84 results on '"Rafael Elias Marques"'
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2. Two +ssRNA mycoviruses cohabiting the fungal cultivar of leafcutter ants
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Asta Rødsgaard-Jørgensen, Caio Ambrosio Leal-Dutra, Sabrina Ferreira de Santana, Asger Roland Jensen, Rafael Elias Marques, Eric Roberto Guimarães Rocha Aguiar, and Jonathan Zvi Shik
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Botourmiaviridae ,Fungus-farming ant ,Leafcutter ant ,Leucoagaricus gongylophorus ,Magoulivirus ,Ourmia-like virus ,Infectious and parasitic diseases ,RC109-216 - Abstract
Abstract Leafcutter ants are dominant herbivores in the Neotropics and rely on a fungus (Leucoagaricus gongylophorus) to transform freshly gathered leaves into a source of nourishment rather than consuming the vegetation directly. Here we report two virus-like particles that were isolated from L. gongylophorus and observed using transmission electron microscopy. RNA sequencing identified two +ssRNA mycovirus strains, Leucoagaricus gongylophorus tymo-like virus 1 (LgTlV1) and Leucoagaricus gongylophorus magoulivirus 1 (LgMV1). Genome annotation of LgTlV1 (7401 nt) showed conserved domains for methyltransferase, endopeptidase, viral RNA helicase, and RNA-dependent RNA polymerase (RdRp). The smaller genome of LgMV1 (2636 nt) contains one open reading frame encoding an RdRp. While we hypothesize these mycoviruses function as symbionts in leafcutter farming systems, further study will be needed to test whether they are mutualists, commensals, or parasites.
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- 2024
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3. The seroconversion history to SARS-CoV-2 in Indigenous people from Brazil – the interplay between exposure, vaccination, and tuberculosis
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Alice Nagai, Renan Barbosa Lemes, José Geraldo Mill, Alexandre Costa Pereira, Rafael Elias Marques, and Tábita Hünemeier
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COVID-19 ,humoral response ,tuberculosis ,vaccination ,Brazilians ,Indigenous ,Immunologic diseases. Allergy ,RC581-607 - Abstract
The COVID-19 pandemic caused a significant loss of human lives and a worldwide decline in quality of life. Although our understanding of the pandemic has improved significantly since the beginning, the natural history of COVID-19 and its impacts on under-represented populations, such as Indigenous people from America, remain largely unknown. We performed a retrospective serological survey with two Brazilian Indigenous populations (n=624), Tupiniquim and Guarani-Mbyá. Samples were collected between September 2020 and July 2021: a period comprising the dissemination of SARS-CoV-2 variants and the beginning of COVID-19 vaccination in Brazil. Seroconversions against S and N antigens were assessed using three different commercially available ELISA kits. Samples were also used to assess the prevalence of tuberculosis (TB) in the same population (n=529). Seroconversion against SARS-CoV-2 antigens was considered positive if at least one of the three ELISA kits detected levels of specific antibodies above the threshold specified by the manufacturer. In this sense, we report 56.0% (n=349/623) of seroconverted individuals. Relative seroconversion peaked after introduction of the Coronavac vaccine in February 2021. Vaccination increased the production of anti-S IgG from 3.9% to 48.6%. Our results also indicated that 11.0% (n=46/417) of all individuals were positive for TB. Seroconversion to SARS-CoV-2 was similar between individuals with positive tuberculosis test results to those with negative test results. Most vaccinated individuals seroconverted to SARS-CoV-2, indicating that Coronavac may be as protective in individuals from these indigenous groups as observed in the general Brazilian population. COVID-19 severity was minimal regardless of incomplete vaccine coverage, suggesting that vaccination may not be the only factor protecting individuals from severe COVID-19. Tuberculosis is highly prevalent and not associated with increased seroconversion to SARS-CoV-2.
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- 2024
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4. Structural Insights into Plasticity and Discovery of Flavonoid Allosteric Inhibitors of Flavivirus NS2B–NS3 Protease
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Marielena Vogel Saivish, Gabriela de Lima Menezes, Vivaldo Gomes da Costa, Liliane Nebo, Gislaine Celestino Dutra da Silva, Carolina Colombelli Pacca, Rafael Elias Marques, Maurício Lacerda Nogueira, and Roosevelt Alves Da Silva
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flaviviral protease inhibitors ,Rocio virus ,Ilhéus virus ,Saint Louis encephalitis virus ,yellow fever virus ,3D pharmacophores ,Biology (General) ,QH301-705.5 - Abstract
Flaviviruses are among the most critical pathogens in tropical regions; they cause various severe diseases in developing countries but are not restricted to these countries. The development of antiviral therapeutics is crucial for managing flavivirus outbreaks. Ten proteins are encoded in the flavivirus RNA. The N2B–NS3pro protein complex plays a fundamental role in flavivirus replication and is a promising drug target; however, no flavivirus protease inhibitors have progressed to the preclinical stage. This study analyzed the structural models and plasticity of the NS2B–NS3pro protein complex of five medically important non-dengue flaviviruses (West Nile, Rocio, Ilhéus, yellow fever, and Saint Louis encephalitis). The flavonoids amentoflavone, tetrahydrorobustaflavone, and quercetin were selected for their exceptional binding energies as potential inhibitors of the NS2B–NS3pro protein complex. AutoDock Vina results ranged from −7.0 kcal/mol to −11.5 kcal/mol and the compounds preferentially acted non-competitively. Additionally, the first structural model for the NS2B–NS3pro protein complex was proposed for Ilhéus and Rocio viruses. The NS2B–NS3pro protease is an attractive molecular target for drug development. The three identified natural flavonoids showed great inhibitory potential against the viral species. Nevertheless, further in silico and in vitro studies are required to obtain more information regarding NS2B–NS3pro inhibition by these flavonoids and their therapeutic potential.
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- 2023
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5. Identification and characterization of the anti-SARS-CoV-2 activity of cationic amphiphilic steroidal compounds
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Alexandre Borin, Laís D. Coimbra, Karina Bispo-dos-Santos, Fabrício F. Naciuk, Marina Fontoura, Camila L. Simeoni, Giovanni F. Gomes, Mariene R. Amorim, Humberto D. Gravina, Jacqueline Farinha Shimizu, Amanda S. C. Passos, Isadora M. de Oliveira, Ana Carolina de Carvalho, Alisson Campos Cardoso, Pierina L. Parise, Daniel A. Toledo-Teixeira, Giuliana E. Sotorilli, Gabriela F. Persinoti, Ingra Morales Claro, Ester C. Sabino, Marcos R. Alborghetti, Silvana A. Rocco, Kleber G. Franchini, William M. de Souza, Paulo S. L. Oliveira, Thiago M. Cunha, Fabiana Granja, José Luiz Proença-Módena, Daniela B.B. Trivella, Marjorie Bruder, Artur T. Cordeiro, and Rafael Elias Marques
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SARS-CoV-2 ,drug discovery ,steroidal compounds ,antiviral activity ,Infectious and parasitic diseases ,RC109-216 - Abstract
The ongoing COVID-19 pandemic caused a significant loss of human lives and a worldwide decline in quality of life. Treatment of COVID-19 patients is challenging, and specific treatments to reduce COVID-19 aggravation and mortality are still necessary. Here, we describe the discovery of a novel class of epiandrosterone steroidal compounds with cationic amphiphilic properties that present antiviral activity against SARS-CoV-2 in the low micromolar range. Compounds were identified in screening campaigns using a cytopathic effect-based assay in Vero CCL81 cells, followed by hit compound validation and characterization. Compounds LNB167 and LNB169 were selected due to their ability to reduce the levels of infectious viral progeny and viral RNA levels in Vero CCL81, HEK293, and HuH7.5 cell lines. Mechanistic studies in Vero CCL81 cells indicated that LNB167 and LNB169 inhibited the initial phase of viral replication through mechanisms involving modulation of membrane lipids and cholesterol in host cells. Selection of viral variants resistant to steroidal compound treatment revealed single mutations on transmembrane, lipid membrane-interacting Spike and Envelope proteins. Finally, in vivo testing using the hACE2 transgenic mouse model indicated that SARS-CoV-2 infection could not be ameliorated by LNB167 treatment. We conclude that anti-SARS-CoV-2 activities of steroidal compounds LNB167 and LNB169 are likely host-targeted, consistent with the properties of cationic amphiphilic compounds that modulate host cell lipid biology. Although effective in vitro, protective effects were cell-type specific and did not translate to protection in vivo, indicating that subversion of lipid membrane physiology is an important, yet complex mechanism involved in SARS-CoV-2 replication and pathogenesis.
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- 2022
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6. Discovery and structural characterization of chicoric acid as a SARS-CoV-2 nucleocapsid protein ligand and RNA binding disruptor
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Gustavo Fernando Mercaldi, Eduardo Henrique Salviano Bezerra, Fernanda Aparecida Heleno Batista, Celisa Caldana Costa Tonoli, Adriana Santos Soprano, Jacqueline Farinha Shimizu, Alice Nagai, Jaqueline Cristina da Silva, Helder Veras Ribeiro Filho, Jéssica do Nascimento Faria, Marcos Guilherme da Cunha, Ana Carolina Mattos Zeri, Andrey Fabricio Ziem Nascimento, José Luiz Proenca-Modena, Marcio Chaim Bajgelman, Silvana Aparecida Rocco, Paulo Sérgio Lopes-de-Oliveira, Artur Torres Cordeiro, Marjorie Bruder, Rafael Elias Marques, Mauricio Luis Sforça, Kleber Gomes Franchini, Celso Eduardo Benedetti, Ana Carolina Migliorini Figueira, and Daniela Barretto Barbosa Trivella
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Medicine ,Science - Abstract
Abstract The nucleocapsid (N) protein plays critical roles in coronavirus genome transcription and packaging, representing a key target for the development of novel antivirals, and for which structural information on ligand binding is scarce. We used a novel fluorescence polarization assay to identify small molecules that disrupt the binding of the N protein to a target RNA derived from the SARS-CoV-2 genome packaging signal. Several phenolic compounds, including L-chicoric acid (CA), were identified as high-affinity N-protein ligands. The binding of CA to the N protein was confirmed by isothermal titration calorimetry, 1H-STD and 15N-HSQC NMR, and by the crystal structure of CA bound to the N protein C-terminal domain (CTD), further revealing a new modulatory site in the SARS-CoV-2 N protein. Moreover, CA reduced SARS-CoV-2 replication in cell cultures. These data thus open venues for the development of new antivirals targeting the N protein, an essential and yet underexplored coronavirus target.
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- 2022
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7. SARS-CoV-2 infects adipose tissue in a fat depot- and viral lineage-dependent manner
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Tatiana Dandolini Saccon, Felippe Mousovich-Neto, Raissa Guimarães Ludwig, Victor Corasolla Carregari, Ana Beatriz dos Anjos Souza, Amanda Stephane Cruz dos Passos, Matheus Cavalheiro Martini, Priscilla Paschoal Barbosa, Gabriela Fabiano de Souza, Stéfanie Primon Muraro, Julia Forato, Mariene Ribeiro Amorim, Rafael Elias Marques, Flavio Protasio Veras, Ester Barreto, Tiago Tomazini Gonçalves, Isadora Marques Paiva, Narayana P. B. Fazolini, Carolina Mie Kawagosi Onodera, Ronaldo Bragança Martins Junior, Paulo Henrique Cavalcanti de Araújo, Sabrina Setembre Batah, Rosa Maria Mendes Viana, Danilo Machado de Melo, Alexandre Todorovic Fabro, Eurico Arruda, Fernando Queiroz Cunha, Thiago Mattar Cunha, Marco Antônio M. Pretti, Bradley Joseph Smith, Henrique Marques-Souza, Thiago L. Knittel, Gabriel Palermo Ruiz, Gerson S. Profeta, Tereza Cristina Minto Fontes-Cal, Mariana Boroni, Marco Aurélio Ramirez Vinolo, Alessandro S. Farias, Pedro Manoel M. Moraes-Vieira, Joyce Maria Annichino Bizzacchi, Tambet Teesalu, Felipe David Mendonça Chaim, Everton Cazzo, Elinton Adami Chaim, José Luiz Proença-Módena, Daniel Martins-de-Souza, Mariana Kiomy Osako, Luiz Osório Leiria, and Marcelo A. Mori
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Science - Abstract
Visceral adiposity is a risk factor for severe COVID-19, and infection of adipose tissue by SARS-CoV-2 has been reported. Here the authors confirm that human adipose tissue is a possible site for SARS-CoV-2 infection, but the degree of adipose tissue infection and the way adipocytes respond to the virus depend on the adipose tissue depot and the viral strain.
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- 2022
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8. The E2 glycoprotein holds key residues for Mayaro virus adaptation to the urban Aedes aegypti mosquito.
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Chelsea Cereghino, Ferdinand Roesch, Lucía Carrau, Alexandra Hardy, Helder V Ribeiro-Filho, Annabelle Henrion-Lacritick, Cassandra Koh, Jeffrey M Marano, Tyler A Bates, Pallavi Rai, Christina Chuong, Shamima Akter, Thomas Vallet, Hervé Blanc, Truitt J Elliott, Anne M Brown, Pawel Michalak, Tanya LeRoith, Jesse D Bloom, Rafael Elias Marques, Maria-Carla Saleh, Marco Vignuzzi, and James Weger-Lucarelli
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Immunologic diseases. Allergy ,RC581-607 ,Biology (General) ,QH301-705.5 - Abstract
Adaptation to mosquito vectors suited for transmission in urban settings is a major driver in the emergence of arboviruses. To better anticipate future emergence events, it is crucial to assess their potential to adapt to new vector hosts. In this work, we used two different experimental evolution approaches to study the adaptation process of an emerging alphavirus, Mayaro virus (MAYV), to Ae. aegypti, an urban mosquito vector of many other arboviruses. We identified E2-T179N as a key mutation increasing MAYV replication in insect cells and enhancing transmission after escaping the midgut of live Ae. aegypti. In contrast, this mutation decreased viral replication and binding in human fibroblasts, a primary cellular target of MAYV in humans. We also showed that MAYV E2-T179N generates reduced viremia and displays less severe tissue pathology in vivo in a mouse model. We found evidence in mouse fibroblasts that MAYV E2-T179N is less dependent on the Mxra8 receptor for replication than WT MAYV. Similarly, exogenous expression of human apolipoprotein receptor 2 and Mxra8 enhanced WT MAYV replication compared to MAYV E2-T179N. When this mutation was introduced in the closely related chikungunya virus, which has caused major outbreaks globally in the past two decades, we observed increased replication in both human and insect cells, suggesting E2 position 179 is an important determinant of alphavirus host-adaptation, although in a virus-specific manner. Collectively, these results indicate that adaptation at the T179 residue in MAYV E2 may result in increased vector competence-but coming at the cost of optimal replication in humans-and may represent a first step towards a future emergence event.
