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  1. Article ; Online: A Simple Luminescent Adenylate-Cyclase Functional Assay for Evaluation of Bacillus anthracis Edema Factor Activity

    Ma’ayan Israeli / Shahar Rotem / Uri Elia / Erez Bar-Haim / Ofer Cohen / Theodor Chitlaru

    Toxins, Vol 8, Iss 8, p

    2016  Volume 243

    Abstract: Edema Factor (EF), the toxic sub-unit of the Bacillus anthracis Edema Toxin (ET) is a calmodulin-dependent adenylate cyclase whose detrimental activity in the infected host results in severe edema. EF is therefore a major virulence factor of B. anthracis. ...

    Abstract Edema Factor (EF), the toxic sub-unit of the Bacillus anthracis Edema Toxin (ET) is a calmodulin-dependent adenylate cyclase whose detrimental activity in the infected host results in severe edema. EF is therefore a major virulence factor of B. anthracis. We describe a simple, rapid and reliable functional adenylate-cyclase assay based on inhibition of a luciferase-mediated luminescence reaction. The assay exploits the efficient adenylate cyclase-mediated depletion of adenosine tri-phosphate (ATP), and the strict dependence on ATP of the light-emitting luciferase-catalyzed luciferin-conversion to oxyluciferin, which can be easily visualized. The assay exhibits a robust EF-dose response decrease in luminescence, which may be specifically reverted by anti-EF antibodies. The application of the assay is exemplified in: (a) determining the presence of EF in B. anthracis cultures, or its absence in cultures of EF-defective strains; (b) evaluating the anti-EF humoral response in experimental animals infected/vaccinated with B. anthracis; and (c) rapid discrimination between EF producing and non-producing bacterial colonies. Furthermore, the assay may be amenable with high-throughput screening for EF inhibitory molecules.
    Keywords Bacillus anthracis ; Edema Factor ; adenylate cyclase ; luminescent assay ; luciferase ; ATP-depletion ; Medicine ; R
    Subject code 570
    Language English
    Publishing date 2016-08-01T00:00:00Z
    Publisher MDPI AG
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  2. Article ; Online: Comparative Analysis of the Global Transcriptomic Response to Oxidative Stress of Bacillus anthracis htrA -Disrupted and Parental Wild Type Strains

    Galia Zaide / Uri Elia / Inbar Cohen-Gihon / Ma’ayan Israeli / Shahar Rotem / Ofir Israeli / Sharon Ehrlich / Hila Cohen / Shirley Lazar / Adi Beth-Din / Avigdor Shafferman / Anat Zvi / Ofer Cohen / Theodor Chitlaru

    Microorganisms, Vol 8, Iss 1896, p

    2020  Volume 1896

    Abstract: We previously demonstrated that the HtrA (High Temperature Requirement A) protease/chaperone active in the quality control of protein synthesis, represents an important virulence determinant of Bacillus anthracis . Virulence attenuation of htrA - ... ...

    Abstract We previously demonstrated that the HtrA (High Temperature Requirement A) protease/chaperone active in the quality control of protein synthesis, represents an important virulence determinant of Bacillus anthracis . Virulence attenuation of htrA -disrupted Bacillus anthracis strains was attributed to susceptibility of Δ htrA strains to stress insults, as evidenced by affected growth under various stress conditions. Here, we report a comparative RNA-seq transcriptomic study generating a database of differentially expressed genes in the B. anthracis htrA -disrupted and wild type parental strains under oxidative stress. The study demonstrates that, apart from protease and chaperone activities, HtrA exerts a regulatory role influencing expression of more than 1000 genes under stress. Functional analysis of groups or individual genes exhibiting strain-specific modulation, evidenced (i) massive downregulation in the Δ htrA and upregulation in the WT strains of various transcriptional regulators, (ii) downregulation of translation processes in the WT strain, and (iii) downregulation of metal ion binding functions and upregulation of sporulation-associated functions in the Δ htrA strain. These modulated functions are extensively discussed. Fifteen genes uniquely upregulated in the wild type strain were further interrogated for their modulation in response to other stress regimens. Overexpression of one of these genes, encoding for MazG (a nucleoside triphosphate pyrophosphohydrolase involved in various stress responses in other bacteria), in the Δ htrA strain resulted in partial alleviation of the H 2 O 2 -sensitive phenotype.
    Keywords Bacillus anthracis ; anthrax ; oxidative stress ; HtrA ; transcriptomics ; RNA-seq ; Biology (General) ; QH301-705.5
    Subject code 570
    Language English
    Publishing date 2020-11-01T00:00:00Z
    Publisher MDPI AG
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  3. Article ; Online: Implementation of Adenovirus-Mediated Pulmonary Expression of Human ACE2 in HLA Transgenic Mice Enables Establishment of a COVID-19 Murine Model for Assessment of Immune Responses to SARS-CoV-2 Infection

