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1. T cell response against SARS-CoV-2 persists after one year in patients surviving severe COVID-19

Author

Fabienne Venet, Morgane Gossez, Frank Bidar, Maxime Bodinier, Rémy Coudereau, Anne-Claire Lukaszewicz, Claire Tardiveau, Karen Brengel-Pesce, Valérie Cheynet, Marie-Angélique Cazalis, Rémi Pescarmona, Lorna Garnier, Marine Ortillon, Marielle Buisson, Maude Bouscambert-Duchamp, Florence Morfin-Sherpa, Jean-Sébastien Casalegno, Filippo Conti, Thomas Rimmelé, Laurent Argaud, Martin Cour, Mitra Saadatian-Elahi, Laetitia Henaff, Philippe Vanhems, and Guillaume Monneret

Subjects
Immune memory, T lymphocyte, SARS-CoV-2, Critically ill patients, HLA-DR, Sepsis, Medicine, Medicine (General), R5-920
Abstract

Summary: Background: In critically ill COVID-19 patients, the initial response to SARS-CoV-2 infection is characterized by major immune dysfunctions. The capacity of these severe patients to mount a robust and persistent SARS-CoV-2 specific T cell response despite the presence of severe immune alterations during the ICU stay is unknown. Methods: Critically ill COVID-19 patients were sampled five times during the ICU stay and 9 and 13 months afterwards. Immune monitoring included counts of lymphocyte subpopulations, HLA-DR expression on monocytes, plasma IL-6 and IL-10 concentrations, anti-SARS-CoV-2 IgG levels and T cell proliferation in response to three SARS-CoV-2 antigens. Findings: Despite the presence of major lymphopenia and decreased monocyte HLA-DR expression during the ICU stay, convalescent critically ill COVID-19 patients consistently generated adaptive and humoral immune responses against SARS-CoV-2 maintained for more than one year after hospital discharge. Patients with long hospital stays presented with stronger anti-SARS-CoV-2 specific T cell response but no difference in anti-SARS-CoV2 IgG levels. Interpretation: Convalescent critically ill COVID-19 patients consistently generated a memory immune response against SARS-CoV-2 maintained for more than one year after hospital discharge. In recovered individuals, the intensity of SARS-CoV-2 specific T cell response was dependent on length of hospital stay. Funding: This observational study was supported by funds from the Hospices Civils de Lyon, Fondation HCL, Claude Bernard Lyon 1 University and Région Auvergne Rhône-Alpes and by partial funding by REACTing (Research and ACTion targeting emerging infectious diseases) INSERM, France and a donation from Fondation AnBer (http://fondationanber.fr/).

Published
2022
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2. Protocol for the DisCoVeRy trial: multicentre, adaptive, randomised trial of the safety and efficacy of treatments for COVID-19 in hospitalised adults

Author

Florence Ader, Jean-François Timsit, Yazdan Yazdanpanah, François-Xavier Lescure, Laetitia Moinot, Christelle Delmas, Hélène Espérou, Laurent Abel, Benjamin Leveau, Vinca Icard, Toni Alfaiate, Linda Wittkop, Charles Burdet, France Mentré, Marion Schneider, Nathan Peiffer-Smadja, Julien Poissy, Lila Bouadma, Bruno Lina, Florence Morfin-Sherpa, Maude Bouscambert, Alexandre Gaymard, Gilles Peytavin, Jeremie Guedj, Claire Andrejak, Cedric Laouenan, Drifa Belhadi, Axelle Dupont, Basma Basli, Anissa Chair, Samira Laribi, Julie Level, Marie-Capucine Tellier, Aline Dechanet, Dominique Costagliola, Sandrine Couffin-Cadiergues, Juliette Saillard, Claire Fougerou, Carole Cagnot, Soizic Le Mestre, Delphine Lebrasseur-Longuet, Ventzislava Petrov-Sanchez, Alpha Diallo, NoéMie Mercier, Sarah Tubiana, Benjamin Hamze, Ambre Gelley, Marion Noret, Eric D’Ortenzio, Oriane Puechal, and Caroline Semaille