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- 2023
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9. Phenotypical Screening of an MMV Open Box Library and Identification of Compounds with Antiviral Activity against St. Louis Encephalitis Virus
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Giuliana Eboli Sotorilli, Humberto Doriguetto Gravina, Ana Carolina de Carvalho, Jacqueline Farinha Shimizu, Marina Alves Fontoura, Talita Diniz Melo-Hanchuk, Artur Torres Cordeiro, and Rafael Elias Marques
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St. Louis encephalitis virus ,cell culture-based screening ,drug repurposing ,Flavivirus ,cationic amphiphilic compounds (CAD) ,Microbiology ,QR1-502 - Abstract
St. Louis encephalitis virus (SLEV) is a neglected mosquito-borne Flavivirus that may cause severe neurological disease in humans and other animals. There are no specific treatments against SLEV infection or disease approved for human use, and drug repurposing may represent an opportunity to accelerate the development of treatments against SLEV. Here we present a scalable, medium-throughput phenotypic cell culture-based screening assay on Vero CCL81 cells to identify bioactive compounds that could be repurposed against SLEV infection. We screened eighty compounds from the Medicines for Malaria Venture (MMV) COVID Box library to identify nine (11%) compounds that protected cell cultures from SLEV-induced cytopathic effects, with low- to mid-micromolar potencies. We validated six hit compounds using viral plaque-forming assays to find that the compounds ABT-239, Amiodarone, Fluphenazine, Posaconazole, Triparanol, and Vidofludimus presented varied levels of antiviral activity and selectivity depending on the mammalian cell type used for testing. Importantly, we identified and validated the antiviral activity of the anti-flavivirus nucleoside analog 7DMA against SLEV. Triparanol and Fluphenazine reduced infectious viral loads in both Vero CCL81 and HBEC-5i cell cultures and, similar to the other validated compounds, are likely to exert antiviral activity through a molecular target in the host.
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- 2023
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10. Competing interests during the key N-glycosylation of 6-chloro-7-deaza-7-iodopurine for the synthesis of 7-deaza-2′-methyladenosine using Vorbrüggen conditions
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Fabrício Fredo Naciuk, Andrey Fabricio Ziem Nascimento, Rebeca Paiva Froes Rocha, Joane Kathelen Rustiguel, Lais Durço Coimbra, Rafael Elias Marques, and Marjorie Bruder
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7DMA ,Vorbrüggen ,N-glycosylation ,TMSOTf ,N-silyl ketene imine formation ,emerging virus ,Chemistry ,QD1-999 - Abstract
A short 3-step synthesis of the antiviral agent 7DMA is described herein. The nature of a major by-product formed during the key N-glycosylation of 6-chloro-7-deaza-7-iodopurine with perbenzoylated 2-methyl-ribose under Vorbrüggen conditions was also investigated. Spectroscopic analyses support that the solvent itself is converted into a nucleophilic species competing with the nucleobase and further reacting with the activated riboside in an unanticipated fashion. These findings call for a revision of reaction conditions when working with weakly reactive nucleobases in the presence of Lewis acids. 7DMA thus obtained was evaluated for its efficacy against an emerging flavivirus in vitro.
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- 2023
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11. Type I interferons are essential while type II interferon is dispensable for protection against St. Louis encephalitis virus infection in the mouse brain
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Rebeca Froes Rocha, Juliana L. Del Sarto, Giovanni F. Gomes, Mariana P. Gonçalves, Milene A. Rachid, Juliana H. C. Smetana, Daniele G. Souza, Mauro Martins Teixeira, and Rafael Elias Marques
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neglected arbovirus ,flavivirus ,st. louis encephalitis ,interferons ,mouse model ,Infectious and parasitic diseases ,RC109-216 - Abstract
St. Louis encephalitis virus (SLEV) is a neglected mosquito-borne flavivirus that causes severe neurological disease in humans. SLEV replication in the central nervous system (CNS) induces the local production of interferons (IFNs), which are attributed to host protection. The antiviral response to SLEV infection in the CNS is not completely understood, which led us to characterize the roles of IFNs using mouse models of St. Louis encephalitis. We infected mice deficient in type I IFN receptor (ABR−/-) or deficient in Type II IFN (IFNγ−/-) and assessed the contribution of each pathway to disease development. We found that type I and II IFNs play different roles in SLEV infection. Deficiency in type I IFN signaling was associated to an early and increased mortality, uncontrolled SLEV replication and impaired ISG expression, leading to increased proinflammatory cytokine production and brain pathology. Conversely, IFNγ−/- mice were moderately resistant to SLEV infection. IFNγ deficiency caused no changes to viral load or SLEV-induced encephalitis and did not change the expression of ISGs in the brain. We found that type I IFN is essential for the control of SLEV replication whereas type II IFN was not associated with protection in this model.
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- 2021
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12. Pediatric COVID-19 patients in South Brazil show abundant viral mRNA and strong specific anti-viral responses
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Tiago Fazolo, Karina Lima, Julia C. Fontoura, Priscila Oliveira de Souza, Gabriel Hilario, Renata Zorzetto, Luiz Rodrigues Júnior, Veridiane Maria Pscheidt, Jayme de Castilhos Ferreira Neto, Alisson F. Haubert, Izza Gambin, Aline C. Oliveira, Raissa S. Mello, Matheus de Bastos Balbe e Gutierres, Rodrigo Benedetti Gassen, Lais Durço Coimbra, Alexandre Borin, Rafael Elias Marques, Ivaine Tais Sauthier Sartor, Gabriela Oliveira Zavaglia, Ingrid Rodrigues Fernandes, Helder I. Nakaya, Fernanda Hammes Varela, Márcia Polese-Bonatto, Thiago J. Borges, Sidia Maria Callegari-Jacques, Marcela Santos Correa da Costa, Jaqueline de Araujo Schwartz, Marcelo Comerlato Scotta, Renato T. Stein, and Cristina Bonorino
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Science - Abstract
Children often show milder COVID-19 symptoms, but the underlying mechanistic insights are still lacking. Here the authors profile both pediatric and adult cohorts of COVID-19 patients in Brazil to find that children exhibit higher viral load but stronger and biased cellular immunity, thereby serving clues for the differential responses in children.
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- 2021
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13. SARS-CoV-2 infected children form early immune memory responses dominated by nucleocapsid-specific CD8+ T cells and antibodies
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Karina Lima, Julia C. Fontoura, Priscila Oliveira de Souza, Tiago Fazolo, Gabriel Hilario, Renata Zorzetto, Luiz C Rodrigues Junior, Lais D. Coimbra, Alexandre Borin, Karina Bispo-dos-Santos, Fabiana Granja, Rafael Elias Marques, Gabriela Oliveira Zavaglia, Ingrid Rodrigues Fernandes, Fernanda Hammes Varela, Marcia Polese-Bonatto, Maiko Luís Tonini, Greice Madeleine Ikeda do Carmo, Walquiria Aparecida Ferreira de Almeida, Thiago J. Borges, Helder I. Nakaya, José Luiz Proenca-Modena, Sidia Maria Callegari-Jacques, Marcelo Comerlato Scotta, Renato T. Stein, and Cristina Bonorino
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Memory T cell ,antibodies ,neutralizing antibodies ,variants of concern ,N protein ,COVID-19 ,Immunologic diseases. Allergy ,RC581-607 - Abstract
This is the third year of the SARS-CoV-2 pandemic, and yet most children remain unvaccinated. COVID-19 in children manifests as mostly mild or asymptomatic, however high viral titers and strong cellular and humoral responses are observed upon acute infection. It is still unclear how long these responses persist, and if they can protect from re-infection and/or disease severity. Here, we analyzed immune memory responses in a cohort of children and adults with COVID-19. Important differences between children and adults are evident in kinetics and profile of memory responses. Children develop early N-specific cytotoxic T cell responses, that rapidly expand and dominate their immune memory to the virus. Children’s anti-N, but not anti-S, antibody titers increase over time. Neutralization titers correlate with N-specific antibodies and CD8+T cells. However, antibodies generated by infection do not efficiently cross-neutralize variants Gamma or Delta. Our results indicate that mechanisms that protect from disease severity are possibly different from those that protect from reinfection, bringing novel insights for pediatric vaccine design. They also underline the importance of vaccination in children, who remain at risk for COVID-19 despite having been previously infected.
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- 2022
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14. Cryo-EM structure of the mature and infective Mayaro virus at 4.4 Å resolution reveals features of arthritogenic alphaviruses
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Helder V. Ribeiro-Filho, Lais D. Coimbra, Alexandre Cassago, Rebeca P. F. Rocha, João Victor da Silva Guerra, Rafael de Felicio, Carolina Moretto Carnieli, Luiza Leme, Antonio Cláudio Padilha, Adriana F. Paes Leme, Daniela B. B. Trivella, Rodrigo Villares Portugal, Paulo Sérgio Lopes-de-Oliveira, and Rafael Elias Marques
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Science - Abstract
Mayaro virus (MAYV) is an emerging arbovirus in Central and South America that is transmitted by mosquitoes and causes arthritogenic disease. Here, the authors present the 4.4 Å resolution cryo-EM structure of MAYV and describe specific features of the virus, which could be exploited for the design of MAYV-specific diagnostics and therapeutics.
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- 2021
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15. Structural dynamics of SARS-CoV-2 nucleocapsid protein induced by RNA binding.
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Helder Veras Ribeiro-Filho, Gabriel Ernesto Jara, Fernanda Aparecida Heleno Batista, Gabriel Ravanhani Schleder, Celisa Caldana Costa Tonoli, Adriana Santos Soprano, Samuel Leite Guimarães, Antonio Carlos Borges, Alexandre Cassago, Marcio Chaim Bajgelman, Rafael Elias Marques, Daniela Barretto Barbosa Trivella, Kleber Gomes Franchini, Ana Carolina Migliorini Figueira, Celso Eduardo Benedetti, and Paulo Sergio Lopes-de-Oliveira
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Biology (General) ,QH301-705.5 - Abstract
The nucleocapsid (N) protein of the SARS-CoV-2 virus, the causal agent of COVID-19, is a multifunction phosphoprotein that plays critical roles in the virus life cycle, including transcription and packaging of the viral RNA. To play such diverse roles, the N protein has two globular RNA-binding modules, the N- (NTD) and C-terminal (CTD) domains, which are connected by an intrinsically disordered region. Despite the wealth of structural data available for the isolated NTD and CTD, how these domains are arranged in the full-length protein and how the oligomerization of N influences its RNA-binding activity remains largely unclear. Herein, using experimental data from electron microscopy and biochemical/biophysical techniques combined with molecular modeling and molecular dynamics simulations, we show that, in the absence of RNA, the N protein formed structurally dynamic dimers, with the NTD and CTD arranged in extended conformations. However, in the presence of RNA, the N protein assumed a more compact conformation where the NTD and CTD are packed together. We also provided an octameric model for the full-length N bound to RNA that is consistent with electron microscopy images of the N protein in the presence of RNA. Together, our results shed new light on the dynamics and higher-order oligomeric structure of this versatile protein.
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- 2022
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16. Identification of Compounds With Antiviral Activity Against SARS-CoV-2 in the MMV Pathogen Box Using a Phenotypic High-Throughput Screening Assay
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Laís D. Coimbra, Alexandre Borin, Marina Fontoura, Humberto D. Gravina, Alice Nagai, Jacqueline Farinha Shimizu, Karina Bispo-dos-Santos, Fabiana Granja, Paulo S. L. Oliveira, Kleber G. Franchini, Kirandeep Samby, Marjorie Bruder, José Luiz Proença-Módena, Daniela B. B. Trivella, Juliana H. C. Smetana, Artur T. Cordeiro, and Rafael Elias Marques
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SARS-CoV-2 ,biosafety level 3 ,MMV pathogen box ,antivirals ,drug repositioning ,high-throughput screening ,Microbiology ,QR1-502 - Abstract
Until December 2021, the COVID-19 pandemic has caused more than 5.5 million deaths. Vaccines are being deployed worldwide to mitigate severe disease and death, but continued transmission and the emergence of SARS-CoV-2 variants indicate that specific treatments against COVID-19 are still necessary. We screened 400 compounds from the Medicines for Malaria Venture (MMV) Pathogen Box seeking for molecules with antiviral activity against SARS-CoV-2 by using a high-throughput screening (HTS) infection assay in Vero CCL81 cells. On resupply of 15 selected hit compounds, we confirmed that 7 of them presented a dose-dependent cytoprotective activity against SARS-CoV-2-induced cytopathic effect (CPE) in the micromolar range. They were validated in low-throughput infection assays using four different cell lines, including the human lung Calu-3 cell line. MMV000063, MMV024937, MMV688279, and MMV688991 reduced viral load in cell culture, assessed by RT-qPCR and viral plaque assay, while MMV688279 and MMV688991 (also known as nitazoxanide) were the most promising, reducing SARS-CoV-2 load by at least 100-fold at 20 µM in almost all cell types tested. Our results indicate that active anti-SARS-CoV-2 molecules exist within the repertoire of antiviral, antiparasitic and antimicrobial compounds available to date. Although the mode of action by which MMV688279 and MMV688991 reduce SARS-CoV-2 replication is yet unknown, the fact that they were active in different cell types holds promise not only for the discovery of new therapeutic targets, but also for the development of novel antiviral medicines against COVID-19.
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- 2022
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17. Atypical response to bacterial coinfection and persistent neutrophilic bronchoalveolar inflammation distinguish critical COVID-19 from influenza
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Seppe Cambier, Mieke Metzemaekers, Ana Carolina de Carvalho, Amber Nooyens, Cato Jacobs, Lore Vanderbeke, Bert Malengier-Devlies, Mieke Gouwy, Elisabeth Heylen, Philippe Meersseman, Greet Hermans, Els Wauters, Alexander Wilmer, the CONTAGIOUS Consortium, Dominique Schols, Patrick Matthys, Ghislain Opdenakker, Rafael Elias Marques, Joost Wauters, Jennifer Vandooren, and Paul Proost
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COVID-19 ,Immunology ,Medicine - Abstract
Neutrophils are recognized as important circulating effector cells in the pathophysiology of severe coronavirus disease 2019 (COVID-19). However, their role within the inflamed lungs is incompletely understood. Here, we collected bronchoalveolar lavage (BAL) fluids and parallel blood samples of critically ill COVID-19 patients requiring invasive mechanical ventilation and compared BAL fluid parameters with those of mechanically ventilated patients with influenza, as a non–COVID-19 viral pneumonia cohort. Compared with those of patients with influenza, BAL fluids of patients with COVID-19 contained increased numbers of hyperactivated degranulating neutrophils and elevated concentrations of the cytokines IL-1β, IL-1RA, IL-17A, TNF-α, and G-CSF; the chemokines CCL7, CXCL1, CXCL8, CXCL11, and CXCL12α; and the protease inhibitors elafin, secretory leukocyte protease inhibitor, and tissue inhibitor of metalloproteinases 1. In contrast, α-1 antitrypsin levels and net proteolytic activity were comparable in COVID-19 and influenza BAL fluids. During antibiotic treatment for bacterial coinfections, increased BAL fluid levels of several activating and chemotactic factors for monocytes, lymphocytes, and NK cells were detected in patients with COVID-19 whereas concentrations tended to decrease in patients with influenza, highlighting the persistent immunological response to coinfections in COVID-19. Finally, the high proteolytic activity in COVID-19 lungs suggests considering protease inhibitors as a treatment option.
