93 results on '"Trimpert, J."'
Search Results
2. Interspecies analysis to dissect cellular transcriptomic signatures of humans and hamsters in COVID-19
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Pennitz, P, additional, Friedrich, V, additional, Wyler, E, additional, Adler, J, additional, Postmus, D, additional, Andreotti, S, additional, Teixeira-Alves, L, additional, Kazmierski, J, additional, Pott, F, additional, Hoefler, T, additional, Goekeri, C, additional, Landthaler, M, additional, Goffinet, C, additional, Trimpert, J, additional, Scholz, M, additional, Witzenrath, M, additional, Kirsten, H, additional, and Nouailles, G, additional
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- 2023
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3. De novo whole genome assembly of the Roborovski dwarf hamster (Phodopus roborovskii) genome, an animal model for severe/critical COVID-19
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Andreotti, S., Altmüller, J., Quedenau, C., Borodina, T., Nouailles, G., Teixeira Alves, L.G., Landthaler, M., Bieniara, M., Trimpert, J., and Wyler, E.
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Cancer Research ,Technology Platforms - Abstract
The Roborovski dwarf hamster Phodopus roborovskii belongs to the Phodopus genus, one of seven within Cricetinae subfamily. Like other rodents such as mice, rats or ferrets, hamsters can be important animal models for a range of diseases. Whereas the Syrian hamster from the genus Mesocricetus is now widely used as a model for mild to moderate COVID-19, Roborovski dwarf hamster show a severe to lethal course of disease upon infection with the novel human coronavirus SARS-CoV-2.
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- 2022
4. Key benefits of dexamethasone and antibody treatment in COVID-19 hamster models revealed by single cell transcriptomics
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Wyler, E., Adler, J.M., Eschke, K., Teixeira Alves, G., Peidli, S., Pott, F., Kazmierski, J., Michalick, L., Kershaw, O., Bushe, J., Andreotti, S., Pennitz, P., Abdelgawad, A., Postmus, D., Goffinet, C., Kreye, J., Reincke, S.M., Prüss, H., Blüthgen, N., Gruber, A.D., Kuebler, W.M., Witzenrath, M., Landthaler, M., Nouailles, G., and Trimpert, J.
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Cancer Research - Abstract
For COVID-19, effective and well-understood treatment options are still scarce. Since vaccine efficacy is challenged by novel variants, short-lasting immunity and vaccine hesitancy, understanding and optimizing therapeutic options remains essential. We aimed at better understanding the effects of two standard-of-care drugs, dexamethasone and anti-SARS-CoV-2 antibodies, on infection and host responses. By using two COVID-19 hamster models, pulmonary immune responses were analyzed to characterize effects of single or combinatorial treatments. Pulmonary viral burden was reduced by anti-SARS-CoV-2 antibody treatment, and similar or increased by dexamethasone alone. Dexamethasone exhibited strong anti-inflammatory effects and prevented fulminant disease in a severe disease model. Combination therapy showed additive benefits with both anti-viral and anti-inflammatory potency. Bulk and single-cell transcriptomic analyses confirmed dampened inflammatory cell recruitment into lungs upon dexamethasone treatment, and identified a specifically responsive subpopulation of neutrophils, thereby indicating a potential mechanism of action. Our analyses confirm the anti-inflammatory properties of dexamethasone and suggest possible mechanisms, validate anti-viral effects of anti-SARS-CoV-2 antibody treatment, and reveal synergistic effects of a combination therapy, thus informing more effective COVID-19 therapies.
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- 2022
5. Integrational approaches for cross-species analysis of lung pathologies at single-cell resolution
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Pennitz, P, additional, Kirsten, H, additional, Wyler, E, additional, Goekeri, C, additional, Obermayer, B, additional, Heinz, G A, additional, Mashreghi, M F, additional, Trimpert, J, additional, Landthaler, M, additional, Suttorp, N, additional, Hocke, A C, additional, Hippenstiel, S, additional, Tönnies, M, additional, Scholz, M, additional, Kuebler, W M, additional, Witzenrath, M, additional, Hoenzke, K, additional, and Nouailles, G, additional
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- 2022
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6. A pulmonologist's guide to perform and analyse cross-species single lung cell transcriptomics
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Pennitz, P., Kirsten, H., Friedrich, V.D., Wyler, E., Goekeri, C., Obermayer, B., Heinz, G.A., Mashreghi, M.F., Büttner, M., Trimpert, J., Landthaler, M., Suttorp, N., Hocke, A.C., Hippenstiel, S., Tönnies, M., Scholz, M., Kuebler, W.M., Witzenrath, M., Hoenzke, K., and Nouailles, G.
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Single lung cell transcriptomics ,Cancer Research ,Single-cell RNA sequencing (scRNA-seq) ,Base Sequence ,Swine ,Rats ,Mice ,Pulmonologists ,Species Specificity ,Cricetinae ,Chlorocebus aethiops ,Pulmonologist’s guide ,Animals ,Humans ,ddc:610 ,Technology Platforms ,Transcriptome ,Lung ,600 Technik, Medizin, angewandte Wissenschaften::610 Medizin und Gesundheit::610 Medizin und Gesundheit - Abstract
Single-cell ribonucleic acid sequencing is becoming widely employed to study biological processes at a novel resolution depth. The ability to analyse transcriptomes of multiple heterogeneous cell types in parallel is especially valuable for cell-focused lung research where a variety of resident and recruited cells are essential for maintaining organ functionality. We compared the single-cell transcriptomes from publicly available and unpublished datasets of the lungs in six different species: human (Homo sapiens), African green monkey (Chlorocebus sabaeus), pig (Sus domesticus), hamster (Mesocricetus auratus), rat (Rattus norvegicus) and mouse (Mus musculus) by employing RNA velocity and intercellular communication based on ligand-receptor co-expression, among other techniques. Specifically, we demonstrated a workflow for interspecies data integration, applied a single unified gene nomenclature, performed cell-specific clustering and identified marker genes for each species. Overall, integrative approaches combining newly sequenced as well as publicly available datasets could help identify species-specific transcriptomic signatures in both healthy and diseased lung tissue and select appropriate models for future respiratory research.
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- 2022
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7. Virus-induced senescence is driver and therapeutic target in COVID-19
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Lee, S., Yu, Y., Trimpert, J., Benthani, F., Mairhofer, M., Richter-Pechanska, P., Wyler, E., Belenki, D., Kaltenbrunner, S., Pammer, M., Kausche, L., Firsching, T.C., Dietert, K., Schotsaert, M., Martínez-Romero, C., Singh, G., Kunz, S., Niemeyer, D., Ghanem, R., Salzer, H.J.F., Paar, C., Mülleder, M., Uccellini, M., Michaelis, E.G., Khan, A., Lau, A., Schönlein, M., Habringer, A., Tomasits, J., Adler, J.M., Kimeswenger, S., Gruber, A.D., Hoetzenecker, W., Steinkellner, H., Purfuerst, B., Motz, R., Di Pierro, F., Lamprecht, B., Osterrieder, N., Landthaler, M., Drosten, C., García-Sastre, A., Langer, R., Ralser, M., Eils, R., Reimann, M., Fan, D.N.Y., and Schmitt, C.A.
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Cancer Research ,Technology Platforms - Abstract
Derailed cytokine and immune cell networks account for organ damage and clinical severity of COVID-19. Here we show that SARS-CoV-2, like other viruses, evokes cellular senescence as a primary stress response in infected cells. Virus-induced senescence (VIS) is indistinguishable from other forms of cellular senescence and accompanied by a senescence-associated secretory phenotype (SASP), composed of pro-inflammatory cytokines, extracellular matrix-active factors and pro-coagulatory mediators. COVID-19 patients displayed markers of senescence in their airway mucosa in situ and elevated serum levels of SASP factors. Mirroring COVID-19 hallmark features such as macrophage and neutrophil infiltration, endothelial damage and widespread thrombosis in affected lung tissue in vitro assays demonstrated macrophage activation with SASP-reminiscent secretion, complement lysis and SASP-amplifying secondary senescence of endothelial cells, neutrophil extracellular trap (NET) formation as well as activation of platelets and the clotting cascade in response to supernatant of VIS cells, including SARS-CoV-2-induced senescence. Senolytics such as Navitoclax and Dasatinib/Quercetin selectively eliminated VIS cells, mitigated COVID-19-reminiscent lung disease and reduced inflammation in SARS-CoV-2-driven hamster and mouse models. Our findings mark VIS as pathogenic trigger of COVID-19-related cytokine escalation and organ damage, and suggest senolytic targeting of virus-infected cells as a novel treatment option against SARS-CoV-2 and perhaps other viral infections.
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- 2021
8. Epithelial response to IFN-γ promotes SARS-CoV-2 infection
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Heuberger, J., Trimpert, J., Vladimirova, D., Goosmann, C., Lin, M., Schmuck, R., Mollenkopf, H.J., Brinkmann, V., Tacke, F., Osterrieder, N., and Sigal, M.
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Cancer Research ,hormones, hormone substitutes, and hormone antagonists - Abstract
SARS-CoV-2, the agent that causes COVID-19, invades epithelial cells, including those of the respiratory and gastrointestinal mucosa, using angiotensin-converting enzyme-2 (ACE2) as a receptor. Subsequent inflammation can promote rapid virus clearance, but severe cases of COVID-19 are characterized by an inefficient immune response that fails to clear the infection. Using primary epithelial organoids from human colon, we explored how the central antiviral mediator IFN-γ, which is elevated in COVID-19, affects epithelial cell differentiation, ACE2 expression, and susceptibility to infection with SARS-CoV-2. In mouse and human colon, ACE2 is mainly expressed by surface enterocytes. Inducing enterocyte differentiation in organoid culture resulted in increased ACE2 production. IFN-γ treatment promoted differentiation into mature KRT20+ enterocytes expressing high levels of ACE2, increased susceptibility to SARS-CoV-2 infection and resulted in enhanced virus production in infected cells. Similarly, infection-induced epithelial interferon signaling promoted enterocyte maturation and enhanced ACE2 expression. We here reveal a mechanism by which IFN-γ-driven inflammatory responses induce a vulnerable epithelial state with robust replication of SARS-CoV-2, which may have an impact on disease outcome and virus transmission.
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- 2021
9. Epithelial response to IFN-c promotes SARS-CoV-2 infection
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Heuberger, J., Trimpert, J., Vladimirova, D., Goosmann, C., Lin, M., Schmuck, R., Mollenkopf, H., Brinkmann, V., Tacke, F., Osterrieder, N., and Sigal, M.
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- 2021
10. ICTV Virus Taxonomy Profile: Herpesviridae 2021
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Gatherer, D, Depledge, DP, Hartley, CA, Szpara, ML, Vaz, PK, Benko, M, Brandt, CR, Bryant, NA, Dastjerdi, A, Doszpoly, A, Gompels, UA, Inoue, N, Jarosinski, KW, Kaul, R, Lacoste, V, Norberg, P, Origgi, FC, Orton, RJ, Pellett, PE, Schmid, DS, Spatz, SJ, Stewart, JP, Trimpert, J, Waltzek, TB, Davison, AJ, Gatherer, D, Depledge, DP, Hartley, CA, Szpara, ML, Vaz, PK, Benko, M, Brandt, CR, Bryant, NA, Dastjerdi, A, Doszpoly, A, Gompels, UA, Inoue, N, Jarosinski, KW, Kaul, R, Lacoste, V, Norberg, P, Origgi, FC, Orton, RJ, Pellett, PE, Schmid, DS, Spatz, SJ, Stewart, JP, Trimpert, J, Waltzek, TB, and Davison, AJ
- Abstract
Members of the family Herpesviridae have enveloped, spherical virions with characteristic complex structures consisting of symmetrical and non-symmetrical components. The linear, double-stranded DNA genomes of 125-241 kbp contain 70-170 genes, of which 43 have been inherited from an ancestral herpesvirus. In general, herpesviruses have coevolved with and are highly adapted to their hosts, which comprise many mammalian, avian and reptilian species. Following primary infection, they are able to establish lifelong latent infection, during which there is limited viral gene expression. Severe disease is usually observed only in the foetus, the very young, the immunocompromised or following infection of an alternative host. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Herpesviridae, which is available at ictv.global/report/herpesviridae.
