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1. HBV and HBsAg strongly reshape the phenotype, function, and metabolism of DCs according to patients’ clinical stage

Author

Lucile Dumolard, Theophile Gerster, Florent Chuffart, Thomas Decaens, Marie-Noelle Hilleret, Sylvie Larrat, Philippe Saas, Evelyne Jouvin-Marche, David Durantel, Patrice N. Marche, Zuzana Macek Jilkova, and Caroline Aspord

Subjects
Diseases of the digestive system. Gastroenterology, RC799-869
Abstract

Background:. Hepatitis B is a liver infection caused by HBV. Infected individuals who fail to control the viral infection develop chronic hepatitis B and are at risk of developing life-threatening liver diseases, such as cirrhosis or liver cancer. Dendritic cells (DCs) play important roles in the immune response against HBV but are functionally impaired in patients with chronic hepatitis B. The underlying mechanisms involved in HBV-induced DC dysfunctions remain to be elucidated. Methods:. We explored DC modulations by HBV and HBsAg by exposing blood-derived cDC1s, cDC2s, and plasmacytoid DCs from healthy donors to HBV or HBsAg and stimulating them with toll-like receptor ligand. Their phenotypic and functional features, as well as their metabolic profile, were analyzed through multiparametric flow cytometry and multiplex assays and further explored on patients’ samples. Results:. We found that HBV deeply reshaped the DC secretome in response to toll-like receptor ligand. Strikingly, we observed that HBV-exposed DCs secrete high levels of CX3CL1 (fractalkine), a chemokine responsible for attracting antiviral effectors to the site of infection. HBsAg exposure favored DC activation while drastically altering TRAIL expression in response to toll-like receptor ligand and increasing the secretion of cytokines/chemokines involved in immune tolerance. HBsAg further dampened the metabolism of DC subsets while driving metabolic switches. Notably, the relevance of the CX3CL1/CX3CR1 axis, TGF-β, and metabolic disturbances was demonstrated within intrahepatic DC subsets in patients according to disease stage. Conclusions:. Our work brings new insights into the immunomodulation induced by HBV on DCs, which contribute to impaired antiviral responses and progression toward chronicity.

Published
2025
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3. Effect of a multimodal strategy for prevention of nosocomial influenza: a retrospective study at Grenoble Alpes University Hospital from 2014 to 2019

Author

Meghann Gallouche, Hugo Terrisse, Sylvie Larrat, Sylvie Marfaing, Christelle Di Cioccio, Bruno Verit, Patrice Morand, Vincent Bonneterre, Jean-Luc Bosson, and Caroline Landelle

Subjects
Healthcare-associated infection, Nosocomial influenza, Infection control, Multimodal strategy, Prevention, Infectious and parasitic diseases, RC109-216
Abstract

Abstract Background A multimodal strategy to prevent nosocomial influenza was implemented in 2015–2016 in Grenoble Alpes University Hospital. Three modalities were implemented in all units: promotion of vaccination among healthcare workers, epidemiologic surveillance and communication campaigns. Units receiving a high number of patients with influenza implemented 2 additional modalities: improvement of diagnosis capacities and systematic surgical mask use. The main objective was to assess the effectiveness of the strategy for reducing the risk of nosocomial influenza. Methods A study was conducted retrospectively investigating 5 epidemic seasons (2014–2015 to 2018–2019) including all patients hospitalized with a positive influenza test at Grenoble Alpes University Hospital. The weekly number of nosocomial influenza cases was analyzed by Poisson regression and incidence rate ratios (IRR) were estimated. Results A total of 1540 patients, resulting in 1559 stays, were included. There was no significant difference between the 5 influenza epidemic seasons in the units implementing only 3 measures. In the units implementing the 5 measures, there was a reduction of nosocomial influenza over the seasons when the strategy was implemented compared to the 2014–2015 epidemic season (IRR = 0.56, 95% CI = 0.23–1.34 in 2015–2016; IRR = 0.39, 95% CI = 0.19–0.81 in 2016–2017; IRR = 0.50, 95% CI = 0.24–1.03 in 2017–2018; IRR = 0.48, 95% CI = 0.23–0.97 in 2018–2019). Conclusions Our data mainly suggested that the application of the strategy with 5 modalities, including systematic surgical mask use and rapid diagnosis, seemed to reduce by half the risk of nosocomial influenza. Further data, including medico-economic studies, are necessary to determine the opportunity of extending these measures at a larger scale.

Published
2022
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4. SARS-CoV-2 Variants in Immunocompromised Patient Given Antibody Monotherapy

Author

Aurélie Truffot, Julien Andréani, Marion Le Maréchal, Alban Caporossi, Olivier Epaulard, Raphaele Germi, Pascal Poignard, and Sylvie Larrat

Subjects
coronavirus disease, COVID-19, severe acute respiratory syndrome coronavirus 2, SARS-CoV-2, coronaviruses, viruses, Medicine, Infectious and parasitic diseases, RC109-216
Abstract

A 72-year-old immunocompromised man infected with severe acute respiratory syndrome coronavirus 2 received bamlanivimab monotherapy. Viral evolution was monitored in nasopharyngeal and blood samples by melting curve analysis of single-nucleotide polymorphisms and whole-genome sequencing. Rapid emergence of spike receptor binding domain mutations was found, associated with a compartmentalization of viral populations.

Published
2021
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5. Screening of hepatitis E in patients presenting for acute neurological disorders

Author

Aude Belbézier, Alban Deroux, Françoise Sarrot-Reynauld, Barbara Colombe, Annick Bosseray, Claire Wintenberger, Perrine Dumanoir, Maxime Lugosi, Isabelle Boccon-Gibod, Vincent Leroy, Maxime Maignan, Roselyne Collomb-Muret, Damien Viglino, Mathieu Vaillant, Lorella Minotti, Emeline Lagrange, Olivier Epaulard, Chantal Dumestre-Perard, Sébastien Lhomme, Julien Lupo, Sylvie Larrat, Patrice Morand, Carole Schwebel, Antoine Vilotitch, Jean-Luc Bosson, and Laurence Bouillet

Subjects
Hepatitis E, Neurological disorder, Acute hepatitis, Hepatitis, Infectious and parasitic diseases, RC109-216, Public aspects of medicine, RA1-1270
Abstract

Introduction: Hepatitis E virus (HEV) infection has been reported to be associated with neurological disorders. However, the real prevalence of acute hepatitis E in those diseases is still unknown. We determined the prevalence of anti-HEV IgM antibody in a population with acute non-traumatic, non-metabolic, non-vascular neurological injury. Method: A registry was created in Grenoble Hospital University from 2014 to 2018 to collect data on patients with acute (

Published
2020
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6. Added Value of Next-Generation Sequencing for Multilocus Sequence Typing Analysis of a Pneumocystis jirovecii Pneumonia Outbreak

Author

Elena Charpentier, Cécile Garnaud, Claire Wintenberger, Sébastien Bailly, Jean-Benjamin Murat, John Rendu, Patricia Pavese, Thibault Drouet, Caroline Augier, Paolo Malvezzi, Anne Thiébaut-Bertrand, Marie-Reine Mallaret, Olivier Epaulard, Muriel Cornet, Sylvie Larrat, and Danièle Maubon

Subjects
Pneumocystis jirovecii, pneumocystosis, pneumonia, disease outbreaks, infectious diseases, high-throughput nucleotide sequencing, Medicine, Infectious and parasitic diseases, RC109-216
Abstract

Pneumocystis jirovecii is a major threat for immunocompromised patients, and clusters of pneumocystis pneumonia (PCP) have been increasingly described in transplant units during the past decade. Exploring an outbreak transmission network requires complementary spatiotemporal and strain-typing approaches. We analyzed a PCP outbreak and demonstrated the added value of next-generation sequencing (NGS) for the multilocus sequence typing (MLST) study of P. jirovecii strains. Thirty-two PCP patients were included. Among the 12 solid organ transplant patients, 5 shared a major and unique genotype that was also found as a minor strain in a sixth patient. A transmission map analysis strengthened the suspicion of nosocomial acquisition of this strain for the 6 patients. NGS-MLST enables accurate determination of subpopulation, which allowed excluding other patients from the transmission network. NGS-MLST genotyping approach was essential to deciphering this outbreak. This innovative approach brings new insights for future epidemiologic studies on this uncultivable opportunistic fungus.

Published
2017
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7. Diagnostic accuracy of a rapid RT-PCR assay for point-of-care detection of influenza A/B virus at emergency department admission: A prospective evaluation during the 2017/2018 influenza season.

Author

Maxime Maignan, Damien Viglino, Maud Hablot, Nicolas Termoz Masson, Anne Lebeugle, Roselyne Collomb Muret, Prudence Mabiala Makele, Valérie Guglielmetti, Patrice Morand, Julien Lupo, Virginie Forget, Caroline Landelle, and Sylvie Larrat

Subjects
Medicine, Science
Abstract

Study objectiveTo investigate the performance of a rapid RT-PCR assay to detect influenza A/B at emergency department admission.MethodsThis single-center prospective study recruited adult patients attending the emergency department for influenza-like illness. Triage nurses performed nasopharyngeal swab samples and ran rapid RT-PCR assays using a dedicated device (cobas Liat, Roche Diagnostics, Meylan, France) located at triage. The same swab sample was also analyzed in the department of virology using conventional RT-PCR techniques. Patients were included 24 hours-a-day, 7 days-a-week. The primary outcome was the diagnostic accuracy of the rapid RT-PCR assay performed at triage.ResultsA total of 187 patients were included over 11 days in January 2018. Median age was 70 years (interquartile range 44 to 84) and 95 (51%) were male. Nine (5%) assays had to be repeated due to failure of the first assay. The sensitivity of the rapid RT-PCR assay performed at triage was 0.98 (95% confidence interval (CI): 0.91-1.00) and the specificity was 0.99 (95% CI: 0.94-1.00). A total of 92 (49%) assays were performed at night-time or during the weekend. The median time from patient entry to rapid RT-PCR assay results was 46 [interquartile range 36-55] minutes.ConclusionRapid RT-PCR assay performed by nurses at triage to detect influenza A/B is feasible and highly accurate.

Published
2019
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8. Hidden Selection of Bacterial Resistance to Fluoroquinolones In Vivo: The Case of Legionella pneumophila and Humans

Author

Lubana Shadoud, Iyad Almahmoud, Sophie Jarraud, Jérôme Etienne, Sylvie Larrat, Carole Schwebel, Jean-François Timsit, Dominique Schneider, and Max Maurin

Subjects
Legionella pneumophila, Legionellosis, Fluoroquinolone resistance, In vivo selection, Resistance detection, Next generation sequencing, Medicine, Medicine (General), R5-920
Abstract

Background: Infectious diseases are the leading cause of human morbidity and mortality worldwide. One dramatic issue is the emergence of microbial resistance to antibiotics which is a major public health concern. Surprisingly however, such in vivo adaptive ability has not been reported yet for many intracellular human bacterial pathogens such as Legionella pneumophila. Methods: We examined 82 unrelated patients with Legionnaire's disease from which 139 respiratory specimens were sampled during hospitalization and antibiotic therapy. We both developed a real time PCR assay and used deep-sequencing approaches to detect antibiotic resistance mutations in L. pneumophila and follow their selection and fate in these samples. Findings: We identified the in vivo selection of fluoroquinolone resistance mutations in L. pneumophila in two infected patients treated with these antibiotics. By investigating the mutational dynamics in patients, we showed that antibiotic resistance occurred during hospitalization most likely after fluoroquinolone treatment. Interpretation: In vivo selection of antibiotic resistances in L. pneumophila may be associated with treatment failures and poor prognosis. This hidden resistance must be carefully considered in the therapeutic management of legionellosis patients and in the control of the gradual loss of effectiveness of antibiotics.

Published
2015
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9. Evaluation of the cobas® GT hepatitis C virus genotyping assay in G1-6 viruses including low viral loads and LiPA failures.

Author

Benjamin Némoz, Léa Roger, Vincent Leroy, Jean-Dominique Poveda, Patrice Morand, and Sylvie Larrat

Subjects
Medicine, Science
Abstract

Direct-acting antiviral (DAA) drug performances depend on the viral genotype. So international recommendations give typing of the virus a prerequisite for treatment choice and patient management. Commercially available HCV genotyping kits are scarce and this analysis is often in-house using tedious PCRs and Sanger sequencing, leading to a lack of standardization. A newly commercialized HCV genotyping assay based on real-time PCR has been developed by Roche Diagnostics (Mannheim, Germany). We compared this new assay with our in-house PCRs -sequencing technique on 101 regular samples and 81 LiPA failures or low viral load samples. No genotype or 1a/1b subtype mismatch was observed. Two samples were misidentified at the subtype level without clinical impact. Three genotype 1b and two genotype 1a samples with low viral load could not be subtyped. Nevertheless, 13 (13%) samples from the regular panel and 35 (43%) from the more difficult-to-type panels failed to give results on first pass with the Roche kit. Failures were mostly associated with genotype 3 subtype a, with genotype 4 subtype non-a, or with viral loads

Published
2018
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10. Natural non-homologous recombination led to the emergence of a duplicated V3-NS5A region in HCV-1b strains associated with hepatocellular carcinoma.

Author

Hélène Le Guillou-Guillemette, Adeline Pivert, Elise Bouthry, Cécile Henquell, Odile Petsaris, Alexandra Ducancelle, Pascal Veillon, Sophie Vallet, Sophie Alain, Vincent Thibault, Florence Abravanel, Arielle A Rosenberg, Elisabeth André-Garnier, Jean-Baptiste Bour, Yazid Baazia, Pascale Trimoulet, Patrice André, Catherine Gaudy-Graffin, Dominique Bettinger, Sylvie Larrat, Anne Signori-Schmuck, Hénia Saoudin, Bruno Pozzetto, Gisèle Lagathu, Sophie Minjolle-Cha, Françoise Stoll-Keller, Jean-Michel Pawlotsky, Jacques Izopet, Christopher Payan, Françoise Lunel-Fabiani, and Christophe Lemaire

Subjects
Medicine, Science
Abstract

BACKGROUND:The emergence of new strains in RNA viruses is mainly due to mutations or intra and inter-genotype homologous recombination. Non-homologous recombinations may be deleterious and are rarely detected. In previous studies, we identified HCV-1b strains bearing two tandemly repeated V3 regions in the NS5A gene without ORF disruption. This polymorphism may be associated with an unfavorable course of liver disease and possibly involved in liver carcinogenesis. Here we aimed at characterizing the origin of these mutant strains and identifying the evolutionary mechanism on which the V3 duplication relies. METHODS:Direct sequencing of the entire NS5A and E1 genes was performed on 27 mutant strains. Quasispecies analyses in consecutive samples were also performed by cloning and sequencing the NS5A gene for all mutant and wild strains. We analyzed the mutant and wild-type sequence polymorphisms using Bayesian methods to infer the evolutionary history of and the molecular mechanism leading to the duplication-like event. RESULTS:Quasispecies were entirely composed of exclusively mutant or wild-type strains respectively. Mutant quasispecies were found to have been present since contamination and had persisted for at least 10 years. This V3 duplication-like event appears to have resulted from non-homologous recombination between HCV-1b wild-type strains around 100 years ago. The association between increased liver disease severity and these HCV-1b mutants may explain their persistence in chronically infected patients. CONCLUSIONS:These results emphasize the possible consequences of non-homologous recombination in the emergence and severity of new viral diseases.