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- 2022
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18. Caffeic Acid Has Antiviral Activity against Ilhéus Virus In Vitro
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Marielena Vogel Saivish, Carolina Colombelli Pacca, Vivaldo Gomes da Costa, Gabriela de Lima Menezes, Roosevelt Alves da Silva, Liliane Nebo, Gislaine Celestino Dutra da Silva, Bruno Henrique Gonçalves de Aguiar Milhim, Igor da Silva Teixeira, Tiago Henrique, Natalia Franco Bueno Mistrão, Victor Miranda Hernandes, Nathalia Zini, Ana Carolina de Carvalho, Marina Alves Fontoura, Paula Rahal, Lívia Sacchetto, Rafael Elias Marques, and Maurício Lacerda Nogueira
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Ilhéus virus ,caffeic acid ,antiviral activity ,inhibition ,in silico analysis ,Microbiology ,QR1-502 - Abstract
Ilhéus virus (ILHV) is a neglected mosquito-borne flavivirus. ILHV infection may lead to Ilhéus fever, an emerging febrile disease like dengue fever with the potential to evolve into a severe neurological disease characterized by meningoencephalitis; no specific treatments are available for this disease. This study assessed the antiviral properties of caffeic acid, an abundant component of plant-based food products that is also compatible with the socioeconomic limitations associated with this neglected infectious disease. The in vitro activity of caffeic acid on ILHV replication was investigated in Vero and A549 cell lines using plaque assays, quantitative RT-PCR, and immunofluorescence assays. We observed that 500 µM caffeic acid was virucidal against ILHV. Molecular docking indicated that caffeic acid might interact with an allosteric binding site on the envelope protein.
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- 2023
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19. Kinetics of peripheral blood neutrophils in severe coronavirus disease 2019
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Mieke Metzemaekers, Seppe Cambier, Marfa Blanter, Jennifer Vandooren, Ana Carolina deCarvalho, Bert Malengier‐Devlies, Lore Vanderbeke, Cato Jacobs, Sofie Coenen, Erik Martens, Noëmie Pörtner, Lotte Vanbrabant, Pierre Van Mol, Yannick Van Herck, Nathalie Van Aerde, Greet Hermans, Jan Gunst, Alexandre Borin, Bruna Toledo N Pereira, Arilson Bernardo dosSP Gomes, Stéfanie Primon Muraro, Gabriela Fabiano de Souza, Alessandro S Farias, José Luiz Proenca‐Modena, Marco Aurélio R Vinolo, the CONTAGIOUS Consortium, Pedro Elias Marques, Carine Wouters, Els Wauters, Sofie Struyf, Patrick Matthys, Ghislain Opdenakker, Rafael Elias Marques, Joost Wauters, Mieke Gouwy, and Paul Proost
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neutrophil ,COVID‐19 ,chemokine ,cytokine ,protease ,emergency myelopoiesis ,Immunologic diseases. Allergy ,RC581-607 - Abstract
Abstract Objectives Emerging evidence of dysregulation of the myeloid cell compartment urges investigations on neutrophil characteristics in coronavirus disease 2019 (COVID‐19). We isolated neutrophils from the blood of COVID‐19 patients receiving general ward care and from patients hospitalised at intensive care units (ICUs) to explore the kinetics of circulating neutrophils and factors important for neutrophil migration and activation. Methods Multicolour flow cytometry was exploited for the analysis of neutrophil differentiation and activation markers. Multiplex and ELISA technologies were used for the quantification of protease, protease inhibitor, chemokine and cytokine concentrations in plasma. Neutrophil polarisation responses were evaluated microscopically. Gelatinolytic and metalloproteinase activity in plasma was determined using a fluorogenic substrate. Co‐culturing healthy donor neutrophils with severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) allowed us to investigate viral replication in neutrophils. Results Upon ICU admission, patients displayed high plasma concentrations of granulocyte–colony‐stimulating factor (G‐CSF) and the chemokine CXCL8, accompanied by emergency myelopoiesis as illustrated by high levels of circulating CD10−, immature neutrophils with reduced CXCR2 and C5aR expression. Neutrophil elastase and non‐metalloproteinase‐derived gelatinolytic activity were increased in plasma from ICU patients. Significantly higher levels of circulating tissue inhibitor of metalloproteinase 1 (TIMP‐1) in patients at ICU admission yielded decreased total MMP proteolytic activity in blood. COVID‐19 neutrophils were hyper‐responsive to CXCL8 and CXCL12 in shape change assays. Finally, SARS‐CoV‐2 failed to replicate inside human neutrophils. Conclusion Our study provides detailed insights into the kinetics of neutrophil phenotype and function in severe COVID‐19 patients, and supports the concept of an increased neutrophil activation state in the circulation.
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- 2021
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20. A Chimeric Japanese Encephalitis Vaccine Protects against Lethal Yellow Fever Virus Infection without Inducing Neutralizing Antibodies
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Niraj Mishra, Robbert Boudewijns, Michael Alexander Schmid, Rafael Elias Marques, Sapna Sharma, Johan Neyts, and Kai Dallmeier
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flavivirus ,chimeric YFV-17D vaccine ,chimeric flavivirus vaccine ,cross-protection ,dual protection ,antibody-dependent enhancement ,Microbiology ,QR1-502 - Abstract
ABSTRACT Recent outbreaks of yellow fever virus (YFV) in West Africa and Brazil resulted in rapid depletion of global vaccine emergency stockpiles and raised concerns about being unprepared against future YFV epidemics. Here we report that a live attenuated virus similar to the Japanese encephalitis virus (JEV) vaccine JE-CVax/Imojev that consists of YFV-17D vaccine from which the structural (prM/E) genes have been replaced with those of the JEV SA14-14-2 vaccine strain confers full protection in mice against lethal YFV challenge. In contrast to the YFV-17D-mediated protection against YFV, this protection is not mediated by neutralizing antibodies but correlates with YFV-specific nonneutralizing antibodies and T cell responses against cell-associated YFV NS1 and other YFV nonstructural (NS) proteins. Our findings reveal the potential of YFV NS proteins to mediate protection and demonstrate that chimeric flavivirus vaccines, such as Imojev, could confer protection against two flaviviruses. This dual protection may have implications for the possible off-label use of JE-CVax in case of emergency and vaccine shortage during YFV outbreaks. In addition, populations in Asia that have been vaccinated with Imojev may already be protected against YFV should outbreaks ever occur on that continent, as several countries/regions in the Asia-Pacific are vulnerable to international spread of the YFV. IMPORTANCE Efficient and safe vaccines against yellow fever (e.g., YFV-17D) that provide long-lasting protection by rapidly inducing neutralizing antibody responses exist. However, the vaccine supply cannot cope with an increasing demand posed by urban outbreaks in recent years. Here we report that JE-CVax/Imojev, a YFV-17D-based chimeric Japanese encephalitis vaccine, also efficiently protects against YFV infection in mice. In case of shortage of the YFV vaccine during yellow fever outbreaks, (off-label) use of JE-CVax/Imojev may be considered. Moreover, wider use of JE-CVax/Imojev in Asia may lower the risk of the much-feared YFV spillover to the continent. More generally, chimeric vaccines that combine surface antigens and replication machineries of two distinct flaviviruses may be considered dual vaccines for the latter pathogen without induction of surface-specific antibodies. Following this rationale, novel flavivirus vaccines that do not hold a risk for antibody-dependent enhancement (ADE) of infection (inherent to current dengue vaccines and dengue vaccine candidates) could be designed.
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- 2020
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21. Neutrophil Recruitment and Participation in Severe Diseases Caused by Flavivirus Infection
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Marina Alves Fontoura, Rebeca Fróes Rocha, and Rafael Elias Marques
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neutrophils ,flavivirus ,encephalitis ,hemorrhagic fever ,pregnancy ,Science - Abstract
Neutrophils are first-line responders to infections and are recruited to target tissues through the action of chemoattractant molecules, such as chemokines. Neutrophils are crucial for the control of bacterial and fungal infections, but their role in the context of viral infections has been understudied. Flaviviruses are important human viral pathogens transmitted by arthropods. Infection with a flavivirus may result in a variety of complex disease manifestations, including hemorrhagic fever, encephalitis or congenital malformations. Our understanding of flaviviral diseases is incomplete, and so is the role of neutrophils in such diseases. Here we present a comprehensive overview on the participation of neutrophils in severe disease forms evolving from flavivirus infection, focusing on the role of chemokines and their receptors as main drivers of neutrophil function. Neutrophil activation during viral infection was shown to interfere in viral replication through effector functions, but the resulting inflammation is significant and may be detrimental to the host. For congenital infections in humans, neutrophil recruitment mediated by CXCL8 would be catastrophic. Evidence suggests that control of neutrophil recruitment to flavivirus-infected tissues may reduce immunopathology in experimental models and patients, with minimal loss to viral clearance. Further investigation on the roles of neutrophils in flaviviral infections may reveal unappreciated functions of this leukocyte population while increasing our understanding of flaviviral disease pathogenesis in its multiple forms.
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- 2021
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22. Hydrocephalus and arthrogryposis in an immunocompetent mouse model of ZIKA teratogeny: A developmental study.
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Jose Xavier-Neto, Murilo Carvalho, Bruno Dos Santos Pascoalino, Alisson Campos Cardoso, Ângela Maria Sousa Costa, Ana Helena Macedo Pereira, Luana Nunes Santos, Ângela Saito, Rafael Elias Marques, Juliana Helena Costa Smetana, Silvio Roberto Consonni, Carla Bandeira, Vivian Vasconcelos Costa, Marcio Chaim Bajgelman, Paulo Sérgio Lopes de Oliveira, Marli Tenorio Cordeiro, Laura Helena Vega Gonzales Gil, Bianca Alves Pauletti, Daniela Campos Granato, Adriana Franco Paes Leme, Lucio Freitas-Junior, Carolina Borsoi Moraes Holanda de Freitas, Mauro Martins Teixeira, Estela Bevilacqua, and Kleber Franchini
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Arctic medicine. Tropical medicine ,RC955-962 ,Public aspects of medicine ,RA1-1270 - Abstract
The teratogenic mechanisms triggered by ZIKV are still obscure due to the lack of a suitable animal model. Here we present a mouse model of developmental disruption induced by ZIKV hematogenic infection. The model utilizes immunocompetent animals from wild-type FVB/NJ and C57BL/6J strains, providing a better analogy to the human condition than approaches involving immunodeficient, genetically modified animals, or direct ZIKV injection into the brain. When injected via the jugular vein into the blood of pregnant females harboring conceptuses from early gastrulation to organogenesis stages, akin to the human second and fifth week of pregnancy, ZIKV infects maternal tissues, placentas and embryos/fetuses. Early exposure to ZIKV at developmental day 5 (second week in humans) produced complex manifestations of anterior and posterior dysraphia and hydrocephalus, as well as severe malformations and delayed development in 10.5 days post-coitum (dpc) embryos. Exposure to the virus at 7.5-9.5 dpc induces intra-amniotic hemorrhage, widespread edema, and vascular rarefaction, often prominent in the cephalic region. At these stages, most affected embryos/fetuses displayed gross malformations and/or intrauterine growth restriction (IUGR), rather than isolated microcephaly. Disrupted conceptuses failed to achieve normal developmental landmarks and died in utero. Importantly, this is the only model so far to display dysraphia and hydrocephalus, the harbinger of microcephaly in humans, as well as arthrogryposis, a set of abnormal joint postures observed in the human setting. Late exposure to ZIKV at 12.5 dpc failed to produce noticeable malformations. We have thus characterized a developmental window of opportunity for ZIKV-induced teratogenesis encompassing early gastrulation, neurulation and early organogenesis stages. This should not, however, be interpreted as evidence for any safe developmental windows for ZIKV exposure. Late developmental abnormalities correlated with damage to the placenta, particularly to the labyrinthine layer, suggesting that circulatory changes are integral to the altered phenotypes.
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- 2017
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23. Exploring the homeostatic and sensory roles of the immune system
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Rafael Elias Marques, Pedro Elias eMarques, Rodrigo eGuabiraba, and Mauro Martins Teixeira
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Homeostasis ,Immune System ,Inflammation ,Nervous System ,information ,Physiological role ,Immunologic diseases. Allergy ,RC581-607 - Abstract
Immunology developed under the notion the immune system exists to fight pathogens. Recently, the discovery of interactions with commensal microbiota that are essential to human health initiated a change in this old paradigm. Here, we argue the immune system has major physiological roles extending far beyond defending the host. Immune and inflammatory responses share the core property of sensing, defining the immune system also as a sensory system. The inference the immune system collects, interprets and store information, while creating an identity of self, places it in close relationship to the nervous system, which suggests these systems may have a profound evolutionary connection.
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- 2016
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24. Isolation of saint louis encephalitis virus from a horse with neurological disease in Brazil.
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Roberta Rosa, Erica Azevedo Costa, Rafael Elias Marques, Taismara Simas Oliveira, Ronaldo Furtini, Maria Rosa Quaresma Bomfim, Mauro Martins Teixeira, Tatiane Alves Paixão, and Renato Lima Santos
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Arctic medicine. Tropical medicine ,RC955-962 ,Public aspects of medicine ,RA1-1270 - Abstract
St. Louis encephalitis virus (SLEV) is a causative agent of encephalitis in humans in the Western hemisphere. SLEV is a positive-sense RNA virus that belongs to the Flavivirus genus, which includes West Nile encephalitis virus, Japanese encephalitis virus, Dengue virus and other medically important viruses. Recently, we isolated a SLEV strain from the brain of a horse with neurological signs in the countryside of Minas Gerais, Brazil. The SLEV isolation was confirmed by reverse-transcription RT-PCR and sequencing of the E protein gene. Virus identity was also confirmed by indirect immunofluorescence using commercial antibodies against SLEV. To characterize this newly isolated strain in vivo, serial passages in newborn mice were performed and led to hemorrhagic manifestations associated with recruitment of inflammatory cells into the central nervous system of newborns. In summary this is the first isolation of SLEV from a horse with neurological signs in Brazil.
