) secrete needle components, pore-forming translocators, and the translocated effectors. In part, effector recognition by a T3SS involves their N-terminal amino acids and their 5′ mRNA. To investigate whether similar molecular constraints influence translocator secretion, we scrutinized this region within YopD from Yersinia pseudotuberculosis. Mutations in the 5′ end of yopD that resulted in specific disruption of the mRNA sequence did not affect YopD secretion. On the other hand, a few mutations affecting the protein sequence reduced secretion. Translational reporter fusions identified the first five codons as a minimal N-terminal secretion signal and also indicated that the YopD N terminus might be important for yopD translation control. Hybrid proteins in which the N terminus of YopD was exchanged with the equivalent region of the YopE effector or the YopB translocator were also constructed. While the in vitro secretion profile was unaltered, these modified bacteria were all compromised with respect to T3SS activity in the presence of immune cells. Thus, the YopD N terminus does harbor a secretion signal that may also incorporate mechanisms of yopD translation control. This signal tolerates a high degree of variation while still maintaining secretion competence suggestive of inherent structural peculiarities that make it distinct from secretion signals of other T3SS substrates.

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  • 36. Amon, Jeremy D.
    et al.
    Yadav, Akhilesh K.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Analytical Chemistry Division, CSIR-Indian Institute of Toxicology Research, Lucknow, India.
    Ramirez-Guadiana, Fernando H.
    Meeske, Alexander J.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Rudner, David Z.
    SwsB and SafA Are Required for CwlJ-Dependent Spore Germination in Bacillus subtilis2020Inngår i: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 202, nr 6, artikkel-id e00668-19Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    When Bacillus subtilis spores detect nutrients, they exit dormancy through the processes of germination and outgrowth. A key step in germination is the activation of two functionally redundant cell wall hydrolases (SleB and CwlJ) that degrade the specialized cortex peptidoglycan that surrounds the spore. How these enzymes are regulated remains poorly understood. To identify additional factors that affect their activity, we used transposon sequencing to screen for synthetic germination defects in spores lacking SleB or CwlJ. Other than the previously characterized protein YpeB, no additional factors were found to be specifically required for SleB activity. In contrast, our screen identified SafA and YlxY (renamed SwsB) in addition to the known factors GerQ and CotE as proteins required for CwlJ function. SafA is a member of the spore's proteinaceous coat and we show that, like GerQ and CotE, it is required for accumulation and retention of CwlJ in the dormant spore. SwsB is broadly conserved among spore formers, and we show that it is required for CwlJ to efficiently degrade the cortex during germination. Intriguingly, SwsB resembles polysaccharide deacetylases, and its putative catalytic residues are required for its role in germination. However, we find no chemical signature of its activity on the spore cortex or in vitro. While the precise, mechanistic role of SwsB remains unknown, we explore and discuss potential activities. IMPORTANCE Spore formation in Bacillus subtilis has been studied for over half a century, and virtually every step in this developmental process has been characterized in molecular detail. In contrast, how spores exit dormancy remains less well understood. A key step in germination is the degradation of the specialized cell wall surrounding the spore called the cortex. Two enzymes (SleB and CwlJ) specifically target this protective layer, but how they are regulated and whether additional factors promote their activity are unknown. Here, we identified the coat protein SafA and a conserved but uncharacterized protein YlxY as additional factors required for CwlJ-dependent degradation of the cortex. Our analysis provides a more complete picture of this essential step in the exit from dormancy.

  • 37.
    Andersson, Christopher
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Regulatory pathways and virulence inhibition in Listeria monocytogenes2016Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Listeria monocytogenes is a rod-shaped Gram positive bacterium. It generally exist ubiquitously in nature, where it lives as a saprophyte. Occasionally it however enters the food chain, from where it can be ingested by humans and cause gastro-intestinal distress. In immunocompetent individuals L. monocytogenes is generally cleared within a couple of weeks, but in immunocompromised patients it can progress to listeriosis, a potentially life-threatening infection in the central nervous system. If the infected individual is pregnant, the bacteria can cross the placental barrier and infect the fetus, possibly leading to spontaneous abortion.