Subjects
Medicine
Abstract

Introduction To find effective and safe treatments for COVID-19, the WHO recommended to systemically evaluate experimental therapeutics in collaborative randomised clinical trials. As COVID-19 was spreading in Europe, the French national institute for Health and Medical Research (Inserm) established a transdisciplinary team to develop a multi-arm randomised controlled trial named DisCoVeRy. The objective of the trial is to evaluate the clinical efficacy and safety of different investigational re-purposed therapeutics relative to Standard of Care (SoC) in patients hospitalised with COVID-19.Methods and analysis DisCoVeRy is a phase III, open-label, adaptive, controlled, multicentre clinical trial in which hospitalised patients with COVID-19 in need of oxygen therapy are randomised between five arms: (1) a control group managed with SoC and four therapeutic arms with re-purposed antiviral agents: (2) remdesivir + SoC, (3) lopinavir/ritonavir + SoC, (4) lopinavir/ritonavir associated with interferon (IFN)-β−1a + SoC and (5) hydroxychloroquine + SoC. The primary endpoint is the clinical status at Day 15 on the 7-point ordinal scale of the WHO Master Protocol (V.3.0, 3 March 2020). This trial involves patients hospitalised in conventional departments or intensive care units both from academic or non-academic hospitals throughout Europe. A sample size of 3100 patients (620 patients per arm) is targeted. This trial has begun on 22 March 2020. Since 5 April 2020, DisCoVeRy has been an add-on trial of the Solidarity consortium of trials conducted by the WHO in Europe and worldwide. On 8 June 2020, 754 patients have been included.Ethics and dissemination Inserm is the sponsor of DisCoVeRy. Ethical approval has been obtained from the institutional review board on 13 March 2020 (20.03.06.51744) and from the French National Agency for Medicines and Health Products (ANSM) on 9 March 2020. Results will be submitted for publication in peer-reviewed journals.Trial registration number NCT04315948 Eudra-CT 2020-000936-23.

Published
2020
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3. Update of European Society of Clinical Microbiology and Infectious Diseases coronavirus disease 2019 guidelines: diagnostic testing for severe acute respiratory syndrome coronavirus 2

Author

Paraskevi C. Fragkou, Giulia De Angelis, Giulia Menchinelli, Fusun Can, Federico Garcia, Florence Morfin-Sherpa, Dimitra Dimopoulou, Konstantina Dimopoulou, Silvia Zelli, Adolfo de Salazar, Rieke Reiter, Hannah Janocha, Adriano Grossi, Jimmy Omony, and Chrysanthi Skevaki

Subjects
Microbiology (medical), Infectious Diseases, General Medicine
Published
2023
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4. Evaluation of High-Throughput SARS-CoV-2 Serological Assays in a Longitudinal Cohort of Patients with Mild COVID-19: Clinical Sensitivity, Specificity, and Association with Virus Neutralization Test

Author

Florence Morfin-Sherpa, Adèle Paul, Carole Langlois-Jacques, Sophie Trouillet-Assant, Virginie Pitiot, Constance D’Aubarède-Frieh, Bruno Pozzetto, Nicolas Guibert, David Goncalves, Jean-Baptiste Fassier, Nicole Fabien, François Gueyffier, Antonin Bal, Mary-Anne Trabaud, Amélie Massardier-Pilonchery, Bruno Lina, Muriel Rabilloud, Vanessa Escuret, and André Boibieux

Subjects
Adult, Male, 0301 basic medicine, Coronavirus disease 2019 (COVID-19), Health-care workers, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Concordance, 030106 microbiology, Clinical Biochemistry, Virus Neutralization, Antibodies, Viral, Sensitivity and Specificity, Article, COVID-19 Serological Testing, Serology, Cohort Studies, 03 medical and health sciences, Neutralization Tests, Humans, Medicine, Longitudinal Studies, Prospective Studies, Neutralizing antibody, Aged, Biochemistry, medical, biology, SARS-CoV-2, business.industry, Biochemistry (medical), COVID-19, Virus neutralization, Antibodies, Neutralizing, Virology, Titer, 030104 developmental biology, biology.protein, Female, Antibody, business, Serological assays
Abstract