Published
2017
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13. Hepatitis E Virus Infection after Platelet Transfusion in an Immunocompetent Trauma Patient

Author

Emmanuelle Loyrion, Thibaut Trouve-Buisson, Patricia Pouzol, Sylvie Larrat, Thomas Decaens, and Jean-Francois Payen

Subjects
hepatitis E virus, platelet transfusion, trauma, injury, HEV, opportunistic infection, Medicine, Infectious and parasitic diseases, RC109-216
Abstract

Hepatitis E virus (HEV) infection causes acute liver disease, but severe infections are rare in immunocompetent patients. We describe a case of HEV infection in a previously healthy male trauma patient in France who received massive transfusions. Genotyping confirmed HEV in a transfused platelet pool and the donor.

Published
2017
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14. Hepatitis E Virus Infection in Sheltered Homeless Persons, France

Author

Mamadou Kaba, Philippe Colson, Sylvie Larrat, Stéphanie Gaillard, Monique Baccard, Lionel Piroth, Patrice Cacoub, Stanislas Pol, Christian Perronne, Fabrice Carrat, and Patrice Morand

Subjects
Hepatitis E virus, homeless persons, HEV transmission, autochthonous hepatitis, injection drug use, France, Medicine, Infectious and parasitic diseases, RC109-216
Published
2012
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16. Persistence at one year of neutralizing antibodies after SARS-CoV-2 infection: Influence of initial severity and steroid use

Author

Myriam Blanc, Sylvie Larrat, Jean-François Payen, Isabelle Pierre, Marion Le Maréchal, Pascal Poignard, Patricia Pavese, Marlyse Buisson, Olivier Epaulard, Nicolas Terzi, Raphaele Germi, Benjamin Nemoz, Laurence Grossi, Marie Froidure, Julien Lupo, Fanny Quénard, and Anne-Laure Mounayar

Subjects
Microbiology (medical), 2019-20 coronavirus outbreak, biology, Coronavirus disease 2019 (COVID-19), business.industry, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Virology, Persistence (computer science), Infectious Diseases, Steroid use, biology.protein, Medicine, Antibody, business
Published
2022
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17. The onset of late severe lung impairment in COVID-19 is associated with high inflammation markers at admission and metabolic syndrome markers

Author

Olivier Epaulard, Audrey Le Gouellec, Marion Le Marechal, Benjamin Nemoz, Anne-Laure Borel, Anaïs Dartevel, Hubert Gheerbrant, Marie-Christine Herault, Annick Bosseray, Giovanna Clavarino, Julien Lupo, Damien Viglino, Fanny Quenard, Clara Candille, Boubou Camara, Michel Durand, Patrice Faure, Dorra Guergour, Elena Chidlovski, Marie-Christine Jacob, Sylvie Larrat, Marie Froidure, Nicolas Terzi, Sébastien Quetant, Jean-François Payen, Barbara Colombe, Tatiana Raskovalova, Patrice Morand, Isabelle Pierre, Carole Schwebel, Rebecca Hamidfar, Laurence Bouillet, Jean-Paul Brion, Candice Trocme, Sylvie Berthier, Carole Chirica, Anne-Laure Mounayar, Myriam Blanc, Patricia Pavese, and Bertrand Toussaint

Abstract

BackgroundCOVID-19 severity is mainly related to lung impairment. However, preexisting patient characteristics and biomarkers at admission associated with this event are not precisely known.MethodsWe report 205 patients admitted for a proven COVID-19 in our institution between March 7 and April 22, 2020, particularly their comorbidities, respiratory severity, immune profile, and metabolic profile.FindingsMedian age was 70 years [interquartile range (IQR) 25-75: 60;79]; 115 (56·1%) patients were men. Oxygen supplementation of >2L/min was required in 107 patients (52·2%) after a median time of 8 days [IQR: 6;10] after the first symptoms; 67 (32·7%) patients were admitted to the intensive care unit (ICU), almost exclusively due to severe hypoxia. Patients requiring >2L/min oxygen therapy and/or ICU admission were older and more frequently males, with a significantly higher body mass index (BMI), a significantly higher total cholesterol (TC) / HDL cholesterol ratio, and higher triglycerides. They also had higher plasma levels of C-reactive protein (CRP) and interleukin 6 (IL-6); IL-6 >20 ng/L and CRP >70 mg/L were significantly associated with ICU admission and/or (for patients with a decision of limitation of life-support therapy) death. Higher BMI and TC/HDL-c ratio were associated with higher CRP and IL-6 levels. Steroid therapy was performed in 61 patients; while its clinical impact was inconclusive due to heterogeneous situations, IL-6 levels decreased significantly more in these patients.InterpretationSevere COVID-19 mostly relates to late-onset pneumonia associated with preexisting metabolic syndrome markers and a surge in inflammatory markers, allowing the early identification of at-risk patients.FundingThis work was supported by Foundation University of Grenoble Alpes.

Published
2022
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18. COVID-19 Geographical Maps and Clinical Data Warehouse PREDIMED

Author

Svetlana, Artemova, Alban, Caporossi, Christophe, Cancé, Pierre-Ephrem, Madiot, Benjamin, Nemoz, Sylvie, Larrat, Alexandre, Moreau-Gaudry, and Jean-Luc, Bosson

Subjects
Hospitals, University, Geography, Data Warehousing, COVID-19, Humans
Abstract

PREDIMED, Clinical Data Warehouse of Grenoble Alps University Hospital, is currently participating in daily COVID-19 epidemic follow-up via spatial and chronological analysis of geographical maps. This monitoring is aimed for cluster detection and vulnerable population discovery. Our real-time geographical representations allow us to track the epidemic both inside and outside the hospital.

Published
2022

19. SARS-CoV-2 Variants in Immunocompromised Patient Given Antibody Monotherapy

Author

Sylvie Larrat, Pascal Poignard, Julien Andreani, Aurélie Truffot, Raphaële Germi, Alban Caporossi, Olivier Epaulard, and Marion Le Maréchal

Subjects
Microbiology (medical), Male, Epidemiology, medicine.drug_class, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), medicine.medical_treatment, viruses, Expedited, coronaviruses, selection, Infectious and parasitic diseases, RC109-216, Monoclonal antibody, Antibodies, Viral, Melting curve analysis, SARS-CoV-2 Variants in Immunocompromised Patient Given Antibody Monotherapy, respiratory infections, Immunocompromised Host, Research Letter, Medicine, Humans, Aged, Whole genome sequencing, variants, biology, business.industry, SARS-CoV-2, COVID-19, Immunocompromised patient, Immunotherapy, Virology, compartmentalization, zoonoses, Infectious Diseases, coronavirus disease, monoclonal antibody, whole-genome sequencing, Viral evolution, monotherapy, Spike Glycoprotein, Coronavirus, biology.protein, immunotherapy, bamlanivimab, Antibody, business, severe acute respiratory syndrome coronavirus 2
Abstract

A 72-year-old immunocompromised man infected with severe acute respiratory syndrome coronavirus 2 received bamlanivimab monotherapy. Viral evolution was monitored in nasopharyngeal and blood samples by melting curve analysis of single-nucleotide polymorphisms and whole-genome sequencing. Rapid emergence of spike receptor binding domain mutations was found, associated with a compartmentalization of viral populations.

Published
2021

20. Risk factors of nosocomial Covid-19 in a French university hospital: a case- control study

Author

Claire Dinh, Meghann Gallouche, Hugo Terrisse, Kamal Gam, Céline Giner, Benjamin Nemoz, Sylvie Larrat, Joris Giai, Jean-Luc Bosson, and Caroline Landelle

Abstract

Background: During the Covid-19 pandemic, prevention strategies implemented by hospitals to reduce nosocomial transmission sometimes failed and determining transmission risk factors remains crucial. Our objective was to determine the risk factors of nosocomial Covid-19.Methods: A case-control study was conducted in a French hospital between 09/01/2020 and 01/31/2021. Adult patients hospitalized in medical or surgical units were included. Infants or patients hospitalized in ICU were excluded. Cases were patients with a nosocomial Covid-19 (clinical symptoms and RT-PCR positive for SARS-CoV-2 or RT-PCR positive for SARS-CoV-2 with Ct ≤28 more than five days after admission); controls were patients without infection (RT-PCR negative for SARS-CoV-2 more than 5 days after admission). They were matched according to length of stay before diagnosis and period of admission. Analyses were performed with a conditional logistic regression.Results: A total of 281 cases and 441 controls were included. In the bivariate analysis, cases were older (OR per 10 years: 1.22; CI95% [1.10; 1.36]), had more often shared a room (OR: 1.74; CI95% [1.25; 2.43]), had more often a risk factor of severe Covid-19 (OR: 1.94; CI95% [1.09; 3.45]), were more often hospitalized in medical units [OR: 1.59; CI95% [1.12; 2.25]), had a higher exposure to contagious health care workers (HCW; OR per 1 person.days: 1.12; CI95% [1.08; 1.17]) and contagious patients (OR per 1 person.days: 1.11; CI95% [1.08; 1.14]) than controls. In an adjusted model, risk factors of nosocomial Covid-19 were exposure to contagious HCW (aOR per 1 person.days: 1.08; CI95% [1.03; 1.14]) and exposure to contagious patients (aOR per 1 person.days: 1.10; CI95% [1.07; 1.13]).Conclusions: Exposure to contagious professionals and contagious patients are the main risk factors for nosocomial Covid-19, outweighing all other potential risk factors including the hospitalisation in double room. Prevention strategies need to be adjusted according to these results to decrease the risk of nosocomial COVID-19.Trial registrationStudy ethics approval was obtained retrospectively on 22 December 2021 (CECIC Rhône-Alpes-Auvergne, Clermont-Ferrand, IRB 5891).

Published
2022
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21. Use of RT-PCR thermocycling program for the quantification of DNA viruses in a single run with RNA viruses: Example of Altona RealStar® HSV or VZV PCR kits

Author

Julien Andreani, Julien Lupo, Benjamin Nemoz, Aurelie Truffot, Sylvie Larrat, Patrice Morand, and Raphaële Germi

Subjects
Infectious Diseases, Reverse Transcriptase Polymerase Chain Reaction, DNA Viruses, Humans, RNA Viruses, Real-Time Polymerase Chain Reaction, Sensitivity and Specificity
Abstract

Real-time PCR plays a key role in the diagnosis of viral infections. Multiple kits can detect or quantify genomes of various viruses with the same thermocycling program. Detection of RNA viruses includes an additional step of reverse transcription and challenge their detection in a single run with DNA viruses. We investigated the analytical performance of HSV-1, HSV-2 and VZV DNA quantification with Altona RealStar® PCR kits using the RT-PCR program for RNA viruses instead of the PCR program for DNA viruses. For each three viruses, Bland-Altman distribution did not show differences between both programs, and quantification curves generated with both thermocycling programs confirmed high correlation (R

Published
2022

23. Evaluation of six commercial SARS-CoV-2 rapid antigen tests in nasopharyngeal swabs: better knowledge for better patient management?

Author

Julien Lupo, Julien Andreani, Marie Roccon, Benjamin Nemoz, Raphaële Germi, Christel Laugier, Patrice Morand, and Sylvie Larrat

Subjects
Rapid Antigen tests, medicine.medical_specialty, Coronavirus disease 2019 (COVID-19), Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Concordance, RT-PCR, Sensitivity and Specificity, Article, Antigen, Nasopharynx, Virology, Pandemic, medicine, Humans, Viral shedding, Intensive care medicine, Antigens, Viral, Pandemics, Retrospective Studies, business.industry, SARS-CoV-2, COVID-19, Retrospective cohort study, Triage, Infectious Diseases, business
Abstract

Robust antigen point-of-care SARS-CoV-2 tests have been proposed as an efficient tool to address the COVID-19 pandemic. This requirement was raised after acknowledging the constraints that are brought by molecular biology. However, worldwide markets have been flooded with cheap and potentially underperforming lateral flow assays. Herein we retrospectively compared the overall performance of five qualitative rapid antigen SARS-CoV-2 assays and one quantitative automated test on 239 clinical swabs. While the overall sensitivity and specificity are relatively similar for all tests, concordance with molecular based methods varies, ranging from 75,7% to 83,3% among evaluated tests. Sensitivity is greatly improved when considering patients with higher viral excretion (Ct≤33), proving that antigen tests accurately distinguish infectious patients from viral shedding. These results should be taken into consideration by clinicians involved in patient triage and management, as well as by national authorities in public health strategies and for mass campaign approaches.