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- 2021
11. Interspecies analysis to dissect transcriptomic signatures of humans and hamsters in CVID19.
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Pennitz, P, Friedrich, V, Wyler, E, Adler, J, Postmus, D, Andreotti, S, Teixeira Alves, L, Kazmierski, J, Pott, F, Hoefler, T, Goekeri, C, Landthaler, M, Goffinet, C, Trimpert, J, Scholz, M, Witzenrath, M, Kirsten, H, and Nouailles, G
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- 2024
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12. Fatal Elephant Endotheliotropic Herpesvirus Infection of Two Young Asian Elephants
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Pavulraj, S, Eschke, K, Prahl, A, Flugger, M, Trimpert, J, van den Doel, Petra, Andreotti, S, Kaessmeyer, S, Osterrieder, N, Azab, W, Pavulraj, S, Eschke, K, Prahl, A, Flugger, M, Trimpert, J, van den Doel, Petra, Andreotti, S, Kaessmeyer, S, Osterrieder, N, and Azab, W
- Published
- 2019
13. Vergleichende Räumliche Proteomanalysen SARS-CoV-2-infizierter lungen von Goldhamstern und Roborowski Zwerghamstern
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Kunder, S., Voß, A., Hempel, B.-F., Trimpert, J., Dietert, K., and Gruber, A. D.
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- 2024
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14. The FDA-Approved Drug Cobicistat Synergizes with Remdesivir to Inhibit SARS-CoV-2 Replication in Vitro and Decreases Viral Titers and Disease Progression in Syrian Hamsters
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Iart Luca Shytaj, Mohamed Fares, Lara Gallucci, Bojana Lucic, Mahmoud M. Tolba, Liv Zimmermann, Julia M. Adler, Na Xing, Judith Bushe, Achim D. Gruber, Ina Ambiel, Ahmed Taha Ayoub, Mirko Cortese, Christopher J. Neufeldt, Bettina Stolp, Mohamed Hossam Sobhy, Moustafa Fathy, Min Zhao, Vibor Laketa, Ricardo Sobhie Diaz, Richard E. Sutton, Petr Chlanda, Steeve Boulant, Ralf Bartenschlager, Megan L. Stanifer, Oliver T. Fackler, Jakob Trimpert, Andrea Savarino, Marina Lusic, Shytaj, I. L., Fares, M., Gallucci, L., Lucic, B., Tolba, M. M., Zimmermann, L., Adler, J. M., Xing, N., Bushe, J., Gruber, A. D., Ambiel, I., Ayoub, A. T., Cortese, M., Neufeldt, C. J., Stolp, B., Sobhy, M. H., Fathy, M., Zhao, M., Laketa, V., Diaz, R. S., Sutton, R. E., Chlanda, P., Boulant, S., Bartenschlager, R., Stanifer, M. L., Fackler, O. T., Trimpert, J., Savarino, A., and Lusic, M.
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direct-acting antiviral ,Mesocricetus ,drug repurposing ,SARS-CoV-2 ,COVID-19 ,remdesivir ,Alanine/analogs & derivatives ,Antiviral Agents/pharmacology ,Hepatitis C, Chronic ,Viral Load ,cobicistat ,spike protein ,COVID-19/drug therapy ,Microbiology ,Adenosine Monophosphate/analogs & derivatives ,Cricetinae ,Virology ,Disease Progression ,Animals ,Humans ,Cobicistat ,Pandemics - Abstract
The lack of effective antiviral treatments against SARS-CoV-2 is a significant limitation in the fight against the COVID-19 pandemic. Single-drug regimens have so far yielded limited results, indicating that combinations of antivirals might be required, as previously seen for other RNA viruses.Combinations of direct-acting antivirals are needed to minimize drug resistance mutations and stably suppress replication of RNA viruses. Currently, there are limited therapeutic options against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and testing of a number of drug regimens has led to conflicting results. Here, we show that cobicistat, which is an FDA-approved drug booster that blocks the activity of the drug-metabolizing proteins cytochrome P450-3As (CYP3As) and P-glycoprotein (P-gp), inhibits SARS-CoV-2 replication. Two independent cell-to-cell membrane fusion assays showed that the antiviral effect of cobicistat is exerted through inhibition of spike protein-mediated membrane fusion. In line with this, incubation with low-micromolar concentrations of cobicistat decreased viral replication in three different cell lines including cells of lung and gut origin. When cobicistat was used in combination with remdesivir, a synergistic effect on the inhibition of viral replication was observed in cell lines and in a primary human colon organoid. This was consistent with the effects of cobicistat on two of its known targets, CYP3A4 and P-gp, the silencing of which boosted the in vitro antiviral activity of remdesivir in a cobicistat-like manner. When administered in vivo to Syrian hamsters at a high dose, cobicistat decreased viral load and mitigated clinical progression. These data highlight cobicistat as a therapeutic candidate for treating SARS-CoV-2 infection and as a potential building block of combination therapies for COVID-19. IMPORTANCE The lack of effective antiviral treatments against SARS-CoV-2 is a significant limitation in the fight against the COVID-19 pandemic. Single-drug regimens have so far yielded limited results, indicating that combinations of antivirals might be required, as previously seen for other RNA viruses. Our work introduces the drug booster cobicistat, which is approved by the FDA and typically used to potentiate the effect of anti-HIV protease inhibitors, as a candidate inhibitor of SARS-CoV-2 replication. Beyond its direct activity as an antiviral, we show that cobicistat can enhance the effect of remdesivir, which was one of the first drugs proposed for treatment of SARS-CoV-2. Overall, the dual action of cobicistat as a direct antiviral and a drug booster can provide a new approach to design combination therapies and rescue the activity of compounds that are only partially effective in monotherapy.
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- 2022
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15. A biomathematical model of SARS-CoV-2 in Syrian hamsters.
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Schirm S, Nouailles G, Kirsten H, Trimpert J, Wyler E, Teixeira Alves LG, Landthaler M, Ahnert P, Suttorp N, Witzenrath M, and Scholz M
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- Animals, Cricetinae, Antibodies, Neutralizing immunology, Humans, Antibodies, Viral immunology, Lung virology, Lung immunology, Lung pathology, CD8-Positive T-Lymphocytes immunology, Cytokines metabolism, CD4-Positive T-Lymphocytes immunology, COVID-19 immunology, COVID-19 virology, Mesocricetus, SARS-CoV-2, Disease Models, Animal
- Abstract
When infected with SARS-CoV-2, Syrian hamsters (Mesocricetus auratus) develop moderate disease severity presenting key features of human COVID-19. We here develop a biomathematical model of the disease course by translating known biological mechanisms of virus-host interactions and immune responses into ordinary differential equations. We explicitly describe the dynamics of virus population, affected alveolar epithelial cells, and involved relevant immune cells comprising for example CD4+ T cells, CD8+ T cells, macrophages, natural killer cells and B cells. We also describe the humoral response dynamics of neutralising antibodies and major regulatory cytokines including CCL8 and CXCL10. The model is developed and parametrized based on experimental data collected at days 2, 3, 5, and 14 post infection. Pulmonary cell composition and their transcriptional profiles were obtained by lung single-cell RNA (scRNA) sequencing analysis. Parametrization of the model resulted in a good agreement of model and data. The model can be used to predict, for example, the time course of the virus population, immune cell dynamics, antibody production and regeneration of alveolar cells for different therapy scenarios or after multiple-infection events. We aim to translate this model to the human situation in the future., Competing Interests: Declarations. Competing interests: MS received funding from Pfizer Inc. for a project not related to this research. The other authors have declared that no competing interests exist., (© 2024. The Author(s).)
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- 2024
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16. Sulfated Cellulose Nanofiber Hydrogel with Mucus-Like Activities for Virus Inhibition.
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Long Y, Dimde M, Adler JM, Vidal RM, Povolotsky TL, Nickl P, Achazi K, Trimpert J, Kaufer BB, Haag R, and Nie C
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- Humans, Animals, Vero Cells, Chlorocebus aethiops, COVID-19 virology, Mucus metabolism, Calcium metabolism, Calcium chemistry, Cellulose chemistry, Cellulose pharmacology, Cellulose analogs & derivatives, Nanofibers chemistry, Herpesvirus 1, Human drug effects, Antiviral Agents pharmacology, Antiviral Agents chemistry, SARS-CoV-2 drug effects, Hydrogels chemistry, Hydrogels pharmacology
- Abstract
Mucus is the first defense barrier against viruses in the human immune system. Inspired by the mucus structure, we designed a highly sulfated hydrogel to bind viruses and prevent infection of the underlying cells. The hydrogel was formed by gelation of sulfated cellulose nanofiber (SCNF) with Ca
2+ . SCNF exhibited a mucin-like nanofiber structure with high numbers of sulfated groups. Based on the electrostatic interactions with a virus, SCNF could efficiently inhibit herpes simplex virus-1 (HSV-1) infection with a half-maximal inhibitory concentration (IC50 ) of 0.43 μg/mL, which is comparable to that of heparin (IC50 = 0.30 μg/mL). Benefiting from the multiporous structure and sulfate groups, the Ca2+ -SCNF hydrogel could efficiently trap HSV-1 and inhibit the virus from attacking the underlying cells in a transwell model. Furthermore, SCNF also inhibited SARS-CoV-2 infection in a similar experimental setting. By integrating the advantages of high and broad-spectrum virus inhibitory activity as well as low toxicity, it is believed that the Ca2+ -SCNF hydrogel can promote the development of highly biocompatible and efficient antiviral material with "binding and inhibition" capability and other diverse strategies.- Published
- 2024
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17. Neural network-assisted humanisation of COVID-19 hamster transcriptomic data reveals matching severity states in human disease.