Background The association between SARS-CoV-2 commercial serological assays and virus neutralization test (VNT) has been poorly explored in mild patients with COVID-19. Methods 439 serum specimens were longitudinally collected from 76 healthcare workers with RT-PCR-confirmed COVID-19. The clinical sensitivity (determined weekly) of 9 commercial serological assays were evaluated. Clinical specificity was assessed using 69 pre-pandemic sera. Correlation, agreement, and concordance with the VNT were also assessed on a subset of 170 samples. Area under the ROC curve (AUC) was estimated at 2 neutralizing antibody titers. Results The Wantai Total Ab assay targeting the receptor binding domain (RBD) within the S protein presented the best sensitivity at different times during the course of disease. The clinical specificity was greater than 95% for all tests except for the Euroimmun IgA assay. The overall agreement with the presence of neutralizing antibodies ranged from 62.2% (95%CI; 56.0–68.1) for bioMérieux IgM to 91.2% (87.0–94.2) for Siemens. The lowest negative percent agreement (NPA) was found with the Wantai Total Ab assay (NPA 33% (21.1–48.3)). The NPA for other total Ab or IgG assays targeting the S or the RBD was 80.7% (66.7–89.7), 90.3% (78.1–96.1), and 96.8% (86.8–99.3) for Siemens, bioMérieux IgG, and DiaSorin, respectively. None of the commercial assays have sufficient performance to detect a neutralizing titer of 80 (AUC Conclusions Although some assays show a better agreement with VNT than others, the present findings emphasize that commercialized serological tests, including those targeting the RBD, cannot substitute a VNT for the assessment of functional antibody response.

Published
2021
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5. Six-month antibody response to SARS-CoV-2 in healthcare workers assessed by virus neutralization and commercial assays

Author

Fascia Pascal, Adnot Jérôme, Fontaine Juliette, Massardier-Pilonchéry Amélie, Boibieux André, Josset Laurence, Nicolas Guibert, Oria Fatima, Nazare Julie-Anne, Chiarello Pierre, Samperiz Audrey, Antonin Bal, Prudent Mélanie, Simon Chantal, Dulce Alfaiate, Fassier Jean-Baptiste, Florence Morfin-Sherpa, Constance d'Aubarede, Brunel-Dalmas Florence, Lahousse Matthieu, Pitiot Virginie, Trabaud Mary-Anne, Gaillot-Durand Lucie, Bal Antonin, Lina Bruno, Trouillet-Assant Sophie, Gueyffier François, Perry Marielle, Gillet Myriam, Adèle Paul, Jean-Baptiste Fassier, Valette Martine, Bourgeois Gaëlle, Lozano Hélène, Kahina Saker, Alfaiate Dulce, Morfin Florence, Mary-Anne Trabaud, Rabilloud Muriel, Schlienger Isabelle, Narbey David, Durupt François, Makhloufi Djamila, Bruno Pozzetto, Paul Adèle, Caire Eurydice, Bergeret Alain, Godinot Matthieu, Escuret Vanessa, Bonnet Florent, Moll Frédéric, Guibert Nicolas, Gaymard Alexandre, Cotte Laurent, Sophie Trouillet-Assant, Muriel Rabilloud, Virginie Pitiot, Amélie Massardier-Pilonchery, Milon Marie-Paule, Charbotel Barbara, Carole Langlois-Jacques, Bruno Lina, Hospices Civils de Lyon (HCL), Unité Mixte de Recherche Epidémiologique et de Surveillance Transport Travail Environnement (UMRESTTE UMR_T9405), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Gustave Eiffel, Université de Lyon, Institut des Agents Infectieux [Lyon] (IAI), Service de médecine et santé au travail, hospices civils de Lyon, Laboratoire de virologie, Physiopathologie et biothérapies des infections muqueuses (GIMAP), Centre International de Recherche en Infectiologie - UMR (CIRI), Institut National de la Santé et de la Recherche Médicale (INSERM)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Université Jean Monnet [Saint-Étienne] (UJM), Centre International de Recherche en Infectiologie (CIRI), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Jean Monnet [Saint-Étienne] (UJM)