    The infectivity of L. monocytogenes requires a certain set of genes, and the majority of them is dependent on the transcriptional regulator PrfA. The expression and activity of PrfA is controlled at several levels, and has traditionally been viewed to be active at 37 °C (virulence conditions) where it bind as a homodimer to a “PrfA-box” and induces the expression of the downstream gene.

    One of these genes is ActA, which enables intracellular movement by recruiting an actin polymerizing protein complex. When studying the effects of a blue light receptor we surprisingly found an effect of ActA at non-virulent conditions, where it is required for the bacteria to properly react to light exposure.

    To further study the PrfA regulon we tested deletion mutants of several PrfA-regulated virulence genes in chicken embryo infection studies. Based on these studies we could conclude that the chicken embryo model is a viable complement to traditional murine models, especially when investigating non-traditional internalin pathogenicity pathways. We have also studied the effects of small molecule virulence inhibitors that, by acting on PrfA, can inhibit L. monocytogenes infectivity in cell cultures with concentrations in the low micro-molar range.

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  • 38.
    Andersson, Christopher
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Gripenland, Jonas
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Johansson, Jörgen
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Using the chicken embryo to assess virulence of Listeria monocytogenes and to model other microbial infections2015Inngår i: Nature Protocols, ISSN 1754-2189, E-ISSN 1750-2799, Vol. 10, nr 8, s. 1155-1164Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Microbial infections are a global health problem, particularly as microbes are continually developing resistance to antimicrobial treatments. An effective and reliable method for testing the virulence of different microbial pathogens is therefore a useful research tool. This protocol describes how the chicken embryo can be used as a trustworthy, inexpensive, ethically desirable and quickly accessible model to assess the virulence of the human bacterial pathogen Listeria monocytogenes, which can also be extended to other microbial pathogens. We provide a step-by-step protocol and figures and videos detailing the method, including egg handling, infection strategies, pathogenicity screening and isolation of infected organs. From the start of incubation of the fertilized eggs, the protocol takes <4 weeks to complete, with the infection part taking only 3 d. We discuss the appropriate controls to use and potential adjustments needed for adapting the protocol for other microbial pathogens.

  • 39.
    Andresen, Liis
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Tenson, Tanel
    Hauryliuk, Vasili
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). University of Tartu, Institute of Technology, Nooruse 1, 50411 Tartu, Estonia.
    Cationic bactericidal peptide 1018 does not specifically target the stringent response alarmone (p)ppGpp2016Inngår i: Scientific Reports, E-ISSN 2045-2322, Vol. 6, artikkel-id 36549Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The bacterial stringent response is a key regulator of bacterial virulence, biofilm formation and antibiotic tolerance, and is a promising target for the development of new antibacterial compounds. The intracellular nucleotide (p)ppGpp acts as a messenger orchestrating the stringent response. A synthetic peptide 1018 was recently proposed to specifically disrupt biofilms by inhibiting the stringent response via direct interaction with (p) ppGpp (de la Fuente-Nunez et al. (2014) PLoS Pathogens). We have interrogated the specificity of the proposed molecular mechanism. When inhibition of Pseudomonas aeruginosa planktonic and biofilm growth is tested simultaneously in the same assay, peptides 1018 and the control peptide 8101 generated by an inversion of the amino acid sequence of 1018 are equally potent, and, importantly, do not display a preferential activity against biofilm. 1018 inhibits planktonic growth of Escherichia coli equally efficiently either when the alleged target, (p) ppGpp, is essential (MOPS media lacking amino acid L-valine), or dispensable for growth (MOPS media supplemented with L-valine). Genetic disruption of the genes relA and spoT responsible for (p) ppGpp synthesis moderately sensitizes-rather than protects-E. coli to 1018. We suggest that the antimicrobial activity of 1018 does not rely on specific recognition of the stringent response messenger (p) ppGpp.