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Friedrich VD, Pennitz P, Wyler E, Adler JM, Postmus D, Müller K, Teixeira Alves LG, Prigann J, Pott F, Vladimirova D, Hoefler T, Goekeri C, Landthaler M, Goffinet C, Saliba AE, Scholz M, Witzenrath M, Trimpert J, Kirsten H, and Nouailles G
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- Animals, Humans, Cricetinae, Neural Networks, Computer, Single-Cell Analysis methods, Gene Expression Profiling methods, Mesocricetus, COVID-19 genetics, COVID-19 virology, Transcriptome, SARS-CoV-2 genetics, Severity of Illness Index, Disease Models, Animal
- Abstract
Background: Translating findings from animal models to human disease is essential for dissecting disease mechanisms, developing and testing precise therapeutic strategies. The coronavirus disease 2019 (COVID-19) pandemic has highlighted this need, particularly for models showing disease severity-dependent immune responses., Methods: Single-cell transcriptomics (scRNAseq) is well poised to reveal similarities and differences between species at the molecular and cellular level with unprecedented resolution. However, computational methods enabling detailed matching are still scarce. Here, we provide a structured scRNAseq-based approach that we applied to scRNAseq from blood leukocytes originating from humans and hamsters affected with moderate or severe COVID-19., Findings: Integration of data from patients with COVID-19 with two hamster models that develop moderate (Syrian hamster, Mesocricetus auratus) or severe (Roborovski hamster, Phodopus roborovskii) disease revealed that most cellular states are shared across species. A neural network-based analysis using variational autoencoders quantified the overall transcriptomic similarity across species and severity levels, showing highest similarity between neutrophils of Roborovski hamsters and patients with severe COVID-19, while Syrian hamsters better matched patients with moderate disease, particularly in classical monocytes. We further used transcriptome-wide differential expression analysis to identify which disease stages and cell types display strongest transcriptional changes., Interpretation: Consistently, hamsters' response to COVID-19 was most similar to humans in monocytes and neutrophils. Disease-linked pathways found in all species specifically related to interferon response or inhibition of viral replication. Analysis of candidate genes and signatures supported the results. Our structured neural network-supported workflow could be applied to other diseases, allowing better identification of suitable animal models with similar pathomechanisms across species., Funding: This work was supported by German Federal Ministry of Education and Research, (BMBF) grant IDs: 01ZX1304B, 01ZX1604B, 01ZX1906A, 01ZX1906B, 01KI2124, 01IS18026B and German Research Foundation (DFG) grant IDs: 14933180, 431232613., Competing Interests: Declaration of interests MS received funding from Pfizer Inc. for a project related to pneumococcal vaccination. MS receives funding from Owkin for a project not related to this research. MW reports grants and personal fees from Biotest, grants and personal fees from Pantherna, grants and personal fees from Vaxxilon, personal fees from Aptarion, personal fees from Astra Zeneca, personal fees from Chiesi, personal fees from Insmed, personal fees from Gilead, outside the submitted work. GN reports grants from Biotest AG outside the submitted work. Unrelated to this work, Freie Universität Berlin has filed a patent application (PCT/EP2022/051215) for SARS-CoV-2 vaccines. JT is named as inventor on this application and receives remuneration in accordance with German law (“Gesetz über Arbeitnehmererfindungen”). Freie Universität Berlin is collaborating with RocketVax Inc. for further development of SARS-CoV-2 vaccines and receives funding for research. The other authors declare no competing interest., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)
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- 2024
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18. Fatal infection caused by a genetically distinct elephant endotheliotropic herpesvirus type 5 in a captive Asian elephant in Germany.
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Abdelgawad A, Nascimento M, Prahl A, Flügger M, Szentiks CA, Holtze S, Hildebrandt TB, and Trimpert J
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- Animals, Germany, Male, Fatal Outcome, DNA, Viral genetics, Genome, Viral genetics, Phylogeny, Sequence Analysis, DNA, Genetic Variation, Whole Genome Sequencing, Elephants virology, Herpesviridae Infections veterinary, Herpesviridae Infections virology, Herpesviridae genetics, Herpesviridae isolation & purification, Herpesviridae classification
- Abstract
Background: Elephant endotheliotropic herpesvirus (EEHV) infection is the most common cause for lethal hemorrhagic disease in captive juvenile Asian elephants (Elephas maximus). Although EEHV1 is known as the most likely cause of fatal haemorrhagic disease in Asian elephants, EEHV5 was lately involved in lethal cases of haemorrhagic disease in captive elephants., Case Presentation: Here we report the first death of a four-year old Asian elephant diagnosed with EEHV5 in Germany. Molecular diagnosis yielded detection of EEHV5 DNA in all tested tissues. Histopathological examination revealed typical features of hemorrhagic disease in all examined organs. EEHV5 was sequenced from total DNA isolated from heart tissue by Illumina and Nanopore sequencing. Sequencing data showed 3,881 variants, distributed across the entire genome, compared to the published EEHV5 sequence., Conclusions: We have detected EEHV5 in a fatal disease case of a male Asian elephant. Whole genome sequencing revealed substantial differences of our DNA isolate compared to available EEHV5 sequences. This report of fatal haemorrhagic disease associated with EEHV5 infection should raise awareness for EEHV5 as an important elephant pathogen. Genome sequencing and downstream SNPs analysis will further encourage future research to understand genetic diversity, pathogenesis and virulence of EEHVs with respect to developing new diagnostic methods, prophylactic strategies, and implementation of surveillance and control measures., (© 2024. The Author(s).)
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- 2024
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19. Antigenic cartography using variant-specific hamster sera reveals substantial antigenic variation among Omicron subvariants.
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Mühlemann B, Trimpert J, Walper F, Schmidt ML, Jansen J, Schroeder S, Jeworowski LM, Beheim-Schwarzbach J, Bleicker T, Niemeyer D, Richter A, Adler JM, Vidal RM, Langner C, Vladimirova D, Wilks SH, Smith DJ, Voß M, Paltzow L, Martínez Christophersen C, Rose R, Krumbholz A, Jones TC, Corman VM, and Drosten C
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- Animals, Cricetinae, Antigens, Viral immunology, Antigens, Viral genetics, Antibodies, Viral blood, Antibodies, Viral immunology, Humans, Immune Sera immunology, SARS-CoV-2 immunology, SARS-CoV-2 genetics, COVID-19 immunology, COVID-19 virology, Antigenic Variation immunology, Antigenic Variation genetics, Mesocricetus, Spike Glycoprotein, Coronavirus immunology, Spike Glycoprotein, Coronavirus genetics
- Abstract
Severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) has developed substantial antigenic variability. As the majority of the population now has pre-existing immunity due to infection or vaccination, the use of experimentally generated animal immune sera can be valuable for measuring antigenic differences between virus variants. Here, we immunized Syrian hamsters by two successive infections with one of nine SARS-CoV-2 variants. Their sera were titrated against 16 SARS-CoV-2 variants, and the resulting titers were visualized using antigenic cartography. The antigenic map shows a condensed cluster containing all pre-Omicron variants (D614G, Alpha, Delta, Beta, Mu, and an engineered B.1+E484K variant) and considerably more diversity among a selected panel of Omicron subvariants (BA.1, BA.2, BA.4/BA.5, the BA.5 descendants BF.7 and BQ.1.18, the BA.2.75 descendant BN.1.3.1, the BA.2-derived recombinants XBB.2 and EG.5.1, and the BA.2.86 descendant JN.1). Some Omicron subvariants were as antigenically distinct from each other as the wildtype is from the Omicron BA.1 variant. Compared to titers measured in human sera, titers in hamster sera are of higher magnitude, show less fold change, and result in a more compact antigenic map topology. The results highlight the potential of sera from hamsters for the continued antigenic characterization of SARS-CoV-2., Competing Interests: Competing interests statement:V.M.C. has his name on patents regarding SARS-CoV-2 serological testing and monoclonal antibodies.
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- 2024
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20. Mucus-Inspired Self-Healing Hydrogels: A Protective Barrier for Cells against Viral Infection.
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Bej R, Stevens CA, Nie C, Ludwig K, Degen GD, Kerkhoff Y, Pigaleva M, Adler JM, Bustos NA, Page TM, Trimpert J, Block S, Kaufer BB, Ribbeck K, and Haag R
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- Humans, Mucins chemistry, Mucins metabolism, Antiviral Agents pharmacology, Antiviral Agents chemistry, Polymers chemistry, Polymers pharmacology, Animals, Disulfides chemistry, Polyethylene Glycols chemistry, Cryoelectron Microscopy, COVID-19 virology, Glycerol, Hydrogels chemistry, Hydrogels pharmacology, Herpesvirus 1, Human drug effects, Mucus metabolism, SARS-CoV-2 drug effects
- Abstract
Mucus is a dynamic biological hydrogel, composed primarily of the glycoprotein mucin, exhibits unique biophysical properties and forms a barrier protecting cells against a broad-spectrum of viruses. Here, this work develops a polyglycerol sulfate-based dendronized mucin-inspired copolymer (MICP-1) with ≈10% repeating units of activated disulfide as cross-linking sites. Cryo-electron microscopy (Cryo-EM) analysis of MICP-1 reveals an elongated single-chain fiber morphology. MICP-1 shows potential inhibitory activity against many viruses such as herpes simplex virus 1 (HSV-1) and SARS-CoV-2 (including variants such as Delta and Omicron). MICP-1 produces hydrogels with viscoelastic properties similar to healthy human sputum and with tuneable microstructures using linear and branched polyethylene glycol-thiol (PEG-thiol) as cross-linkers. Single particle tracking microrheology, electron paramagnetic resonance (EPR) and cryo-scanning electron microscopy (Cryo-SEM) are used to characterize the network structures. The synthesized hydrogels exhibit self-healing properties, along with viscoelastic properties that are tuneable through reduction. A transwell assay is used to investigate the hydrogel's protective properties against viral infection against HSV-1. Live-cell microscopy confirms that these hydrogels can protect underlying cells from infection by trapping the virus, due to both network morphology and anionic multivalent effects. Overall, this novel mucin-inspired copolymer generates mucus-mimetic hydrogels on a multi-gram scale. These hydrogels can be used as models for disulfide-rich airway mucus research, and as biomaterials., (© 2024 The Author(s). Advanced Materials published by Wiley‐VCH GmbH.)
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- 2024
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21. Evolutionary Dynamics of Accelerated Antiviral Resistance Development in Hypermutator Herpesvirus.
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Höfler T, Nascimento MM, Zeitlow M, Kim JY, and Trimpert J
- Subjects
- Mutation, Mutation Rate, Biological Evolution, Humans, Drug Resistance, Viral genetics, Antiviral Agents pharmacology, Herpesvirus 1, Human genetics, Herpesvirus 1, Human drug effects, Evolution, Molecular
- Abstract
Antiviral therapy is constantly challenged by the emergence of resistant pathogens. At the same time, experimental approaches to understand and predict resistance are limited by long periods required for evolutionary processes. Here, we present a herpes simplex virus 1 mutant with impaired proofreading capacity and consequently elevated mutation rates. Comparing this hypermutator to parental wild type virus, we study the evolution of antiviral drug resistance in vitro. We model resistance development and elucidate underlying genetic changes against three antiviral substances. Our analyzes reveal no principle difference in the evolutionary behavior of both viruses, adaptive processes are overall similar, however significantly accelerated for the hypermutator. We conclude that hypermutator viruses are useful for modeling adaptation to antiviral therapy. They offer the benefit of expedited adaptation without introducing apparent bias and can therefore serve as an accelerator to predict natural evolution., Competing Interests: Conflict of Interest The authors declare no conflict of interest., (© The Author(s) 2024. Published by Oxford University Press on behalf of Society for Molecular Biology and Evolution.)
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- 2024
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22. Single-cell-resolved interspecies comparison shows a shared inflammatory axis and a dominant neutrophil-endothelial program in severe COVID-19.
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Peidli S, Nouailles G, Wyler E, Adler JM, Kunder S, Voß A, Kazmierski J, Pott F, Pennitz P, Postmus D, Teixeira Alves LG, Goffinet C, Gruber AD, Blüthgen N, Witzenrath M, Trimpert J, Landthaler M, and Praktiknjo SD
- Subjects
- Animals, Humans, Cricetinae, Inflammation pathology, Mesocricetus, Disease Models, Animal, Male, Species Specificity, COVID-19 immunology, COVID-19 virology, COVID-19 pathology, Neutrophils immunology, SARS-CoV-2 immunology, Single-Cell Analysis, Lung pathology, Lung virology, Lung immunology, Endothelial Cells virology, Endothelial Cells pathology
- Abstract
A key issue for research on COVID-19 pathogenesis is the lack of biopsies from patients and of samples at the onset of infection. To overcome these hurdles, hamsters were shown to be useful models for studying this disease. Here, we further leverage the model to molecularly survey the disease progression from time-resolved single-cell RNA sequencing data collected from healthy and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected Syrian and Roborovski hamster lungs. We compare our data to human COVID-19 studies, including bronchoalveolar lavage, nasal swab, and postmortem lung tissue, and identify a shared axis of inflammation dominated by macrophages, neutrophils, and endothelial cells, which we show to be transient in Syrian and terminal in Roborovski hamsters. Our data suggest that, following SARS-CoV-2 infection, commitment to a type 1- or type 3-biased immunity determines moderate versus severe COVID-19 outcomes, respectively., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)
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- 2024
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23. Computationally Designed Epitope-Mediated Imprinted Polymers versus Conventional Epitope Imprints for the Detection of Human Adenovirus in Water and Human Serum Samples.