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- 2013
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25. Role of the chemokine receptors CCR1, CCR2 and CCR4 in the pathogenesis of experimental dengue infection in mice.
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Rodrigo Guabiraba, Rafael Elias Marques, Anne-Gaëlle Besnard, Caio T Fagundes, Danielle G Souza, Bernhard Ryffel, and Mauro M Teixeira
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Medicine ,Science - Abstract
Dengue virus (DENV), a mosquito-borne flavivirus, is a public health problem in many tropical countries. Recent clinical data have shown an association between levels of different chemokines in plasma and severity of dengue. We evaluated the role of CC chemokine receptors CCR1, CCR2 and CCR4 in an experimental model of DENV-2 infection in mice. Infection of mice induced evident clinical disease and tissue damage, including thrombocytopenia, hemoconcentration, lymphopenia, increased levels of transaminases and pro-inflammatory cytokines, and lethality in WT mice. Importantly, infected WT mice presented increased levels of chemokines CCL2/JE, CCL3/MIP-1α and CCL5/RANTES in spleen and liver. CCR1⁻/⁻ mice had a mild phenotype with disease presentation and lethality similar to those of WT mice. In CCR2⁻/⁻ mice, lethality, liver damage, levels of IL-6 and IFN-γ, and leukocyte activation were attenuated. However, thrombocytopenia, hemoconcentration and systemic TNF-α levels were similar to infected WT mice. Infection enhanced levels of CCL17/TARC, a CCR4 ligand. In CCR4⁻/⁻ mice, lethality, tissue injury and systemic inflammation were markedly decreased. Despite differences in disease presentation in CCR-deficient mice, there was no significant difference in viral load. In conclusion, activation of chemokine receptors has discrete roles in the pathogenesis of dengue infection. These studies suggest that the chemokine storm that follows severe primary dengue infection associates mostly to development of disease rather than protection.
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- 2010
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26. Antiviral Activity of Quercetin Hydrate against Zika Virus
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Marielena Vogel Saivish, Gabriela de Lima Menezes, Roosevelt Alves da Silva, Marina Alves Fontoura, Jacqueline Farinha Shimizu, Gislaine Celestino Dutra da Silva, Igor da Silva Teixeira, Natalia Franco Bueno Mistrão, Victor Miranda Hernandes, Paula Rahal, Lívia Sacchetto, Carolina Colombelli Pacca, Rafael Elias Marques, and Maurício Lacerda Nogueira
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Inorganic Chemistry ,Zika virus ,natural products ,antiviral activity ,inhibition ,in silico analysis ,Organic Chemistry ,General Medicine ,Physical and Theoretical Chemistry ,Molecular Biology ,Spectroscopy ,Catalysis ,Computer Science Applications - Abstract
Zika virus (ZIKV) has re-emerged in recent decades, leading to outbreaks of Zika fever in Africa, Asia, and Central and South America. Despite its drastic re-emergence and clinical impact, no vaccines or antiviral compounds are available to prevent or control ZIKV infection. This study evaluated the potential antiviral activity of quercetin hydrate against ZIKV infection and demonstrated that this substance inhibits virus particle production in A549 and Vero cells under different treatment conditions. In vitro antiviral activity was long-lasting (still observed 72 h post-infection), suggesting that quercetin hydrate affects multiple rounds of ZIKV replication. Molecular docking indicates that quercetin hydrate can efficiently interact with the specific allosteric binding site cavity of the NS2B-NS3 proteases and NS1-dimer. These results identify quercetin as a potential compound to combat ZIKV infection in vitro.
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- 2023
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27. The E2 glycoprotein holds key residues for Mayaro virus adaptation to the urban Aedes aegypti mosquito
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Ferdinand Roesch, Chelsea Cereghino, Lucia Carrau, Alexandra Hardy, Helder Ribeiro-Filho, Annabelle Henrion Lacritick, Cassandra Koh, Jeffrey Marano, Tyler Bates, Pallavi Rai, Christina Chuong, Shamima Akter, Thomas Vallet, Hervé Blanc, Truitt Elliot, Anne M. Brown, Pawel Michalak, Tanya LeRoith, Jesse Bloom, Rafael Elias Marques, Maria-Carla Saleh, Marco Vignuzzi, James Weger-Lucarelli, Virginia Tech [Blacksburg], Populations virales et Pathogenèse - Viral Populations and Pathogenesis, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), New York University Langone Medical Center (NYU Langone Medical Center), NYU System (NYU), Centro Nacional de Pesquisa em Energia e Materiais = Brazilian Center for Research in Energy and Materials (CNPEM), Virus et Interférence ARN - Viruses and RNA Interference, George Mason University [Fairfax], Program of Genetics, Bioinformatics, and Computational Biology [Blacksburg] (GBCB), University of Haifa [Haifa], Fred Hutchinson Cancer Research Center [Seattle] (FHCRC), Howard Hughes Medical Institute [Chevy Chase] (HHMI), Howard Hughes Medical Institute (HHMI), This work was funded by the DARPA PREEMPT program administered through DARPA Cooperative Agreement HR001118S0017, this funding was awarded to M-C.S., M.V, and J.W-L. This work also received funding from Laboratoire d'Excellence Integrative Biology of Emerging Infectious Diseases (grant ANR-10-LABX-62-IBEID) to M-C.S. and M.V. Further support was provided by startup funds awarded to J.W-L by the Virginia-Maryland College of Veterinary Medicine and a grant from the One Health Research Funding Program awarded to J.W-L and P.M., and ANR-10-LABX-0062,IBEID,Integrative Biology of Emerging Infectious Diseases(2010)
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Virology ,Immunology ,[SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology ,Genetics ,Parasitology ,Molecular Biology ,Microbiology - Abstract
International audience; Adaptation to mosquito vectors suited for transmission in urban settings is a major driver in the emergence of arboviruses. To better anticipate future emergence events, it is crucial to assess their potential to adapt to new vector hosts. In this work, we used two different experimental evolution approaches to study the adaptation process of an emerging alphavirus, Mayaro virus (MAYV), to Ae. aegypti, an urban mosquito vector of many other arboviruses. We identified E2-T179N as a key mutation increasing MAYV replication in insect cells and enhancing transmission after escaping the midgut of live Ae. aegypti. In contrast, this mutation decreased viral replication and binding in human fibroblasts, a primary cellular target of MAYV in humans. We also showed that MAYV E2-T179N generates reduced viremia and displays less severe tissue pathology in vivo in a mouse model. We found evidence in mouse fibroblasts that MAYV E2-T179N is less dependent on the Mxra8 receptor for replication than WT MAYV. Similarly, exogenous expression of human apolipoprotein receptor 2 and Mxra8 enhanced WT MAYV replication compared to MAYV E2-T179N. When this mutation was introduced in the closely related chikungunya virus, which has caused major outbreaks globally in the past two decades, we observed increased replication in both human and insect cells, suggesting E2 position 179 is an important determinant of alphavirus host-adaptation, although in a virus-specific manner. Collectively, these results indicate that adaptation at the T179 residue in MAYV E2 may result in increased vector competence-but coming at the cost of optimal replication in humans-and may represent a first step towards a future emergence event.Author summary: Mosquito-borne viruses must replicate in both mosquito and vertebrate hosts to be maintained in nature successfully. When viruses that are typically transmitted by forest dwelling mosquitoes enter urban environments due to deforestation or travel, they must adapt to urban mosquito vectors to transmit effectively. For mosquito-borne viruses, the need to also replicate in a vertebrate host like humans constrains this adaptation process. Towards understanding how the emerging alphavirus, Mayaro virus, might adapt to transmission by the urban mosquito vector, Ae. aegypti, we used natural evolution approaches to identify several viral mutations that impacted replication in both mosquito and vertebrate hosts. We show that a single mutation in the receptor binding domain of E2 increased transmission by Ae. aegypti after bypassing the midgut infection barrier but simultaneously reduced replication and pathology in a mouse model. Mechanistic studies suggested that this mutation decreases the dependence of MAYV on human Mxra8 and the putative MAYV receptor human ApoER2 during replication. This suggests MAYV with this mutation alone is unlikely to be maintained in a natural transmission cycle between mosquitoes and humans. Understanding the adaptive potential of emerging viruses is critical to preventing future pandemics.
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- 2023
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28. Type I interferons are essential while type II interferon is dispensable for protection against St. Louis encephalitis virus infection in the mouse brain
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Juliana Helena Costa Smetana, Giovanni Freitas Gomes, Mariana Piccoli Gonçalves, Rafael Elias Marques, Mauro M. Teixeira, Daniele G. Souza, Milene Alvarenga Rachid, Rebeca Rocha, and Juliana L. Del Sarto
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Microbiology (medical) ,viruses ,mouse model ,Immunology ,Central nervous system ,Encephalitis Virus, St. Louis ,neglected arbovirus ,Disease ,Infectious and parasitic diseases ,RC109-216 ,Biology ,Virus Replication ,Microbiology ,Virus ,03 medical and health sciences ,Interferon-gamma ,Mice ,flavivirus ,medicine ,Animals ,St. Louis encephalitis virus infection ,030304 developmental biology ,0303 health sciences ,Encephalitis, St. Louis ,030306 microbiology ,St louis encephalitis ,Brain ,Viral Load ,biology.organism_classification ,medicine.disease ,Virology ,st. louis encephalitis ,Type II interferon ,Mice, Inbred C57BL ,interferons ,Flavivirus ,Disease Models, Animal ,Infectious Diseases ,medicine.anatomical_structure ,Interferon Type I ,Parasitology ,Encephalitis ,Research Article ,Research Paper - Abstract
St. Louis encephalitis virus (SLEV) is a neglected mosquito-borne flavivirus that causes severe neurological disease in humans. SLEV replication in the central nervous system (CNS) induces the local production of interferons (IFNs), which are attributed to host protection. The antiviral response to SLEV infection in the CNS is not completely understood, which led us to characterize the roles of IFNs using mouse models of St. Louis encephalitis. We infected mice deficient in type I IFN receptor (ABR−/-) or deficient in Type II IFN (IFNγ−/-) and assessed the contribution of each pathway to disease development. We found that type I and II IFNs play different roles in SLEV infection. Deficiency in type I IFN signaling was associated to an early and increased mortality, uncontrolled SLEV replication and impaired ISG expression, leading to increased proinflammatory cytokine production and brain pathology. Conversely, IFNγ−/- mice were moderately resistant to SLEV infection. IFNγ deficiency caused no changes to viral load or SLEV-induced encephalitis and did not change the expression of ISGs in the brain. We found that type I IFN is essential for the control of SLEV replication whereas type II IFN was not associated with protection in this model.
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- 2021
29. Predicting Antigenic Peptides from Rocio Virus NS1 Protein for Immunodiagnostic Testing Using Immunoinformatics and Molecular Dynamics Simulation
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Marielena Vogel Saivish, Gabriela de Lima Menezes, Vivaldo Gomes da Costa, Gislaine Celestino Dutra da Silva, Rafael Elias Marques, Maurício Lacerda Nogueira, and Roosevelt Alves Da Silva
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Flavivirus ,Organic Chemistry ,General Medicine ,Immunologic Tests ,Molecular Dynamics Simulation ,Viral Nonstructural Proteins ,Catalysis ,Computer Science Applications ,Inorganic Chemistry ,Molecular Docking Simulation ,Animals ,antigenic epitopes ,Rocio virus ,immunodiagnostics ,nonstructural protein ,linear B-cell epitope prediction ,Physical and Theoretical Chemistry ,Peptides ,Molecular Biology ,Spectroscopy - Abstract
The mosquito-borne disease caused by the Rocio virus is a neglected threat, and new immune inputs for serological testing are urgently required for diagnosis in low-resource settings and epidemiological surveillance. We used in silico approaches to identify a specific antigenic peptide (p_ROCV2) in the NS1 protein of the Rocio virus that was theoretically predicted to be stable and exposed on its surface, where it demonstrated key properties allowing it to interact with antibodies. These findings related to the molecular dynamics of this peptide provide important insights for advancing diagnostic platforms and investigating therapeutic alternatives.
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- 2022
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30. Different biological effects of exposure to far-UVC (222 nm) and near-UVC (254 nm) irradiation
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Renata Spagolla Napoleão Tavares, Douglas Adamoski, Alessandra Girasole, Ellen Nogueira Lima, Amauri da Silva Justo-Junior, Romênia Domingues, Ana Clara Caznok Silveira, Rafael Elias Marques, Murilo de Carvalho, Andre Luis Berteli Ambrosio, Adriana Franco Paes Leme, and Sandra Martha Gomes Dias
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Radiation ,Radiological and Ultrasound Technology ,Biophysics ,Radiology, Nuclear Medicine and imaging - Abstract
Ultraviolet C (UVC) light has long been used as a sterilizing agent, primarily through devices that emit at 254 nm. Depending on the dose and duration of exposure, UV 254 nm can cause erythema and photokeratitis and potentially cause skin cancer since it directly modifies nitrogenated nucleic acid bases. Filtered KrCl excimer lamps (emitting mainly at 222 nm) have emerged as safer germicidal tools and have even been proposed as devices to sterilize surgical wounds. All the studies that showed the safety of 222 nm analyzed cell number and viability, erythema generation, epidermal thickening, the formation of genetic lesions such as cyclobutane pyrimidine dimers (CPDs) and pyrimidine-(6-4)-pyrimidone photoproducts (6-4PPs) and cancer-inducing potential. Although nucleic acids can absorb and be modified by both UV 254 nm and UV 222 nm equally, compared to UV 254 nm, UV 222 nm is more intensely absorbed by proteins (especially aromatic side chains), causing photooxidation and cross-linking. Here, in addition to analyzing DNA lesion formation, for the first time, we evaluated changes in the proteome and cellular pathways, reactive oxygen species formation, and metalloproteinase (MMP) levels and activity in full-thicknessin vitroreconstructed human skin (RHS) exposed to UV 222 nm. We also performed the longest (40 days)in vivostudy of UV 222 nm exposure in the HRS/J mouse model at the occupational threshold limit value (TLV) for indirect exposure (25 mJ/cm2) and evaluated overall skin morphology, cellular pathological alterations, CPD and 6-4PP formation and MMP-9 activity. Our study showed that processes related to reactive oxygen species and inflammatory responses were more altered by UV 254 nm than by UV 222 nm. Our chronicin vivoexposure assay using the TLV confirmed that UV 222 nm causes minor damage to the skin. However, alterations in pathways related to skin regeneration raise concerns about direct exposure to UV 222 nm.