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  • 40.
    Andresen, Liis
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Varik, Vallo
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). University of Tartu, Institute of Technology, Nooruse 1, 50411 Tartu, Estonia.
    Tozawa, Yuzuru
    Jimmy, Steffi
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Lindberg, Stina
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Tenson, Tanel
    Hauryliuk, Vasili
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). University of Tartu, Institute of Technology, Nooruse 1, 50411 Tartu, Estonia.
    Auxotrophy-based High Throughput Screening assay for the identification of Bacillus subtilis stringent response inhibitors2016Inngår i: Scientific Reports, E-ISSN 2045-2322, Vol. 6, artikkel-id 35824Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The stringent response is a central adaptation mechanism that allows bacteria to adjust their growth and metabolism according to environmental conditions. The functionality of the stringent response is crucial for bacterial virulence, survival during host invasion as well as antibiotic resistance and tolerance. Therefore, specific inhibitors of the stringent response hold great promise as molecular tools for disarming and pacifying bacterial pathogens. By taking advantage of the valine amino acid auxotrophy of the Bacillus subtilis stringent response-deficient strain, we have set up a High Throughput Screening assay for the identification of stringent response inhibitors. By screening 17,500 compounds, we have identified a novel class of antibacterials based on the 4-(6-(phenoxy) alkyl)-3,5-dimethyl-1H-pyrazole core. Detailed characterization of the hit compounds as well as two previously identified promising stringent response inhibitors-a ppGpp-mimic nucleotide Relacin and cationic peptide 1018 - showed that neither of the compounds is sufficiently specific, thus motivating future application of our screening assay to larger and more diverse molecular libraries.

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  • 41.
    Angelin, Martin
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Infektionssjukdomar.
    Forsell, Joakim
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi.
    Granlund, Margareta
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi.
    Evengård, Birgitta
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Infektionssjukdomar.
    Palmgren, Helena
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Infektionssjukdomar.
    Johansson, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Risk factors for colonization with extended-spectrum beta-lactamase producing Enterobacteriaceae in healthcare students on clinical assignment abroad: A prospective study2015Inngår i: Travel Medicine and Infectious Disease, ISSN 1477-8939, E-ISSN 1873-0442, Vol. 13, nr 3, s. 223-229Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: The increase of antibiotic resistance in clinically important bacteria is a worldwide threat, especially in healthcare environments. International travel is a risk factor for gut colonization with extended-spectrum beta-lactamase-producing Enterobacteriaceae (ESBL-PE). The risk for healthcare students of being colonized with ESBL-PE when participating in patient-related work abroad has not been previously investigated. Methods: Swedish healthcare students travelling for pre-clinical and clinical courses outside Scandinavia submitted faecal samples and survey data before and after travel. The faecal samples were screened for ESBL-PE and carbapenemase-producing Enterobacteriaceae (CPE). Screening results and survey data were analysed to identify risk factors for colonization. Results: In the 99 subjects who submitted a full set of samples, 35% were colonized with a new ESBL-PE strain during travel. No CPE was found. The most important risk factor for ESBL-PE colonization was travel destination, and the highest colonization rate was found in the South East Asia region. Antibiotic treatment during travel was an independent risk factor for ESBL-PE colonization but patient-related work was not significantly associated with an increased risk. Conclusions: Patient-related work abroad was not a risk factor for ESBL-PE suggesting that transmission from patients is uncommon. Pre-travel advice on avoiding unnecessary antibiotic treatment during travel is recommended.

  • 42.
    Antti, Henrik
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Fahlgren, Anna
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Näsström, Elin
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Kouremenos, Konstantinos
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Sundén-Cullberg, Jonas
    Guo, Yongzhi
    Moritz, Thomas
    Wolf-Watz, Hans
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Johansson, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Infektionssjukdomar.
    Fällman, Maria
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Metabolic profiling for detection of staphylococcus aureus infection and antibiotic resistance2013Inngår i: PLOS ONE, E-ISSN 1932-6203, Vol. 8, nr 2, artikkel-id e56971Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Due to slow diagnostics, physicians must optimize antibiotic therapies based on clinical evaluation of patients without specific information on causative bacteria. We have investigated metabolomic analysis of blood for the detection of acute bacterial infection and early differentiation between ineffective and effective antibiotic treatment. A vital and timely therapeutic difficulty was thereby addressed: the ability to rapidly detect treatment failures because of antibiotic-resistant bacteria. Methicillin-resistant (MRSA) and methicillin-sensitive (MSSA) were used and for infecting mice, while natural MSSA infection was studied in humans. Samples of bacterial growth media, the blood of infected mice and of humans were analyzed with combined Gas Chromatography/Mass Spectrometry. Multivariate data analysis was used to reveal the metabolic profiles of infection and the responses to different antibiotic treatments. experiments resulted in the detection of 256 putative metabolites and mice infection experiments resulted in the detection of 474 putative metabolites. Importantly, ineffective and effective antibiotic treatments were differentiated already two hours after treatment start in both experimental systems. That is, the ineffective treatment of MRSA using cloxacillin and untreated controls produced one metabolic profile while all effective treatment combinations using cloxacillin or vancomycin for MSSA or MRSA produced another profile. For further evaluation of the concept, blood samples of humans admitted to intensive care with severe sepsis were analyzed. One hundred thirty-three putative metabolites differentiated severe MSSA sepsis (n = 6) from severe sepsis (n = 10) and identified treatment responses over time. Combined analysis of human, , and mice samples identified 25 metabolites indicative of effective treatment of sepsis. Taken together, this study provides a proof of concept of the utility of analyzing metabolite patterns in blood for early differentiation between ineffective and effective antibiotic treatment in acute infections.