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Sehit E, Yao G, Battocchio G, Radfar R, Trimpert J, Mroginski MA, Süssmuth R, and Altintas Z
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- Humans, Biosensing Techniques methods, Polymers chemistry, Molecular Dynamics Simulation, Molecularly Imprinted Polymers chemistry, Molecular Imprinting methods, Limit of Detection, SARS-CoV-2 immunology, SARS-CoV-2 isolation & purification, SARS-CoV-2 chemistry, Adenoviruses, Human immunology, Adenoviruses, Human chemistry, Epitopes immunology, Epitopes chemistry
- Abstract
Detection of pathogenic viruses for point-of-care applications has attracted great attention since the COVID-19 pandemic. Current virus diagnostic tools are laborious and expensive, while requiring medically trained staff. Although user-friendly and cost-effective biosensors are utilized for virus detection, many of them rely on recognition elements that suffer major drawbacks. Herein, computationally designed epitope-imprinted polymers (eIPs) are conjugated with a portable piezoelectric sensing platform to establish a sensitive and robust biosensor for the human pathogenic adenovirus (HAdV). The template epitope is selected from the knob part of the HAdV capsid, ensuring surface accessibility. Computational simulations are performed to evaluate the conformational stability of the selected epitope. Further, molecular dynamics simulations are executed to investigate the interactions between the epitope and the different functional monomers for the smart design of eIPs. The HAdV epitope is imprinted via the solid-phase synthesis method to produce eIPs using in silico-selected ingredients. The synthetic receptors show a remarkable detection sensitivity (LOD: 10
2 pfu mL-1 ) and affinity (dissociation constant ( Kd ): 6.48 × 10-12 M) for HAdV. Moreover, the computational eIPs lead to around twofold improved binding behavior than the eIPs synthesized with a well-established conventional recipe. The proposed computational strategy holds enormous potential for the intelligent design of ultrasensitive imprinted polymer binders.- Published
- 2024
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24. Protective role of the HSP90 inhibitor, STA-9090, in lungs of SARS-CoV-2-infected Syrian golden hamsters.
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Teixeira Alves LG, Baumgardt M, Langner C, Fischer M, Maria Adler J, Bushe J, Firsching TC, Mastrobuoni G, Grobe J, Hoenzke K, Kempa S, Gruber AD, Hocke AC, Trimpert J, Wyler E, and Landthaler M
- Subjects
- Cricetinae, Animals, Humans, Mesocricetus, Lung pathology, SARS-CoV-2, COVID-19 pathology, Triazoles
- Abstract
Introduction: The emergence of new SARS-CoV-2 variants, capable of escaping the humoral immunity acquired by the available vaccines, together with waning immunity and vaccine hesitancy, challenges the efficacy of the vaccination strategy in fighting COVID-19. Improved therapeutic strategies are urgently needed to better intervene particularly in severe cases of the disease. They should aim at controlling the hyperinflammatory state generated on infection, reducing lung tissue pathology and inhibiting viral replication. Previous research has pointed to a possible role for the chaperone HSP90 in SARS-CoV-2 replication and COVID-19 pathogenesis. Pharmacological intervention through HSP90 inhibitors was shown to be beneficial in the treatment of inflammatory diseases, infections and reducing replication of diverse viruses., Methods: In this study, we investigated the effects of the potent HSP90 inhibitor Ganetespib (STA-9090) in vitro on alveolar epithelial cells and alveolar macrophages to characterise its effects on cell activation and viral replication. Additionally, the Syrian hamster animal model was used to evaluate its efficacy in controlling systemic inflammation and viral burden after infection., Results: In vitro, STA-9090 reduced viral replication on alveolar epithelial cells in a dose-dependent manner and lowered significantly the expression of proinflammatory genes, in both alveolar epithelial cells and alveolar macrophages. In vivo, although no reduction in viral load was observed, administration of STA-9090 led to an overall improvement of the clinical condition of infected animals, with reduced oedema formation and lung tissue pathology., Conclusion: Altogether, we show that HSP90 inhibition could serve as a potential treatment option for moderate and severe cases of COVID-19., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)
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- 2024
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25. Improving the Antigenicity of SARS-CoV-2 Vaccine Genes by Merging Mutations from Different Variants of Concern.
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Herwig S, Adler JM, Vladimirova D, Trimpert J, Sehouli J, and Cichon G
- Abstract
During the COVID-19 pandemic, the early emergence of viral variants repeatedly undermined the effects of vaccination. Our aim here is to explore strategies for improving spike vaccine gene antigenicity by merging mutations from different variants of concern (VOCs) in a single vaccine gene. To this end, newly developed recombinant vaccine genes were designed, cloned into adenoviral vectors, and applied to C57BL/6 mice; then, serum-neutralizing antibodies against the wildtype SARS-CoV-2 strains were determined in neutralization assays. The merger of mutations from different variants of concern (alpha, beta, gamma, and delta) in a single recombinant spike-based vaccine gene provided a substantial improvement in neutralizing immunity to all variants of concern, including the omicron strains. To date, only unmodified spike genes of the original SARS-CoV-2 Wuhan strain (B.1) or dominant variants (BA.1, BA.5, and XBB.1.5) have been used as vaccine genes. The employment of unmodified vaccine genes is afflicted by limited cross-protection among variant strains. In contrast, recombinant vaccine genes that combine mutations from different strains in a single gene hold the potential to broaden and improve immune protection and might help to reduce the need for frequent vaccine adaptations in the future.
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- 2024
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26. An intranasal live-attenuated SARS-CoV-2 vaccine limits virus transmission.
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Adler JM, Martin Vidal R, Langner C, Vladimirova D, Abdelgawad A, Kunecova D, Lin X, Nouailles G, Voss A, Kunder S, Gruber AD, Wu H, Osterrieder N, Kunec D, and Trimpert J
- Subjects
- Animals, Cricetinae, Male, Humans, BNT162 Vaccine, mRNA Vaccines, SARS-CoV-2, Mesocricetus, Antibodies, Viral, Antibodies, Neutralizing, COVID-19 Vaccines, COVID-19 prevention & control
- Abstract
The development of effective SARS-CoV-2 vaccines has been essential to control COVID-19, but significant challenges remain. One problem is intramuscular administration, which does not induce robust mucosal immune responses in the upper airways-the primary site of infection and virus shedding. Here we compare the efficacy of a mucosal, replication-competent yet fully attenuated virus vaccine, sCPD9-ΔFCS, and the monovalent mRNA vaccine BNT162b2 in preventing transmission of SARS-CoV-2 variants B.1 and Omicron BA.5 in two scenarios. Firstly, we assessed the protective efficacy of the vaccines by exposing vaccinated male Syrian hamsters to infected counterparts. Secondly, we evaluated transmission of the challenge virus from vaccinated and subsequently challenged male hamsters to naïve contacts. Our findings demonstrate that the live-attenuated vaccine (LAV) sCPD9-ΔFCS significantly outperformed the mRNA vaccine in preventing virus transmission in both scenarios. Our results provide evidence for the advantages of locally administered LAVs over intramuscularly administered mRNA vaccines in preventing infection and reducing virus transmission., (© 2024. The Author(s).)
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- 2024
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27. A systematic analysis of anthocyanins inhibiting human, murine, and equine herpesviruses.
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Roll V, Diesendorf V, Roewer N, Abdelgawad A, Roewer J, Trimpert J, and Bodem J
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- Child, Humans, Animals, Horses, Mice, Anthocyanins pharmacology, Anthocyanins analysis, Antiviral Agents pharmacology, Plant Extracts pharmacology, Mammals, Hepatitis C, Chronic, Herpesvirus 1, Human, Elaeocarpaceae, Cytomegalovirus Infections
- Abstract
Background: Herpesviruses are common animal and human pathogens that cause severe health problems in children, immunocompromised patients, and infected animals with a host range from fish to mammals. Anthocyanin-containing plant extracts have been described as potent antivirals, which might cause fewer harmful side effects than direct-acting antivirals. Here, we report that an extract of Aristotelia chilensis (Molina) Stuntz (Elaeocarpaceae) (MBE) with a high content of the anthocyanin delphinidin suppresses lytic replication of equine, murine and human herpesviruses of replication in vitro., Methods: We treated cultured cells with MBE and purified individual anthocyanins present in the extract to determine the most active compound at different concentrations. We subsequently infected the cultures with human herpesviruses 1 (HSV-1) or 8 (HHV-8), murine cytomegalovirus (CMV), or equine herpesviruses 1 (EHV-1) and determined the number of infected cells and viral infectivity., Results: MBE inhibited the HSV-1, murine CMV, and EHV-1 by up to 2 orders of magnitude. In the presence of the stabilizing randomly methylated-beta-cyclodextrin, the inhibitory concentration could be lowered significantly. We identified delphinidin as an active antiviral compound and showed that the non-glycosylated delphinidin solved and stabilized with sulfobutylether-beta-cyclodextrin allowed usage of approximately 50 times lower concentrations., Conclusion: Glycosylated delphinidin derivatives were identified as active antiviral compounds of MBE. This suggests that plant extracts rich in delphinidin-anthocyanins have potent antiviral properties that could be used in treatment and prevention., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier GmbH. All rights reserved.)
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- 2024
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28. A non-transmissible live attenuated SARS-CoV-2 vaccine.
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Adler JM, Martin Vidal R, Voß A, Kunder S, Nascimento M, Abdelgawad A, Langner C, Vladimirova D, Osterrieder N, Gruber AD, Kunec D, and Trimpert J
- Subjects
- Animals, Cricetinae, Humans, Pandemics, SARS-CoV-2, Vaccines, Attenuated, Antibodies, Viral, Antibodies, Neutralizing, COVID-19 Vaccines, COVID-19 prevention & control
- Abstract
Live attenuated vaccines (LAVs) administered via the mucosal route may offer better control of the COVID-19 pandemic than non-replicating vaccines injected intramuscularly. Conceptionally, LAVs have several advantages, including presentation of the entire antigenic repertoire of the virus, and the induction of strong mucosal immunity. Thus, immunity induced by LAV could offer superior protection against future surges of COVID-19 cases caused by emerging SARS-CoV-2 variants. However, LAVs carry the risk of unintentional transmission. To address this issue, we investigated whether transmission of a SARS-CoV-2 LAV candidate can be blocked by removing the furin cleavage site (FCS) from the spike protein. The level of protection and immunity induced by the attenuated virus with the intact FCS was virtually identical to the one induced by the attenuated virus lacking the FCS. Most importantly, removal of the FCS completely abolished horizontal transmission of vaccine virus between cohoused hamsters. Furthermore, the vaccine was safe in immunosuppressed animals and showed no tendency to recombine in vitro or in vivo with a SARS-CoV-2 field strain. These results indicate that removal of the FCS from SARS-CoV-2 LAV is a promising strategy to increase vaccine safety and prevent vaccine transmission without compromising vaccine efficacy., Competing Interests: Declaration of interests Related to this work, Freie Universität Berlin has filed a patent application for the use of sCPD9 and sCPD9-ΔFCS as vaccine. In this application, J.T., N.O., and D.K. are named as inventors of sCPD9. Freie Universität Berlin is collaborating with RocketVax AG for further development of sCPD9-ΔFCS as vaccine and receives funding for research., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)
- Published
- 2023
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29. In vitro and in vivo effects of Pelargonium sidoides DC. root extract EPs ® 7630 and selected constituents against SARS-CoV-2 B.1, Delta AY.4/AY.117 and Omicron BA.2.