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- 2023
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31. UV 254 nm is more efficient than UV 222 nm in inactivating SARS-CoV-2 present in human saliva
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Renata Sesti-Costa, Cyro von Zuben Negrão, Jacqueline Farinha Shimizu, Alice Nagai, Renata Spagolla Napoleão Tavares, Douglas Adamoski, Wanderley Costa, Marina Alves Fontoura, Thiago Jasso da Silva, Adriano de Barros, Alessandra Girasole, Murilo de Carvalho, Veronica de Carvalho Teixeira, Andre Luis Berteli Ambrosio, Fabiana Granja, José Luiz Proença-Módena, Rafael Elias Marques, and Sandra Martha Gomes Dias
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Disinfection ,Oncology ,Photochemotherapy ,SARS-CoV-2 ,Ultraviolet Rays ,Biophysics ,COVID-19 ,Humans ,Pharmacology (medical) ,Dermatology ,Saliva ,RADIAÇÃO ULTRAVIOLETA - Abstract
Ultraviolet (UV) light can inactivate SARS-CoV-2. However, the practicality of UV light is limited by the carcinogenic potential of mercury vapor-based UV lamps. Recent advances in the development of krypton chlorine (KrCl) excimer lamps hold promise, as these emit a shorter peak wavelength (222 nm), which is highly absorbed by the skin's stratum corneum and can filter out higher wavelengths. In this sense, UV 222 nm irradiation for the inactivation of virus particles in the air and surfaces is a potentially safer option as a germicidal technology. However, these same physical properties make it harder to reach microbes present in complex solutions, such as saliva, a critical source of SARS-CoV-2 transmission. We provide the first evaluation for using a commercial filtered KrCl excimer light source to inactivate SARS-CoV-2 in saliva spread on a surface. A conventional germicidal lamp (UV 254 nm) was also evaluated under the same condition. Using plaque-forming units (PFU) and Median Tissue Culture Infectious Dose (TCID
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- 2022
32. Structural dynamics of SARS-CoV-2 nucleocapsid protein induced by RNA binding
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Celisa Caldana Costa Tonoli, Kleber G. Franchini, Marcio C. Bajgelman, Gabriel Ernesto Jara, Celso Eduardo Benedetti, Antonio Carlos Borges, Ana Carolina Migliorini Figueira, Adriana Santos Soprano, Gabriel Ravanhani Schleder, Fernanda Aparecida Heleno Batista, Helder Veras Ribeiro-Filho, Alexandre Cassago, Samuel Leite Guimarães, Rafael Elias Marques, Paulo Sérgio Lopes-de-Oliveira, and Daniela Barrreto Barbosa Trivella
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Molecular model ,Molecular Dynamics Simulation ,Virus ,Molecular dynamics ,Cellular and Molecular Neuroscience ,Viral life cycle ,Transcription (biology) ,Genetics ,Coronavirus Nucleocapsid Proteins ,Humans ,Molecular Biology ,Ecology, Evolution, Behavior and Systematics ,Ecology ,Chemistry ,SARS-CoV-2 ,RNA ,COVID-19 ,Nucleocapsid Proteins ,Phosphoproteins ,Microscopy, Electron ,Computational Theory and Mathematics ,Phosphoprotein ,Modeling and Simulation ,Biophysics ,RNA, Viral ,CTD ,Protein Binding - Abstract
The nucleocapsid (N) protein of the SARS-CoV-2 virus, the causal agent of COVID-19, is a multifunction phosphoprotein that plays critical roles in the virus life cycle, including transcription and packaging of the viral RNA. To play such diverse roles, the N protein has two globular RNA-binding modules, the N-(NTD) and C-terminal (CTD) domains, which are connected by an intrinsically disordered region. Despite the wealth of structural data available for the isolated NTD and CTD, how these domains are arranged in the full-length protein and how the oligomerization of N influences its RNA-binding activity remains largely unclear. Herein, using experimental data from electron microscopy and biochemical/biophysical techniques combined with molecular modeling and molecular dynamics simulations, we showed that, in the absence of RNA, the N protein formed structurally dynamic dimers, with the NTD and CTD arranged in extended conformations. However, in the presence of RNA, the N protein assumed a more compact conformation where the NTD and CTD are packed together. We also provided an octameric model for the full-length N bound to RNA that was consistent with electron microscopy images of the N protein in the presence of RNA. Together, our results shed new light on the dynamics and higher-order oligomeric structure of this versatile protein.
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- 2022
33. Atypical response to bacterial coinfection and persistent neutrophilic bronchoalveolar inflammation distinguish critical COVID-19 from influenza
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Lore Vanderbeke, Elisabeth Heylen, Paul Proost, Seppe Cambier, Joost Wauters, Mieke Gouwy, Rafael Elias Marques, Jennifer Vandooren, Patrick Matthys, Mieke Metzemaekers, Cato Jacobs, Greet Hermans, Ana Carolina de Carvalho, Philippe Meersseman, Dominique Schols, Bert Malengier-Devlies, Alexander Wilmer, Els Wauters, Ghislain Opdenakker, and Amber Nooyens
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Adult ,Male ,Chemokine ,Neutrophils ,medicine.medical_treatment ,Immunology ,Inflammation ,Influenza, Human ,Medicine ,Humans ,Protease inhibitor (pharmacology) ,Interleukin 8 ,Lung ,Aged ,Protease ,biology ,business.industry ,Coinfection ,COVID-19 ,General Medicine ,Bacterial Infections ,Middle Aged ,medicine.disease ,Influenza ,respiratory tract diseases ,Viral pneumonia ,biology.protein ,Cytokines ,Female ,medicine.symptom ,business ,Bronchoalveolar Lavage Fluid ,Elafin ,SLPI ,Research Article - Abstract
Neutrophils are recognized as important circulating effector cells in the pathophysiology of severe coronavirus disease 2019 (COVID-19). However, their role within the inflamed lungs is incompletely understood. Here, we collected bronchoalveolar lavage (BAL) fluids and parallel blood samples of critically ill COVID-19 patients requiring invasive mechanical ventilation and compared BAL fluid parameters with those of mechanically ventilated patients with influenza, as a non-COVID-19 viral pneumonia cohort. Compared with those of patients with influenza, BAL fluids of patients with COVID-19 contained increased numbers of hyperactivated degranulating neutrophils and elevated concentrations of the cytokines IL-1β, IL-1RA, IL-17A, TNF-α, and G-CSF; the chemokines CCL7, CXCL1, CXCL8, CXCL11, and CXCL12α; and the protease inhibitors elafin, secretory leukocyte protease inhibitor, and tissue inhibitor of metalloproteinases 1. In contrast, α-1 antitrypsin levels and net proteolytic activity were comparable in COVID-19 and influenza BAL fluids. During antibiotic treatment for bacterial coinfections, increased BAL fluid levels of several activating and chemotactic factors for monocytes, lymphocytes, and NK cells were detected in patients with COVID-19 whereas concentrations tended to decrease in patients with influenza, highlighting the persistent immunological response to coinfections in COVID-19. Finally, the high proteolytic activity in COVID-19 lungs suggests considering protease inhibitors as a treatment option. ispartof: JCI INSIGHT vol:7 issue:1 ispartof: location:United States status: published
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- 2022
34. Pediatric COVID-19 patients in South Brazil show abundant viral mRNA and strong specific anti-viral responses
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Julia C Fontoura, Jayme de Castilhos Ferreira Neto, Alexandre Borin, Lais D. Coimbra, Marcela Santos Correa da Costa, Ivaine Tais Sauthier Sartor, Cristina Bonorino, Marcelo Comerlato Scotta, Luiz Carlos Rodrigues Junior, Rafael Elias Marques, Ingrid Rodrigues Fernandes, Sidia M. Callegari-Jacques, Alisson F Haubert, Tiago Fazolo, Karina Lima, Márcia Polese-Bonatto, Raissa S Mello, Gabriel Hilario, Thiago J Borges, Priscila Oliveira de Souza, Veridiane Maria Pscheidt, Matheus de Bastos Balbe E Gutierres, Renata Zorzetto, Rodrigo Benedetti Gassen, Renato T. Stein, Jaqueline de Araujo Schwartz, Fernanda Hammes Varela, Helder I. Nakaya, Gabriela Oliveira Zavaglia, Aline C Oliveira, and Izza Gambin
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Adult ,CD4-Positive T-Lymphocytes ,Male ,COVID-19 Vaccines ,Adolescent ,T-Lymphocytes ,Science ,T cell ,General Physics and Astronomy ,Disease ,CD8-Positive T-Lymphocytes ,Biology ,Antibodies, Viral ,Article ,Antibodies ,General Biochemistry, Genetics and Molecular Biology ,Virus ,Young Adult ,Immune system ,medicine ,Humans ,RNA, Messenger ,Child ,Aged ,Viral Structural Proteins ,Vaccines, Synthetic ,Messenger RNA ,Multidisciplinary ,SARS-CoV-2 ,Transmission (medicine) ,COVID-19 ,Antimicrobial responses ,General Chemistry ,Middle Aged ,Immunity, Innate ,Immunity, Humoral ,medicine.anatomical_structure ,Child, Preschool ,Spike Glycoprotein, Coronavirus ,Immunology ,Infectious diseases ,Cytokines ,RNA, Viral ,Female ,Tumor necrosis factor alpha ,Brazil ,CD8 - Abstract
COVID-19 manifests as a milder disease in children than adults, but the underlying mechanisms are not fully characterized. Here we assess the difference in cellular or humoral immune responses of pediatric and adult COVID-19 patients to see if these factors contribute to the severity dichotomy. Children’s non-specific immune profile is dominated by naive lymphocytes and HLA-DRhighCX3CR1low dendritic cells; meanwhile, children show strong specific antibody and T cell responses for viral structural proteins, with their T cell responses differing from adults by having weaker CD8+TNF+ T cells responses to S peptide pool but stronger responses to N and M peptide pools. Finally, viral mRNA is more abundant in pediatric patients. Our data thus support a scenario in which SARS-CoV-2 infected children contribute to transmission yet are less susceptible to COVID-19 symptoms due to strong and differential responses to the virus., Children often show milder COVID-19 symptoms, but the underlying mechanistic insights are still lacking. Here the authors profile both pediatric and adult cohorts of COVID-19 patients in Brazil to find that children exhibit higher viral load but stronger and biased cellular immunity, thereby serving clues for the differential responses in children.
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- 2021
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35. Neutralisation of SARS-CoV-2 lineage P.1 by antibodies elicited through natural SARS-CoV-2 infection or vaccination with an inactivated SARS-CoV-2 vaccine: an immunological study
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Maria Luiza Moretti, Rafael Elias Marques, Mariene R. Amorim, Cecilia da C. Camilo, Gabriela Fabiano de Souza, Pamela S Andrade, Mariana C. Pinho, Audrey Basso Zangirolami, Pierina Lorencini Parise, Carolina Costa-Lima, Lais D. Coimbra, Marcelo A. Mori, Luciana S. Mofatto, Ingra Morales Claro, Nuno R. Faria, Grazielle C. Maktura, Clarice Weis Arns, Myuki A E Crispim, Bruno Deltreggia Benites, Magnun N. N. Santos, Erika R. Manuli, Lucas A M Franco, Mariana S. Ramundo, Darlan da Silva Candido, Camila L. Simeoni, Natalia S Brunetti, José Luiz Proença-Módena, Marcelo Addas-Carvalho, Christopher Dye, Marco Aurélio Ramirez Vinolo, Alessandro S. Farias, Esmenia C. Rocha, Oliver G. Pybus, Thais M. Coletti, Nelson Gaburo, Adriana S. S. Duarte, Stéfanie Primon Muraro, Ester Cerdeira Sabino, Arilson Bernardo dos Santos Pereira Gomes, Michael S. Diamond, Priscilla P. Barbosa, Fernando Rosado Spilki, Karina Bispo-dos-Santos, Fabiana Granja, Flavia C. S. Sales, Daniel A. Toledo-Teixeira, Vitor A. Costa, Rodrigo Nogueira Angerami, William Marciel de Souza, Renata Sesti-Costa, Jaqueline Goes de Jesus, Henrique Marques-Souza, Leandro Marques de Souza, Giulia M. Ferreira, Camila A. M. Silva, Lucas I. Buscaratti, Medical Research Council-São Paulo Research Foundation (FAPESP), Wellcome Trust, and Medical Research Council (MRC)
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Microbiology (medical) ,COVID-19 Vaccines ,Lineage (genetic) ,biology ,SARS-CoV-2 ,Vaccination ,COVID-19 ,Articles ,Antibodies, Viral ,Antibodies, Neutralizing ,Microbiology ,Virology ,United States ,Virus ,Neutralization ,Titer ,Infectious Diseases ,Immune system ,Polyclonal antibodies ,biology.protein ,Humans ,Antibody ,Brazil - Abstract
Background Mutations accrued by SARS-CoV-2 lineage P.1—first detected in Brazil in early January, 2021—include amino acid changes in the receptor-binding domain of the viral spike protein that also are reported in other variants of concern, including B.1.1.7 and B.1.351. We aimed to investigate whether isolates of wild-type P.1 lineage SARS-CoV-2 can escape from neutralising antibodies generated by a polyclonal immune response. Methods We did an immunological study to assess the neutralising effects of antibodies on lineage P.1 and lineage B isolates of SARS-CoV-2, using plasma samples from patients previously infected with or vaccinated against SARS-CoV-2. Two specimens (P.1/28 and P.1/30) containing SARS-CoV-2 lineage P.1 (as confirmed by viral genome sequencing) were obtained from nasopharyngeal and bronchoalveolar lavage samples collected from patients in Manaus, Brazil, and compared against an isolate of SARS-CoV-2 lineage B (SARS.CoV2/SP02.2020) recovered from a patient in Brazil in February, 2020. Isolates were incubated with plasma samples from 21 blood donors who had previously had COVID-19 and from a total of 53 recipients of the chemically inactivated SARS-CoV-2 vaccine CoronaVac: 18 individuals after receipt of a single dose and an additional 20 individuals (38 in total) after receipt of two doses (collected 17–38 days after the most recent dose); and 15 individuals who received two doses during the phase 3 trial of the vaccine (collected 134–230 days after the second dose). Antibody neutralisation of P.1/28, P.1/30, and B isolates by plasma samples were compared in terms of median virus neutralisation titre (VNT50, defined as the reciprocal value of the sample dilution that showed 50% protection against cytopathic effects). Findings In terms of VNT50, plasma from individuals previously infected with SARS-CoV-2 had an 8·6 times lower neutralising capacity against the P.1 isolates (median VNT50 30 [IQR Interpretation SARS-CoV-2 lineage P.1 might escape neutralisation by antibodies generated in response to polyclonal stimulation against previously circulating variants of SARS-CoV-2. Continuous genomic surveillance of SARS-CoV-2 combined with antibody neutralisation assays could help to guide national immunisation programmes. Funding São Paulo Research Foundation, Brazilian Ministry of Science, Technology and Innovation and Funding Authority for Studies, Medical Research Council, National Council for Scientific and Technological Development, National Institutes of Health. Translation For the Portuguese translation of the abstract see Supplementary Materials section.