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  • 43. Arenz, Stefan
    et al.
    Abdelshahid, Maha
    Sohmen, Daniel
    Payoe, Roshani
    Starosta, Agata L.
    Berninghausen, Otto
    Hauryliuk, Vasili
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). University of Tartu, Institute of Technology, Tartu, Estonia.
    Beckmann, Roland
    Wilson, Daniel N.
    The stringent factor RelA adopts an open conformation on the ribosome to stimulate ppGpp synthesis2016Inngår i: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 44, nr 13, s. 6471-6481Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Under stress conditions, such as nutrient starvation, deacylated tRNAs bound within the ribosomal A-site are recognized by the stringent factor RelA, which converts ATP and GTP/GDP to (p)ppGpp. The signaling molecules (p) ppGpp globally rewire the cellular transcriptional program and general metabolism, leading to stress adaptation. Despite the additional importance of the stringent response for regulation of bacterial virulence, antibiotic resistance and persistence, structural insight into how the ribosome and deacylated-tRNA stimulate RelA-mediated (p)ppGpp has been lacking. Here, we present a cryo-EM structure of RelA in complex with the Escherichia coli 70S ribosome with an average resolution of 3.7 angstrom and local resolution of 4 to > 10 angstrom for RelA. The structure reveals that RelA adopts a unique 'open' conformation, where the C-terminal domain (CTD) is intertwined around an A/T-like tRNA within the intersubunit cavity of the ribosome and the N-terminal domain (NTD) extends into the solvent. We propose that the open conformation of RelA on the ribosome relieves the autoinhibitory effect of the CTD on the NTD, thus leading to stimulation of (p)ppGpp synthesis by RelA.

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  • 44. Asghar, Naveed
    et al.
    Lee, Yi-Ping
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Nilsson, Emma
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Lindqvist, Richard
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Melik, Wessam
    Kröger, Andrea
    Överby, Anna K.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Johansson, Magnus
    The role of the poly(A) tract in the replication and virulence of tick-borne encephalitis virus2016Inngår i: Scientific Reports, E-ISSN 2045-2322, Vol. 6, artikkel-id 39265Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The tick-borne encephalitis virus (TBEV) is a flavivirus transmitted to humans, usually via tick bites. The virus causes tick-borne encephalitis (TBE) in humans, and symptoms range from mild flu-like symptoms to severe and long-lasting sequelae, including permanent brain damage. It has been suggested that within the population of viruses transmitted to the mammalian host, quasispecies with neurotropic properties might become dominant in the host resulting in neurological symptoms. We previously demonstrated the existence of TBEV variants with variable poly(A) tracts within a single blood-fed tick. To characterize the role of the poly(A) tract in TBEV replication and virulence, we generated infectious clones of Toro-2003 with the wild-type (A)(3)C(A)(6) sequence (Toro-6A) or with a modified (A)(3)C(A)(38) sequence (Toro-38A). Toro-38A replicated poorly compared to Toro-6A in cell culture, but Toro-38A was more virulent than Toro-6A in a mouse model of TBE. Next-generation sequencing of TBEV genomes after passaging in cell culture and/or mouse brain revealed mutations in specific genomic regions and the presence of quasispecies that might contribute to the observed differences in virulence. These data suggest a role for quasispecies development within the poly(A) tract as a virulence determinant for TBEV in mice.