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Emanuel J, Papies J, Galander C, Adler JM, Heinemann N, Eschke K, Merz S, Pischon H, Rose R, Krumbholz A, Kulić Ž, Lehner MD, Trimpert J, and Müller MA
- Abstract
The occurrence of immune-evasive SARS-CoV-2 strains emphasizes the importance to search for broad-acting antiviral compounds. Our previous in vitro study showed that Pelargonium sidoides DC. root extract EPs
® 7630 has combined antiviral and immunomodulatory properties in SARS-CoV-2-infected human lung cells. Here we assessed in vivo effects of EPs® 7630 in SARS-CoV-2-infected hamsters, and investigated properties of EPs® 7630 and its functionally relevant constituents in context of phenotypically distinct SARS-CoV-2 variants. We show that EPs® 7630 reduced viral load early in the course of infection and displayed significant immunomodulatory properties positively modulating disease progression in hamsters. In addition, we find that EPs® 7630 differentially inhibits SARS-CoV-2 variants in nasal and bronchial human airway epithelial cells. Antiviral effects were more pronounced against Omicron BA.2 compared to B.1 and Delta, the latter two preferring TMPRSS2-mediated fusion with the plasma membrane for cell entry instead of receptor-mediated low pH-dependent endocytosis. By using SARS-CoV-2 Spike VSV-based pseudo particles (VSVpp), we confirm higher EPs® 7630 activity against Omicron Spike-VSVpp, which seems independent of the serine protease TMPRSS2, suggesting that EPs® 7630 targets endosomal entry. We identify at least two molecular constituents of EPs® 7630, i.e., (-)-epigallocatechin and taxifolin with antiviral effects on SARS-CoV-2 replication and cell entry. In summary, our study shows that EPs® 7630 ameliorates disease outcome in SARS-CoV-2-infected hamsters and has enhanced activity against Omicron, apparently by limiting late endosomal SARS-CoV-2 entry., Competing Interests: Authors ŽK and ML were employees by Dr. Willmar Schwabe GmbH and Co. KG. The funder had the following involvement with the study: ML contributed to the design of the study, analyzed data, provided material, wrote and edited the main text. ŽK provided material, analyzed data, wrote and edited the main text. SM and HP were employed by IDEXX Laboratories. AK was employed by Labor Dr. Krause und Kollegen MVZ GmbH. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Emanuel, Papies, Galander, Adler, Heinemann, Eschke, Merz, Pischon, Rose, Krumbholz, Kulić, Lehner, Trimpert and Müller.)- Published
- 2023
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30. Mucin-Inspired Single-Chain Polymer (MIP) Fibers as Potent SARS-CoV-2 Inhibitors.
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Bej R, Nie C, Ludwig K, Ahmadi V, Trimpert J, Adler JM, Povolotsky TL, Achazi K, Kagelmacher M, Vidal RM, Dernedde J, Kaufer BB, and Haag R
- Subjects
- Humans, Mucins, SARS-CoV-2, Polymers pharmacology, Polymers chemistry, Molecular Imprinting methods, COVID-19
- Abstract
Mucins are the key component of the defensive mucus barrier. They are extended fibers of very high molecular weight with diverse biological functions depending strongly on their specific structural parameters. Here, we present a mucin-inspired nanostructure, produced via a synthetic methodology to prepare methacrylate-based dendronized polysulfates (MIP-1) on a multi gram-scale with high molecular weight (MW=450 kDa) and thiol end-functionalized mucin-inspired polymer (MIP) via RAFT polymerization. Cryo-electron tomography (Cryo-ET) analysis of MIP-1 confirmed a mucin-mimetic wormlike single-chain fiber structure (length=144±59 nm) in aqueous solution. This biocompatible fiber showed promising activity against SARS-CoV-2 and its mutant strain, with a remarkable low half maximal (IC
50 ) inhibitory concentration (IC50 =10.0 nM). Additionally, we investigate the impact of fiber length on SARS-CoV-2 inhibition by testing other functional polymers (MIPs) of varying fiber lengths., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)- Published
- 2023
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31. Early protective effect of a ("pan") coronavirus vaccine (PanCoVac) in Roborovski dwarf hamsters after single-low dose intranasal administration.
- Author
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Abdelaziz MO, Raftery MJ, Weihs J, Bielawski O, Edel R, Köppke J, Vladimirova D, Adler JM, Firsching T, Voß A, Gruber AD, Hummel LV, Fernandez Munoz I, Müller-Marquardt F, Willimsky G, Elleboudy NS, Trimpert J, and Schönrich G
- Subjects
- Cricetinae, Humans, Animals, Mice, SARS-CoV-2, COVID-19 Vaccines, Epitopes, T-Lymphocyte, Administration, Intranasal, Antibodies, Neutralizing, HLA-A Antigens, COVID-19 prevention & control, Viral Vaccines
- Abstract
Introduction: The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has highlighted the danger posed by human coronaviruses. Rapid emergence of immunoevasive variants and waning antiviral immunity decrease the effect of the currently available vaccines, which aim at induction of neutralizing antibodies. In contrast, T cells are marginally affected by antigen evolution although they represent the major mediators of virus control and vaccine protection against virus-induced disease., Materials and Methods: We generated a multi-epitope vaccine (PanCoVac) that encodes the conserved T cell epitopes from all structural proteins of coronaviruses. PanCoVac contains elements that facilitate efficient processing and presentation of PanCoVac-encoded T cell epitopes and can be uploaded to any available vaccine platform. For proof of principle, we cloned PanCoVac into a non-integrating lentivirus vector (NILV-PanCoVac). We chose Roborovski dwarf hamsters for a first step in evaluating PanCoVac in vivo . Unlike mice, they are naturally susceptible to SARS-CoV-2 infection. Moreover, Roborovski dwarf hamsters develop COVID-19-like disease after infection with SARS-CoV-2 enabling us to look at pathology and clinical symptoms., Results: Using HLA-A
* 0201-restricted reporter T cells and U251 cells expressing a tagged version of PanCoVac, we confirmed in vitro that PanCoVac is processed and presented by HLA-A* 0201. As mucosal immunity in the respiratory tract is crucial for protection against respiratory viruses such as SARS-CoV-2, we tested the protective effect of single-low dose of NILV-PanCoVac administered via the intranasal (i.n.) route in the Roborovski dwarf hamster model of COVID-19. After infection with ancestral SARS-CoV-2, animals immunized with a single-low dose of NILV-PanCoVac i.n. did not show symptoms and had significantly decreased viral loads in the lung tissue. This protective effect was observed in the early phase (2 days post infection) after challenge and was not dependent on neutralizing antibodies., Conclusion: PanCoVac, a multi-epitope vaccine covering conserved T cell epitopes from all structural proteins of coronaviruses, might protect from severe disease caused by SARS-CoV-2 variants and future pathogenic coronaviruses. The use of (HLA-) humanized animal models will allow for further efficacy studies of PanCoVac-based vaccines in vivo ., Competing Interests: MA, MR and GS hold a patent EP4176898 related to the PanCoVac vaccine described herein. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Abdelaziz, Raftery, Weihs, Bielawski, Edel, Köppke, Vladimirova, Adler, Firsching, Voß, Gruber, Hummel, Fernandez Munoz, Müller-Marquardt, Willimsky, Elleboudy, Trimpert and Schönrich.)- Published
- 2023
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32. The NSP4 T492I mutation increases SARS-CoV-2 infectivity by altering non-structural protein cleavage.
- Author
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Lin X, Sha Z, Trimpert J, Kunec D, Jiang C, Xiong Y, Xu B, Zhu Z, Xue W, and Wu H
- Subjects
- Animals, Cricetinae, Humans, SARS-CoV-2 genetics, Biological Evolution, Mutation, Spike Glycoprotein, Coronavirus, COVID-19
- Abstract
The historically dominant SARS-CoV-2 Delta variant and the currently dominant Omicron variants carry a T492I substitution within the non-structural protein 4 (NSP4). Based on in silico analyses, we hypothesized that the T492I mutation increases viral transmissibility and adaptability, which we confirmed with competition experiments in hamster and human airway tissue culture models. Furthermore, we showed that the T492I mutation increases the replication capacity and infectiveness of the virus and improves its ability to evade host immune responses. Mechanistically, the T492I mutation increases the cleavage efficiency of the viral main protease NSP5 by enhancing enzyme-substrate binding, which increases production of nearly all non-structural proteins processed by NSP5. Importantly, the T492I mutation suppresses viral-RNA-associated chemokine production in monocytic macrophages, which may contribute to the attenuated pathogenicity of Omicron variants. Our results highlight the importance of NSP4 adaptation in the evolutionary dynamics of SARS-CoV-2., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)
- Published
- 2023
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33. Live-attenuated vaccine sCPD9 elicits superior mucosal and systemic immunity to SARS-CoV-2 variants in hamsters.
- Author
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Nouailles G, Adler JM, Pennitz P, Peidli S, Teixeira Alves LG, Baumgardt M, Bushe J, Voss A, Langenhagen A, Langner C, Martin Vidal R, Pott F, Kazmierski J, Ebenig A, Lange MV, Mühlebach MD, Goekeri C, Simmons S, Xing N, Abdelgawad A, Herwig S, Cichon G, Niemeyer D, Drosten C, Goffinet C, Landthaler M, Blüthgen N, Wu H, Witzenrath M, Gruber AD, Praktiknjo SD, Osterrieder N, Wyler E, Kunec D, and Trimpert J
- Subjects
- Animals, Cricetinae, Humans, Vaccines, Attenuated, COVID-19 Vaccines, BNT162 Vaccine, Pandemics, Mesocricetus, SARS-CoV-2, COVID-19 prevention & control
- Abstract
Vaccines play a critical role in combating the COVID-19 pandemic. Future control of the pandemic requires improved vaccines with high efficacy against newly emerging SARS-CoV-2 variants and the ability to reduce virus transmission. Here we compare immune responses and preclinical efficacy of the mRNA vaccine BNT162b2, the adenovirus-vectored spike vaccine Ad2-spike and the live-attenuated virus vaccine candidate sCPD9 in Syrian hamsters, using both homogeneous and heterologous vaccination regimens. Comparative vaccine efficacy was assessed by employing readouts from virus titrations to single-cell RNA sequencing. Our results show that sCPD9 vaccination elicited the most robust immunity, including rapid viral clearance, reduced tissue damage, fast differentiation of pre-plasmablasts, strong systemic and mucosal humoral responses, and rapid recall of memory T cells from lung tissue after challenge with heterologous SARS-CoV-2. Overall, our results demonstrate that live-attenuated vaccines offer advantages over currently available COVID-19 vaccines., (© 2023. The Author(s).)
- Published
- 2023
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34. Preclinical safety and efficacy of a therapeutic antibody that targets SARS-CoV-2 at the sotrovimab face but is escaped by Omicron.