Published
2021

24. Is it possible to hospitalize patients in multiple-bed room without increasing the risk of hospital-acquired influenza? Description of a pragmatic preventive strategy in a French university hospital

Author

Othmane Azzam, Caroline Landelle, Sylvie Larrat, Meghann Gallouche, Marie Reine Mallaret, Patrice Morand, Céline Giner, Fabiana Cazzorla, and Emeline Buet

Subjects
Oseltamivir, Epidemiology, Health Personnel, Detection bias, Beds, Antiviral Agents, Hospitals, University, chemistry.chemical_compound, Health care, Influenza, Human, medicine, Humans, Preventive strategy, business.industry, Health Policy, Public Health, Environmental and Occupational Health, virus diseases, Emergency department, University hospital, medicine.disease, Surgical mask, Infectious Diseases, Physical Barrier, chemistry, Medical emergency, business
Abstract

Background Large inrush of patients through Emergency Department during influenza season can be dramatic. The purpose of this study was to evaluate the impact of an emergency preventive strategy, namely admission of patients with influenza in multiple-bed room with patients free from influenza, on the occurrence of hospital-acquired influenza (HAI). Methods When a patient with an influenza RT-PCR diagnosis was hospitalized in a multiple-bed room, the emergency preventive strategy was applied: selection of non-immunocompromised neighbor, implementation of physical barriers (rigid screen pulled between beds, surgical mask for healthcare workers and visitors), preemptive Oseltamivir therapy for the neighbor. Results From 29/11/2017 to 10/05/2018 a total of 464 hospitalized influenza patients were included; 318 were placed in multiple-bed room and 141 in single room. Emergency preventive strategy was correctly applied for 75.1% of patients in multiple-bed room. A total of 8 exposed neighbors matched HAI definition despite strategy. 7 were already exposed to the case before the set-up of the strategy. Only one case of documented transmission of influenza occurred after application of an incorrect emergency preventive strategy: preventive posology of Oseltamivir was not correct. Conclusions These preliminary results suggest that the occurrence of HAI in multiple-bed rooms can be limited by the implementation of maximum precautions and urge us to promote further evaluation of the strategy. A detection bias should be considered without a systematic neighbors monitoring.

Published
2021

25. SARS-CoV-2 nosocomial infection acquired in a French university hospital during the 1st wave of the Covid-19 pandemic, a prospective study

Author

Benjamin Nemoz, C. Giner, Meghann Gallouche, A Landoas, Sylvie Larrat, Olivier Epaulard, Patricia Pavese, M.R. Mallaret, Patrice Morand, F Cazzorla, Caroline Landelle, and M Le Maréchal

Subjects
Microbiology (medical), Male, medicine.medical_specialty, media_common.quotation_subject, Health Personnel, Infectious and parasitic diseases, RC109-216, Hospitals, University, COVID-19 Testing, Hygiene, Interquartile range, Pandemic, medicine, Infection control, Humans, Pharmacology (medical), Hand Hygiene, Prospective Studies, Healthcare-associated infection, Prospective cohort study, Pandemics, media_common, Aged, Retrospective Studies, Aged, 80 and over, Cross Infection, Infection Control, Mask, business.industry, SARS-CoV-2, Research, Public Health, Environmental and Occupational Health, Masks, COVID-19, Retrospective cohort study, Outbreak, Middle Aged, Surgical mask, Infectious Diseases, Emergency medicine, Observational study, Female, France, business
Abstract

Background In healthcare facilities, nosocomial transmissions of respiratory viruses are a major issue. SARS-CoV-2 is not exempt from nosocomial transmission. Our goals were to describe COVID-19 nosocomial cases during the first pandemic wave among patients in a French university hospital and compliance with hygiene measures. Methods We conducted a prospective observational study in Grenoble Alpes University Hospital from 01/03/2020 to 11/05/2020. We included all hospitalised patients with a documented SARS-CoV-2 diagnosis. Nosocomial case was defined by a delay of 5 days between hospitalisation and first symptoms. Hygiene measures were evaluated between 11/05/2020 and 22/05/2020. Lockdown measures were effective in France on 17/03/2020 and ended on 11/05/2020. Systematic wearing of mask was mandatory for all healthcare workers (HCW) and visits were prohibited in our institution from 13/03/2021 and for the duration of the lockdown period. Results Among 259 patients included, 14 (5.4%) were considered as nosocomial COVID-19. Median time before symptom onset was 25 days (interquartile range: 12–42). Eleven patients (79%) had risk factors for severe COVID-19. Five died (36%) including 4 deaths attributable to COVID-19. Two clusters were identified. The first cluster had 5 cases including 3 nosocomial acquisitions and no tested HCWs were positive. The second cluster had 3 cases including 2 nosocomial cases and 4 HCWs were positive. Surgical mask wearing and hand hygiene compliance were adequate for 95% and 61% of HCWs, respectively. Conclusions The number of nosocomial COVID-19 cases in our hospital was low. Compliance regarding mask wearing, hand hygiene and lockdown measures drastically reduced transmission of the virus. Monitoring of nosocomial COVID-19 cases during the first wave enabled us to determine to what extent the hygiene measures taken were effective and patients protected. Trial registration Study ethics approval was obtained retrospectively on 30 September 2020 (CECIC Rhône-Alpes-Auvergne, Clermont-Ferrand, IRB 5891).

Published
2020

26. Screening of hepatitis E in patients presenting for acute neurological disorders

Author

Sébastien Lhomme, Chantal Dumestre-Pérard, Vincent Leroy, Lorella Minotti, Emeline Lagrange, Patrice Morand, Jean-Luc Bosson, M. Lugosi, Claire Wintenberger, Roselyne Collomb-Muret, Aude Belbézier, Maxime Maignan, Damien Viglino, Françoise Sarrot-Reynauld, Sylvie Larrat, Perrine Dumanoir, Isabelle Boccon-Gibod, Alban Deroux, Carole Schwebel, A. Bosseray, Antoine Vilotitch, Mathieu Vaillant, Julien Lupo, Laurence Bouillet, Olivier Epaulard, B. Colombe, Clinique de médecine interne, Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble, Centre Hospitalier Universitaire [Grenoble] (CHU), Pôle Psychiatrie et Neurologie [Grenoble], Service d'immunologie [CHU Grenoble], CHU de Grenoble, Centre de Physiopathologie Toulouse Purpan (CPTP), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Virologie [Toulouse], CHU Toulouse [Toulouse], Département de virologie [Grenoble], Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Radiopharmaceutiques biocliniques (LRB), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA), Translational Innovation in Medicine and Complexity / Recherche Translationnelle et Innovation en Médecine et Complexité - UMR 5525 (TIMC ), VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP ), Université Grenoble Alpes (UGA), Biologie du Cancer et de l'Infection (BCI ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Laboratoire Virologie [CHU Toulouse], Institut Fédératif de Biologie (IFB), Centre Hospitalier Universitaire de Toulouse (CHU Toulouse)-Centre Hospitalier Universitaire de Toulouse (CHU Toulouse)-Pôle Biologie [CHU Toulouse], Centre Hospitalier Universitaire de Toulouse (CHU Toulouse), and Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications Grenoble - UMR 5525 (TIMC-IMAG)

Subjects
0301 basic medicine, Male, viruses, Neurological disorder, medicine.disease_cause, Serology, Hepatitis, 0302 clinical medicine, Hepatitis E virus, Seroepidemiologic Studies, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, 030212 general & internal medicine, Prospective Studies, Acute hepatitis, MESH: Acute Disease / epidemiology, Adult, Aged, Aged, 80 and over, Brachial Plexus Neuritis / epidemiology, Female, France / epidemiology, Guillain-Barre Syndrome / epidemiology, Hepatitis Antibodies / blood, Hepatitis E / blood, Hepatitis E / diagnosis, Hepatitis E / epidemiology, Hepatitis E / immunology, Hepatitis E virus / immunology, Humans Immunoglobulin M / blood, Middle Aged, Nervous System Diseases / epidemiology, Nervous System Diseases / immunology, RNA, Viral / blood, Reverse Transcriptase Polymerase Chain Reaction, Transaminases / blood, Young Adult, education.field_of_study, lcsh:Public aspects of medicine, virus diseases, General Medicine, Hepatitis E, 3. Good health, Infectious Diseases, Acute Disease, RNA, Viral, France, medicine.medical_specialty, 030106 microbiology, Population, Guillain-Barre Syndrome, lcsh:Infectious and parasitic diseases, 03 medical and health sciences, Internal medicine, medicine, Seroprevalence, Brachial Plexus Neuritis, Humans, lcsh:RC109-216, Hepatitis Antibodies, education, Transaminases, business.industry, Public Health, Environmental and Occupational Health, lcsh:RA1-1270, medicine.disease, Immunoglobulin M, Elevated transaminases, Nervous System Diseases, business
Abstract

International audience; Introduction: Hepatitis E virus (HEV) infection has been reported to be associated with neurological disor-ders. However, the real prevalence of acute hepatitis E in those diseases is still unknown. We determinedthe prevalence of anti-HEV IgM antibody in a population with acute non-traumatic, non-metabolic,non-vascular neurological injury.Method: A registry was created in Grenoble Hospital University from 2014 to 2018 to collect data onpatients with acute (

Published
2020
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27. Baseline and Post-Treatment Hepatitis C NS5A Resistance in Relapsed Patients from a Multicentric Real-Life Cohort

Author

Philippe Halfon, Jacques Izopet, Pascale Trimoulet, Caroline Scholtes, Sofiane Mohamed, Denis Ouzan, Laurent Chiche, Victor de Ledinghen, Hacène Khiri, Marc Bourlière, M.A. Thelu, Sophie Metivier, Guillaume Penaranda, Anne Plauzolles, Laurent Alric, Vincent Leroy, Fabien Zoulim, Sylvie Larrat, Florence Abravanel, ALPHABIO - Laboratoire de biologie médicale, Centre de Recherche en Cancérologie de Lyon (CRCL), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre Léon Bérard [Lyon]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Hôpital Purpan [Toulouse], CHU Toulouse [Toulouse], Institut de biologie structurale (IBS - UMR 5075), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS), Service de virologie et d'immunologie biologique, CHU Bordeaux [Bordeaux]-Groupe hospitalier Pellegrin, Service d'Hépatologie et de Gastroentérologie [Lyon], Hospices Civils de Lyon (HCL), Institut de Recherche en Santé Digestive (IRSD ), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Recherche Agronomique (INRA)-Ecole Nationale Vétérinaire de Toulouse (ENVT), Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM), Service de Gastro-entérologie - Hépatologie [Purpan], CHU Toulouse [Toulouse]-Hôpital Purpan [Toulouse], Hôpital Michallon, Institut Arnault Tzanck, Service d'Hépato-Gastro-Entérologie, CHU Bordeaux [Bordeaux]-Hôpital Saint-André, Institut d'oncologie/développement Albert Bonniot de Grenoble (INSERM U823), Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble-EFS-Institut National de la Santé et de la Recherche Médicale (INSERM), Service de médecine interne [Hôpital Nord - APHM], Aix Marseille Université (AMU)-Assistance Publique - Hôpitaux de Marseille (APHM)- Hôpital Nord [CHU - APHM], Service d'hépatologie et de gastroentérologie [Hôpital Saint-Joseph - Marseille], Aix Marseille Université (AMU)-Hôpital Saint-Joseph [Marseille], Centre de Physiopathologie Toulouse Purpan ex IFR 30 et IFR 150 (CPTP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Hôpital Européen [Fondation Ambroise Paré - Marseille], Centre Hospitalier Universitaire de Lyon (CHU Lyon), Centre Hospitalier Universitaire de Toulouse (CHU Toulouse), Centre Hospitalier Universitaire [Grenoble] (CHU), CHU Bordeaux [Bordeaux], Pharmacochimie et Biologie pour le Développement (PHARMA-DEV), Institut de Recherche pour le Développement (IRD)-Institut de Chimie de Toulouse (ICT), Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université de Toulouse (UT)-Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT), Hôpital Haut-Lévêque [CHU Bordeaux], Hôpital Saint-Joseph [Marseille], Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut de Chimie de Toulouse (ICT-FR 2599), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Institut de Chimie du CNRS (INC)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche pour le Développement (IRD)-Institut de Chimie du CNRS (INC)-Institut de Recherche pour le Développement (IRD), Centre de Recherche en Cancérologie de Lyon (UNICANCER/CRCL), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre Léon Bérard [Lyon], Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Institut de Recherche pour le Développement (IRD)-Institut de Chimie de Toulouse (ICT-FR 2599), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées, Physiopathologie du cancer du foie, Université Bordeaux Segalen - Bordeaux 2-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut National de la Santé et de la Recherche Médicale (INSERM)-EFS-CHU Grenoble-Université Joseph Fourier - Grenoble 1 (UJF), Laboratoire de Virologie [CHU Purpan, Toulouse], Institut Fédératif de Biologie (IFB) - Hôpital Purpan, CHU Toulouse [Toulouse]-CHU Toulouse [Toulouse]-Hôpital Purpan [Toulouse], CHU Toulouse [Toulouse]-CHU Toulouse [Toulouse], Centre Léon Bérard [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 National de la Recherche Agronomique (INRA)-Université Toulouse III - Paul Sabatier (UT3), and Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Ecole Nationale Vétérinaire de Toulouse (ENVT)

Subjects
Male, 0301 basic medicine, DNA Mutational Analysis, Viral Nonstructural Proteins / genetics, Drug Resistance, Hepacivirus, Drug resistance, Viral Nonstructural Proteins, Virological response, Viral Nonstructural Proteins / antagonists & inhibitors, 0302 clinical medicine, Medicine, Pharmacology (medical), Viral, Treatment Failure, Hepatitis C, Viral Load, [SDV.SP]Life Sciences [q-bio]/Pharmaceutical sciences, 3. Good health, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Treatment Outcome, Infectious Diseases, Antiviral Agents / pharmacology, Cohort, Drug Therapy, Combination, Female, 030211 gastroenterology & hepatology, Viral load, Hepacivirus / genetics, medicine.medical_specialty, Genotype, Antiviral Agents / therapeutic use, Antiviral Agents, 03 medical and health sciences, Pharmacotherapy, Drug Therapy, Internal medicine, Drug Resistance, Viral, Humans, NS5A, Pharmacology, business.industry, medicine.disease, 030104 developmental biology, Hepatitis C / virology, Hepacivirus / drug effects, Post treatment, Hepatitis C / drug therapy, business
Abstract

Background Recent data have suggested that failure to achieve sustained virological response with direct-acting antiviral therapy is usually due to relapse and is primarily associated with the emergence of resistance-associated substitutions. The aim of this study was to investigate the prevalence and characterization of non-structural-5A resistance-associated substitutions in patients infected with HCV genotypes 1, 3 and 4 treated by direct-acting antiviral therapy, including anti-non-structural-5A, and to characterize the pre-existing resistance-associated substitutions in subjects treated with anti-non-structural-5A inhibitors. Methods From January 2014 to March 2016, 2,995 patients infected with HCV genotypes 1, 3 and 4 were exposed to non-structural-5A inhibitors. Sequencing results at the time of virological failure were available for 61 patients; sequencing at baseline was available for 35 of these patients. Results Among the 35 patients with sequencing results available at baseline, 15 had no resistance-associated substitution, 16 had only one resistance-associated substitution, and 4 had more than one resistance-associated substitution. Resistance-associated substitutions were harbored in 57% of the sequences in the non-structural-5A region. Among the 61 patients sequenced at virological failure, 50 (82%) patients presented at least one resistance-associated substitutions inducing a high level of resistance to non-structural-5A inhibitors (>10-fold resistance). Conclusions This pooled analysis suggests that non-structural-5A resistance-associated substitutions screening should be recommended when considering retreatment with a non-structural-5A inhibitor regimen in patients who have previously experienced failed non-structural-5A treatment.