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  • 45. Askarian, Fatemeh
    et al.
    Lapek, John D., Jr.
    Dongre, Mitesh
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Tsai, Chih-Ming
    Kumaraswamy, Monika
    Kousha, Armin
    Valderrama, J. Andres
    Ludviksen, Judith A.
    Cavanagh, Jorunn P.
    Uchiyama, Satoshi
    Mollnes, Tom E.
    Gonzalez, David J.
    Wai, Sun N.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Nizet, Victor
    Johannessen, Mona
    Staphylococcus aureus Membrane-Derived Vesicles Promote Bacterial Virulence and Confer Protective Immunity in Murine Infection Models2018Inngår i: Frontiers in Microbiology, E-ISSN 1664-302X, Vol. 9, artikkel-id 262Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Staphylococcus aureus produces membrane-derived vesicles (MVs), which share functional properties to outer membrane vesicles. Atomic force microscopy revealed that S. aureus-derived MVs are associated with the bacterial surface or released into the surrounding environment depending on bacterial growth conditions. By using a comparative proteomic approach, a total of 131 and 617 proteins were identified in MVs isolated from S. aureus grown in Luria-Bertani and brain-heart infusion broth, respectively. Purified S. aureus MVs derived from the bacteria grown in either media induced comparable levels of cytotoxicity and neutrophil-activation. Administration of exogenous MVs increased the resistance of S. aureus to killing by whole blood or purified human neutrophils ex vivo and increased S. aureus survival in vivo. Finally, immunization of mice with S. aureus-derived MVs induced production of IgM, total IgG, IgG1, IgG2a, and IgG2b resulting in protection against subcutaneous and systemic S. aureus infection. Collectively, our results suggest S. aureus MVs can influence bacterial-host interactions during systemic infections and provide protective immunity in murine models of infection.

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  • 46.
    Atkinson, Gemma C.
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). University of Tartu, Institute of Technology, Nooruse 1, 50411 Tartu, Estonia .
    Kuzmenko, Anton
    University of Tartu, Institute of Technology, Nooruse 1, 50411 Tartu, Estonia & Department of Molecular Biology, Faculty of Biology, Moscow State University, Moscow, Russia .
    Chicherin, Ivan
    Department of Molecular Biology, Faculty of Biology, Moscow State University, Moscow, Russia.
    Soosaar, Axel
    University of Tartu, Institute of Technology, Nooruse 1, 50411 Tartu, Estonia .
    Tenson, Tanel
    University of Tartu, Institute of Technology, Nooruse 1, 50411 Tartu, Estonia .
    Carr, Martin
    School of Applied Sciences, University of Huddersfield, Queensgate, Huddersfield, UK.
    Kamenski, Piotr
    Department of Molecular Biology, Faculty of Biology, Moscow State University, Moscow, Russia .
    Hauryliuk, Vasili
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). University of Tartu, Institute of Technology, Nooruse 1, 50411 Tartu, Estonia .
    An evolutionary ratchet leading to loss of elongation factors in eukaryotes2014Inngår i: BMC Evolutionary Biology, E-ISSN 1471-2148, Vol. 14, s. 35-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: The GTPase eEF1A is the eukaryotic factor responsible for the essential, universal function of aminoacyl-tRNA delivery to the ribosome. Surprisingly, eEF1A is not universally present in eukaryotes, being replaced by the paralog EFL independently in multiple lineages. The driving force behind this unusually frequent replacement is poorly understood. Results: Through sequence searching of genomic and EST databases, we find a striking association of eEF1A replacement by EFL and loss of eEF1A's guanine exchange factor, eEF1Ba, suggesting that EFL is able to spontaneously recharge with GTP. Sequence conservation and homology modeling analyses indicate several sequence regions that may be responsible for EFL's lack of requirement for eEF1Ba. Conclusions: We propose that the unusual pattern of eEF1A, eEF1Ba and EFL presence and absence can be explained by a ratchet-like process: if either eEF1A or eEF1Ba diverges beyond functionality in the presence of EFL, the system is unable to return to the ancestral, eEF1A:eEFBa-driven state.