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Kreye J, Reincke SM, Edelburg S, Jeworowski LM, Kornau HC, Trimpert J, Hombach P, Halbe S, Nölle V, Meyer M, Kattenbach S, Sánchez-Sendin E, Schmidt ML, Schwarz T, Rose R, Krumbholz A, Merz S, Adler JM, Eschke K, Abdelgawad A, Schmitz D, Sander LE, Janssen U, Corman VM, and Prüss H
- Abstract
The recurrent emerging of novel viral variants of concern (VOCs) with evasion of preexisting antibody immunity upholds severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) case numbers and maintains a persistent demand for updated therapies. We selected the patient-derived antibody CV38-142 based on its potency and breadth against the VOCs Alpha, Beta, Gamma, and Delta for preclinical development into a therapeutic. CV38-142 showed in vivo efficacy in a Syrian hamster VOC infection model after post-exposure and therapeutic application and revealed a favorable safety profile in a human protein library screen and tissue cross-reactivity study. Although CV38-142 targets the same viral surface as sotrovimab, which maintains activity against Omicron, CV38-142 did not neutralize the Omicron lineages BA.1 and BA.2. These results highlight the contingencies of developing antibody therapeutics in the context of antigenic drift and reinforce the need to develop broadly neutralizing variant-proof antibodies against SARS-CoV-2., Competing Interests: The German Center for Neurodegenerative Diseases (DZNE) and Charité-Universitätsmedizin Berlin have filed a patent application (application number: PCT/EP2021/064,352) on antibodies for the treatment of SARS-CoV-2 infection on which J.K., S.M.R., H.-C.K., E.S.-S., L.E.S., V.M.C., and H.P. are named as inventors. V.M.C. is named together with Euroimmun GmbH on a patent application filed recently regarding the diagnostic of SARS-CoV-2 by antibody testing. S.E., V.N., M.M., S.K., S.H., P.H., and U.J. are employees of Miltenyi Biotec B.V. & Co. KG., (© 2023 The Author(s).)
- Published
- 2023
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35. Functionalized Fullerene for Inhibition of SARS-CoV-2 Variants.
- Author
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Page TM, Nie C, Neander L, Povolotsky TL, Sahoo AK, Nickl P, Adler JM, Bawadkji O, Radnik J, Achazi K, Ludwig K, Lauster D, Netz RR, Trimpert J, Kaufer B, Haag R, and Donskyi IS
- Subjects
- Humans, SARS-CoV-2, Protein Binding, COVID-19, Fullerenes pharmacology
- Abstract
As virus outbreaks continue to pose a challenge, a nonspecific viral inhibitor can provide significant benefits, especially against respiratory viruses. Polyglycerol sulfates recently emerge as promising agents that mediate interactions between cells and viruses through electrostatics, leading to virus inhibition. Similarly, hydrophobic C
60 fullerene can prevent virus infection via interactions with hydrophobic cavities of surface proteins. Here, two strategies are combined to inhibit infection of SARS-CoV-2 variants in vitro. Effective inhibitory concentrations in the millimolar range highlight the significance of bare fullerene's hydrophobic moiety and electrostatic interactions of polysulfates with surface proteins of SARS-CoV-2. Furthermore, microscale thermophoresis measurements support that fullerene linear polyglycerol sulfates interact with the SARS-CoV-2 virus via its spike protein, and highlight importance of electrostatic interactions within it. All-atom molecular dynamics simulations reveal that the fullerene binding site is situated close to the receptor binding domain, within 4 nm of polyglycerol sulfate binding sites, feasibly allowing both portions of the material to interact simultaneously., (© 2023 The Authors. Small published by Wiley-VCH GmbH.)- Published
- 2023
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36. Protocol to dissociate healthy and infected murine- and hamster-derived lung tissue for single-cell transcriptome analysis.
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Pennitz P, Goekeri C, Trimpert J, Wyler E, Ebenig A, Weissfuss C, Mühlebach MD, Witzenrath M, and Nouailles G
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- Cricetinae, Animals, Mice, Cell Death, Dissection, Lung, Endothelial Cells, Single-Cell Gene Expression Analysis
- Abstract
In infectious disease research, single-cell RNA sequencing allows dissection of host-pathogen interactions. As a prerequisite, we provide a protocol to transform solid and complex organs such as lungs into representative diverse, viable single-cell suspensions. Our protocol describes performance of vascular perfusion, pneumonectomy, enzymatic digestion, and mechanical dissociation of lung tissue, as well as red blood cell lysis and counting of isolated cells. A challenge remains, however, to further increase the proportion of pulmonary endothelial cells without compromising on viability. For complete details on the use and execution of this protocol, please refer to Nouailles et al. (2021),
1 Wyler et al. (2022),2 and Ebenig et al. (2022).3 ., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)- Published
- 2023
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37. Pet Rats as the Likely Reservoir for Human Seoul Orthohantavirus Infection.
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Heuser E, Drewes S, Trimpert J, Kunec D, Mehl C, de Cock MP, de Vries A, Klier C, Oskamp M, Tenhaken P, Hashemi F, Heinz D, Nascimento M, Boelhauve M, Petraityte-Burneikiene R, Raafat D, Maas M, Krüger DH, Latz A, Hofmann J, Heckel G, Dreesman J, and Ulrich RG
- Subjects
- Humans, Animals, Rats, Europe, Breeding, Exons, France, RNA, Zoonoses, Seoul virus genetics
- Abstract
Seoul orthohantavirus (SEOV) is a rat-associated zoonotic pathogen with an almost worldwide distribution. In 2019, the first autochthonous human case of SEOV-induced hemorrhagic fever with renal syndrome was reported in Germany, and a pet rat was identified as the source of the zoonotic infection. To further investigate the SEOV reservoir, additional rats from the patient and another owner, all of which were purchased from the same vendor, were tested. SEOV RNA and anti-SEOV antibodies were found in both of the patient's rats and in two of the three rats belonging to the other owner. The complete coding sequences of the small (S), medium (M), and large (L) segments obtained from one rat per owner exhibited a high sequence similarity to SEOV strains of breeder rat or human origin from the Netherlands, France, the USA, and Great Britain. Serological screening of 490 rats from breeding facilities and 563 wild rats from Germany (2007-2020) as well as 594 wild rats from the Netherlands (2013-2021) revealed 1 and 6 seropositive individuals, respectively. However, SEOV RNA was not detected in any of these animals. Increased surveillance of pet, breeder, and wild rats is needed to identify the origin of the SEOV strain in Europe and to develop measures to prevent transmission to the human population.
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- 2023
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38. Suicidal Phenotype of Proofreading-Deficient Herpes Simplex Virus 1 Polymerase Mutants.
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Brunialti M, Höfler T, Nascimento M, and Trimpert J
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- DNA Replication genetics, Exonucleases genetics, Exonucleases metabolism, Mutation, Phenotype, Herpesvirus 1, Human enzymology, Herpesvirus 1, Human genetics, Herpesvirus 1, Human metabolism
- Abstract
Herpes simplex virus 1 (HSV-1) encodes a family B DNA polymerase (Pol) capable of exonucleolytic proofreading whose functions have been extensively studied in the past. Early studies on the in vitro activity of purified Pol protein found that the enzymatic functions of the holoenzyme are largely separate. Consequently, exonuclease activity can be reduced or abolished by certain point mutations within catalytically important regions, with no or only minor effects on polymerase activity. Despite unimpaired polymerase activity, the recovery of HSV-1 mutants with a catalytically inactive exonuclease has been so far unsuccessful. Hence, mutations such as D368A, which abolish exonuclease activity, are believed to be lethal. Here, we show that HSV-1 can be recovered in the absence of Pol intrinsic exonuclease activity and demonstrate that a lack of proofreading causes the rapid accumulation of likely detrimental mutations. Although mutations that abolish exonuclease activity do not appear to be lethal, the lack of proofreading yields viruses with a suicidal phenotype that cease to replicate within few passages following reconstitution. Hence, we conclude that high replication fidelity conferred by proofreading is essential to maintain HSV-1 genome integrity and that a lack of exonuclease activity produces an initially viable but rapidly suicidal phenotype. However, stably replicating viruses with reduced exonuclease activity and therefore elevated mutation rates can be generated by mutating a catalytically less important site located within a conserved exonuclease domain. IMPORTANCE Recovery of fully exonuclease-deficient herpes simplex virus 1 (HSV-1) DNA polymerase mutants has been so far unsuccessful. However, exonuclease activity is not known to be directly essential for virus replication, and the lethal phenotype of certain HSV-1 polymerase mutants is thus attributed to factors other than exonuclease activity. Here, we showed that the recovery of a variety of exonuclease-deficient HSV-1 polymerase mutants is possible and that these mutants are initially replication competent. We, however, observed a progressive loss of mutant viability upon cell culture passaging, which coincided with the rapid accumulation of mutations in exonuclease-deficient viruses. We thus concluded that a lack of DNA proofreading in exonuclease-deficient viruses causes an initially viable but rapidly suicidal hypermutator phenotype and, consequently, the extinction of mutant viruses within few generations following recovery. This would make the absence of exonuclease activity the primary reason for the long-reported difficulties in culturing exonuclease-deficient HSV-1 mutants.
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- 2023
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39. Unconventional secretion of unglycosylated ORF8 is critical for the cytokine storm during SARS-CoV-2 infection.
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Lin X, Fu B, Xiong Y, Xing N, Xue W, Guo D, Zaky M, Pavani K, Kunec D, Trimpert J, and Wu H
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- Humans, Inflammation, Open Reading Frames, COVID-19 pathology, Cytokine Release Syndrome pathology, SARS-CoV-2 physiology, Viral Proteins metabolism
- Abstract
Coronavirus disease 2019 is a respiratory infectious disease caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Evidence on the pathogenesis of SARS-CoV-2 is accumulating rapidly. In addition to structural proteins such as Spike and Envelope, the functional roles of non-structural and accessory proteins in regulating viral life cycle and host immune responses remain to be understood. Here, we show that open reading frame 8 (ORF8) acts as messenger for inter-cellular communication between alveolar epithelial cells and macrophages during SARS-CoV-2 infection. Mechanistically, ORF8 is a secretory protein that can be secreted by infected epithelial cells via both conventional and unconventional secretory pathways. Conventionally secreted ORF8 is glycosylated and loses the ability to recognize interleukin 17 receptor A of macrophages, possibly due to the steric hindrance imposed by N-glycosylation at Asn78. However, unconventionally secreted ORF8 does not undergo glycosylation without experiencing the ER-Golgi trafficking, thereby activating the downstream NF-κB signaling pathway and facilitating a burst of cytokine release. Furthermore, we show that ORF8 deletion in SARS-CoV-2 attenuates inflammation and yields less lung lesions in hamsters. Our data collectively highlights a role of ORF8 protein in the development of cytokine storms during SARS-CoV-2 infection., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Lin et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)
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- 2023
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40. The trispecific DARPin ensovibep inhibits diverse SARS-CoV-2 variants.