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- 2021
36. Neutrophil Recruitment and Participation in Severe Diseases Caused by Flavivirus Infection
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Rafael Elias Marques, Marina Alves Fontoura, and Rebeca Rocha
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0301 basic medicine ,Chemokine ,viruses ,encephalitis ,Science ,Population ,Inflammation ,Context (language use) ,Review ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,0302 clinical medicine ,neutrophils ,flavivirus ,Immunopathology ,medicine ,education ,Ecology, Evolution, Behavior and Systematics ,education.field_of_study ,biology ,business.industry ,Paleontology ,biology.organism_classification ,medicine.disease ,Flavivirus ,030104 developmental biology ,Viral replication ,Space and Planetary Science ,Immunology ,biology.protein ,pregnancy ,medicine.symptom ,business ,030217 neurology & neurosurgery ,Encephalitis ,hemorrhagic fever - Abstract
Neutrophils are first-line responders to infections and are recruited to target tissues through the action of chemoattractant molecules, such as chemokines. Neutrophils are crucial for the control of bacterial and fungal infections, but their role in the context of viral infections has been understudied. Flaviviruses are important human viral pathogens transmitted by arthropods. Infection with a flavivirus may result in a variety of complex disease manifestations, including hemorrhagic fever, encephalitis or congenital malformations. Our understanding of flaviviral diseases is incomplete, and so is the role of neutrophils in such diseases. Here we present a comprehensive overview on the participation of neutrophils in severe disease forms evolving from flavivirus infection, focusing on the role of chemokines and their receptors as main drivers of neutrophil function. Neutrophil activation during viral infection was shown to interfere in viral replication through effector functions, but the resulting inflammation is significant and may be detrimental to the host. For congenital infections in humans, neutrophil recruitment mediated by CXCL8 would be catastrophic. Evidence suggests that control of neutrophil recruitment to flavivirus-infected tissues may reduce immunopathology in experimental models and patients, with minimal loss to viral clearance. Further investigation on the roles of neutrophils in flaviviral infections may reveal unappreciated functions of this leukocyte population while increasing our understanding of flaviviral disease pathogenesis in its multiple forms.
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- 2021
37. Clusters of SARS-CoV-2 Lineage B.1.1.7 Infection After Vaccination With Adenovirus-Vectored and Inactivated Vaccines: A Cohort Study
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Pierina Lorencini Parise, Scott C. Weaver, Stéfanie Primon Muraro, Renata Sesti-Costa, Maria Luiza Moretti, Julia Forato, Gabriela Felix Marchesi, Silvia de Barros-Mazon, Andrea Paula Bruno von Zuben, Vitor A. Costa, Lair Zambon, Bruno Deltreggia Benites, José Luiz Proença-Módena, André Schwambach Vieira, Luciana S. Mofatto, Daniel A. Toledo-Teixeira, Valeria Correia Almeida, Ester Cerdeira Sabino, Priscilla P. Barbosa, Fernando Rosado Spilki, Karina Bispo-dos-Santos, Josélia Cristina de Oliveira Moreira, Nuno R. Faria, Rodrigo Nogueira Angerami, Marcelo Addas-Carvalho, William Marciel de Souza, Daniela Maira Cardozo, Fabiana Granja, Alessandro S. Farias, Christiane Ambrosio, Rafael Elias Marques, Mariene R. Amorim, Gabriela Fabiano de Souza, and Natalia S Brunetti
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Vaccination ,Immunization ,business.industry ,Inactivated vaccine ,Outbreak ,Medicine ,Breakthrough infection ,Context (language use) ,business ,Virology ,Viral load ,Herd immunity - Abstract
Background: Some studies have determined that the SARS-CoV-2 B.1.1.7 variant of concern (VOC) has been associated with increased transmissibility, hospitalization, and mortality. This study aimed to explore the factors associated with B.1.1.7 VOC infection in the context of vaccination with adenovirus-vectored and inactivated vaccines. Methods: We analyzed epidemiological, clinical, and laboratory data from simultaneous COVID-19 outbreaks in a convent (Outbreak A) and in a long-term care facility (Outbreak B) in individuals vaccinated with a single dose of ChAdOx1 or two doses of the CoronaVac vaccine, respectively. First, we identified the viral lineages associated with these outbreaks by genome sequencing. Then, we assessed the relationship of viral load, specific immunoglobulin antibody levels, neutralization antibodies, case characteristics (age, vaccine type, and presence or absence of symptoms), and associations with risk of developing COVID-19. Findings: On March 2021, we detected SARS-CoV-2 RNA in nasopharyngeal samples from 14 of 22 of the ChAdOx1 vaccine recipients (Outbreak A, n=26), and 22 of 42 of the CoronaVac-vaccinated individuals (Outbreak B, n=52) for breakthrough infection rates for ChAdOx1 of 63·6% and 52·4% for CoronaVac. Vaccinated individuals from Outbreak A received a single dose at least 23 days before the outbreak, and the Outbreak B was involved virus detection 5 to 27 days after a second dose. In these outbreaks, the median ages were 73 and 77 years old, respectively. The median viral loads were 3·4×101 and 2·3×103 RNA copies per mL in Outbreaks A and B, respectively. In total for both outbreaks, 12 people had mild-COVID-19 while 26 were asymptomatic, but one case involving a person from Outbreak B was fatal. Based on analysis of SARS-CoV-2 genome sequences, we determined that outbreaks A and B were caused by two independent clusters of the B.1.1.7 VOC. Interestingly, the serum of PCR-positive symptomatic SARS-CoV-2-infected individuals had ~1·8 to 3·4-fold more neutralizing capacity against B.1.1.7 compared to the serum of asymptomatic, SARS-CoV-2-infected individuals, indicating that the levels of neutralizing antibodies induced by ChAdOx1 and CoronaVac were not correlated with protection against symptomatic infection. Interpretation: These data suggest that the B.1.1.7 variant can escape from the immune response elicited by immunization with a single dose of an adenovirus-vectored vaccine or two doses of an inactivated vaccine. However, partial immunization with ChAdOx1 and the complete series of CoronaVac seemed to provide protection against severe COVID-19 and death in the large majority of individuals. Continued and rapid immunization combined with nonpharmaceutical interventions (e.g., masking and physical distancing), are necessary to break the SARS-CoV-2 transmission until herd immunity improves. Funding Information: Sao Paulo Research Foundation, Brazilian Ministry of Science, Technology and Innovation and Funding Authority for Studies, Coordination for the Improvement of Higher Education Personnel, Wellcome Trust, Medical Research Council, National Institutes of Health, and Brazilian National Council for Scientific and Technological Development. Declaration of Interests: All other authors declare no competing interests. Ethics Approval Statement: All procedures followed the ethical standards of the responsible committee on human experimentation and were approved by the ethics committees from the University of Campinas, Brazil (Approval number: CAEE 31170720.3.0000.5404).
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- 2021
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38. Kinetics of peripheral blood neutrophils in severe coronavirus disease 2019
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Bert Malengier-Devlies, Lore Vanderbeke, José Luiz Proença-Módena, Seppe Cambier, Paul Proost, Gabriela Fabiano de Souza, Pierre Van Mol, Jennifer Vandooren, Lotte Vanbrabant, Arilson Bernardo Dos Sp Gomes, Jan Gunst, Yannick Van Herck, Alessandro S. Farias, Joost Wauters, Alexandre Borin, Erik Martens, Mieke Metzemaekers, Nathalie Van Aerde, Mieke Gouwy, Carine Wouters, Ghislain Opdenakker, Els Wauters, Bruna Toledo Nunes Pereira, Noëmie Pörtner, Pedro Elias Marques, Marco Aurélio Ramirez Vinolo, Sofie Coenen, Sofie Struyf, Stéfanie Primon Muraro, Cato Jacobs, Greet Hermans, Rafael Elias Marques, Marfa Blanter, Patrick Matthys, and Ana Carolina de Carvalho
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0301 basic medicine ,Chemokine ,Immunology ,03 medical and health sciences ,0302 clinical medicine ,Neutrophil differentiation ,COVID‐19 ,Intensive care ,cytokine ,Immunology and Allergy ,Medicine ,CXC chemokine receptors ,Interleukin 8 ,General Nursing ,biology ,business.industry ,chemokine ,neutrophil ,protease ,emergency myelopoiesis ,Tissue inhibitor of metalloproteinase ,RC581-607 ,030104 developmental biology ,030220 oncology & carcinogenesis ,Neutrophil elastase ,biology.protein ,Original Article ,Myelopoiesis ,Immunologic diseases. Allergy ,business - Abstract
Objectives Emerging evidence of dysregulation of the myeloid cell compartment urges investigations on neutrophil characteristics in coronavirus disease 2019 (COVID‐19). We isolated neutrophils from the blood of COVID‐19 patients receiving general ward care and from patients hospitalised at intensive care units (ICUs) to explore the kinetics of circulating neutrophils and factors important for neutrophil migration and activation. Methods Multicolour flow cytometry was exploited for the analysis of neutrophil differentiation and activation markers. Multiplex and ELISA technologies were used for the quantification of protease, protease inhibitor, chemokine and cytokine concentrations in plasma. Neutrophil polarisation responses were evaluated microscopically. Gelatinolytic and metalloproteinase activity in plasma was determined using a fluorogenic substrate. Co‐culturing healthy donor neutrophils with severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) allowed us to investigate viral replication in neutrophils. Results Upon ICU admission, patients displayed high plasma concentrations of granulocyte–colony‐stimulating factor (G‐CSF) and the chemokine CXCL8, accompanied by emergency myelopoiesis as illustrated by high levels of circulating CD10−, immature neutrophils with reduced CXCR2 and C5aR expression. Neutrophil elastase and non‐metalloproteinase‐derived gelatinolytic activity were increased in plasma from ICU patients. Significantly higher levels of circulating tissue inhibitor of metalloproteinase 1 (TIMP‐1) in patients at ICU admission yielded decreased total MMP proteolytic activity in blood. COVID‐19 neutrophils were hyper‐responsive to CXCL8 and CXCL12 in shape change assays. Finally, SARS‐CoV‐2 failed to replicate inside human neutrophils. Conclusion Our study provides detailed insights into the kinetics of neutrophil phenotype and function in severe COVID‐19 patients, and supports the concept of an increased neutrophil activation state in the circulation., In this study, we found that patients suffering from severe COVID‐19 presented with immature, activated blood neutrophils and were characterized by elevated plasma levels of G‐CSF and CXCL8 that disappeared towards ICU discharge. Elevated neutrophil‐derived proteases in plasma of ICU patients were associated with increased non‐metalloproteinase‐derived gelatinolytic activity but decreased total MMP proteolytic activity. SARS‐CoV‐2 failed to replicate inside human neutrophils, suggesting that changes observed in neutrophils from patients with COVID‐19 are indirect consequences of SARS‐CoV‐2 infection in vivo.
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- 2021
39. Flavonoids from Pterogyne nitens as Zika virus NS2B-NS3 protease inhibitors
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Kimberley M. Zorn, Bruna Katiele de Paula Sousa, Glaucius Oliva, Letícia Ribeiro de Assis, Nathalya Cristina de Moraes Roso Mesquita, Melina Mottin, Karina Bispo dos Santos, Rafael Elias Marques, Carolina Horta Andrade, Lais D. Coimbra, José Luiz Proença-Módena, Sean Ekins, Rafael Victorio Carvalho Guido, Luis Octávio Regasini, Caroline Sprengel Lima, Universidade Estadual Paulista (Unesp), Universidade Federal de Goiás (UFG), Universidade de São Paulo (USP), Brazilian Center for Research in Energy and Materials (CNPEM), Universidade Estadual de Campinas (UNICAMP), and Inc.