    Theodor Chitlaru / Erez Bar-Haim / Liat Bar-On / Shahar Rotem / Hila Cohen / Uri Elia / David Gur / Moshe Aftalion / Ron Alkalay / Efi Makdasi / Yentl Evgy / Reut Falach / Ma’ayan Israeli / Adi Bercovich-Kinori / Hagit Achdout / Yfat Yahalom-Ronen / Ronit Rosenfeld / Ofer Cohen

    Pathogens, Vol 10, Iss 940, p

    2021  Volume 940

    Abstract: HLA transgenic mice are instrumental for evaluation of human-specific immune responses to viral infection. Mice do not develop COVID-19 upon infection with SARS-CoV-2 due to the strict tropism of the virus to the human ACE2 receptor. The aim of the ... ...

    Abstract HLA transgenic mice are instrumental for evaluation of human-specific immune responses to viral infection. Mice do not develop COVID-19 upon infection with SARS-CoV-2 due to the strict tropism of the virus to the human ACE2 receptor. The aim of the current study was the implementation of an adenovirus-mediated infection protocol for human ACE2 expression in HLA transgenic mice. Transient pulmonary expression of the human ACE2 receptor in these mice results in their sensitisation to SARS-CoV-2 infection, consequently providing a valuable animal model for COVID-19. Infection results in a transient loss in body weight starting 3 days post-infection, reaching 20–30% loss of weight at day 7 and full recovery at days 11–13 post-infection. The evolution of the disease revealed high reproducibility and very low variability among individual mice. The method was implemented in two different strains of HLA immunized mice. Infected animals developed strong protective humoral and cellular immune responses specific to the viral spike-protein, strictly depending on the adenovirus-mediated human ACE2 expression. Convalescent animals were protected against a subsequent re-infection with SARS-CoV-2, demonstrating that the model may be applied for assessment of efficacy of anti-viral immune responses.
    Keywords adenovirus vector ; human ACE2 ; SARS-CoV-2 ; COVID-19 ; HLA transgenic mice ; Medicine ; R
    Language English
    Publishing date 2021-07-01T00:00:00Z
    Publisher MDPI AG
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  4. Article ; Online: Genome-wide CRISPR screens identify GATA6 as a proviral host factor for SARS-CoV-2 via modulation of ACE2

    Ma’ayan Israeli / Yaara Finkel / Yfat Yahalom-Ronen / Nir Paran / Theodor Chitlaru / Ofir Israeli / Inbar Cohen-Gihon / Moshe Aftalion / Reut Falach / Shahar Rotem / Uri Elia / Ital Nemet / Limor Kliker / Michal Mandelboim / Adi Beth-Din / Tomer Israely / Ofer Cohen / Noam Stern-Ginossar / Adi Bercovich-Kinori

    Nature Communications, Vol 13, Iss 1, Pp 1-

    2022  Volume 16

    Abstract: Here, Israeli and Finkel et al. perform genome-wide CRISPR knockout screens to identify host factors required for the infection with SARS-CoV-2 and two additionally variants of concern, Alpha and Beta, unveiling shared and differential host pathways ... ...