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Rothenberger S, Hurdiss DL, Walser M, Malvezzi F, Mayor J, Ryter S, Moreno H, Liechti N, Bosshart A, Iss C, Calabro V, Cornelius A, Hospodarsch T, Neculcea A, Looser T, Schlegel A, Fontaine S, Villemagne D, Paladino M, Schiegg D, Mangold S, Reichen C, Radom F, Kaufmann Y, Schaible D, Schlegel I, Zitt C, Sigrist G, Straumann M, Wolter J, Comby M, Sacarcelik F, Drulyte I, Lyoo H, Wang C, Li W, Du W, Binz HK, Herrup R, Lusvarghi S, Neerukonda SN, Vassell R, Wang W, Adler JM, Eschke K, Nascimento M, Abdelgawad A, Gruber AD, Bushe J, Kershaw O, Knutson CG, Balavenkatraman KK, Ramanathan K, Wyler E, Teixeira Alves LG, Lewis S, Watson R, Haeuptle MA, Zürcher A, Dawson KM, Steiner D, Weiss CD, Amstutz P, van Kuppeveld FJM, Stumpp MT, Bosch BJ, Engler O, and Trimpert J
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- Animals, Cricetinae, Humans, Designed Ankyrin Repeat Proteins, Cryoelectron Microscopy, Antibodies, Monoclonal therapeutic use, Combined Antibody Therapeutics, Antibodies, Neutralizing, SARS-CoV-2 genetics, COVID-19
- Abstract
The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants with potential resistance to existing drugs emphasizes the need for new therapeutic modalities with broad variant activity. Here we show that ensovibep, a trispecific DARPin (designed ankyrin repeat protein) clinical candidate, can engage the three units of the spike protein trimer of SARS-CoV-2 and inhibit ACE2 binding with high potency, as revealed by cryo-electron microscopy analysis. The cooperative binding together with the complementarity of the three DARPin modules enable ensovibep to inhibit frequent SARS-CoV-2 variants, including Omicron sublineages BA.1 and BA.2. In Roborovski dwarf hamsters infected with SARS-CoV-2, ensovibep reduced fatality similarly to a standard-of-care monoclonal antibody (mAb) cocktail. When used as a single agent in viral passaging experiments in vitro, ensovibep reduced the emergence of escape mutations in a similar fashion to the same mAb cocktail. These results support further clinical evaluation of ensovibep as a broad variant alternative to existing targeted therapies for Coronavirus Disease 2019 (COVID-19)., (© 2022. The Author(s).)
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- 2022
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41. SARS-CoV-2 variant Alpha has a spike-dependent replication advantage over the ancestral B.1 strain in human cells with low ACE2 expression.
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Niemeyer D, Stenzel S, Veith T, Schroeder S, Friedmann K, Weege F, Trimpert J, Heinze J, Richter A, Jansen J, Emanuel J, Kazmierski J, Pott F, Jeworowski LM, Olmer R, Jaboreck MC, Tenner B, Papies J, Walper F, Schmidt ML, Heinemann N, Möncke-Buchner E, Baumgardt M, Hoffmann K, Widera M, Thao TTN, Balázs A, Schulze J, Mache C, Jones TC, Morkel M, Ciesek S, Hanitsch LG, Mall MA, Hocke AC, Thiel V, Osterrieder K, Wolff T, Martin U, Corman VM, Müller MA, Goffinet C, and Drosten C
- Subjects
- Humans, Angiotensin-Converting Enzyme 2 genetics, Virus Shedding, Antibodies, Blocking, SARS-CoV-2 genetics, COVID-19
- Abstract
Epidemiological data demonstrate that Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) Alpha and Delta are more transmissible, infectious, and pathogenic than previous variants. Phenotypic properties of VOC remain understudied. Here, we provide an extensive functional study of VOC Alpha replication and cell entry phenotypes assisted by reverse genetics, mutational mapping of spike in lentiviral pseudotypes, viral and cellular gene expression studies, and infectivity stability assays in an enhanced range of cell and epithelial culture models. In almost all models, VOC Alpha spread less or equally efficiently as ancestral (B.1) SARS-CoV-2. B.1. and VOC Alpha shared similar susceptibility to serum neutralization. Despite increased relative abundance of specific sgRNAs in the context of VOC Alpha infection, immune gene expression in infected cells did not differ between VOC Alpha and B.1. However, inferior spreading and entry efficiencies of VOC Alpha corresponded to lower abundance of proteolytically cleaved spike products presumably linked to the T716I mutation. In addition, we identified a bronchial cell line, NCI-H1299, which supported 24-fold increased growth of VOC Alpha and is to our knowledge the only cell line to recapitulate the fitness advantage of VOC Alpha compared to B.1. Interestingly, also VOC Delta showed a strong (595-fold) fitness advantage over B.1 in these cells. Comparative analysis of chimeric viruses expressing VOC Alpha spike in the backbone of B.1, and vice versa, showed that the specific replication phenotype of VOC Alpha in NCI-H1299 cells is largely determined by its spike protein. Despite undetectable ACE2 protein expression in NCI-H1299 cells, CRISPR/Cas9 knock-out and antibody-mediated blocking experiments revealed that multicycle spread of B.1 and VOC Alpha required ACE2 expression. Interestingly, entry of VOC Alpha, as opposed to B.1 virions, was largely unaffected by treatment with exogenous trypsin or saliva prior to infection, suggesting enhanced resistance of VOC Alpha spike to premature proteolytic cleavage in the extracellular environment of the human respiratory tract. This property may result in delayed degradation of VOC Alpha particle infectivity in conditions typical of mucosal fluids of the upper respiratory tract that may be recapitulated in NCI-H1299 cells closer than in highly ACE2-expressing cell lines and models. Our study highlights the importance of cell model evaluation and comparison for in-depth characterization of virus variant-specific phenotypes and uncovers a fine-tuned interrelationship between VOC Alpha- and host cell-specific determinants that may underlie the increased and prolonged virus shedding detected in patients infected with VOC Alpha., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: Technische Universität Berlin, Freie Universität Berlin and Charité - Universitätsmedizin have filed a patent application for siRNAs inhibiting SARS-CoV-2 replication with DN as co-author. MAMü and VMC are named together with Charité - Universitätsmedizin Berlin and Euroimmun Medizinische Labordiagnostika AG on a patent application (EP3715847) filed recently regarding the diagnostic of SARS-CoV-2 by antibody testing. The other authors declare no competing interests., (Copyright: © 2022 Niemeyer et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)
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- 2022
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42. Fast-forwarding evolution-Accelerated adaptation in a proofreading-deficient hypermutator herpesvirus.
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Xing N, Höfler T, Hearn CJ, Nascimento M, Camps Paradell G, McMahon DP, Kunec D, Osterrieder N, Cheng HH, and Trimpert J
- Abstract
Evolution relies on the availability of genetic diversity for fitness-based selection. However, most deoxyribonucleic acid (DNA) viruses employ DNA polymerases (Pol) capable of exonucleolytic proofreading to limit mutation rates during DNA replication. The relative genetic stability produced by high-fidelity genome replication can make studying DNA virus adaptation and evolution an intensive endeavor, especially in slowly replicating viruses. Here, we present a proofreading-impaired Pol mutant (Y547S) of Marek's disease virus that exhibits a hypermutator phenotype while maintaining unimpaired growth in vitro and wild-type (WT)-like pathogenicity in vivo . At the same time, mutation frequencies observed in Y547S virus populations are 2-5-fold higher compared to the parental WT virus. We find that Y547S adapts faster to growth in originally non-permissive cells, evades pressure conferred by antiviral inhibitors more efficiently, and is more easily attenuated by serial passage in cultured cells compared to WT. Our results suggest that hypermutator viruses can serve as a tool to accelerate evolutionary processes and help identify key genetic changes required for adaptation to novel host cells and resistance to antiviral therapy. Similarly, the rapid attenuation achieved through adaptation of hypermutators to growth in cell culture enables identification of genetic changes underlying attenuation and virulence, knowledge that could practically exploited, e.g. in the rational design of vaccines., (© The Author(s) 2022. Published by Oxford University Press.)
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- 2022
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43. Prevalence of anti-severe acute respiratory syndrome coronavirus 2 antibodies in cats in Germany and other European countries in the early phase of the coronavirus disease-19 pandemic.
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Adler JM, Weber C, Wernike K, Michelitsch A, Friedrich K, Trimpert J, Beer M, Kohn B, Osterrieder K, and Müller E
- Subjects
- Animals, Antibodies, Viral, Cats, Enzyme-Linked Immunosorbent Assay methods, Enzyme-Linked Immunosorbent Assay veterinary, Humans, Pandemics, Prevalence, SARS-CoV-2, COVID-19 epidemiology, COVID-19 veterinary, Cat Diseases epidemiology
- Abstract
During the first months of the coronavirus disease (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), cases of human-to-cat transmission were reported. Seroconversion was shown in cats infected under experimental and natural conditions. This large-scale survey of 1,005 serum samples was conducted to investigate anti-SARS-CoV-2 antibody prevalence in domestic cats during the first 7 months of the pandemic in Germany and other European countries. In addition, we compared the sensitivity and specificity of two multispecies SARS-CoV-2 antibody enzyme-linked immunosorbent assays (ELISA). Results were confirmed by using an indirect immunofluorescence test (iIFT) and a surrogate virus neutralization test (sVNT). Sera that were highly positive for feline coronavirus (FCoV) antibodies (n = 103) were included to correct for cross-reactivity of the tests used. Our results showed an overall SARS-CoV-2 seropositivity of 1.9% (n = 19) in a receptor-binding domain (RBD)-based ELISA, additional 0.8% (n = 8) were giving inconclusive results. In contrast, a nucleocapsid-based ELISA revealed 0.5% (n = 5) positive and 0.2% (n = 2) inconclusive results. While the iIFT and sVNT confirmed 100% of positive and 50%-57.1% of the doubtful results as determined in the RBD ELISA, the nucleocapsid-based assay showed a high discrepancy and only one of the five positive results could be confirmed. The results indicate significant deficits of the nucleocapsid-based ELISA with respect to sensitivity and specificity. Due to a significantly higher rate (5.8%) of positive results in the group of highly FCoV antibody-positive samples, cross-reactivity of the FCoV-ELISA with SARS-CoV-2 antibodies cannot be excluded. Furthermore, we investigated the impact of direct contact of domestic cats (n = 23) to SARS-CoV-2 positive owners. Considering one inconclusive result, which got confirmed by iIFT, this exposure did not lead to a significantly higher prevalence (4.4%; p = .358) among tested subjects. Overall, we conclude that cats are a negligible entity with respect to virus transmission in Europe., (© 2022 The Authors. Zoonoses and Public Health published by Wiley-VCH GmbH.)
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- 2022
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44. Engineering and Characterization of Avian Coronavirus Mutants Expressing Fluorescent Reporter Proteins from the Replicase Gene.
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Xing N, Wang Z, Wang J, Nascimento M, Jongkaewwattana A, Trimpert J, Osterrieder N, and Kunec D
- Subjects
- Animals, Chickens, Genes, Reporter, Green Fluorescent Proteins, Peptide Hydrolases, Polyproteins, RNA, Viral genetics, Coronavirus Infections veterinary, Infectious bronchitis virus genetics, Reverse Genetics
- Abstract
Infectious bronchitis virus (IBV) is an avian coronavirus that causes infectious bronchitis, an acute and highly contagious respiratory disease of chickens. IBV evolution under the pressure of comprehensive and widespread vaccination requires surveillance for vaccine resistance, as well as periodic vaccine updates. Reverse genetics systems are very valuable tools in virology, as they facilitate rapid genetic manipulation of viral genomes, thereby advancing basic and applied research. We report here the construction of an infectious clone of IBV strain Beaudette as a bacterial artificial chromosome (BAC). The engineered full-length IBV clone allowed the rescue of an infectious virus that was phenotypically indistinguishable from the parental virus. We used the infectious IBV clone and examined whether an enhanced green fluorescent protein (EGFP) can be produced by the replicase gene ORF1 and autocatalytically released from the replicase polyprotein through cleavage by the main coronavirus protease. We show that IBV tolerates insertion of the EGFP ORF at the 3' end of the replicase gene, between the sequences encoding nsp13 and nsp16 (helicase, RNA exonuclease, RNA endonuclease, and RNA methyltransferase). We further show that EGFP is efficiently cleaved from the replicase polyprotein and can be localized in double-membrane vesicles along with viral RNA polymerase and double-stranded RNA, an intermediate of IBV genome replication. One of the engineered reporter EGFP viruses were genetically stable during passage in cultured cells. We demonstrate that the reporter EGFP viruses can be used to study virus replication in host cells and for antiviral drug discovery and development of diagnostic assays. IMPORTANCE Reverse genetics systems based on bacterial artificial chromosomes (BACs) are the most valuable systems in coronavirus research. Here, we describe the establishment of a reverse genetics system for the avian coronavirus strain Beaudette, the most intensively studied strain. We cloned a copy of the avian coronavirus genome into a BAC vector and recovered infectious virus in permissive cells. We used the new system to construct reporter viruses that produce enhanced green fluorescent protein (EGFP). The EGFP coding sequence was inserted into 11 known cleavage sites of the major coronavirus protease in the replicase gene ORF1. Avian coronavirus tolerated the insertion of the EGFP coding sequence at three sites. The engineered reporter viruses replicated with parental efficiency in cultured cells and were sufficiently genetically stable. The new system facilitates functional genomics of the avian coronavirus genome but can also be used for the development of novel vaccines and anticoronaviral drugs.