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Models, Molecular ,Protein Conformation ,medicine.medical_treatment ,Rutin ,Viral Nonstructural Proteins ,01 natural sciences ,Biochemistry ,Zika virus ,Machine Learning ,Docking (dog) ,Drug Discovery ,Chlorocebus aethiops ,Enzyme Inhibitors ,biology ,Chemistry ,Drug discovery ,Serine Endopeptidases ,Enzyme inhibitors ,Molecular Docking Simulation ,Quercetin ,Virtual screening ,medicine.drug_class ,Cell Survival ,In silico ,Antiviral Agents ,FARMACOLOGIA ,Article ,Viral Proteins ,medicine ,Animals ,NS3 protein ,Antiviral ,Molecular Biology ,Vero Cells ,NS3 ,Protease ,010405 organic chemistry ,Organic Chemistry ,Zika Virus ,biology.organism_classification ,Flavones ,Virology ,0104 chemical sciences ,010404 medicinal & biomolecular chemistry ,Vero cell ,Flavonoid ,Pterogyne nitens ,Antiviral drug ,Emerging arboviruses - Abstract
Made available in DSpace on 2021-06-25T10:23:35Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-04-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Fundação de Amparo à Pesquisa do Estado de Goiás National Institute of General Medical Sciences Although the widespread epidemic of Zika virus (ZIKV) and its neurological complications are well-known there are still no approved drugs available to treat this arboviral disease or vaccine to prevent the infection. Flavonoids from Pterogyne nitens have already demonstrated anti-flavivirus activity, although their target is unknown. In this study, we virtually screened an in-house database of 150 natural and semi-synthetic compounds against ZIKV NS2B-NS3 protease (NS2B-NS3p) using docking-based virtual screening, as part of the OpenZika project. As a result, we prioritized three flavonoids from P. nitens, quercetin, rutin and pedalitin, for experimental evaluation. We also used machine learning models, built with Assay Central® software, for predicting the activity and toxicity of these flavonoids. Biophysical and enzymatic assays generally agreed with the in silico predictions, confirming that the flavonoids inhibited ZIKV protease. The most promising hit, pedalitin, inhibited ZIKV NS2B-NS3p with an IC50 of 5 μM. In cell-based assays, pedalitin displayed significant activity at 250 and 500 µM, with slight toxicity in Vero cells. The results presented here demonstrate the potential of pedalitin as a candidate for hit-to-lead (H2L) optimization studies towards the discovery of antiviral drug candidates to treat ZIKV infections. Laboratory of Antibiotics and Chemotherapeutics (LAQ) Institute of Biosciences Humanities and Exact Sciences São Paulo State University (Unesp) Laboratory of Molecular Modeling and Drug Design (LabMol) Faculdade de Farmácia Universidade Federal de Goiás Institute of Physics of São Carlos University of São Paulo Brazilian Biosciences National Laboratory (LNBio) Brazilian Center for Research in Energy and Materials (CNPEM) Laboratory of Emerging Viruses (LEVE) Department of Genetics Evolution Microbiology and Immunology Institute of Biology University of Campinas (UNICAMP) Collaborations Pharmaceuticals Inc. Laboratory of Antibiotics and Chemotherapeutics (LAQ) Institute of Biosciences Humanities and Exact Sciences São Paulo State University (Unesp) CAPES: 130767/2016-01 CNPq: 150759/2017-7 FAPESP: 2013/07600-3 FAPESP: 2014/18330-0 FAPESP: 2016/00194-8 Fundação de Amparo à Pesquisa do Estado de Goiás: 20171026700006 FAPESP: 2018/03917-6 FAPESP: 2018/15083-2 Fundação de Amparo à Pesquisa do Estado de Goiás: 300508/2017-4 CNPq: 306251/2016-7 CNPq: 309957/2019-2 CNPq: 429322/2018-6 CNPq: 471129/2013-5 National Institute of General Medical Sciences: R43AT010585-01S1
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- 2021
40. Cryo-EM structure of the mature and infective Mayaro virus at 4.4 Å resolution reveals features of arthritogenic alphaviruses
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Adriana Franco Paes Leme, Helder Veras Ribeiro-Filho, Luiza Leme, Lais D. Coimbra, João Victor da Silva Guerra, Carolina Moretto Carnieli, A. C. M. Padilha, Alexandre Cassago, Rafael de Felício, Daniela B. B. Trivella, Rodrigo Villares Portugal, Rebeca Rocha, Rafael Elias Marques, and Paulo Sérgio Lopes-de-Oliveira
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0301 basic medicine ,Glycosylation ,Cryo-electron microscopy ,Science ,viruses ,Alphaviruses ,General Physics and Astronomy ,Immunoglobulins ,Alphavirus ,Arbovirus ,General Biochemistry, Genetics and Molecular Biology ,Epitope ,Virus ,Article ,03 medical and health sciences ,chemistry.chemical_compound ,Chlorocebus aethiops ,medicine ,Animals ,Humans ,Vero Cells ,Multidisciplinary ,030102 biochemistry & molecular biology ,biology ,Mass spectrometry ,Alphavirus Infections ,Cryoelectron Microscopy ,Membrane Proteins ,General Chemistry ,biology.organism_classification ,medicine.disease ,Virology ,Antibodies, Neutralizing ,030104 developmental biology ,chemistry ,Structural biology - Abstract
Mayaro virus (MAYV) is an emerging arbovirus of the Americas that may cause a debilitating arthritogenic disease. The biology of MAYV is not fully understood and largely inferred from related arthritogenic alphaviruses. Here, we present the structure of MAYV at 4.4 Å resolution, obtained from a preparation of mature, infective virions. MAYV presents typical alphavirus features and organization. Interactions between viral proteins that lead to particle formation are described together with a hydrophobic pocket formed between E1 and E2 spike proteins and conformational epitopes specific of MAYV. We also describe MAYV glycosylation residues in E1 and E2 that may affect MXRA8 host receptor binding, and a molecular “handshake” between MAYV spikes formed by N262 glycosylation in adjacent E2 proteins. The structure of MAYV is suggestive of structural and functional complexity among alphaviruses, which may be targeted for specificity or antiviral activity., Mayaro virus (MAYV) is an emerging arbovirus in Central and South America that is transmitted by mosquitoes and causes arthritogenic disease. Here, the authors present the 4.4 Å resolution cryo-EM structure of MAYV and describe specific features of the virus, which could be exploited for the design of MAYV-specific diagnostics and therapeutics.
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- 2020
41. Cryo-EM structure of the mature and infective Mayaro virus at 4.4 Å resolution reveals new features of arthritogenic alphaviruses
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Lais D. Coimbra, Rafael Elias Marques, Paulo Sérgio Lopes-de-Oliveira, Helder Ribeiro Filho, Rebeca Rocha, Rodrigo Villares Portugal, Luiza Leme, João Victor da Silva Guerra, A. C. M. Padilha, Daniela B. B. Trivella, and Alexandre Cassago
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Glycosylation ,Cryo-electron microscopy ,viruses ,Alphavirus ,Biology ,medicine.disease ,biology.organism_classification ,Arbovirus ,Virology ,Virus ,Epitope ,chemistry.chemical_compound ,chemistry ,medicine - Abstract
Mayaro virus (MAYV) is an emerging arbovirus of the Americas that may cause a debilitating arthritogenic disease. The biology of MAYV is not fully understood and largely inferred from related arthritogenic alphaviruses. Here we present the structure of MAYV at 4.4 Å resolution, obtained from a preparation of mature, infective virions. MAYV presents typical alphavirus features and organization. Interactions between viral proteins that lead to particle formation are described together with a hydrophobic pocket formed between E1 and E2 spike proteins and conformational epitopes specific of MAYV. We also describe MAYV glycosylation residues in E1 and E2 that may affect MXRA8 host receptor binding, and a molecular “handshake” between MAYV spikes formed by N262 glycosylation in adjacent E2 proteins. The structure of MAYV is suggestive of structural and functional complexity among alphaviruses, which may be targeted for specificity or antiviral activity.
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- 2020
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42. Early use of nitazoxanide in mild Covid-19 disease: randomized, placebo-controlled trial
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Pedro Augusto Alves, Daniela B. B. Trivella, Patricia R. M. Rocco, Ana Paula Morais Fernandes, Kleber G. Franchini, Artur T. Cordeiro, Raissa Prado Rocha, Erick Magri, Natalia de Fatima Paes, Paolo Pelosi, Nara Franzin de Moraes, Ivonise Sampaio Dos Santos, José Luiz Proença Modena, Paulo Fernando Guimarães Morando Marzocchi Tierno, Melanie Nogueira Carbonieri, Paula Veronica Martini Maciel, Fernanda F. Cruz, José Roberto Lapa e Silva, Luis Frederico Gerbase De Oliveira, Walter Freitas Junior, Ronir Raggio Luiz, Cristiano Cleidson Lima, Ezequiel Aparecido Dos Santos, Marco Antonio C. M. Junior, Pedro L. Silva, Alex Fiorini de Carvalho, Marcos de Assis Moura, Jose Mario de Jesus Goncalves, and Rafael Elias Marques
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medicine.medical_specialty ,Coronavirus disease 2019 (COVID-19) ,business.industry ,Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ,Placebo-controlled study ,Nitazoxanide ,Disease ,Placebo ,Internal medicine ,medicine ,Adverse effect ,business ,Viral load ,medicine.drug - Abstract
The antiparasitic drug nitazoxanide is widely available and exerts broad-spectrum antiviral activity in vitro. However, there is no evidence of its impact on SARS-CoV-2 infection.In a multicenter, randomized, double-blind, placebo-controlled trial, adult patients who presented up to 3 days after onset of Covid-19 symptoms (dry cough, fever, and/or fatigue) were enrolled. After confirmation of SARS-CoV2 infection by RT-PCR on nasopharyngeal swab, patients were randomized 1:1 to receive either nitazoxanide (500 mg) or placebo, TID, for 5 days. The primary outcome was complete resolution of symptoms. Secondary outcomes were viral load, general laboratory tests, serum biomarkers of inflammation, and hospitalization rate. Adverse events were also assessed.From June 8 to August 20, 2020, 1,575 patients were screened. Of these, 392 (198 placebo, 194 nitazoxanide) were analyzed. Median time from symptom onset to first dose of study drug was 5 (4-5) days. At the 5-day study visit, symptom resolution did not differ between the nitazoxanide and placebo arms. However, at the 1-week follow-up, 78% in the nitazoxanide arm and 57% in the placebo arm reported complete resolution of symptoms (p=0.048). Swabs collected were negative for SARS-CoV-2 in 29.9% of patients in the nitazoxanide arm versus 18.2% in the placebo arm (p=0.009). Viral load was also reduced after nitazoxanide compared to placebo (p=0.006). No serious adverse events were observed.In patients with mild Covid-19, symptom resolution did not differ between the nitazoxanide and placebo groups after 5 days of therapy. However, early nitazoxanide therapy was safe and reduced viral load significantly.Take home messageThis was the first study to evaluate the effect of early nitazoxanide therapy in mild Covid-19. Nitazoxanide did not accelerate symptom resolution after 5 days of therapy; however, reduced viral load significantly with no serious adverse events.
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- 2020
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43. Elevated Glucose Levels Favor SARS-CoV-2 Infection and Monocyte Response through a HIF-1α/Glycolysis-Dependent Axis
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Carlos Alberto Oliveira de Biagi Junior, Natalia S Brunetti, Robson Francisco Carvalho, André Schwambach Vieira, Victor Corasolla Carregari, Ana Campos Codo, Eli Mansour, Maria Luiza Moretti, Raisa G. Ulaf, Lais D. Coimbra, Stéfanie Primon Muraro, Licio A. Velloso, Juliana Silveira Prodonoff, Thyago A. Nunes, Lauar de Brito Monteiro, Karina Bispo dos Santos, Alexandre Borin, Guilherme Reis-de-Oliveira, André Damasio, Andrei C. Sposito, Marcus V. Agrela, Gabriela Fabiano de Souza, Fernanda Crunfli, Daniel A. Toledo-Teixeira, Helder I. Nakaya, Gustavo Gastão Davanzo, Daniel Martins-de-Souza, Pierina Lorencini Parise, Pedro M. Moraes-Vieira, Jeffersson Leandro Jimenez Restrepo, Vinícius O. Boldrini, Rafael Elias Marques, Alessandro S. Farias, João Victor Virgilio-da-Silva, Andre C. Palma, A. F. Bernardes, Fabrício Bíscaro Pereira, Helison R. Carmo, Marcelo A. Mori, Luciana C. Ribeiro, José Luiz Proença-Módena, Pedro Henrique Vendramini, Marco Aurélio Ramirez Vinolo, M. C. Martini, Universidade Estadual de Campinas (UNICAMP), Universidade de São Paulo (USP), Brazilian Biosciences National Laboratory (LNBio), Universidade Estadual Paulista (Unesp), D'Or Institute for Research and Education (IDOR), and Conselho Nacional de Desenvolvimento Científico e Tecnológico
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0301 basic medicine ,Mitochondrial ROS ,Blood Glucose ,Male ,Physiology ,Mitochondrion ,Monocytes ,0302 clinical medicine ,Medicine ,diabetes ,HIF-1alpha ,Endocrine system and metabolic diseases ,interferon ,glycolysis ,Middle Aged ,Research Highlight ,mitochondria ,medicine.anatomical_structure ,monocyte ,Female ,medicine.symptom ,Signal transduction ,Covid-19 ,Coronavirus Infections ,Glycolysis ,Signal Transduction ,Adult ,Pneumonia, Viral ,Inflammation ,Lung injury ,Cell Line ,Diabetes Complications ,03 medical and health sciences ,Betacoronavirus ,Immune system ,Diabetes Mellitus ,Humans ,Pandemics ,Molecular Biology ,business.industry ,SARS-CoV-2 ,Monocyte ,Correction ,COVID-19 ,Cell Biology ,medicine.disease ,Hypoxia-Inducible Factor 1, alpha Subunit ,030104 developmental biology ,inflammation ,Immunology ,business ,Cytokine storm ,Reactive Oxygen Species ,metabolism ,030217 neurology & neurosurgery - Abstract
Made available in DSpace on 2020-12-12T02:25:21Z (GMT). No. of bitstreams: 0 Previous issue date: 2020-09-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Fundo de Apoio ao Ensino, à Pesquisa e Extensão, Universidade Estadual de Campinas Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) COVID-19 can result in severe lung injury. It remained to be determined why diabetic individuals with uncontrolled glucose levels are more prone to develop the severe form of COVID-19. The molecular mechanism underlying SARS-CoV-2 infection and what determines the onset of the cytokine storm found in severe COVID-19 patients are unknown. Monocytes and macrophages are the most enriched immune cell types in the lungs of COVID-19 patients and appear to have a central role in the pathogenicity of the disease. These cells adapt their metabolism upon infection and become highly glycolytic, which facilitates SARS-CoV-2 replication. The infection triggers mitochondrial ROS production, which induces stabilization of hypoxia-inducible factor-1α (HIF-1α) and consequently promotes glycolysis. HIF-1α-induced changes in monocyte metabolism by SARS-CoV-2 infection directly inhibit T cell response and reduce epithelial cell survival. Targeting HIF-1ɑ may have great therapeutic potential for the development of novel drugs to treat COVID-19. Diabetic people with uncontrolled blood glucose levels have a greater risk to develop severe COVID-19 disease. Codo et al. show that elevated glucose levels and glycolysis promote SARS-CoV-2 (CoV-2) replication and cytokine production in monocytes through a mitochondrial ROS/hypoxia-inducible factor-1α dependent pathway, resulting in T cell dysfunction and epithelial cell death. Laboratory of Immunometabolism Department of Genetics Evolution Microbiology and Immunology Institute of Biology University of Campinas Department of Genetics Evolution Microbiology and Immunology Institute of Biology University of Campinas Department of Biochemistry and Tissue Biology Institute of Biology University of Campinas Department of Genetics at Ribeirao Preto Medical School University of Sao Paulo, Ribeirao Preto Department of Clinical and Toxicological analyses School of Pharmaceutical Sciences University of São Paulo Brazilian Biosciences National Laboratory (LNBio), Campinas Department of Animal Biology Institute of Biology University of Campinas, Campinas Department of Internal Medicine School of Medical Sciences University of Campinas, Campinas Department of Clinical Medicine School of Medical Sciences University of Campinas, Campinas Hematology and Hemotherapy Center University of Campinas, Campinas Obesity and Comorbidities Research Center (OCRC) University of Campinas Experimental Medicine Research Cluster (EMRC) University of Campinas Department of Structural and Functional Biology Institute of Biosciences São Paulo State University (UNESP), Botucatu D'Or Institute for Research and Education (IDOR) Instituto Nacional de Biomarcadores em Neuropsiquiatria Conselho Nacional de Desenvolvimento Científico e Tecnológico Department of Structural and Functional Biology Institute of Biosciences São Paulo State University (UNESP), Botucatu FAPESP: 20/04579-7 FAPESP: 2015/15626-8 FAPESP: 2016/18031-8 FAPESP: 2016/23328-0 FAPESP: 2017/01184-9 FAPESP: 2018/22505-0 FAPESP: 2019/00098-7 FAPESP: 2019/06372-3 FAPESP: 2020/04522-5 FAPESP: 2020/04558-0 FAPESP: 2020/04583-4 FAPESP: 2020/04746-0 FAPESP: 2020/04919-2 Fundo de Apoio ao Ensino, à Pesquisa e Extensão, Universidade Estadual de Campinas: 2274/20
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- 2020
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44. Biological and social challenges of human reproduction in a long-term Mars base
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Kateryna Lysenko-Ryba, Steven Abood, Aleksandra Kędzior, Dobrochna Minich, Rafael Elias Marques, and Konrad Szocik
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0301 basic medicine ,Sociology and Political Science ,media_common.quotation_subject ,Reproduction (economics) ,Subject (philosophy) ,Outer space ,Environmental ethics ,Mars Exploration Program ,Development ,Space (commercial competition) ,03 medical and health sciences ,030104 developmental biology ,Multidisciplinary approach ,Human settlement ,Political science ,Business and International Management ,Settlement (litigation) ,media_common - Abstract
A manned mission to Mars and the establishment of the first human settlement in outer space was once a mere figment of science fiction but is now being planned and expected to take place in the following twenty years. Mars is under consideration as the next planet beyond Earth to support continuous human exploration. Unfortunately, such an endeavor comes with titanic challenges in various disciplines, from space travel technology to medical, biological, social and ethical challenges. Reproduction involves many aspects of human life, and is also the subject of various disciplines. We assume that human reproduction in a Mars settlement will be necessary for the long-term success of an outer space mission. Thus, here we explore and discuss the challenges involving the likely scenario of human reproduction in Mars. To anticipate the many issues associated with human reproduction outside Earth, we applied multidisciplinary approaches to discuss possible social, ethical, medical and biological challenges of human reproduction on Mars.