    Abstract Here, Israeli and Finkel et al. perform genome-wide CRISPR knockout screens to identify host factors required for the infection with SARS-CoV-2 and two additionally variants of concern, Alpha and Beta, unveiling shared and differential host pathways required by the variants, and demonstrate GATA6 is critical for SARS-CoV-2 viral entry through modulation of ACE2 expression.
    Keywords Science ; Q
    Language English
    Publishing date 2022-04-01T00:00:00Z
    Publisher Nature Portfolio
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  5. Article: A novel live attenuated anthrax spore vaccine based on an acapsular Bacillus anthracis Sterne strain with mutations in the htrA, lef and cya genes

    Chitlaru, Theodor / Avigdor Shafferman / Erez Bar-Haim / Ma'ayan Israeli / Ofer Cohen / Shahar Rotem / Sharon Ehrlich / Uri Elia

    Vaccine. 2017,

    2017  

    Abstract: We recently reported the development of a novel, next-generation, live attenuated anthrax spore vaccine based on disruption of the htrA (High Temperature Requirement A) gene in the Bacillus anthracis Sterne veterinary vaccine strain. This vaccine ... ...

    Abstract We recently reported the development of a novel, next-generation, live attenuated anthrax spore vaccine based on disruption of the htrA (High Temperature Requirement A) gene in the Bacillus anthracis Sterne veterinary vaccine strain. This vaccine exhibited a highly significant decrease in virulence in murine, guinea pig and rabbit animal models yet preserved the protective value of the parental Sterne strain. Here, we report the evaluation of additional mutations in the lef and cya genes, encoding for the toxin components lethal factor (LF) and edema factor (EF), to further attenuate the SterneΔhtrA strain and improve its compatibility for human use. Accordingly, we constructed seven B. anthracis Sterne-derived strains exhibiting different combinations of mutations in the htrA, cya and lef genes. The various strains were indistinguishable in growth in vitro and in their ability to synthesise the protective antigen (PA, necessary for the elicitation of protection). In the sensitive murine model, we observed a gradual increase (ΔhtrA<ΔhtrAΔcya<ΔhtrAΔlef<ΔhtrAΔlefΔcya) in attenuation – up to 108-fold relative to the parental Sterne vaccine strain. Most importantly, all various SterneΔhtrA derivative strains did not differ in their ability to elicit protective immunity in guinea pigs. Immunisation of guinea pigs with a single dose (109 spores) or double doses (>107spores) of the most attenuated triple mutant strain SterneΔhtrAlefMUTΔcya induced a robust immune response, providing complete protection against a subsequent respiratory lethal challenge. Partial protection was observed in animals vaccinated with a double dose of as few as 105spores. Furthermore, protective immune status was maintained in all vaccinated guinea pigs and rabbits for at least 40 and 30weeks, respectively.
    Keywords animal models ; anthrax ; anthrax toxin ; Bacillus anthracis ; genes ; guinea pigs ; humans ; immune response ; immunization ; mice ; mutants ; mutation ; rabbits ; spores ; temperature ; vaccines ; virulence
    Language English
    Size p. .
    Publishing place Elsevier Ltd
    Document type Article
    Note Pre-press version
    ZDB-ID 605674-x
    ISSN 1873-2518 ; 0264-410X
    ISSN (online) 1873-2518
    ISSN 0264-410X
    DOI 10.1016/j.vaccine.2017.03.033
    Database NAL-Catalogue (AGRICOLA)

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  6. Article ; Online: Early Diagnosis of Pathogen Infection by Cell-Based Activation Immunoassay

    Erez Bar-Haim / Shahar Rotem / Uri Elia / Adi Bercovich-Kinori / Ma’ayan Israeli / Inbar Cohen-Gihon / Ofir Israeli / Noam Erez / Hagit Achdout / Ayelet Zauberman / Moshe Aftalion / Emanuelle Mamroud / Theodor Chitlaru / Ofer Cohen

    Cells, Vol 8, Iss 9, p

    2019  Volume 952

    Abstract: Diagnostic identification of pathogens is usually accomplished by isolation of the pathogen or its substances, and should correlate with the time and site of infection. Alternatively, immunoassays such as enzyme-linked immunosorbent assay (ELISA) tests ... ...