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- 2022
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45. Body temperature variation controls pre-mRNA processing and transcription of antiviral genes and SARS-CoV-2 replication.
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Los B, Preußner M, Eschke K, Vidal RM, Abdelgawad A, Olofsson D, Keiper S, Paulo-Pedro M, Grindel A, Meinke S, Trimpert J, and Heyd F
- Subjects
- Child, Humans, Aged, RNA Precursors genetics, Body Temperature, SARS-CoV-2 genetics, COVID-19 genetics
- Abstract
Antiviral innate immunity represents the first defense against invading viruses and is key to control viral infections, including SARS-CoV-2. Body temperature is an omnipresent variable but was neglected when addressing host defense mechanisms and susceptibility to SARS-CoV-2 infection. Here, we show that increasing temperature in a 1.5°C window, between 36.5 and 38°C, strongly increases the expression of genes in two branches of antiviral immunity, nitric oxide production and type I interferon response. We show that alternative splicing coupled to nonsense-mediated decay decreases STAT2 expression in colder conditions and suggest that increased STAT2 expression at elevated temperature induces the expression of diverse antiviral genes and SARS-CoV-2 restriction factors. This cascade is activated in a remarkably narrow temperature range below febrile temperature, which reflects individual, circadian and age-dependent variation. We suggest that decreased body temperature with aging contributes to reduced expression of antiviral genes in older individuals. Using cell culture and in vivo models, we show that higher body temperature correlates with reduced SARS-CoV-2 replication, which may affect the different vulnerability of children versus seniors toward severe SARS-CoV-2 infection. Altogether, our data connect body temperature and pre-mRNA processing to provide new mechanistic insight into the regulation of antiviral innate immunity., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)
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- 2022
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46. De Novo-Whole Genome Assembly of the Roborovski Dwarf Hamster (Phodopus roborovskii) Genome: An Animal Model for Severe/Critical COVID-19.
- Author
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Andreotti S, Altmüller J, Quedenau C, Borodina T, Nouailles G, Teixeira Alves LG, Landthaler M, Bieniara M, Trimpert J, and Wyler E
- Subjects
- Animals, Cricetinae, Ferrets, Humans, Mice, Models, Animal, Rats, COVID-19 genetics, Phodopus
- Abstract
The Roborovski dwarf hamster Phodopus roborovskii belongs to the Phodopus genus, one of the seven within Cricetinae subfamily. Like other rodents such as mice, rats, or ferrets, hamsters can be important animal models for a range of diseases. Whereas the Syrian hamster from the genus Mesocricetus is now widely used as a model for mild-to-moderate coronavirus disease 2019, Roborovski dwarf hamster shows a severe-to-lethal course of disease upon infection with the novel human coronavirus severe acute respiratory syndrome coronavirus 2., (© The Author(s) 2022. Published by Oxford University Press on behalf of Society for Molecular Biology and Evolution.)
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- 2022
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47. Hamster models of COVID-19 pneumonia reviewed: How human can they be?
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Gruber AD, Firsching TC, Trimpert J, and Dietert K
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- Animals, Cricetinae, Humans, Mice, Disease Models, Animal, Lung, Mesocricetus, Pandemics, Respiratory System, SARS-CoV-2, COVID-19 veterinary
- Abstract
The dramatic global consequences of the coronavirus disease 2019 (COVID-19) pandemic soon fueled quests for a suitable model that would facilitate the development and testing of therapies and vaccines. In contrast to other rodents, hamsters are naturally susceptible to infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and the Syrian hamster ( Mesocricetus auratus ) rapidly developed into a popular model. It recapitulates many characteristic features as seen in patients with a moderate, self-limiting course of the disease such as specific patterns of respiratory tract inflammation, vascular endothelialitis, and age dependence. Among 4 other hamster species examined, the Roborovski dwarf hamster ( Phodopus roborovskii ) more closely mimics the disease in highly susceptible patients with frequent lethal outcome, including devastating diffuse alveolar damage and coagulopathy. Thus, different hamster species are available to mimic different courses of the wide spectrum of COVID-19 manifestations in humans. On the other hand, fewer diagnostic tools and information on immune functions and molecular pathways are available than in mice, which limits mechanistic studies and inference to humans in several aspects. Still, under pandemic conditions with high pressure on progress in both basic and clinically oriented research, the Syrian hamster has turned into the leading non-transgenic model at an unprecedented pace, currently used in innumerable studies that all aim to combat the impact of the virus with its new variants of concern. As in other models, its strength rests upon a solid understanding of its similarities to and differences from the human disease, which we review here.
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- 2022
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48. ADAM10 and ADAM17 promote SARS-CoV-2 cell entry and spike protein-mediated lung cell fusion.
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Jocher G, Grass V, Tschirner SK, Riepler L, Breimann S, Kaya T, Oelsner M, Hamad MS, Hofmann LI, Blobel CP, Schmidt-Weber CB, Gokce O, Jakwerth CA, Trimpert J, Kimpel J, Pichlmair A, and Lichtenthaler SF
- Subjects
- ADAM10 Protein genetics, ADAM17 Protein, Amyloid Precursor Protein Secretases genetics, Angiotensin-Converting Enzyme 2, Cell Fusion, Humans, Lung, Membrane Proteins genetics, Membrane Proteins metabolism, Metalloproteases, Spike Glycoprotein, Coronavirus genetics, Spike Glycoprotein, Coronavirus metabolism, Virus Internalization, COVID-19, SARS-CoV-2
- Abstract
The severe-acute-respiratory-syndrome-coronavirus-2 (SARS-CoV-2) is the causative agent of COVID-19, but host cell factors contributing to COVID-19 pathogenesis remain only partly understood. We identify the host metalloprotease ADAM17 as a facilitator of SARS-CoV-2 cell entry and the metalloprotease ADAM10 as a host factor required for lung cell syncytia formation, a hallmark of COVID-19 pathology. ADAM10 and ADAM17, which are broadly expressed in the human lung, cleave the SARS-CoV-2 spike protein (S) in vitro, indicating that ADAM10 and ADAM17 contribute to the priming of S, an essential step for viral entry and cell fusion. ADAM protease-targeted inhibitors severely impair lung cell infection by the SARS-CoV-2 variants of concern alpha, beta, delta, and omicron and also reduce SARS-CoV-2 infection of primary human lung cells in a TMPRSS2 protease-independent manner. Our study establishes ADAM10 and ADAM17 as host cell factors for viral entry and syncytia formation and defines both proteases as potential targets for antiviral drug development., (© 2022 The Authors. Published under the terms of the CC BY 4.0 license.)
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- 2022
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49. The Diverse Major Histocompatibility Complex Haplotypes of a Common Commercial Chicken Line and Their Effect on Marek's Disease Virus Pathogenesis and Tumorigenesis.
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Bertzbach LD, Tregaskes CA, Martin RJ, Deumer US, Huynh L, Kheimar AM, Conradie AM, Trimpert J, Kaufman J, and Kaufer BB
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- Animals, Carcinogenesis genetics, Chickens genetics, Disease Resistance genetics, Haplotypes, Histocompatibility Antigens, Major Histocompatibility Complex genetics, Herpesvirus 2, Gallid genetics, Marek Disease
- Abstract
The major histocompatibility complex (MHC) is crucial for appropriate immune responses against invading pathogens. Chickens possess a single predominantly-expressed class I molecule with strong associations between disease resistance and MHC haplotype. For Marek's disease virus (MDV) infections of chickens, the MHC haplotype is one of the major determinants of genetic resistance and susceptibility. VALO specific pathogen free (SPF) chickens are widely used in biomedical research and vaccine production. While valuable findings originate from MDV infections of VALO SPF chickens, their MHC haplotypes and associated disease resistance remained elusive. In this study, we used several typing systems to show that VALO SPF chickens possess MHC haplotypes that include B9, B9:02, B15, B19 and B21 at various frequencies. Moreover, we associate the MHC haplotypes to MDV-induced disease and lymphoma formation and found that B15 homozygotes had the lowest tumor incidence while B21 homozygotes had the lowest number of organs with tumors. Finally, we found transmission at variable levels to all contact birds except B15/B21 heterozygotes. These data have immediate implications for the use of VALO SPF chickens and eggs in the life sciences and add another piece to the puzzle of the chicken MHC complex and its role in infections with this oncogenic herpesvirus., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Bertzbach, Tregaskes, Martin, Deumer, Huynh, Kheimar, Conradie, Trimpert, Kaufman and Kaufer.)
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- 2022
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50. Key benefits of dexamethasone and antibody treatment in COVID-19 hamster models revealed by single-cell transcriptomics.
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Wyler E, Adler JM, Eschke K, Teixeira Alves G, Peidli S, Pott F, Kazmierski J, Michalick L, Kershaw O, Bushe J, Andreotti S, Pennitz P, Abdelgawad A, Postmus D, Goffinet C, Kreye J, Reincke SM, Prüss H, Blüthgen N, Gruber AD, Kuebler WM, Witzenrath M, Landthaler M, Nouailles G, and Trimpert J
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- Animals, Anti-Inflammatory Agents pharmacology, Anti-Inflammatory Agents therapeutic use, Antibodies, Viral, Antiviral Agents, Cricetinae, Dexamethasone pharmacology, SARS-CoV-2, Transcriptome, COVID-19 Drug Treatment
- Abstract
For coronavirus disease 2019 (COVID-19), effective and well-understood treatment options are still scarce. Since vaccine efficacy is challenged by novel variants, short-lasting immunity, and vaccine hesitancy, understanding and optimizing therapeutic options remains essential. We aimed at better understanding the effects of two standard-of-care drugs, dexamethasone and anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies, on infection and host responses. By using two COVID-19 hamster models, pulmonary immune responses were analyzed to characterize effects of single or combinatorial treatments. Pulmonary viral burden was reduced by anti-SARS-CoV-2 antibody treatment and unaltered or increased by dexamethasone alone. Dexamethasone exhibited strong anti-inflammatory effects and prevented fulminant disease in a severe disease model. Combination therapy showed additive benefits with both anti-viral and anti-inflammatory potency. Bulk and single-cell transcriptomic analyses confirmed dampened inflammatory cell recruitment into lungs upon dexamethasone treatment and identified a specifically responsive subpopulation of neutrophils, thereby indicating a potential mechanism of action. Our analyses confirm the anti-inflammatory properties of dexamethasone and suggest possible mechanisms, validate anti-viral effects of anti-SARS-CoV-2 antibody treatment, and reveal synergistic effects of a combination therapy, thus informing more effective COVID-19 therapies., Competing Interests: Declaration of interests Related to this work, the German Center for Neurodegenerative Diseases (DZNE) and Charité-Universitätsmedizin Berlin have filed a patent application on which J. Kreye., S.M.R., and H.P. are named as inventors of mAb CV07-209., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)
- Published
- 2022
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