Published
2017
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28. Different precore/core mutations of hepatitis B interact with, limit, or favor liver fibrosis severity

Author

Pascal Veillon, Evelyne Schvoerer, Joël Gozlan, Hélène Le Guillou-Guillemette, Sylvie Larrat, Françoise Lunel-Fabiani, Adeline Pivert, Vincent Mackiewicz, Véronique Brodard, Véronique Loustaud-Ratti, Vincent Thibault, Emmanuel Gordien, Sophie Alain, Alexandra Ducancelle, Jérôme Boursier, Sandrine Castelain, Sandrine Bertrais, Marianne Coste-Burel, Bénédicte Roquebert, Paul Calès, Sarah Maylin, Stéphane Chevaliez, and Viorica Balan

Subjects
Hepatitis B virus, medicine.medical_specialty, HBsAg, Mutation, Hepatology, business.industry, Gastroenterology, Hepatitis B, medicine.disease, medicine.disease_cause, Virology, 3. Good health, 03 medical and health sciences, Liver disease, 0302 clinical medicine, Fibrosis, 030220 oncology & carcinogenesis, Internal medicine, Medicine, 030211 gastroenterology & hepatology, Hepatic fibrosis, business, Viral hepatitis
Abstract

BACKGROUND AND AIM The impact of basal core promoter (BCP) and precore (PC) mutants of the hepatitis B virus (HBV) on liver disease severity remains controversial. The aim of the present study was to screen BCP and PC mutations in 252 HBV surface antigen (HBsAg) positive carriers in France and to assess relationships between these mutations and severe fibrosis. METHODS Direct sequencing of the precore/core gene was used to detect A1762T/G1764A and G1757A mutations in the BCP and G1896A and G1899A mutations in the PC region. RESULTS The prevalences of A1762T/G1764A, G1757A, G1896A, and G1899A mutations were 34.1%, 38.7%, 54.9%, and 29.3% (P

Published
2016
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29. Hepatitis C virus whole genome sequencing: Current methods/issues and future challenges

Author

Alban Caporossi, Pauline Tremeaux, Michael G. B. Blum, Patrice Morand, Vincent Leroy, M.A. Thelu, Sylvie Larrat, Université Grenoble Alpes - UFR Pharmacie (UGA UFRP), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Université Grenoble Alpes - UFR Médecine (UGA UFRM), Biologie Computationnelle et Mathématique (TIMC-IMAG-BCM), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institut d'oncologie/développement Albert Bonniot de Grenoble (INSERM U823), Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble-EFS-Institut National de la Santé et de la Recherche Médicale (INSERM), As Scalable As Possible: foundations of large scale dynamic distributed systems (ASAP), Inria Rennes – Bretagne Atlantique, Institut National de Recherche en Informatique et en Automatique (Inria)-Institut National de Recherche en Informatique et en Automatique (Inria)-SYSTÈMES LARGE ÉCHELLE (IRISA-D1), Institut de Recherche en Informatique et Systèmes Aléatoires (IRISA), Université de Rennes (UR)-Institut National des Sciences Appliquées - Rennes (INSA Rennes), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université de Bretagne Sud (UBS)-École normale supérieure - Rennes (ENS Rennes)-Institut National de Recherche en Informatique et en Automatique (Inria)-Télécom Bretagne-CentraleSupélec-Centre National de la Recherche Scientifique (CNRS)-Université de Rennes (UR)-Institut National des Sciences Appliquées - Rennes (INSA Rennes), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université de Bretagne Sud (UBS)-École normale supérieure - Rennes (ENS Rennes)-Institut National de Recherche en Informatique et en Automatique (Inria)-Télécom Bretagne-CentraleSupélec-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche en Informatique et Systèmes Aléatoires (IRISA), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université de Bretagne Sud (UBS)-École normale supérieure - Rennes (ENS Rennes)-Télécom Bretagne-CentraleSupélec-Centre National de la Recherche Scientifique (CNRS), Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Institut National de la Santé et de la Recherche Médicale (INSERM)-EFS-CHU Grenoble-Université Joseph Fourier - Grenoble 1 (UJF), SYSTÈMES LARGE ÉCHELLE (IRISA-D1), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Institut National des Sciences Appliquées - Rennes (INSA Rennes), Institut National des Sciences Appliquées (INSA)-Université de Rennes (UNIV-RENNES)-Institut National des Sciences Appliquées (INSA)-Université de Bretagne Sud (UBS)-École normale supérieure - Rennes (ENS Rennes)-Institut National de Recherche en Informatique et en Automatique (Inria)-Télécom Bretagne-CentraleSupélec-Centre National de la Recherche Scientifique (CNRS)-Université de Rennes 1 (UR1), Institut National des Sciences Appliquées (INSA)-Université de Rennes (UNIV-RENNES)-Institut National des Sciences Appliquées (INSA)-Université de Bretagne Sud (UBS)-École normale supérieure - Rennes (ENS Rennes)-Institut National de Recherche en Informatique et en Automatique (Inria)-Télécom Bretagne-CentraleSupélec-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche en Informatique et Systèmes Aléatoires (IRISA), Institut National des Sciences Appliquées (INSA)-Université de Rennes (UNIV-RENNES)-Institut National des Sciences Appliquées (INSA)-Université de Bretagne Sud (UBS)-École normale supérieure - Rennes (ENS Rennes)-Institut National de Recherche en Informatique et en Automatique (Inria)-Télécom Bretagne-CentraleSupélec-Centre National de la Recherche Scientifique (CNRS)-Inria Rennes – Bretagne Atlantique, Institut National de Recherche en Informatique et en Automatique (Inria), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Cancer genome sequencing, Sanger sequencing, Hepatitis C virus, Clinical Biochemistry, Genome, Viral, Hepacivirus, MESH: Molecular Diagnostic Techniques, Biology, medicine.disease_cause, General Biochemistry, Genetics and Molecular Biology, DNA sequencing, 03 medical and health sciences, symbols.namesake, medicine, Humans, MESH: Hepacivirus, MESH: High-Throughput Nucleotide Sequencing, Genotyping, Exome sequencing, MESH: Hepatitis C, Whole genome sequencing, Genetics, MESH: Humans, Whole Genome Sequencing, MESH: Molecular Typing, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], resistance-associated variants, Biochemistry (medical), High-Throughput Nucleotide Sequencing, Hepatitis C, 3. Good health, Molecular Typing, 030104 developmental biology, Molecular Diagnostic Techniques, genotyping, symbols, next-generation sequencing, MESH: Genome, Viral, MESH: Whole Genome Sequencing, Personal genomics
Abstract

International audience; Therapy for hepatitis C is currently undergoing a revolution. The arrival of new antiviral agents targeting viral proteins reinforces the need for a better knowledge of the viral strains infecting each patient. Hepatitis C virus (HCV) whole genome sequencing provides essential information for precise typing, study of the viral natural history or identification of resistance-associated variants. First performed with Sanger sequencing, the arrival of next-generation sequencing (NGS) has simplified the technical process and provided more detailed data on the nature and evolution of viral quasi-species. We will review the different techniques used for HCV complete genome sequencing and their applications, both before and after the apparition of NGS. The progress brought by new and future technologies will also be discussed, as well as the remaining difficulties, largely due to the genomic variability.

Published
2016
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30. Diagnostic accuracy of a rapid RT-PCR assay for point-of-care detection of influenza A/B virus at emergency department admission: A prospective evaluation during the 2017/2018 influenza season

Author

Roselyne Collomb Muret, Anne Lebeugle, Valérie Guglielmetti, Sylvie Larrat, Virginie Forget, Prudence Mabiala Makele, Maxime Maignan, Julien Lupo, Damien Viglino, Maud Hablot, Patrice Morand, Nicolas Termoz Masson, Caroline Landelle, Hypoxie : Physiopathologie Respiratoire et Cardiovasculaire (HP2 ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Emergency department [Grenoble], Centre Hospitalier Universitaire [Grenoble] (CHU), Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Laboratoire de Virologie [Grenoble], Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Infection Control Unit [Grenoble], Techniques pour l'Evaluation et la Modélisation des Actions de la Santé (TIMC-IMAG-ThEMAS), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), This work was partly funded by Roche Diagnostics (http://www.roche-diagnostics.fr/), the industrial company that markets the cobas Liat system. MM received this grant., Laboratoire Hypoxie et Physiopathologies cardiovasculaires et respiratoires [Grenoble] (HP2), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), and Bodescot, Myriam

Subjects
0301 basic medicine, Male, RNA viruses, Viral Diseases, Influenza Viruses, Critical Care and Emergency Medicine, Health Care Providers, Nurses, Artificial Gene Amplification and Extension, medicine.disease_cause, Roche Diagnostics, Pathology and Laboratory Medicine, Polymerase Chain Reaction, Biochemistry, 0302 clinical medicine, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Interquartile range, Nucleic Acids, Influenza A virus, Medicine and Health Sciences, Medicine, 030212 general & internal medicine, Prospective Studies, Medical Personnel, Prospective cohort study, Aged, 80 and over, Multidisciplinary, Reverse Transcriptase Polymerase Chain Reaction, Middle Aged, 3. Good health, Professions, Infectious Diseases, Molecular Diagnostic Techniques, Medical Microbiology, Viral Pathogens, Viruses, [SDV.MHEP.MI] Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Female, France, Seasons, Pathogens, Emergency Service, Hospital, Research Article, Adult, medicine.medical_specialty, Science, Point-of-Care Systems, 030106 microbiology, Research and Analysis Methods, Microbiology, Specimen Handling, 03 medical and health sciences, Internal medicine, Virology, Influenza, Human, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Molecular Biology Techniques, Molecular Biology, Microbial Pathogens, Point of care, Aged, business.industry, Organisms, Biology and Life Sciences, Emergency department, Reverse Transcriptase-Polymerase Chain Reaction, Triage, Confidence interval, Influenza, Health Care, Influenza B virus, People and Places, Population Groupings, business, Orthomyxoviruses
Abstract

Study objectiveTo investigate the performance of a rapid RT-PCR assay to detect influenza A/B at emergency department admission.MethodsThis single-center prospective study recruited adult patients attending the emergency department for influenza-like illness. Triage nurses performed nasopharyngeal swab samples and ran rapid RT-PCR assays using a dedicated device (cobas Liat, Roche Diagnostics, Meylan, France) located at triage. The same swab sample was also analyzed in the department of virology using conventional RT-PCR techniques. Patients were included 24 hours-a-day, 7 days-a-week. The primary outcome was the diagnostic accuracy of the rapid RT-PCR assay performed at triage.ResultsA total of 187 patients were included over 11 days in January 2018. Median age was 70 years (interquartile range 44 to 84) and 95 (51%) were male. Nine (5%) assays had to be repeated due to failure of the first assay. The sensitivity of the rapid RT-PCR assay performed at triage was 0.98 (95% confidence interval (CI): 0.91-1.00) and the specificity was 0.99 (95% CI: 0.94-1.00). A total of 92 (49%) assays were performed at night-time or during the weekend. The median time from patient entry to rapid RT-PCR assay results was 46 [interquartile range 36-55] minutes.ConclusionRapid RT-PCR assay performed by nurses at triage to detect influenza A/B is feasible and highly accurate.

Published
2019
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31. Hepatitis C subtype distribution in chronically infected patients with mild liver fibrosis in France: the GEMHEP study

Author

Vincent Thibault, Benjamin Nemoz, Magali Bouvier-Alias, P. Marcellin, Françoise Lunel-Fabiani, Vincent Leroy, C. Henquell, Etienne Brochot, Patrice Morand, T. Semenova, Christopher Payan, Gilles Duverlie, S. Vallet, Pascale Trimoulet, Sylvie Larrat, L. Izquierdo, Gisèle Lagathu, A. M. Roque-Afonso, Sarah Maylin, Stéphane Chevaliez, Centre Hospitalier Universitaire [Grenoble] (CHU), CHU Pontchaillou [Rennes], Institut de biologie et chimie des protéines [Lyon] (IBCP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Service de virologie et d'immunologie biologique, CHU Bordeaux [Bordeaux]-Groupe hospitalier Pellegrin, Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Laboratoire Universitaire de Biodiversité et Ecologie Microbienne (LUBEM), Université de Brest (UBO), CHU Clermont-Ferrand, Laboratoire Microorganismes : Génome et Environnement (LMGE), Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Centre National de la Recherche Scientifique (CNRS), Hôpital Henri Mondor, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Henri Mondor-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Institut Mondor de Recherche Biomédicale (IMRB), Institut National de la Santé et de la Recherche Médicale (INSERM)-IFR10-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Laboratoire de Virologie [CHU Saint-Louis], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Hémodynamique, Interaction Fibrose et Invasivité tumorales Hépatiques (HIFIH), Université d'Angers (UA), Service d'hépatologie, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Diderot - Paris 7 (UPD7)-Hôpital Beaujon [AP-HP], Agents infectieux, résistance et chimiothérapie - UR UPJV 4294 (AGIR ), Université de Picardie Jules Verne (UPJV)-CHU Amiens-Picardie, Université Grenoble Alpes (UGA), Laboratoire Microorganismes : Génome et Environnement - Clermont Auvergne (LMGE), Université Clermont Auvergne (UCA)-Centre National de la Recherche Scientifique (CNRS), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpital Henri Mondor-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Université Paris Diderot - Paris 7 (UPD7)-Hôpital Beaujon, Institut de biologie structurale (IBS - UMR 5075), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Liver Cirrhosis, Male, Databases, Factual, Epidemiology, Hepacivirus, medicine.disease_cause, Severity of Illness Index, Tertiary Care Centers, 0302 clinical medicine, Genotype, Prevalence, ComputingMilieux_MISCELLANEOUS, education.field_of_study, [SDE.IE]Environmental Sciences/Environmental Engineering, Incidence (epidemiology), Hepatitis C, 3. Good health, Infectious Diseases, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, RNA, Viral, 030211 gastroenterology & hepatology, Female, France, Viral hepatitis, Adult, treatment-naive HCV, medicine.medical_specialty, Hepatitis C virus, [SDE.MCG]Environmental Sciences/Global Changes, Population, HCV genotypes, Statistics, Nonparametric, 03 medical and health sciences, Viral Proteins, Internal medicine, medicine, Humans, education, Retrospective Studies, Hepatitis, Original Paper, business.industry, fibrosis, Hepatitis C, Chronic, medicine.disease, [SDE.ES]Environmental Sciences/Environmental and Society, 030104 developmental biology, Logistic Models, Multivariate Analysis, [SDE.BE]Environmental Sciences/Biodiversity and Ecology, business
Abstract