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  • 47.
    Atkinson, Gemma Catherine
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Institute of Technology, University of Tartu, Nooruse 1, Tartu, 50411, Estonia.
    The evolutionary and functional diversity of classical and lesser-known cytoplasmic and organellar translational GTPases across the tree of life2015Inngår i: BMC Genomics, E-ISSN 1471-2164, Vol. 16, artikkel-id 78Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: The ribosome translates mRNA to protein with the aid of a number of accessory protein factors. Translational GTPases (trGTPases) are an integral part of the 'core set' of essential translational factors, and are some of the most conserved proteins across life. This study takes advantage of the wealth of available genomic data, along with novel functional information that has come to light for a number of trGTPases to address the full evolutionary and functional diversity of this superfamily across all domains of life. 

    Results: Through sensitive sequence searching combined with phylogenetic analysis, 57 distinct subfamilies of trGTPases are identified: 14 bacterial, 7 archaeal and 35 eukaryotic (of which 21 are known or predicted to be organellar). The results uncover the functional evolution of trGTPases from before the last common ancestor of life on earth to the current day. 

    Conclusions: While some trGTPases are universal, others are limited to certain taxa, suggesting lineage-specific translational control mechanisms that exist on a base of core factors. These lineage-specific features may give organisms the ability to tune their translation machinery to respond to their environment. Only a fraction of the diversity of the trGTPase superfamily has been subjected to experimental analyses; this comprehensive classification brings to light novel and overlooked translation factors that are worthy of further investigation.

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  • 48.
    Aung, Kyaw Min
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Sjöström, Annika E
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    von Pawel-Rammingen, Ulrich
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Riesbeck, Kristian
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Wai, Sun Nyunt
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Naturally Occurring IgG Antibodies Provide Innate Protection against Vibrio cholerae Bacteremia by Recognition of the Outer Membrane Protein U2016Inngår i: Journal of Innate Immunity, ISSN 1662-811X, E-ISSN 1662-8128, Vol. 8, nr 3, s. 269-283Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Cholera epidemics are caused by Vibrio cholerae serogroups O1 and O139, whereas strains collectively known as non-O1/non-O139 V. cholerae are found in cases of extraintestinal infections and bacteremia. The mechanisms and factors influencing the occurrence of bacteremia and survival of V. cholerae in normal human serum have remained unclear. We found that naturally occurring IgG recognizing V. cholerae outer membrane protein U (OmpU) mediates a serum-killing effect in a complement C1q-dependent manner. Moreover, outer membrane vesicles (OMVs) containing OmpU caused enhanced survival of highly serum-sensitive classical V. cholerae in a dose-dependent manner. OMVs from wild-type and ompU mutant V. cholerae thereby provided a novel means to verify by extracellular transcomplementation the involvement of OmpU. Our data conclusively indicate that loss, or reduced expression, of OmpU imparts resistance to V. cholerae towards serum killing. We propose that the difference in OmpU protein levels is a plausible reason for differences in serum resistance and the ability to cause bacteremia observed among V. cholerae biotypes. Our findings provide a new perspective on how naturally occurring antibodies, perhaps induced by members of the microbiome, may play a role in the recognition of pathogens and the provocation of innate immune defense against bacteremia.