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- 2018
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45. Clusters of SARS-CoV-2 Lineage B.1.1.7 Infection after Vaccination with Adenovirus-Vectored and Inactivated Vaccines
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José Luiz Proença-Módena, Natalia S Brunetti, André Schwambach Vieira, Bruno Deltreggia Benites, Gabriela F. P. de Souza, Gabriela Felix Marchesi, Rodrigo Nogueira Angerami, Josélia Cristina de Oliveira Moreira, William Marciel de Souza, Maria Luiza Moretti, Daniel A. Toledo-Teixeira, Stéfanie Primon Muraro, Andrea B Von Zuben, Luciana S. Mofatto, Ester Cerdeira Sabino, Nuno R. Faria, Pierina Lorencini Parise, Fernando Rosado Spilki, Karina Bispo-dos-Santos, Valeria Correia Almeida, Scott C. Weaver, Silvia de Barros-Mazon, Priscilla P. Barbosa, Marcelo Addas-Carvalho, Lair Zambon, Alessandro S. Farias, Vitor A. Costa, Mariene R. Amorim, Christiane Ambrosio, Rafael Elias Marques, Renata Sesti-Costa, Fabiana Granja, Daniela Maira Cardozo, and Julia Forato
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Adult ,Male ,COVID-19 Vaccines ,Lineage (genetic) ,Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ,Genetic Vectors ,Context (language use) ,Microbiology ,Asymptomatic ,Article ,Adenoviridae ,COVID-19 Serological Testing ,Disease Outbreaks ,Cohort Studies ,Young Adult ,vaccine ,Virology ,medicine ,Humans ,IMUNIZAÇÃO ,B.1.1.7 ,Aged ,Aged, 80 and over ,Whole Genome Sequencing ,outbreak ,SARS-CoV-2 ,business.industry ,Vaccination ,COVID-19 ,Outbreak ,Breakthrough infection ,Middle Aged ,Antibodies, Neutralizing ,QR1-502 ,Infectious Diseases ,Vaccines, Inactivated ,Immunoglobulin G ,Inactivated vaccine ,RNA, Viral ,Female ,medicine.symptom ,business ,variant of concern ,Brazil - Abstract
A SARS-CoV-2 B.1.1.7 variant of concern (VOC) has been associated with increased transmissibility, hospitalization, and mortality. This study aimed to explore the factors associated with B.1.1.7 VOC infection in the context of vaccination. On March 2021, we detected SARS-CoV-2 RNA in nasopharyngeal samples from 14 of 22 individuals vaccinated with a single-dose of ChAdOx1 (outbreak A, n = 26), and 22 of 42 of individuals with two doses of the CoronaVac vaccine (outbreak B, n = 52) for breakthrough infection rates for ChAdOx1 of 63.6% and 52.4% for CoronaVac. The outbreaks were caused by two independent clusters of the B.1.1.7 VOC. The serum of PCR-positive symptomatic SARS-CoV-2-infected individuals had ~1.8–3.4-fold more neutralizing capacity against B.1.1.7 compared to the serum of asymptomatic individuals. These data based on exploratory analysis suggest that the B.1.1.7 variant can infect individuals partially immunized with a single dose of an adenovirus-vectored vaccine or fully immunized with two doses of an inactivated vaccine, although the vaccines were able to reduce the risk of severe disease and death caused by this VOC, even in the elderly.
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- 2021
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46. Thiosemicarbazones and Phthalyl-Thiazoles compounds exert antiviral activity against yellow fever virus and Saint Louis encephalitis virus
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Rafael Elias Marques, Mauro M. Teixeira, Gevânio B.O.Oliveira Filho, Ana Cristina Lima Leite, Carolina Colombelli Pacca, José Wanderlan Pontes Espíndola, and Maurício Lacerda Nogueira
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Thiosemicarbazones ,0301 basic medicine ,Encephalitis Virus, St. Louis ,Biology ,Virus Replication ,Immunofluorescence ,Antiviral Agents ,01 natural sciences ,Virus ,Cell Line ,Microbiology ,03 medical and health sciences ,Chlorocebus aethiops ,Veterinary virology ,medicine ,Animals ,Vero Cells ,Pharmacology ,medicine.diagnostic_test ,010405 organic chemistry ,Yellow fever ,Outbreak ,General Medicine ,medicine.disease ,Virology ,0104 chemical sciences ,Thiazoles ,030104 developmental biology ,Viral replication ,Saint Louis encephalitis ,Sylvatic cycle ,Yellow fever virus ,Brazil - Abstract
Arboviruses, arthropod-borneviruses, are frequency associated to human outbreak and represent a serious health problem. The genus Flavivirus, such as Yellow Fever Virus (YFV) and Saint Louis Encephalitis Virus (SLEV), are important pathogens with high morbidity and mortality worldwide. In Brazil, YFV is maintained in sylvatic cycle, but many cases are notified annually, despite the efficiency of vaccine. SLEV causes an acute encephalitis and is widely distributed in the Americas. There is no specific antiviral drugs for these viruses, only supporting treatment that can alleviate symptoms and prevent complications. Here, we evaluated the potential anti-YFV and SLEV activity of a series of thiosemicarbazones and phthalyl-thiazoles. Plaque reduction assay, flow cytometry, immunofluorescence and cellular viability were used to test the compounds in vitro. Treated cells showed efficient inhibition of the viral replication at concentrations that presented minimal toxicity to cells. The assays showed that phthalyl-thiazole and phenoxymethyl-thiosemicarbazone reduced 60% of YFV replication and 75% of SLEV replication.
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- 2017
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47. Shielding and stealth effects of zwitterion moieties in double-functionalized silica nanoparticles
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Lívia Mesquita Dias Loiola, Marcio C. Bajgelman, Rafael Elias Marques, Marina Batista, Larissa Brentano Capeletti, Mateus Borba Cardoso, Gabriela Borba Mondo, and Rhubia S. M. Rosa
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Nanostructure ,Nanoparticle ,02 engineering and technology ,Adhesion ,010402 general chemistry ,021001 nanoscience & nanotechnology ,01 natural sciences ,Combinatorial chemistry ,0104 chemical sciences ,Surfaces, Coatings and Films ,Electronic, Optical and Magnetic Materials ,Biomaterials ,chemistry.chemical_compound ,Colloid ,Colloid and Surface Chemistry ,chemistry ,Zwitterion ,Moiety ,Surface modification ,0210 nano-technology ,Protein adsorption - Abstract
Surface functionalization of silica nanoparticles (SiO2NPs) has been considered as a promising strategy to develop target-specific nanostructures. However, finding a chemical functionalization that can be used as an active targeting moiety while preserving the nanoparticles colloidal stability in biological fluids is still challenging. We present here a dual surface modification strategy for SiO2NPs where a zwitterion (ZW) and a biologically active group (BAG) (amino, mercapto or carboxylic functionalities) are simultaneously grafted on the nanoparticles' surface. The rationale behind this strategy is to generate colloidally stable nanoparticles and avoid the nonspecific protein adsorption due to ZW groups insertion, while the effective interaction with biosystems is guaranteed by the BAGs presence. The biological efficacy was tested against VERO cells, E. coli bacteria and Zika viruses and a similar trend was observed for all tested particles. The desirable "stealth property" to prevent nonspecific protein adhesion also generated a ZW shielding effect of the BAG functionality hindering their proper interaction and activity in cells, bacteria and viruses.
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- 2019
48. Zika crisis in Brazil: challenges in research and development
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Amélia Ribeiro de Jesus, Lucas S. Ribeiro, Rafael Elias Marques, Roque P. Almeida, and Mauro M. Teixeira
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Primates ,0301 basic medicine ,medicine.medical_specialty ,Microcephaly ,Neurotropism ,Guillain-Barre Syndrome ,Zika virus ,Acute illness ,Mice ,03 medical and health sciences ,0302 clinical medicine ,Pregnancy ,Virology ,Research community ,medicine ,Animals ,Humans ,030212 general & internal medicine ,Intensive care medicine ,Psychiatry ,Guillain-Barre syndrome ,biology ,Zika Virus Infection ,business.industry ,Infant, Newborn ,Zika Virus ,medicine.disease ,biology.organism_classification ,Infant newborn ,Disease Models, Animal ,030104 developmental biology ,Female ,business ,Brazil - Abstract
Infection with the Zika virus (ZIKV) usually causes a mild acute illness, but two major severe syndromes have been described during the epidemic in Brazil: microcephaly and the Guillain-Barré Syndrome. There is now much evidence to show that ZIKV can infect and damage neuronal cells in vitro. In experimental animals, ZIKV has significant neurotropism and can cause brain damage. At present, diagnosis is still a challenge in the field and there is no treatment available. Another major challenge is that one must devise therapies for pregnant women, at all stages of pregnancy. Devising adequate treatment for ZIKV infections represents a challenge that will only be met by the joint effort of the research community.
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- 2016
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49. Establishment and characterization of a model of Mayaro virus infection in immunocompromised mice
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Lais D. Coimbra, Alexandre Borin Pereira, Rafael Elias Marques, Ana Carolina de Carvalho, Sílvio Roberto Consonni, and Maurício Lacerda Nogueira
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biology ,business.industry ,viruses ,virus diseases ,Outbreak ,General Medicine ,Disease ,Alphavirus ,biology.organism_classification ,medicine.disease_cause ,Virology ,Virus ,Chronic disease ,Medicine ,Chikungunya ,business - Abstract
Mayaro virus (MAYV) belongs to the Alphavirus genus along with other important viruses such as Chikungunya virus (CHIKV). The chronic disease caused by MAYV is described by a highly incapacitating joint pain which may endure for several weeks or months in at least 50% of the symptomatic patients. Recently, reports of sporadic outbreaks in humans has increased, however, the dynamic of the Mayaro infection is not completely understood and there is still no treatment or vaccine available. Herein, our goal was to develop an animal immunocompromised model ideal to understand the immunologic dynamic of the disease and perform antiviral tests.
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- 2018
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50. Interleukin-33 contributes to disease severity in Dengue virus infection in mice
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Mauro M. Teixeira, Danielle G. Souza, Isabelle Maillet, Foo Y. Liew, Anne-Gaelle Besnard, Rafael Elias Marques, Caio T. Fagundes, Bernhard Ryffel, Rodrigo Guabiraba, Institute of Infection, Immunity and Inflammation, Glasgow Biomedical Research Centre, University of Glasgow, Immunologie et Neurogénétique Expérimentales et Moléculaires (INEM), Centre National de la Recherche Scientifique (CNRS)-Université d'Orléans (UO), Immunologie et Embryologie Moléculaires (IEM), Université d'Orléans (UO)-Centre National de la Recherche Scientifique (CNRS), INRA, (France), Wellcome Trust, Medical Research Council (UK), Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq, Brazil), Fundacao de Amparo a Pesquisa do Estado de Minas Gerais (FAPEMIG, Brazil), and programme INCT em Dengue (CNPq, Brazil)
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Male ,0301 basic medicine ,medicine.drug_class ,medicine.medical_treatment ,[SDV]Life Sciences [q-bio] ,Immunology ,Inflammation ,Disease ,Dengue virus ,medicine.disease_cause ,Receptors, Interleukin-8B ,Dengue fever ,Dengue ,Mice ,03 medical and health sciences ,0302 clinical medicine ,medicine ,Animals ,Immunology and Allergy ,mouse ,Mice, Knockout ,Sulfonamides ,business.industry ,Granulocytosis ,Original Articles ,Dengue Virus ,Interleukin-33 ,medicine.disease ,Receptor antagonist ,Interleukin-1 Receptor-Like 1 Protein ,Recombinant Proteins ,3. Good health ,Mice, Inbred C57BL ,Interleukin 33 ,030104 developmental biology ,Cytokine ,inflammation ,Disease Progression ,IL-33 ,medicine.symptom ,business ,030215 immunology - Abstract
International audience; The excessive inflammation often present in patients with severe dengue infection is considered both a hallmark of disease and a target for potential treatments. IL-33 is a pleiotropic cytokine with pro-inflammatory effects whose role in dengue has not been fully elucidated. Here we demonstrate that IL-33 plays a disease-exacerbating role during experimental dengue infection in immunocompetent mice. Mice infected with Dengue virus serotype 2 (DENV2) produced high levels of IL-33. DENV2-infected mice treated with recombinant IL-33 developed markedly more severe disease compared to untreated mice as assessed by mortality, granulocytosis, liver damage and pro-inflammatory cytokine production. Conversely, ST2-/- mice (deficient in IL-33 receptor) infected with DENV2 developed significantly less severe disease compared to wild-type (WT) mice. Furthermore, the increased disease severity and the accompanying pathology induced by IL-33 during dengue infection were reversed by the simultaneous treatment with a CXCR2 receptor antagonist (DF2156A). Together, these results indicate that IL-33 plays a disease-exacerbating role in experimental dengue infection, likely driven by CXCR2-expressing cells, leading to elevated pro-inflammatory response-mediated pathology. Our results also indicate that IL-33 is a potential therapeutic target for dengue infection.
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- 2018
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