Treatment options for Hepatitis C infection have greatly improved with direct-acting antiviral (DAA) combinations achieving high cure rates. Nevertheless, the cost of this treatment is still high and access to treatment in many countries has been preferentially reserved for patients with more severe fibrosis (F3 and F4). In this French nationwide study, we investigated the epidemiological characteristics and genotype distribution of hepatitis C virus (HCV) in treatment-naive patients with METAVIR fibrosis stages between F0 and F2 in order to identify patient profiles that became eligible for unrestricted treatment in a second period. Between 2015 and 2016 we collected data from nine French university hospitals on a total of 584 HCV positive patients with absent, mild or moderate liver fibrosis. The most represented genotypes were genotype 1b (159/584; 27.2%), followed by genotype 1a (150/584; 25.7%); genotype 3 (87/584: 14.9%); genotype 4 (80/584; 13.7%). Among genotype 4: 4a was predominantly encountered with 22 patients (27.5% of genotype 4). Genotypes 1b and 1a are currently the most frequent virus types present in treatment-naive patients with mild fibrosis in France. They can be readily cured using the available DAA. Nevertheless, non-a/non-d genotype 4 is also frequent in this population and clinical data on the efficacy of DAA on these subtypes is missing. The GEMHEP is the French group for study and evaluation of viral hepatitis on a national scale. Data collection on epidemiological and molecular aspects of viral hepatitis is performed on a regular basis in all main French teaching hospitals and serves as a basis for surveillance of these infections. Analysis and trends are regularly published on behalf of the GEMHEP group. Data collection was performed retrospectively over the 2015–2016 period, covering nine main university hospitals in France. A total of 584 hepatitis C positive patients were included in this study. Genotyping of the circulating viruses showed a high prevalence of genotypes 1b and 1a in our population. The epidemiology of hepatitis C is slowly changing in France, particularly as a consequence of the rise of ‘non-a non-d’ genotype 4 viruses mainly originating from African populations. More data concerning treatment efficacy of these genotypes is needed in order to guide clinical care.

Published
2019
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32. Using high-throughput sequencing for investigating intra-host hepatitis C evolution over long retrospective periods

Author

Om Kulkarni, V. Leroy, Olivier François, Alban Caporossi, Patrice Morand, Michael G. B. Blum, Sylvie Larrat, Francois, Olivier, Biologie Computationnelle et Mathématique (TIMC-IMAG-BCM), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)

Subjects
0301 basic medicine, Microbiology (medical), Genotype, Secondary infection, Hepatitis C virus, 030106 microbiology, Population, Hepacivirus, Viral Nonstructural Proteins, Biology, medicine.disease_cause, Microbiology, Evolution, Molecular, 03 medical and health sciences, chemistry.chemical_compound, Drug Resistance, Viral, Genetics, medicine, Humans, Selection, Genetic, education, Molecular Biology, NS5B, Phylogeny, Ecology, Evolution, Behavior and Systematics, ComputingMilieux_MISCELLANEOUS, Retrospective Studies, [SDV.MP.VIR] Life Sciences [q-bio]/Microbiology and Parasitology/Virology, education.field_of_study, Genetic diversity, [SDV.GEN.GPO]Life Sciences [q-bio]/Genetics/Populations and Evolution [q-bio.PE], Genetic heterogeneity, Genetic Variation, High-Throughput Nucleotide Sequencing, Sequence Analysis, DNA, Hepatitis C, medicine.disease, 3. Good health, 030104 developmental biology, Infectious Diseases, chemistry, Host-Pathogen Interactions, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, [SDV.GEN.GPO] Life Sciences [q-bio]/Genetics/Populations and Evolution [q-bio.PE], Selective sweep
Abstract

Collections of biological samples held by hospitals represent invaluable resources for conducting retrospective evolutionary studies of chronic infections. Using high-throughput sequencing, those collections permit analysis of within-host genetic diversity over long follow-up periods, and allow a better understanding of resistance to treatment regimes during disease evolution. Here, we studied the evolution of hepatitis C virus (HCV) populations in two patients with an absence of response to dual therapies. We implemented amplicon sequencing to survey genomic variation at the Core and NS5B regions of HCV over a period of 13 years from blood samples obtained at multiple time points. We observed mixed infection by multiple HCV genotypes in both patients. Genetic heterogeneity and sample composition analysis provided information about the changes in viral population over the course of clinical treatment, with NS5B experiencing an increase in diversity after treatment initiation. Secondary infections were estimated to predate treatment year, and our results pointed towards diversifying selection occurring post-treatment, acting on standing genomic variation and maintaining high genetic heterogeneity during infection. For these two patients infected with multiple HCV genotypes, the maintenance of viral diversity was explained with the hypothesis of soft selective sweep started at the same time as antiviral treatment was initiated.

Published
2019

33. Innovative multiplexed point-of-care immunoassay applied to hepatitis B screening

Author

Paul Kauffmann, Pn Marche, Guillaume Blaire, Sarah Delshadi, Thomas Decaens, Patrice Morand, V. Masse, O. Cugat, Sylvie Larrat, Microbiology and Infectious Diseases, Sherbrooke University Hospital, Unit for Virus Host-Cell Interactions [Grenoble] (UVHCI), Université Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Génie Electrique de Grenoble (G2ELab), Université Joseph Fourier - Grenoble 1 (UJF)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-Institut Polytechnique de Grenoble - Grenoble Institute of Technology-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA), Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Chadebec, Olivier, Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)

Subjects
medicine.diagnostic_test, business.industry, [SPI.NRJ]Engineering Sciences [physics]/Electric power, Biochemistry (medical), Clinical Biochemistry, General Medicine, Biochemistry, Virology, 3. Good health, Hepatitis B screening, Immunoassay, medicine, business, ComputingMilieux_MISCELLANEOUS, [SPI.NRJ] Engineering Sciences [physics]/Electric power, Point of care
Abstract

International audience

Published
2019
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34. Hepatitis E Virus in Acute Non-Traumatic, Non-Vascular Neurological Injury: A French Monocentric Prospective Study

Author

aude belbézier, Alban Deroux, Françoise Sarrot-Reynauld, Barbara Colombe, Annick Bosseray, Claire Wintenberger, Perrine Dumanoir, Maxime Lugosi, Isabelle Boccond-Gibod, Vincent Leroy, Maxime Maignan, Roselyne Collomb-Muret, Damien Viglino, Mathieu Vaillant, Emeline Lagrange, Olivier Epaulard, Chantal Dumestre-Perard, Julien Lupo, Sylvie Larrat, Patrice Morand, Carole Schwebel, Antoine Vilotitch, Jean-Luc Bosson, and Laurence Bouillet

Published
2018
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35. Evaluation of the cobas® GT hepatitis C virus genotyping assay in G1-6 viruses including low viral loads and LiPA failures

Author

Sylvie Larrat, Jean-Dominique Poveda, Patrice Morand, Benjamin Nemoz, Vincent Leroy, Léa Roger, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Hôpital Michallon, Laboratoire CERBA [Saint Ouen l'Aumône], Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)

Subjects
RNA viruses, 0301 basic medicine, Genotyping Techniques, MESH: Reagent Kits, Diagnostic, lcsh:Medicine, Artificial Gene Amplification and Extension, Hepacivirus, medicine.disease_cause, Roche Diagnostics, Polymerase Chain Reaction, MESH: Genotype, Genotype, MESH: Sequence Analysis, RNA, Medicine, MESH: Hepacivirus, lcsh:Science, Asymptomatic Infections, Pathology and laboratory medicine, Sanger sequencing, Multidisciplinary, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Hepatitis C virus, MESH: Real-Time Polymerase Chain Reaction, Viral Load, Medical microbiology, Subtyping, MESH: Hepatitis C, Chronic, 3. Good health, MESH: RNA, Viral, Viruses, symbols, RNA, Viral, Pathogens, MESH: Viral Load, Viral load, Research Article, MESH: Antiviral Agents, Genotyping, 030106 microbiology, MESH: Asymptomatic Infections, Research and Analysis Methods, Real-Time Polymerase Chain Reaction, Microbiology, Antiviral Agents, 03 medical and health sciences, symbols.namesake, Extraction techniques, Virology, MESH: Genotyping Techniques, Humans, Serologic Tests, Typing, Molecular Biology Techniques, Sequencing Techniques, Molecular Biology, Retrospective Studies, Medicine and health sciences, MESH: Humans, Flaviviruses, Sequence Analysis, RNA, business.industry, MESH: Serologic Tests, lcsh:R, Organisms, Viral pathogens, Automated Fluorescent Genotyping, Biology and Life Sciences, MESH: Retrospective Studies, Reverse Transcriptase-Polymerase Chain Reaction, Hepatitis C, Chronic, Hepatitis viruses, RNA extraction, Microbial pathogens, lcsh:Q, Reagent Kits, Diagnostic, business, Viral Transmission and Infection
Abstract

International audience; Direct-acting antiviral (DAA) drug performances depend on the viral genotype. So international recommendations give typing of the virus a prerequisite for treatment choice and patient management. Commercially available HCV genotyping kits are scarce and this analysis is often in-house using tedious PCRs and Sanger sequencing, leading to a lack of standardization. A newly commercialized HCV genotyping assay based on real-time PCR has been developed by Roche Diagnostics (Mannheim, Germany). We compared this new assay with our in-house PCRs -sequencing technique on 101 regular samples and 81 LiPA failures or low viral load samples. No genotype or 1a/1b subtype mismatch was observed. Two samples were misidentified at the subtype level without clinical impact. Three genotype 1b and two genotype 1a samples with low viral load could not be subtyped. Nevertheless, 13 (13%) samples from the regular panel and 35 (43%) from the more difficult-to-type panels failed to give results on first pass with the Roche kit. Failures were mostly associated with genotype 3 subtype a, with genotype 4 subtype non-a, or with viral loads

Published
2018
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36. Hepatitis E Virus in Acute Non-Traumatic, Non-Vascular Neurological Injury: A French Monocentric Prospective Study

Author

Claire Wintenberger, Maxime Maignan, Damien Viglino, Olivier Epaulard, Sylvie Larrat, Carole Schwebel, Françoise Sarrot-Reynauld, B. Colombe, Chantal Dumestre-Pérard, Lorella Minotti, Emeline Lagrange, Isabelle Boccond-Gibod, Alban Deroux, Maxime Lugosi, Laurence Bouillet, Perrine Dumanoir, Patrice Morand, A. Bosseray, Roselyne Collomb-Muret, Vincent Leroy, Jean-Luc Bosson, Julien Lupo, Antoine Vilotitch, Mathieu Vaillant, and Aude Belbézier

Subjects
medicine.medical_specialty, Neurological injury, Hepatitis E virus, business.industry, Non traumatic, Internal medicine, Medicine, business, Prospective cohort study, medicine.disease_cause
Published
2018
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37. Ultradeep Pyrosequencing of NS3 To Predict Response to Triple Therapy with Protease Inhibitors in Previously Treated Chronic Hepatitis C Patients

Author

Olivier François, Katia Fusillier, Michael G. B. Blum, Jean-Pierre Zarski, Sylvie Larrat, Pauline Tremeaux, Patrice Morand, Réjane Beugnot, Alice Marlu, Jean-Baptiste Claude, Agnès Plages, Om Kulkarni, Anne Signori-Schmuck, Julien Lupo, Hervé Duborjal, Vincent Leroy, Unit for Virus Host-Cell Interactions [Grenoble] (UVHCI), Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), Biologie Computationnelle et Mathématique (TIMC-IMAG-BCM), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF)-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Genostar, GENOME Express (GENOME Express), GENOME Express, Earth and Environmental, AMEC, Biochimie et Génétique Moléculaire, CHU Grenoble-Hôpital Michallon, Pôle Digidune, Pôle DigiDune, Unité de Biophysique, CRSSA, Département d'hépato-gastroentérologie, Université Grenoble Alpes (UGA)-CHU Grenoble, Matière et Systèmes Complexes (MSC (UMR_7057)), Centre National de la Recherche Scientifique (CNRS)-Université Paris Diderot - Paris 7 (UPD7), CHU Grenoble-Université Grenoble Alpes (UGA), Université Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS), Université Joseph Fourier - Grenoble 1 (UJF)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-IMAG-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Université Joseph Fourier - Grenoble 1 (UJF)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-IMAG-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA), Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble, and Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Adult, Male, Microbiology (medical), NS3, medicine.medical_specialty, Entropy, medicine.medical_treatment, Mutation, Missense, Salvage therapy, Viral quasispecies, Viral Nonstructural Proteins, Biology, Antiviral Agents, Peripheral blood mononuclear cell, Gastroenterology, Young Adult, 03 medical and health sciences, 0302 clinical medicine, Virology, Internal medicine, Drug Resistance, Viral, Genotype, medicine, Humans, Protease Inhibitors, Aged, 030304 developmental biology, Aged, 80 and over, Salvage Therapy, 0303 health sciences, Protease, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Genetic Variation, High-Throughput Nucleotide Sequencing, virus diseases, Odds ratio, Hepatitis C, Chronic, Middle Aged, Prognosis, digestive system diseases, Confidence interval, 3. Good health, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], HCV, Immunology, Drug Therapy, Combination, Female, 030211 gastroenterology & hepatology
Abstract

Despite the gain in sustained virological responses (SVR) provided by protease inhibitors (PIs), failures still occur. The aim of this study was to determine if a baseline analysis of the NS3 region using ultradeep pyrosequencing (UDPS) can help to predict an SVR. Serum samples from 40 patients with previously nonresponding genotype 1 chronic hepatitis C who were retreated with triple therapy, including a PI, were analyzed. Baseline UDPS of the NS3 gene was performed on plasma and peripheral blood mononuclear cells (PBMC). Mutations conferring resistance to PIs were sought. The overall diversity of the quasispecies was evaluated by calculating the Shannon entropy (SE). Resistance mutations were found in plasma and PBMC but were not discriminating enough to predict an SVR. NS3 quasispecies heterogeneity was significantly lower at baseline in patients achieving an SVR than in those not achieving an SVR (SE of 26.98 ±16.64 x10^3 versus 44.93 ±19.58 x10^3, P =0.0047). With multivariate analysis, the independent predictors of an SVR were fibrosis of stage F <2 (odds ratio [OR], 13.3; 95% confidence interval [CI], 1.25 to 141.096; P < 0.03) and SE below the median (OR, 5.4; 95% CI, 1.22 to 23.87; P < 0.03). More than the presence of minor mutations at the baseline in plasma or in PBMC, the NS3 viral heterogeneity determined by UDPS is an independent factor for an SVR in previously treated patients receiving triple therapy that includes a PI.&nbsp