    Abstract Diagnostic identification of pathogens is usually accomplished by isolation of the pathogen or its substances, and should correlate with the time and site of infection. Alternatively, immunoassays such as enzyme-linked immunosorbent assay (ELISA) tests for quantification of serum antibodies are expedient and are usually employed for retrospective diagnostic of a particular infective agent. Here, the potential of cell-based immunoassays for early pathogen detection was evaluated by quantification of specific, antigen-activated, low-frequency IFNγ-secreting cells in mouse spleens following infection with various pathogens. Using enzyme-linked immunospot (ELISPOT) assays, specific responses were observed within 3−6 days following infection with F. tularensis , B. anthracis , Y. pestis , or Influenza virus. Blood samples collected from F. tularensis -infected mice revealed the presence of IFNγ-producing activated cells within one week post infection. When non-human primates were infected with B. anthracis , cellular response was observed in peripheral blood samples as early as five days post infection, 3−5 days earlier than serum antibodies. Finally, the expression pattern of genes in splenocytes of F. tularensis -infected mice was inspected by a transcriptomic approach, enabling the identification of potential host targets for the future development of genetic-based cellular immunoassays. Altogether, the data demonstrate the potential of cell-based immunoassays for early pathogen detection.
    Keywords Early detection ; Francisella tularensis ; Bacillus anthracis ; Yersinia pestis ; Influenza virus ; ELISPOT ; lymphocyte activation ; infection biomarkers ; lymphocyte transcriptomics ; animal models ; Biology (General) ; QH301-705.5
    Language English
    Publishing date 2019-08-01T00:00:00Z
    Publisher MDPI AG
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  7. Article ; Online: A panel of human neutralizing mAbs targeting SARS-CoV-2 spike at multiple epitopes

    Tal Noy-Porat / Efi Makdasi / Ron Alcalay / Adva Mechaly / Yinon Levy / Adi Bercovich-Kinori / Ayelet Zauberman / Hadas Tamir / Yfat Yahalom-Ronen / Ma’ayan Israeli / Eyal Epstein / Hagit Achdout / Sharon Melamed / Theodor Chitlaru / Shay Weiss / Eldar Peretz / Osnat Rosen / Nir Paran / Shmuel Yitzhaki /
    Shmuel C. Shapira / Tomer Israely / Ohad Mazor / Ronit Rosenfeld

    Nature Communications, Vol 11, Iss 1, Pp 1-

    2020  Volume 7

    Abstract: Here, Noy-Porat, Makdasi et al. report the isolation of a panel of neutralizing mAbs selected against SARS-CoV-2 receptor-binding domain (RBD) from a phage display library constructed based on patient samples collected in the acute phase of the disease, ... ...

    Abstract Here, Noy-Porat, Makdasi et al. report the isolation of a panel of neutralizing mAbs selected against SARS-CoV-2 receptor-binding domain (RBD) from a phage display library constructed based on patient samples collected in the acute phase of the disease, which show efficient neutralizing activities against authentic virus in vitro.
    Keywords Science ; Q
    Language English
    Publishing date 2020-08-01T00:00:00Z
    Publisher Nature Portfolio
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  8. Article ; Online: A panel of human neutralizing mAbs targeting SARS-CoV-2 spike at multiple epitopes

    Tal Noy-Porat / Efi Makdasi / Ron Alcalay / Adva Mechaly / Yinon Levy / Adi Bercovich-Kinori / Ayelet Zauberman / Hadas Tamir / Yfat Yahalom-Ronen / Ma’ayan Israeli / Eyal Epstein / Hagit Achdout / Sharon Melamed / Theodor Chitlaru / Shay Weiss / Eldar Peretz / Osnat Rosen / Nir Paran / Shmuel Yitzhaki /
    Shmuel C. Shapira / Tomer Israely / Ohad Mazor / Ronit Rosenfeld

    Nature Communications, Vol 11, Iss 1, Pp 1-

    2020  Volume 7

    Abstract: Here, Noy-Porat, Makdasi et al. report the isolation of a panel of neutralizing mAbs selected against SARS-CoV-2 receptor-binding domain (RBD) from a phage display library constructed based on patient samples collected in the acute phase of the disease, ... ...

    Abstract Here, Noy-Porat, Makdasi et al. report the isolation of a panel of neutralizing mAbs selected against SARS-CoV-2 receptor-binding domain (RBD) from a phage display library constructed based on patient samples collected in the acute phase of the disease, which show efficient neutralizing activities against authentic virus in vitro.
    Keywords Science ; Q ; covid19
    Language English
    Publishing date 2020-08-01T00:00:00Z
    Publisher Nature Publishing Group
    Document type Article ; Online
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