Subjects
Male, Time Factors, Antibodies, Viral, Neutralization, MEDECINE, Health personnel, 0302 clinical medicine, SANTE, SARS-COV2, Medicine, 030212 general & internal medicine, Letter to the Editor, 0303 health sciences, biology, PERSONNEL DE SANTE, virus diseases, General Medicine, Middle Aged, University hospital, TRAVAIL, 3. Good health, Titer, Infectious Diseases, Female, Antibody, Serological assays, ANTICORPS, Adult, Microbiology (medical), 2019-20 coronavirus outbreak, Coronavirus disease 2019 (COVID-19), Health Personnel, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), education, GESTION DU PERSONNEL, Virus Neutralization, Virus, 03 medical and health sciences, Neutralization Tests, Seroprevalence, Humans, Healthcare workers, Virus neutralisation assay, 030304 developmental biology, SARS-CoV-2, business.industry, COVID-19, Antibodies, Neutralizing, Virology, Antibody response, biology.protein, [SDV.SPEE]Life Sciences [q-bio]/Santé publique et épidémiologie, business
Abstract

We conducted a prospective study in healthcare workers (n=296) of the University Hospital of Lyon, France. Serum samples (n=296) collected six months after disease onset were tested using three commercial assays: the Wantai Ab assay detecting total antibodies against the receptor binding domain (RBD) of the S protein, the bioMerieux Vidas assay detecting IgG to the RBD and the Abbott Architect assay detecting IgG to the N protein. The neutralising antibody (NAb) titre was also determined for all samples with a virus neutralisation assay (VNA) using live virus. The positivity rate was 100% with the Wantai assay, 84.8% with the bioMerieux assay and 55.4% with the Abbott assay. Only 51% of HCWs were positive for the presence of NAb. Less than 10 % of HCWs had a NAb titre greater than 80. At a neutralising titre of 80, the area under the curves [IC 95%] was 0.71 [0.62-0.81], 0.75 [0.65-0.85] and 0.95 [0.92-0.97] for Wantai, Abbott and Vidas respectively. The data presented herein suggest that commercial assays detecting antibodies against the N protein must not be used in long-term seroprevalence surveys while the Wantai assay could be useful for this purpose. VNA should remain the gold standard to assess the protective antibody response, but some commercial assays could be used as first-line screening of long-term presence of NAb.

Published
2021
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6. ESCMID COVID-19 guidelines: diagnostic testing for SARS-CoV-2

Author

Paraskevi C. Fragkou, Giulia De Angelis, Giulia Menchinelli, Fusun Can, Federico Garcia, Florence Morfin-Sherpa, Dimitra Dimopoulou, Elisabeth Mack, Adolfo de Salazar, Adriano Grossi, Theodore Lytras, Chrysanthi Skevaki, Can, Füsun (ORCID 0000-0001-9387-2526 & YÖK ID 103165), Fragkou, Paraskevi C., De Angelis, Giulia, Menchinelli, Giulia, Garcia, Federico, Morfin-Sherpa, Florence, Dimopoulou, Dimitra, Mack, Elisabeth, de Salazar, Adolfo, Grossi, Adriano, Lytras, Theodore, Skevaki, Chrysanthi, Koç Üniversitesi İş Bankası Enfeksiyon Hastalıkları Uygulama ve Araştırma Merkezi (EHAM) / Koç University İşbank Center for Infectious Diseases (KU-IS CID), and School of Medicine

Subjects
Microbiology (medical), Infectious Diseases, SARS-CoV-2, Health Personnel, COVID-19, Humans, General Medicine, Nasopharyngeal swabs, Serologic tests, Diagnosis, Guidelines, Testing, Nucleic Acid Amplification Techniques, Diagnostic Techniques and Procedures
Abstract