Published
2015
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38. Added Value of Next-Generation Sequencing for Multilocus Sequence Typing Analysis of a Pneumocystis jirovecii Pneumonia Outbreak1

Author

Patricia Pavese, Sébastien Bailly, Claire Wintenberger, Cécile Garnaud, Caroline Augier, Anne Thiebaut-Bertrand, Sylvie Larrat, Thibault Drouet, Marie-Reine Mallaret, Olivier Epaulard, Danièle Maubon, Eléna Charpentier, John Rendu, Paolo Malvezzi, Muriel Cornet, Jean-Benjamin Murat, Université Grenoble Alpes - UFR Pharmacie (UGA UFRP), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Thérapeutique Recombinante Expérimentale (TIMC-IMAG-TheREx), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Clinique de médecine interne, Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble, Laboratoire Adaptation et pathogénie des micro-organismes [Grenoble] (LAPM), Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), INSERM U836, équipe 4, Muscles et pathologies, Grenoble Institut des Neurosciences (GIN), Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Biochimie et Génétique Moléculaire, CHU Grenoble-Hôpital Michallon-CHU Grenoble-Hôpital Michallon, Service des Maladies Infectieuses, CHU Grenoble, Université Grenoble Alpes - UFR Médecine (UGA UFRM), Clinique Universitaire d'Hématologie [La Tronche, Grenoble], Centre Hospitalier Universitaire [Grenoble] (CHU), Unité d'hygiène Hospitalière, Department of infectious diseases, Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Institute for Advanced Biosciences / Institut pour l'Avancée des Biosciences (Grenoble) (IAB), Centre Hospitalier Universitaire [Grenoble] (CHU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang - Auvergne-Rhône-Alpes (EFS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)

Subjects
Male, 0301 basic medicine, transplants, Epidemiology, multilocus sequence typing, Pneumocystis carinii, Pneumocystis pneumonia, Disease Outbreaks, Genotype, Pneumocystosis, high-throughput nucleotide sequencing, Child, Pneumocystis jirovecii, Phylogeny, ComputingMilieux_MISCELLANEOUS, [SDV.MP.MYC]Life Sciences [q-bio]/Microbiology and Parasitology/Mycology, biology, Pneumonia, Pneumocystis, Middle Aged, 3. Good health, Infectious Diseases, NGS, Child, Preschool, Synopsis, Female, MLST, Adult, Microbiology (medical), Adolescent, 030106 microbiology, Sensitivity and Specificity, DNA sequencing, pneumocystosis, respiratory infections, Young Adult, 03 medical and health sciences, medicine, pneumonia, Humans, Genotyping, Aged, Polymorphism, Genetic, Added Value of Next-Generation Sequencing for Multilocus Sequence Typing Analysis of a Pneumocystis jirovecii Pneumonia Outbreak, Computational Biology, Infant, Outbreak, biology.organism_classification, medicine.disease, Virology, Case-Control Studies, Multilocus sequence typing, next-generation sequencing, fungi
Abstract

Pneumocystis jirovecii is a major threat for immunocompromised patients, and clusters of pneumocystis pneumonia (PCP) have been increasingly described in transplant units during the past decade. Exploring an outbreak transmission network requires complementary spatiotemporal and strain-typing approaches. We analyzed a PCP outbreak and demonstrated the added value of next-generation sequencing (NGS) for the multilocus sequence typing (MLST) study of P. jirovecii strains. Thirty-two PCP patients were included. Among the 12 solid organ transplant patients, 5 shared a major and unique genotype that was also found as a minor strain in a sixth patient. A transmission map analysis strengthened the suspicion of nosocomial acquisition of this strain for the 6 patients. NGS-MLST enables accurate determination of subpopulation, which allowed excluding other patients from the transmission network. NGS-MLST genotyping approach was essential to deciphering this outbreak. This innovative approach brings new insights for future epidemiologic studies on this uncultivable opportunistic fungus.

Published
2017
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39. Hépatite E dans les troubles neurologiques non traumatiques et non vasculaires : étude prospective française monocentrique

Author

M. Maignan, M. Lugosi, Claire Wintenberger, E. Lagrange, Belbezier, P. Dumanoir, B. Colombe, M. Vaillant, Sylvie Larrat, Laurence Bouillet, and Alban Deroux

Subjects
Gastroenterology, Internal Medicine
Abstract

Introduction Le virus de l’hepatite E fut decrit comme associe a differents troubles neurologiques aigus. La prevalence reelle de l’hepatite E dans ces maladies n’est pas encore connue. L’objectif de notre etude est de determiner la prevalence de l’hepatite E dans les troubles neurologiques aigus non vasculaires, non traumatiques. Patients et methodes Cette etude, transversale, prospective fut realisee de 2014 a 2018 dans l’hopital universitaire de Grenoble (Isere, France). Une serologie hepatite E avec dosage des anticorps type immunoglobuline fut realisee chez les patients hospitalises durant cette periode presentant des troubles neurologiques aigus non traumatiques et non vasculaires. Les patients etaient eligibles s’ils etaient hospitalises et presentaient des manifestations neurologiques aigues definies comme des deficits sensitivomoteurs, des crises d’epilepsie febriles, une confusion febrile, des mouvements anormaux ou des symptomes neuromusculaires intermittents. Les patients souffrants d’encephalopathie, d’accident vasculaire cerebral ou de traumatismes etaient exclus. Parmi 210 patients eligibles, 4 ont refuse de participer, 5 n’ont pas ete testes et 42 ont ete exclus. Le diagnostic de trouble neurologique non traumatique, non vasculaire fut realise chez 159 patients. Nous avons mesure la prevalence de l’hepatite E chez les patients souffrant de troubles neurologiques aigus non traumatiques, non vasculaires et compare cette prevalence a celle des donneurs de sang issus de la meme zone geographique. Cette prevalence fut calculee par l’equipe de Mansuy et al. et publiee en 2016 dans Hepatology. Resultats Onze des 159 patients avec troubles neurologiques aigus non traumatiques, non vasculaires presentaient une infection recente par le virus de l’hepatite E (6,9 %, IC95 % [3,5–12]). Six patients avaient une charge virale serique positive, et 2 une charge virale positive dans les selles. Trois patients avaient une infection associee (un patient presentait une infection a Listeria monocytogenes, une patiente une infection a varicella-zoster virus et un a cytomegalovirus). Une elevation des transaminases etait observee chez 64 % des patients avec une hepatite E et 24 % des patients sans hepatite E (p = 0,011), une cholestase etait notee chez 64 % et 28 % respectivement (p = 0,005). Les patients souffrant de syndrome de Parsonage–Turner associe a l’hepatite E presentaient frequemment une forme bilaterale (p Conclusion Dans cette etude prospective, 6,9 % des patients avec un trouble neurologique aigu non traumatique, non vasculaire presentaient une infection aigue par le virus de l’hepatite E. La serologie de l’hepatite E devrait etre realisee devant tout trouble neurologique aigu non traumatique, non vasculaire. La cytolyse hepatique peut etre absente.

Published
2018
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40. Sequencing Assays for Failed Genotyping with the Versant Hepatitis C Virus Genotype Assay (LiPA), Version 2.0

Author

Anne Signori-Schmuck, Katia Fusillier, Jean-Dominique Poveda, Sylvie Larrat, Vincent Thibault, Nelly Magnat, Camille Coudret, and Patrice Morand

Subjects
Adult, Male, Microbiology (medical), Untranslated region, Genotype, Hepacivirus, Hepatitis C virus, Molecular Sequence Data, medicine.disease_cause, chemistry.chemical_compound, Virology, medicine, Humans, False Negative Reactions, NS5B, Genotyping, Aged, Genetics, NS3, Base Sequence, biology, Genetic Variation, Sequence Analysis, DNA, Hepatitis C, Middle Aged, medicine.disease, biology.organism_classification, Molecular Diagnostic Techniques, chemistry, Mutation, RNA, Viral, Female
Abstract

For optimal antiviral therapy, the hepatitis C virus (HCV) genotype needs to be determined, as it remains a strong predictor of sustained viral response. In this study, we assessed the number of HCV genotyping results that could not be determined using the commercially available line probe assay (LiPA) (Versant hepatitis C virus genotype 2.0 assay) in a large international panel of samples from 9,874 HCV-positive patients. In-house sequencing assays targeting the 5′ untranslated region (UTR), core region, NS3 region, and NS5B region of the HCV genome and phylogenetic analyses were used to resolve these LiPA failures. Among all cases, the genotypes of 51 samples (0.52%) could not be determined with the LiPA. These undetermined results were observed more frequently among samples from non-European regions (mainly the Arabian Peninsula). The use of sequencing assays coupled with phylogenetic analysis provided reliable genotype results for 86% of the LiPA failures, which exhibited higher rates of genotypes 4, 5, and 6 than did LiPA-resolved genotypes. As expected, the 5′ UTR was not sufficiently variable for clear discrimination between genotypes 1 and 6, but it also resulted in errors in classification of some genotype 3 and 4 cases using well-known Web-based BLAST programs. This study demonstrates the low frequency of genotyping failures with the Versant hepatitis C virus genotype 2.0 assay (LiPA) and also underlines the need for a complex combination of sequences and phylogenetic analyses in order to genotype these particular HCV strains correctly.

Published
2013
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41. Fatal autochthonous fulminant hepatitis E early after allogeneic stem cell transplantation

Author

Anne Thiebaut-Bertrand, Sylvie Larrat, N Sturm, Sébastien Lhomme, Vincent Leroy, Martin Carre, J. Y. Cahn, Frédéric Garban, Patricia Pouzol, Patrice Morand, Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)

Subjects
MESH: Fatal Outcome, Fatal outcome, medicine.medical_treatment, MESH: Allografts, Hematopoietic stem cell transplantation, Precursor Cell Lymphoblastic Leukemia Lymphoma, 03 medical and health sciences, 0302 clinical medicine, Fatal Outcome, medicine, MESH: Liver Failure, Acute, Humans, 030212 general & internal medicine, Fulminant hepatitis, ComputingMilieux_MISCELLANEOUS, MESH: Hematopoietic Stem Cell Transplantation, MESH: Precursor Cell Lymphoblastic Leukemia-Lymphoma, Transplantation, MESH: Humans, MESH: Middle Aged, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], business.industry, MESH: Hepatitis E, Liver failure, Hematopoietic Stem Cell Transplantation, Hematology, Liver Failure, Acute, Middle Aged, Precursor Cell Lymphoblastic Leukemia-Lymphoma, Hepatitis E, medicine.disease, Allografts, Virology, 3. Good health, Immunology, 030211 gastroenterology & hepatology, Female, Stem cell, business, MESH: Female
Abstract

International audience

Published
2017
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42. Natural non-homologous recombination led to the emergence of a duplicated V3-NS5A region in HCV-1b strains associated with hepatocellular carcinoma

Author

Françoise Stoll-Keller, Henia Saoudin, Arielle A. Rosenberg, Dominique Bettinger, Pascal Veillon, Odile Petsaris, Cécile Henquell, Bruno Pozzetto, Catherine Gaudy-Graffin, Jean-Baptiste Bour, Sophie Minjolle-Cha, Jacques Izopet, Christopher Payan, E. Bouthry, Alexandra Ducancelle, Adeline Pivert, Hélène Le Guillou-Guillemette, Gisèle Lagathu, Sylvie Larrat, P. André, Anne Signori-Schmuck, Sophie Vallet, Françoise Lunel-Fabiani, Jean-Michel Pawlotsky, Yazid Baazia, Elisabeth André-Garnier, Christophe Lemaire, Florence Abravanel, Pascale Trimoulet, Sophie Alain, Vincent Thibault, Hémodynamique, Interaction Fibrose et Invasivité tumorales Hépatiques (HIFIH), Université d'Angers (UA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM), CHU Clermont-Ferrand, Laboratoire Microorganismes : Génome et Environnement (LMGE), Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Centre National de la Recherche Scientifique (CNRS), Laboratoire Universitaire de Biodiversité et Ecologie Microbienne (LUBEM), Université de Brest (UBO), Service de Bactériologie, Virologie, Hygiène [CHU Limoges], CHU Limoges, Anti-infectieux : supports moléculaires des résistances et innovations thérapeutiques (RESINFIT), CHU Limoges-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Génomique, Environnement, Immunité, Santé, Thérapeutique (GEIST), Université de Limoges (UNILIM)-Université de Limoges (UNILIM), CHU Pontchaillou [Rennes], Centre de Physiopathologie Toulouse Purpan (CPTP), Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Virologie de l'hépatite C (EA 4474), Université Paris Descartes - Paris 5 (UPD5), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Hôpital Cochin [AP-HP], Hôtel-Dieu de Nantes, Laboratoire de sérologie-virologie (CHU de Dijon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Service de bactériologie-Virologie-Hygiène [Avicenne], Université Paris 13 (UP13)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Avicenne [AP-HP], Service de virologie et d'immunologie biologique, CHU Bordeaux [Bordeaux]-Groupe hospitalier Pellegrin, Hôpital de la Croix-Rousse [CHU - HCL], Hospices Civils de Lyon (HCL), Morphogénèse et antigénicité du VIH et du virus des Hépatites (MAVIVH - U1259 Inserm - CHRU Tours ), Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Université de Tours (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre Hospitalier Régional Universitaire de Besançon (CHRU Besançon), Université Grenoble Alpes - UFR Pharmacie (UGA UFRP), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Groupe Immunité des Muqueuses et Agents Pathogènes (GIMAP), Université Jean Monnet - Saint-Étienne (UJM), Interaction virus-hôte et maladies du foie, Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Mondor de Recherche Biomédicale (IMRB), Institut National de la Santé et de la Recherche Médicale (INSERM)-IFR10-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Institut de Recherche en Horticulture et Semences (IRHS), Université d'Angers (UA)-Institut National de la Recherche Agronomique (INRA)-AGROCAMPUS OUEST, Laboratoire Microorganismes : Génome et Environnement - Clermont Auvergne (LMGE), Université Clermont Auvergne (UCA)-Centre National de la Recherche Scientifique (CNRS), Centre de Physiopathologie Toulouse Purpan ex IFR 30 et IFR 150 (CPTP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), CHU Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Université Paris 13 (UP13)-Hôpital Avicenne, Morphogénèse et antigénicité du VIH et du virus des Hépatites (MAVIVH), Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Université de Tours-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre Hospitalier Régional Universitaire [Besançon] (CHRU Besançon), Université Grenoble Alpes (UGA), Institut de biologie structurale (IBS - UMR 5075), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS), Unit for Virus Host-Cell Interactions [Grenoble] (UVHCI), Université Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS), Université Jean Monnet [Saint-Étienne] (UJM), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Génomique, Environnement, Immunité, Santé, Thérapeutique (GEIST), Université de Limoges (UNILIM)-Université de Limoges (UNILIM)-CHU Limoges, Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université de Tours-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre Hospitalier Régional Universitaire de Tours (CHRU TOURS), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-IFR10, AGROCAMPUS OUEST, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut National de la Recherche Agronomique (INRA)-Université d'Angers (UA), Université Paris 13 (UP13)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpital Avicenne, Université de Tours-Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Institut National de la Santé et de la Recherche Médicale (INSERM), and AGROCAMPUS OUEST-Institut National de la Recherche Agronomique (INRA)-Université d'Angers (UA)