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  • 49.
    Aurass, Philipp
    et al.
    Department of Enteropathogenic Bacteria and Legionella, Robert Koch Institute, Wernigerode, Germany; Department of Molecular Biology and Microbiology, Tufts University School of Medicine, MA, Boston, United States.
    Kim, Seongok
    Department of Molecular Biology and Microbiology, Tufts University School of Medicine, MA, Boston, United States.
    Pinedo, Victor
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Isberg, Ralph R.
    Department of Molecular Biology and Microbiology, Tufts University School of Medicine, MA, Boston, United States.
    Identification of genes required for long-term survival of Legionella Pneumophila in water2023Inngår i: mSphere, E-ISSN 2379-5042, Vol. 8, nr 2, artikkel-id e0045422Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Long-term survival of Legionella pneumophila in aquatic environments is thought to be important for facilitating epidemic outbreaks. Eliminating bacterial colonization in plumbing systems is the primary strategy that depletes this reservoir and prevents disease. To uncover L. pneumophila determinants facilitating survival in water, a Tn-seq strategy was used to identify survival-defective mutants during 50-day starvation in tap water at 42°C. The mutants with the most drastic survival defects carried insertions in electron transport chain genes, indicating that membrane energy charge and/or ATP synthesis requires the generation of a proton gradient by the respiratory chain to maintain survival in the presence of water stress. In addition, periplasmically localized proteins that are known (EnhC) or hypothesized (lpg1697) to stabilize the cell wall against turnover were essential for water survival. To test that the identified mutations disrupted water survival, candidate genes were knocked down by CRISPRi. The vast majority of knockdown strains with verified transcript depletion showed remarkably low viability after 50-day incubations. To demonstrate that maintenance of cell wall integrity was an important survival determinant, a deletion mutation in lpg1697, in a gene encoding a predicted l,d-transpeptidase domain, was analyzed. The loss of this gene resulted in increased osmolar sensitivity and carbenicillin hypersensitivity relative to the wild type, as predicted for loss of an l,d-transpeptidase. These results indicate that the L. pneumophila envelope has been evolutionarily selected to allow survival under conditions in which the bacteria are subjected to long-term exposure to starvation and low osmolar conditions. IMPORTANCE Water is the primary vector for transmission of L. pneumophila to humans, and the pathogen is adapted to persist in this environment for extended periods of time. Preventing survival of L. pneumophila in water is therefore critical for prevention of Legionnaires' disease. We analyzed dense transposon mutation pools for strains with severe survival defects during a 50-day water incubation at 42°C. By tracking the associated transposon insertion sites in the genome, we defined a distinct essential gene set for water survival and demonstrate that a predicted peptidoglycan cross-linking enzyme, lpg1697, and components of the electron transport chain are required to ensure survival of the pathogen. Our results indicate that select characteristics of the cell wall and components of the respiratory chain of L. pneumophila are primary evolutionary targets being shaped to promote its survival in water.

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  • 50.
    Avican, Kemal
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Persistent infection by Yersinia pseudotuberculosis2015Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Enteropathogenic Yersinia species can infect many mammalian organs such as the small intestine, cecum, Peyer’s patches, liver, spleen, and lung and cause diseases that resemble a typhoid-like syndrome, as seen for other enteropathogens. We found that sublethal infection doses of Y. pseudotuberculosis gave rise to asymptomatic persistent infection in mice and identified the cecal lymphoid follicles as the primary site for colonization during persistence. Persistent Y. pseudotuberculosis is localized in the dome area, often in inflammatory lesions, as foci or as single cells, and also in neutrophil exudates in the cecal lumen. This new mouse model for bacterial persistence in cecum has potential as an investigative tool for deeper understanding of bacterial adaptation and host immune defense mechanisms during persistent infection. Here, we investigated the nature of the persistent infection established by Y. pseudotuberculosis in mouse cecal tissue using in vivo RNA-seq of bacteria during early and persistent stages of infection. Comparative analysis of the bacterial transcriptomes revealed that Y. pseudotuberculosis undergoes transcriptional reprogramming with drastic down-regulation of T3SS virulence genes during persistence in the cecum. At the persistent stage, the expression pattern in many respects resembles the pattern seen in vitro at 26°C. Genes that are up-regulated during persistence are genes involved in anaerobiosis, chemotaxis, and protection against oxidative and acidic stress, which indicates the influence of different environmental cues. We found that the Crp/CsrA/RovA regulatory cascades influence the pattern of bacterial gene expression during persistence. Furthermore, we show that ArcA, Fnr, FrdA, WrbA, RovA, and RfaH play critical roles in persistence. An extended investigation of the transcriptional regulator rfaH employing mouse infection studies, phenotypic characterizations, and RNA-seq transcriptomics analyses indicated that this gene product contributes to establishment of infection and confirmed that it regulates O-antigen biosynthesis genes in Y. pseudotuberculosis. The RNA-seq results also suggest that rfaH has a relatively global effect. Furthermore, we also found that the dynamics of the cecal tissue organization and microbial composition shows changes during different stages of the infection. Taken together, based on our findings, we speculate that this enteropathogen initiates infection by using its virulence factors in meeting the innate immune response in the cecal tissue. Later on, these factors lead to dysbiosis in the local microbiota and altered tissue organization. At later stages of the infection, the pathogen adapts to the environment in the cecum by reprogramming its transcriptome from a highly virulent mode to a more environmentally adaptable mode for survival and shedding. The in vivo transcriptomic analyses for essential genes during infections present strong candidates for novel targets for antimicrobials.

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