Scope: the objective of these guidelines is to identify the most appropriate diagnostic test and/or diagnostic approach for SARS-CoV-2. The recommendations are intended to provide guidance to clinicians, clinical microbiologists, other health care personnel, and decision makers. Methods: an ESCMID COVID-19 guidelines task force was established by the ESCMID Executive Committee. A small group was established, half appointed by the chair and the remaining selected with an open call. Each panel met virtually once a week. For all decisions, a simple majority vote was used. A list of clinical questions using the PICO (population, intervention, comparison, outcome) format was developed at the beginning of the process. For each PICO, two panel members performed a literature search focusing on systematic reviews, with a third panellist involved in case of inconsistent results. Quality of evidence assessment was based on the GRADE-ADOLOPMENT (Grading of Recommendations Assessment, Development and Evaluation - adoption, adaptation, and de novo development of recommendations) approach. Recommendations: a total of 43 PICO questions were selected that involve the following types of populations: (a) patients with signs and symptoms of COVID-19; (b) travellers, healthcare workers, and other individuals at risk for exposure to SARS-CoV-2; (c) asymptomatic individuals, and (d) close contacts of patients infected with SARS-CoV-2. The type of diagnostic test (commercial rapid nucleic acid amplification tests and rapid antigen detection), biomaterial, time since onset of symptoms/contact with an infectious case, age, disease severity, and risk of developing severe disease are also taken into consideration., "Hycor Biomedical; Bencard Allergie; Thermo Fisher Scientific; Mead Johnson Nutrition; Universities Giessen and Marburg Lung Center; German Center for Lung Research; University Hospital Giessen and Marburg; Deutsche Forschungsgemeinschaft-funded SFB 1021; KFO 309 (P10); SK 317/1-1; Foundation for Pathobiochemistry and Molecular Diagnostics; Fundacion Progreso y Salud; Consejería de Salud; Junta de Andalucía; Philipps University Marburg Fundation of the Faculty of Medicine; Deutsche Jose Carreras Leukamie-Stiftung"

Published
2021

7. Impact of Host Immune Status on Discordant Anti-SARS-CoV-2 Circulating B Cell Frequencies and Antibody Levels

Author

Frédéric Coutant, Jean-Jacques Pin, Florence Morfin-Sherpa, Tristan Ferry, Stéphane Paul, Bruno Pozzetto, Myriam Normand, and Pierre Miossec

Subjects
Adult, Aged, 80 and over, Male, B-Lymphocytes, anti-SARS-CoV-2, B cell repertoire, QH301-705.5, SARS-CoV-2, Health Personnel, COVID-19, Comorbidity, Middle Aged, Antibodies, Viral, Antibodies, Neutralizing, Severity of Illness Index, Article, Chemistry, Antibody Formation, Spike Glycoprotein, Coronavirus, Humans, Female, human monoclonal antibody, Biology (General), QD1-999, Aged
Abstract

Individuals with pre-existing chronic systemic low-grade inflammation are prone to develop severe COVID-19 and stronger anti-SARS-CoV-2 antibody responses. Whether this phenomenon reflects a differential expansion of antiviral B cells or a failure to regulate antibody synthesis remains unknown. Here, we compared the antiviral B cell repertoire of convalescent healthcare personnel to that of hospitalized patients with pre-existing comorbidities. Out of 277,500 immortalized B cell clones, antiviral B cell frequencies were determined by indirect immunofluorescence screening on SARS-CoV-2 infected cells. Surprisingly, frequencies of SARS-CoV-2 specific clones from the two groups were not statistically different, despite higher antibody levels in hospitalized patients. Moreover, functional analyses revealed that several B cell clones from healthcare personnel with low antibody levels had neutralizing properties. This study reveals for the first time a key qualitative defect of antibody synthesis in severe patients and calls for caution regarding estimated protective immunity based only on circulating antiviral antibodies.