Subjects
RNA viruses, 0301 basic medicine, Mutant, lcsh:Medicine, Hepacivirus, Viral Nonstructural Proteins, Biochemistry, Cohort Studies, Database and Informatics Methods, Viral Envelope Proteins, Gene Duplication, Gene duplication, Evolutionary Emergence, lcsh:Science, Homologous Recombination, Pathology and laboratory medicine, Phylogeny, Data Management, Recombination, Genetic, Genetics, Multidisciplinary, Hepatitis C virus, Liver Neoplasms, Phylogenetic Analysis, Medical microbiology, Hepatitis C, 3. Good health, Nucleic acids, Phylogenetics, Viruses, Host-Pathogen Interactions, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, Pathogens, Sequence Analysis, Recombination, Research Article, Computer and Information Sciences, Evolutionary Processes, Carcinoma, Hepatocellular, DNA recombination, Bioinformatics, 030106 microbiology, Viral quasispecies, Biology, Research and Analysis Methods, Microbiology, Viral Evolution, Evolution, Molecular, 03 medical and health sciences, Virology, Humans, Evolutionary Systematics, Gene, DNA sequence analysis, Taxonomy, Medicine and health sciences, Cloning, Evolutionary Biology, Polymorphism, Genetic, Biology and life sciences, Flaviviruses, lcsh:R, Organisms, Viral pathogens, RNA, Bayes Theorem, DNA, Hepatitis viruses, Organismal Evolution, Microbial pathogens, 030104 developmental biology, Genetic Loci, Microbial Evolution, Mutation, lcsh:Q, Homologous recombination, Sequence Alignment
Abstract

International audience; Background: The emergence of new strains in RNA viruses is mainly due to mutations or intra and inter-genotype homologous recombination. Non-homologous recombinations may be deleterious and are rarely detected. In previous studies, we identified HCV-1b strains bearing two tandemly repeated V3 regions in the NS5A gene without ORF disruption. This polymorphism may be associated with an unfavorable course of liver disease and possibly involved in liver carcinogenesis. Here we aimed at characterizing the origin of these mutant strains and identifying the evolutionary mechanism on which the V3 duplication relies.Methods: Direct sequencing of the entire NS5A and E1 genes was performed on 27 mutant strains. Quasispecies analyses in consecutive samples were also performed by cloning and sequencing the NS5A gene for all mutant and wild strains. We analyzed the mutant and wild-type sequence polymorphisms using Bayesian methods to infer the evolutionary history of and the molecular mechanism leading to the duplication-like event.Results: Quasispecies were entirely composed of exclusively mutant or wild-type strains respectively. Mutant quasispecies were found to have been present since contamination and had persisted for at least 10 years. This V3 duplication-like event appears to have resulted from non-homologous recombination between HCV-1b wild-type strains around 100 years ago. The association between increased liver disease severity and these HCV-1b mutants may explain their persistence in chronically infected patients.Conclusions: These results emphasize the possible consequences of non-homologous recombination in the emergence and severity of new viral diseases.

Published
2017
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43. New insights into HCV replication in original cells from Aedes mosquitoes

Author

Yassine Rechoum, Emmanuel Drouet, Frederic Garzoni, Pascal Fender, Olivier François, Alban Caporossi, Sylvie Larrat, Catherine Fallecker, Patrice Morand, Marie Anne Petit, Imre Berger, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Institut de Biologie et Pathologie [CHU Grenoble] (IBP), CHU Grenoble-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Biologie Computationnelle et Mathématique (TIMC-IMAG-BCM), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Laboratoire européen de biologie moléculaire - European Molecular Biology Laboratory (EMBL Grenoble), European Molecular Biology Laboratory [Grenoble] (EMBL), The School of Biochemistry [Bristol, U.K.], University of Bristol [Bristol], Centre de Recherche en Cancérologie de Lyon (UNICANCER/CRCL), Centre Léon Bérard [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), C. F. is the recipient of MENRT fellowship (French Ministry of Higher Education and Research). I.B. and F.G. acknowledge funding from the European Commission (EC) Framework Program (FP) 7 through the projects ComplexINC (contract nr. 279,039) and Biostruct-X (contract nr. 283,570). This work used the platforms of the Grenoble Instruct Center (ISBG: UMS 3518 CNRS-CEA-UJF-EMBL) with support from FRISBI (ANR-10-INSB-05-02) and GRAL (ANR-10-LABX-49-01) within the Grenoble Partnership for Structural Biology (PSB)., ANR-10-INBS-0005,FRISBI,Infrastructure Française pour la Biologie Structurale Intégrée(2010), ANR-10-LABX-0049,GRAL,Grenoble Alliance for Integrated Structural Cell Biology(2010), European Project: 279039,EC:FP7:HEALTH,FP7-HEALTH-2011-two-stage,COMPLEXINC(2011), European Project: 283570,EC:FP7:INFRA,FP7-INFRASTRUCTURES-2011-1,BIOSTRUCT-X(2011), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de biologie structurale ( IBS - UMR 5075 ), Université Joseph Fourier - Grenoble 1 ( UJF ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Grenoble Alpes ( UGA ), Institut de Biologie et Pathologie [CHU Grenoble] ( IBP ), CHU Grenoble-Université Grenoble Alpes ( UGA ), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications [Grenoble] ( TIMC-IMAG ), Université Joseph Fourier - Grenoble 1 ( UJF ) -Institut polytechnique de Grenoble - Grenoble Institute of Technology ( Grenoble INP ) -IMAG-Centre National de la Recherche Scientifique ( CNRS ) -Université Grenoble Alpes ( UGA ), Laboratoire européen de biologie moléculaire - European Molecular Biology Laboratory ( EMBL Grenoble ), European Molecular Biology Laboratory [Grenoble] ( EMBL ), University of Bristol [UK], Centre de Recherche en Cancérologie de Lyon ( CRCL ), Centre Léon Bérard [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 ), ANR-10-INSB-05-02,FRISBI, ANR-10-LABX-49-01,Labex GRAL, European Project : 279039,EC:FP7:HEALTH,FP7-HEALTH-2011-two-stage,COMPLEXINC ( 2011 ), European Project : 283570,EC:FP7:INFRA,FP7-INFRASTRUCTURES-2011-1,BIOSTRUCT-X ( 2011 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-CHU Grenoble, BMC, BMC, Infrastructure Française pour la Biologie Structurale Intégrée - - FRISBI2010 - ANR-10-INBS-0005 - INBS - VALID, Grenoble Alliance for Integrated Structural Cell Biology - - GRAL2010 - ANR-10-LABX-0049 - LABX - VALID, New Technologies and Production Tools for Complex Protein Biologics - COMPLEXINC - - EC:FP7:HEALTH2011-11-01 - 2015-10-31 - 279039 - VALID, and Transnational access and enhancement of integrated Biological Structure determination at synchrotron X-ray radiation facilities - BIOSTRUCT-X - - EC:FP7:INFRA2011-09-01 - 2016-02-29 - 283570 - VALID

Subjects
0301 basic medicine, Human hepatocytes, [SDV]Life Sciences [q-bio], Hepacivirus, Virus Replication, Polymerase Chain Reaction, 0302 clinical medicine, Aedes aegypti, Viral Envelope Proteins, Aedes, Genotype, Phylogeny, Mosquito cell, chemistry.chemical_classification, education.field_of_study, biology, virus diseases, Hepatitis C virus (HCV), Aedes albopictus, Hepatitis C, 3. Good health, [SDV] Life Sciences [q-bio], Infectious Diseases, RNA, Viral, 030211 gastroenterology & hepatology, Sequence Analysis, Population, Cell Line, Microbiology, 03 medical and health sciences, Virology, Animals, Humans, education, Polymorphism, Genetic, Insect cell, [ SDV ] Life Sciences [q-bio], Research, fungi, biology.organism_classification, Insect Vectors, 030104 developmental biology, chemistry, Cell culture, Mutation, Hepatocytes, Peptides, Glycoprotein, HCV pseudo particles
Abstract

Background The existing literature about HCV association with, and replication in mosquitoes is extremely poor. To fill this gap, we performed cellular investigations aimed at exploring (i) the capacity of HCV E1E2 glycoproteins to bind on Aedes mosquito cells and (ii) the ability of HCV serum particles (HCVsp) to replicate in these cell lines. Methods First, we used purified E1E2 expressing baculovirus-derived HCV pseudo particles (bacHCVpp) so we could investigate their association with mosquito cell lines from Aedes aegypti (Aag-2) and Aedes albopictus (C6/36). We initiated a series of infections of both mosquito cells (Ae aegypti and Ae albopictus) with the HCVsp (Lat strain - genotype 3) and we observed the evolution dynamics of viral populations within cells over the course of infection via next-generation sequencing (NGS) experiments. Results Our binding assays revealed bacHCVpp an association with the mosquito cells, at comparable levels obtained with human hepatocytes (HepaRG cells) used as a control. In our infection experiments, the HCV RNA (+) were detectable by RT-PCR in the cells between 21 and 28 days post-infection (p.i.). In human hepatocytes HepaRG and Ae aegypti insect cells, NGS experiments revealed an increase of global viral diversity with a selection for a quasi-species, suggesting a structuration of the population with elimination of deleterious mutations. The evolutionary pattern in Ae albopictus insect cells is different (stability of viral diversity and polymorphism). Conclusions These results demonstrate for the first time that natural HCV could really replicate within Aedes mosquitoes, a discovery which may have major consequences for public health as well as in vaccine development. Electronic supplementary material The online version of this article (doi:10.1186/s12985-017-0828-z) contains supplementary material, which is available to authorized users.

Published
2016
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44. First next-generation sequencing full-genome characterization of a hepatitis C virus genotype 7 divergent subtype

Author

M.-A. Thélu, Peter Simmonds, Katia Fusillier, N. De Castro, Maud Salmona, Séverine Mercier-Delarue, Olivier François, Jérôme LeGoff, Sarah Maylin, Marie-Laure Chaix, Alban Caporossi, Patrice Morand, Sylvie Larrat, and François Simon

Subjects
0301 basic medicine, Microbiology (medical), Male, Genotype, Hepacivirus, Hepatitis C virus, Genome, Viral, medicine.disease_cause, Genome, DNA sequencing, Evolution, Molecular, 03 medical and health sciences, Phylogenetics, medicine, Humans, Phylogeny, Whole genome sequencing, Genetics, biology, Strain (biology), High-Throughput Nucleotide Sequencing, General Medicine, Sequence Analysis, DNA, Middle Aged, biology.organism_classification, Virology, 030104 developmental biology, Infectious Diseases
Abstract

We report the near-full-length genome sequence of a hepatitis C virus (HCV) isolate from a man originating from Democratic Republic of Congo, the genotype of which could not be determined by the routinely used sequencing technique. The near-complete genome sequence of this variant BAK1 was obtained by the association of two next-generation sequencing technologies. Evolutionary analysis indicates that this isolate, BAK1, could be the first reported strain belonging to a new HCV-7b subtype. This new subtype has been incorrectly identified as genotype 2 by the Versant HCV Genotype 2.0 assay (LiPA). The requirement of three independent isolates has been filled, and a new subtype can be assigned. More examples of HCV-7 are required to better understand its origin, its pathogenicity and its relationship with genotype 2.

Published
2016

45. Ultradeep sequencing of B and non-B HIV-1 subtypes: Viral diversity and drug resistance mutations before and after one month of antiretroviral therapy in naive patients

Author

Michael G. B. Blum, Myriam Blanc, Om Kulkarni, Olivier Epaulard, Anne Signori-Schmuck, Katia Fusillier, Olivier François, Patrice Morand, Sylvie Larrat, and P Leclercq

Subjects
0301 basic medicine, Adult, Male, Genotype, Anti-HIV Agents, Population, HIV Infections, Drug resistance, Biology, medicine.disease_cause, 03 medical and health sciences, Plasma, immune system diseases, Virology, Antiretroviral Therapy, Highly Active, Genetic variation, Drug Resistance, Viral, medicine, Humans, education, education.field_of_study, Mutation, virus diseases, Genetic Variation, High-Throughput Nucleotide Sequencing, Sequence Analysis, DNA, Middle Aged, Viral Load, Reverse transcriptase, HIV Reverse Transcriptase, CD4 Lymphocyte Count, 030104 developmental biology, Infectious Diseases, HIV-1, Pyrosequencing, RNA, Viral, Reverse Transcriptase Inhibitors, Female, Viral load
Abstract

Background Ultradeep pyrosequencing technologies permit an assessment of the genetic diversity and the presence and frequency of minority variants in a viral population. The effect of these parameters on the outcome of highly active antiretroviral therapy (HAART) in HIV-infected patients is poorly understood. Objectives The present study used the pyrosequencing Roche 454 prototype assay to determine whether antiretroviral efficacy is correlated with viral diversity and minority drug resistance mutations in HIV-infected treatment-naive patients and to compare assay performance in B and non-B subtypes. Study design The study included 30 HIV-1 infected naive patients (20 with subtype non-B and 10 with subtype B). Ultradeep pyrosequencing of protease and reverse transcriptase genes was performed at baseline and 1 month after HAART initiation. Plasma HIV VL was measured at 0 and after 1, 3, and 6 months of HAART. Results Pre-HAART minority drug resistance mutations were observed to NRTI in 4 patients, to NNRTI in 6 patients, and to PI in 1 patient; there was no difference in HAART-induced VL decay between patients. Pre-HAART diversity was significantly correlated with the time elapsed since HIV-1 infection diagnosis, but not with the subtype, VL, or CD4 count. Patients with an undetectable VL after 3 months of HAART had a higher pre-HAART diversity. Pre- and post-HAART diversities were not statistically different. There was no difference in assay performance between subtype B and non-B. Conclusions A high pre-HAART viral diversity might have a positive effect on the outcome of HAART. Pre-therapeutic minority drug resistance mutations are uncommon in naive patients.