Published
2021

8. Clinical management and viral genomic diversity analysis of a child's influenza A(H1N1)pdm09 infection in the context of a severe combined immunodeficiency

Author

Bruno Massenavette, Etienne Javouhey, Laurence Josset, Christophe Malcus, Caroline Picard, Bruno Simon, Florence Morfin-Sherpa, Martine Valette, Guillaume Monneret, Gilles Millat, Alexandre Gaymard, Bruno Lina, Cécile Renard, Maxime Pichon, Vanessa Escuret, Centre National de Référence des virus des infections respiratoires, Laboratoire de Virologie, Institut des Agents Infectieux, Groupement Hospitalier Nord, Hospices Civils de Lyon, Centre International de Recherche en Infectiologie - UMR (CIRI), École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut d'hématologie et d'oncologie pédiatrique [CHU - HCL] (IHOPe), Hospices Civils de Lyon (HCL), Hôpital Femme Mère Enfant [CHU - HCL] (HFME), Hospices Civils de Lyon, Laboratoire d'Immunologie, Groupement Hospitalier Edouard Herriot, 5 Place d'Arsonval, F-69437, Lyon Cedex 03, France, parent, Unité Mixte de Recherche Epidémiologique et de Surveillance Transport Travail Environnement (UMRESTTE UMR T9405), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut Français des Sciences et Technologies des Transports, de l'Aménagement et des Réseaux (IFSTTAR), Hospices Civils de Lyon, Plateforme de séquençage diagnostique, Centre de Biologie et de Pathologie Est, Groupement Hospitalier Est, F-69677, Bron, France, Centre International de Recherche en Infectiologie (CIRI), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, viruses, chemistry.chemical_compound, Influenza A Virus, H1N1 Subtype, NA-E119A SUBSTITUTION, Medicine, Zanamivir, Child, biology, virus diseases, Immunoglobulins, Intravenous, Viral Load, 3. Good health, Cord Blood Stem Cell Transplantation, Viral load, NEXT-GENERATION SEQUENCING, medicine.drug, Oseltamivir, NA-H275Y SUBSTITUTION, Mutation, Missense, Neuraminidase, Context (language use), Viral quasispecies, Real-Time Polymerase Chain Reaction, Antiviral Agents, Virus, 03 medical and health sciences, Viral Proteins, Virology, Drug Resistance, Viral, Influenza, Human, Humans, Pharmacology, Severe combined immunodeficiency, business.industry, OSELTAMIVIR RESISTANCE, Genetic Variation, medicine.disease, INFLUENZA A(H1N1)PDM09VIRUS, 030104 developmental biology, chemistry, INTRAVENOUS ZANAMIVIR, biology.protein, [SDV.SPEE]Life Sciences [q-bio]/Santé publique et épidémiologie, Severe Combined Immunodeficiency, business
Abstract

Introduction A child with severe combined immunodeficiency (SCID) had an influenza A(H1N1)pdm09 infection with viral excretion longer than 6 months, during 2013–2014 influenza season, despite cord blood transplantation and antiviral treatments. Methods Conventional real-time RT-PCR methods were used to estimate viral load and to detect the presence of the common N1 neuraminidase (NA) H275Y substitution responsible for oseltamivir resistance. Next-generation sequencing (NGS) of influenza viruses was performed retrospectively to characterize viral quasispecies in specimens. Results The patient was first treated with oral oseltamivir, leading to detection of low-levels of NA-H275Y substitution. Concomitant cord blood cell transplantation, intravenous administration of zanamivir and immunoglobulins led to an increase in white blood cells and influenza viral load decrease. A viral rebound occurred as soon as the antiviral treatment was discontinued. Eventually, influenza viral load was negated with immune reconstitution. NGS found influenza quasispecies harboring NA-E119A substitution (10.3%). Moreover, NGS showed that viral genomic diversity evolved under antiviral treatment and immune status. Conclusions Conventional virological techniques were sufficient for influenza infection follow-up but NGS performances allowed characterization of viral variants evolution in this specific case of prolonged influenza virus infection. New and efficient treatments against influenza in immunocompromised patients are needed.