Published
2016

46. Performance of an antigen–antibody combined assay for hepatitis C virus testing without venipuncture

Author

Marie-Noelle Hilleret, Cécile Bourdon, Cécile Garnaud, Myriam Blanc, Jean-Louis Quesada, Vincent Leroy, Raphaele Germi, P Leclercq, Jean-Pierre Zarski, Monique Baccard, Sophie Mathieu, Sylvie Larrat, Patrice Morand, and Anne Signori-Schmuck

Subjects
Adult, Male, Fingerstick, Point-of-Care Systems, Hepatitis C virus, medicine.disease_cause, Sensitivity and Specificity, Young Adult, Phlebotomy, Antigen, Virology, medicine, Humans, Saliva, Dried blood, Antigens, Viral, Aged, Immunoassay, Venipuncture, biology, Clinical Laboratory Techniques, business.industry, virus diseases, Hepatitis C Antibodies, Middle Aged, Serum samples, Hepatitis C, digestive system diseases, Blood, Infectious Diseases, biology.protein, Female, Antibody, business
Abstract

Background Hepatitis C virus (HCV) is underdiagnosed and therefore increasing the opportunities for HCV testing without venipuncture may be useful. Objectives We evaluated the analytical performance of a modified, commercially available, combined HCV antigen–antibody assay (cEIA) (Monolisa ® HCV-Ag-Ab-ULTRA) and a commercially available point-of-care (POC) device (OraQuick ® HCV) on fingerstick blood (FSB) and oral mucosal transudate (OMT). Study design FSB, OMT and serum samples were collected from 113 cases of HCV-antibody-positive patients and 88 HCV-antibody-negative controls. The HCV-antibody-positive group included 63 patients with quantifiable HCV-RNA (56%) and 17 HIV/HCV co-infected patients (15%). FSB and OMT specimens were collected as dried blood spots (DBSs) or with the OraSure collection system, before testing with cEIA. Results With FSB specimens, the cEIA and the POC device exhibited 100% specificity and 98.2% and 97.4% sensitivity, respectively. The specificity of the cEIA in FSB sharply decreased if stored 3 days at room temperature. With OMT specimens, the cEIA sensitivity (71.7%) and specificity (94.3%) were significantly lower than the performance of OraQuick ® HCV (sensitivity, 94.6%; specificity, 100%). The optical densities obtained with the cEIA in FSB and OMT were lower in HIV/HCV co-infected patients compared with HCV monoinfected patients. Conclusion The cEIA using FSB specimens collected on DBSs preserved in appropriate storage conditions was a reliable alternative, equivalent to the POC assay, for HCV testing without venipuncture. The cEIA was not adapted for HCV testing on OMT.

Published
2012
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47. Plasmacytoid dendritic cells induce efficient stimulation of antiviral immunity in the context of chronic hepatitis B virus infection

Author

Joel Plumas, Marie-Jeanne Richard, Vincent Leroy, Jérémie Martinet, Tania Dufeu-Duchesne, Fabien Zoulim, Sylvie Larrat, and Caroline Aspord

Subjects
Adult, Male, Hepatitis B virus, Cellular immunity, Mice, SCID, Adaptive Immunity, CD8-Positive T-Lymphocytes, Biology, Transfection, medicine.disease_cause, Interferon-gamma, Mice, Young Adult, Immune system, Antigen, Lysosomal-Associated Membrane Protein 1, Mice, Inbred NOD, Lysosomal-Associated Membrane Protein 2, medicine, Animals, Humans, Cytotoxic T cell, Hepatitis B Vaccines, Hepatitis B e Antigens, Aged, Hepatitis B Surface Antigens, HLA-A Antigens, Hepatology, virus diseases, hemic and immune systems, Dendritic Cells, Hep G2 Cells, Middle Aged, Viral Load, Hepatitis B, Acquired immune system, medicine.disease, Hepatitis B Core Antigens, Virology, digestive system diseases, Disease Models, Animal, HBeAg, Immunology, Hepatocytes, Leukocytes, Mononuclear, Female
Abstract

The immune control of hepatitis B virus (HBV) infection is essential for viral clearance. Therefore, restoring functional anti–HBV immunity is a promising immunotherapeutic approach to treatment of chronic infection. Plasmacytoid dendritic cells (pDCs) play a crucial role in triggering antiviral immunity through their ability to capture and process viral antigens and subsequently induce adaptive immune responses. We investigated the potential of pDCs to trigger antiviral cellular immunity against HBV. We used a human leukocyte antigen A (HLA–A)*0201+ pDC line loaded with HLA–A*0201-restricted peptides derived from hepatitis B core/hepatitis B surface (HBc/HBs) antigens to amplify specific CD8 T cells ex vivo from chronic HBV patients and established a Hepato-HuPBL mouse model to address the therapeutic potential of the strategy in vivo. Stimulation of PBMCs or liver-infiltrating lymphocytes from HLA–A*0201+ chronic HBV patients by HBc peptide-loaded pDCs elicited up to 23.1% and 76.1% HBV-specific CD8 T cells in 45.8% of cases. The specific T cells from the “responder” group secreted interferon-γ, expressed CD107 upon restimulation, and efficiently lysed HBV antigen-expressing hepatocytes. Circulating hepatitis B e antigen (HBeAg) was found to distinguish the group of patients not responding to the pDC stimulation. The therapeutic efficacy of the pDC vaccine was evaluated in immunodeficient NOD-SCID β2m−/− mice reconstituted with HBV patients' PBMCs and xenotransplanted with human HBV-transfected hepatocytes. Vaccination of Hepato–HuPBL mice with the HBc/HBs peptide–loaded pDCs elicited HBV-specific T cells able to specifically lyse the transfected hepatocytes and reduce the systemic viral load. Conclusion: pDCs loaded with HBV–derived peptides can elicit functional virus-specific T cells. HBeAg appears to be critical in determining the outcome of immunotherapies in chronic HBV patients. A pDC-based immunotherapeutic approach could be of interest in attempts to restore functional antiviral immunity, which is critical for the control of the virus in chronic HBV patients. (HEPATOLOGY 2012;56:1706–1718)

Published
2012
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48. Comparison of Commercial Extraction Systems and PCR Assays for Quantification of Epstein-Barr Virus DNA Load in Whole Blood

Author

Sylvie Larrat, Julien Lupo, Nelly Magnat, Jean-Marie Seigneurin, Touyana Semenova, Laurence Grossi, Patrice Morand, and Raphaële Germi

Subjects
Microbiology (medical), Epstein-Barr Virus Infections, Herpesvirus 4, Human, Biology, Real-Time Polymerase Chain Reaction, Statistics, Nonparametric, Virus, chemistry.chemical_compound, Virology, Humans, Whole blood, Automation, Laboratory, Extraction (chemistry), Hematopoietic Stem Cell Transplantation, Organ Transplantation, Reference Standards, Viral Load, Molecular diagnostics, Molecular biology, DNA extraction, Real-time polymerase chain reaction, chemistry, DNA, Viral, Reagent Kits, Diagnostic, Viral load, DNA
Abstract

The automation of DNA extraction and the use of commercial quantitative real-time PCR assays could help obtain more reliable results for the quantification of Epstein-Barr virus DNA loads (EBV VL). This study compared two automated extraction platforms and two commercial PCRs for measurement of EBV VL in 10 EBV specimens from Quality Control for Molecular Diagnostics (QCMD) and in 200 whole-blood (WB) specimens from transplant ( n = 137) and nontransplant ( n = 63) patients. The WB specimens were extracted using the QIAcube or MagNA Pure instrument; VL were quantified with the EBV R-gene quantification kit (Argene) or the artus EBV RG PCR kit (Qiagen) on the Rotor-Gene 6000 real-time analyzer; and the results were compared with those of a laboratory-developed PCR. DNA was extracted from the QCMD specimens by use of the QIAamp DNA minikit and was quantified by the three PCR assays. The extraction platforms and the PCR assays showed good correlation ( R , >0.9; P , 10 copies/ml), and standard deviations reached as high as 0.49 log 10 copy/ml. In WB but not in QCMD samples, Argene PCR tended to give higher VL values than artus PCR or the laboratory-developed PCR (mean difference for the 200 WB VL, −0.42 or −0.36, respectively). In conclusion, the two automated extraction platforms and the two PCRs provided reliable and comparable VL results, but differences greater than 0.5 log 10 copy/ml remained between the two commercial PCRs after common DNA extraction.

Published
2012
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49. Apport du séquençage nouvelle génération au typage MLST de Pneumocysits jirovecii , dans le cadre d’une épidémie chez des patients transplantés d’organe solide

Author

Eléna Charpentier, Danièle Maubon, Jean-Benjamin Murat, Cécile Garnaud, Sébastien Bailly, Patricia Pavese, Thibault Drouet, Paolo Malvezzi, Caroline Augier, Anne Thiebaut-Bertrand, Claire Wintenberger, Muriel Cornet, Olivier Epaulard, Marie-Reine Mallaret, Sylvie Larrat, and John Rendu

Subjects
Infectious Diseases
Abstract

Le typage par multilocus sequence typing (MLST) permet une caracterisation genotypique precise et reproductible des souches, mais les performances du sequencage Sanger (Sseq) pour la detection de souches minoritaires sont limitees. Or, les infections a differentes souches (infections mixtes) sont frequentes au cours des pneumonies a Pneumocystis jirovecii (PCP). Le sequencage nouvelle generation (NGS) permet la mise en evidence a la fois des populations minoritaires et majoritaires. Dans ce travail, nous avons etudie l’apport du NGS pour le typage MLST des souches issues de cas groupes (cluster) de PCP chez des patients transplantes d’organe solide (TOS). Trente-deux patients presentant une PCP au CHU Grenoble-Alpes entre 2012 et 2015 ont ete inclus. Parmi eux, 12 TOS (rein, cœur, foie, poumons) appartenaient a un cluster observe entre mai 2014 et aout 2015 et 20 patients, epidemiologiquement non relies entre eux, representaient le groupe controle. Une strategie MSLT a trois loci ( mtLSU , CYTB , SOD ) a ete adoptee, comprenant, pour des contraintes techniques liees au sequencage NGS, de nouvelles amorces permettant l’obtention d’amplicons d’environ 750 pb. Les souches de 12 des 32 patients ont egalement ete analysees par Sseq en parallele. Les genotypes obtenus ont ete nommes arbitrairement car il n’existait pas d’etudes prealables utilisant des amplicons etendus de 750 pb. Parmi les 12 TOS, 5 partageaient un meme genotype majoritaire (C2a), regroupant 3 patients transplantes renaux, un patient transplante cardiaque et un patient double transplante renal et cardiaque. Ce genotype C2a n’a pas ete retrouve dans le groupe controle. Toutes populations confondues, la proportion d’infections mixtes etait plus elevee avec le NGS (65 %) qu’avec le Sseq (25 %). Si seule la population controle (hors cluster) etait consideree, cette proportion d’infection mixtes augmentait a 85 %. Le NGS a revele des infections avec plus de deux souches (53 %), et la presence de souches minoritaires, parmi lesquelles le genotype C2a. Une carte de transmission a confirme la probable acquisition nosocomiale de PCP. Un sixieme patient qui presentait la souche C2a sous forme minoritaire a ete inclus dans la carte de transmission. Les sequences etendues de mtLSU ont revele 4 nouveaux polymorphismes augmentant significativement le pouvoir discriminant de ce locus. En conclusion, et comme attendu dans ce contexte de PCP, le NGS surpasse les capacites du Sseq pour la detection d’infections mixtes et permet de preciser la carte de transmission. L’utilisation d’un amplicon etendu a 750 bp pour le locus mtLSU est interessante pour augmenter le pouvoir discriminant du schema MLST. Ce travail ouvre de nouvelles perspectives pour les futures etudes epidemiologiques de ce champignon non cultivable.

Published
2017
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50. Inhibition of Epstein–Barr virus replication by small interfering RNA targeting the Epstein–Barr virus protease gene

Author

Patrice Morand, Solenne Vigne, Marlyse Buisson, Wim P. Burmeister, Jean-Marie Seigneurin, Sylvie Larrat, Jean-Marc Crance, Ariane Bas, Véronique Boyer, Raphaële Germi, Laurence Grossi, and Sandrine Nicod

Subjects
Pharmacology, Biology, medicine.disease_cause, biology.organism_classification, Virology, Molecular biology, Epstein–Barr virus, Herpesviridae, Virus, Infectious Diseases, Capsid, Lytic cycle, RNA interference, hemic and lymphatic diseases, medicine, Gene silencing, Gammaherpesvirinae, Pharmacology (medical)
Abstract

Background The Epstein–Barr virus (EBV) protease (PR), coded by the BVRF2 gene, is essential for the maturation of the viral capsid and viral DNA packaging during the late stage of the EBV lytic cycle. Like the other herpesvirus serine PRs, EBV PR could be a target for the inhibition of EBV replication. To date, no data have been reported on the inhibition of EBV PR messenger RNA (mRNA) by small interfering RNA (siRNA). Methods In this study, siRNAs targeting EBV PR were delivered to the epithelial 293 cell line stably transfected with the complete B95-8 EBV episome. EBV DNA and PR mRNA were quantified by real-time PCR in cells and supernatant, protein expression was assessed by immunoblotting, and production of EBV infectious particles in the culture medium was measured by Raji cell superinfection. Results The EBV PR mRNA within the cells was reduced by 73%, the PR protein by 35% and the amount of virus in the cell supernatant was drastically decreased by 86% or 95%, depending on the method. Conclusions The strong effect of the siRNA targeting EBV PR on EBV replication attests to the crucial role played by EBV PR in the production of infectious particles and suggests that targeting this enzyme can be a new strategy against EBV-associated diseases where virus replication occurs.

Published
2008
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