Published
2018
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9. Clinical severity and molecular characteristics of circulating and emerging rotaviruses in young children attending hospital emergency departments in France

Author

Jérôme Kaplon, Florence Morfin-Sherpa, D. Gendrel, Christopher Payan, A. Schnuriger, Anne Decoster, Sylvie Pillet, J.-F. Meritet, Aurélie Guigon, François Dubos, Bruno Pozzetto, M.-C. Legrand-Guillien, Céline Fremy, C. Bouquignaud, Y. Gillet, M. Rodiere, Didier Hober, Y. Mekki, Ronald Colimon, Pierre Lebon, O. Mory, A. Beby-Defaux, Sophie Alain, Stéphane Bonacorsi, Bruno Lina, Gisèle Lagathu, J.-L. Stephan, M. Chouchane, Pierre Pothier, A. Garbarg-Chenon, L. Mendes-Martins, R. Maudinas, Jérôme Guinard, Frédéric Huet, Patricia Mariani, J. Languepin, A. Minoui-Tran, A. de Rougemont, Mouna Lazrek, Véronique Avettand-Fenoel, Vincent Foulongne, Gérard Agius, Astrid Vabret, Denis Oriot, Serge Aho, Mathie Lorrot, UFR des Sciences de Santé (Université de Bourgogne), Université de Bourgogne (UB), Centre National de Référence des virus entériques [CHU de Dijon] (CNR virus entériques), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Laboratoire de sérologie-virologie (CHU de Dijon), Procédés Alimentaires et Microbiologiques [Dijon] (PAM), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Laboratoire Interactions Muqueuses Agents Transmissibles (LIMA), Centre Hospitalier Régional Universitaire de Brest (CHRU Brest), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Laboratoire de Virologie Humaine et Moléculaire [Caen], CHU Caen, Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN), Centre Hospitalier de Charleville-Mezières, Service de pédiatrie (CHU de Dijon), Hôpital du Bocage, Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Université Cadi Ayyad [Marrakech] (UCA), Groupe Hospitalier de l'institut Catholique de Lille (GHICL), Université catholique de Lille (UCL), Teaching hospital, CHU Limoges, Service d'urgence et de réanimation pédiatriques [Hôpital Edouard Herriot - HCL], Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Virologie et pathogenèse virale (VPV), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Hospices Civils de Lyon (HCL), ONERA - The French Aerospace Lab [Palaiseau], ONERA-Université Paris Saclay (COmUE), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Université Pierre et Marie Curie - Paris 6 (UPMC), Laboratoire de Virologie, Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Institut Curie [Paris], Centre hospitalier universitaire de Poitiers (CHU Poitiers), CHU Pontchaillou [Rennes], Centre Hospitalier Universitaire de Saint-Etienne (CHU de Saint-Etienne), University Hospital and University Jean Monnet, Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Université Bourgogne Franche-Comté [COMUE] (UBFC), Université Grenoble Alpes (UGA), Palmipèdes à Foie Gras (UEPFG), Institut National de la Recherche Agronomique (INRA), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon, ONERA, Station de recherches sur la vache laitière, Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), CHU Cochin [AP-HP], Institut Curie, Groupe Hospitalier de l'Institut Catholique de Lille (GHICL), CHU Orléans, Pathogénèse et contrôle des infections chroniques (PCCI), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM)-Centre Hospitalier Universitaire de Montpellier (CHU Montpellier ), Centre Hospitalier Universitaire de Saint-Etienne [CHU Saint-Etienne] (CHU ST-E), and Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre Hospitalier Universitaire de Montpellier (CHU Montpellier )-Université de Montpellier (UM)

Subjects
0301 basic medicine, Male, Rotavirus, Pediatrics, Emerging rotavirus, medicine.disease_cause, Group A, Communicable Diseases, Emerging, Severity of Illness Index, Feces, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Genotype, Prevalence, Clinical severity, Acute gastroenteritis, Phylogeny, ComputingMilieux_MISCELLANEOUS, Incidence (epidemiology), General Medicine, Diarrhoea, 3. Good health, Infectious Diseases, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, Child, Preschool, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, Female, France, Seasons, Emergency Service, Hospital, Reassortant Viruses, Microbiology (medical), medicine.medical_specialty, Genotyping, 030106 microbiology, Rotavirus Infections, Severity, 03 medical and health sciences, medicine, Animals, Humans, business.industry, Infant, Newborn, Infant, Relative stability, 030104 developmental biology, business
Abstract

International audience; Group A rotavirus (RVA) is the leading cause of acute gastroenteritis in young children worldwide. A prospective surveillance network has been set up to investigate the virological and clinical features of RVA infections and to detect the emergence of potentially epidemic strains in France. From 2009 to 2014, RVA-positive stool samples were collected from 4800 children

Published
2016
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10. Genetic reassortment of influenza viruses with different origins or subtypes

Author

Bouscambert-Duchamp , Maude, Virologie et Pathologie Humaine ( VirPath ), École normale supérieure - Lyon ( ENS Lyon ) -Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Université Claude Bernard - Lyon I, Bruno Lina, Florence Morfin-Sherpa, STAR, ABES, Virologie et Pathologie Humaine (VirPath), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
[SDV.SA]Life Sciences [q-bio]/Agricultural sciences, [SDV.SA] Life Sciences [q-bio]/Agricultural sciences, A(H1N1)2009, viruses, Virus influenza, RT-PCR quantitative, virus diseases, Quantitative RT-PCR, Réassortiment génétique, respiratory tract diseases, A(H5N1), Reverse genetic, Genetic reassortment, Influenza virus, [ SDV.SA ] Life Sciences [q-bio]/Agricultural sciences, Génétique inverse
Abstract

In the context of A(H5N1) pandemics threat, an « avian flu and flu pandemics » project was proposed by LyonBioPole to develop influenza viruses characterization tools for vaccine production. To study genetic reassortment between influenza viruses, 3 reverse genetic systems of A(H3N2) human virus and A(H5N2) and A(H5N1) avian viruses were developed and reassortant viruses were produced. Their replicative capacities were evaluated using growth kinetics on MDCK cells with viral production quantification by real-time qRT-PCR. The A(H1N1)2009 emergence raises two questions about the acquisition by genetic reassortment of oseltamivir resistance and/or pathogenicity determinants. A co-infection protocol on MDCK cells was developed to study gene constellations of reassortant viruses like A(H1N1)2009-A(H1N1) H275Y and A(H1N1)2009-A(H5N1). We report here that genetic reassortment is possible between avian, human and swine strains using reverse genetic and viral co-infection and that some specific constellations emerged. We also report, that pandemic A(H1N1)2009 can acquire the H275Y mutated NA from seasonal oseltamivir resistant A(H1N1) viruses without any modifications on replicative capacities. This genetic reassortment is also possible with A(H5N1) viruses. These observations strenght the importance of vaccination against all these influenza strains to reduce the risk of one-individual viral co-infection., Dans le contexte de la menace pandémique liée au virus influenza A(H5N1), un projet «GRIPPE AVIAIRE ET GRIPPE PANDÉMIQUE » a émergé au sein de LyonBioPôle avec comme objectif le développement d’outils de caractérisation des virus influenza pour la production de vaccins. Pour étudier le réassortiment génétique entre virus influenza, nous avons développé 3 systèmes de génétique inverse : virus humain A(H3N2) et aviaires A(H5N2) et A(H5N1) et produit des virus réassortants de composition déterminée. Leurs capacités réplicatives ont été évaluées par cinétiques de croissance virale sur MDCK avec quantification de la production virale par qRT-PCR temps réel. L’émergence du virus influenza A(H1N1)2009 pose deux questions sur l’acquisition par réassortiment génétique, d’une résistance à l’oseltamivir d’une part ou de facteurs de virulence d’autre part. Nous avons donc développé un protocole de co-infection virale de cellules MDCK pour étudier les constellations de gènes des réassortants entre différents virus: A(H1N1)2009-A(H1N1) H275Y et A(H1N1)2009-A(H5N1). Nous montrons par deux approches différentes, génétique inverse et co-infections virales, que le réassortiment génétique entre souches aviaires et humaines et surtout aviaires et porcines est possible, en privilégiant certaines constellations. Nous rapportons que le virus pandémique peut acquérir la NA H275Y des virus A(H1N1) Brisbane-like résistants à l’oseltamivir sans que ses capacités de réplication ne soient altérées. De même nous montrons que son réassortiment avec un virus hautement pathogène A(H5N1) est possible. Ces observations renforcent la nécessité de promouvoir la vaccination afin de limiter les risques de co-infection virale chez un même individu.

Published
2010

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