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2.
Infect Dis Now ; 53(3): 104650, 2023 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-36702307

RESUMO

OBJECTIVES: Experience of Nextstrain [1,2] and its approach adapted to the local context encouraged us to carry out real-time monitoring of COVID-19 nosocomial clusters in our establishment, the Grenoble Alpes University Hospital. PATIENTS AND METHODS, RESULTS: Through identification from electronic health records of nosocomial pathways and clusters and calculation of genetic distances from sequenced samples of COVID-19 patients, we were able to identify potential nosocomial clusters in very close to real time with a significant time saving compared to classical epidemiological surveillance, and to better understand and characterize nosocomial clusters. CONCLUSION: Through early detection and characterization of clusters, we may prevent infection of our patients by further implementing the appropriate measures.


Assuntos
COVID-19 , Infecção Hospitalar , Humanos , COVID-19/diagnóstico , COVID-19/epidemiologia , COVID-19/prevenção & controle , SARS-CoV-2/genética , Infecção Hospitalar/epidemiologia , Hospitais Universitários
3.
Infect Dis Now ; 52(8): 453-455, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-36108970

RESUMO

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 (R2 ≥ 0.9983). Detection of low viral load samples was evaluated, on 10-times repeat-test. All replicate samples were detected with both thermocycling programs and were quantified at similar viral loads (bias in log10 copies/mL: +0.05 (HSV-1), -0.01 (HSV-2) and +0.25 (VZV)). This confirms the feasibility of using the RT-PCR thermocycling program to detect and quantify the genome of RNA and DNA viruses in a single run.


Assuntos
Vírus de RNA , Humanos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sensibilidade e Especificidade , Reação em Cadeia da Polimerase em Tempo Real , Vírus de DNA
4.
Stud Health Technol Inform ; 290: 1046-1047, 2022 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-35673198

RESUMO

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.


Assuntos
COVID-19 , COVID-19/epidemiologia , Data Warehousing , Geografia , Hospitais Universitários , Humanos
5.
Antimicrob Resist Infect Control ; 11(1): 31, 2022 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-35135618

RESUMO

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.


Assuntos
Infecção Hospitalar , Influenza Humana , Infecção Hospitalar/epidemiologia , Infecção Hospitalar/prevenção & controle , Hospitais Universitários , Humanos , Influenza Humana/epidemiologia , Influenza Humana/prevenção & controle , Estudos Retrospectivos , Vacinação
7.
Am J Infect Control ; 50(2): 155-158, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34562527

RESUMO

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.


Assuntos
Influenza Humana , Antivirais/uso terapêutico , Leitos , Pessoal de Saúde , Hospitais Universitários , Humanos , Influenza Humana/epidemiologia , Oseltamivir/uso terapêutico
8.
J Clin Virol ; 143: 104947, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34492569

RESUMO

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.


Assuntos
COVID-19 , SARS-CoV-2 , Antígenos Virais , Humanos , Nasofaringe , Pandemias , Estudos Retrospectivos , Sensibilidade e Especificidade
9.
Emerg Infect Dis ; 27(10): 2725-2728, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34352197

RESUMO

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.


Assuntos
COVID-19 , SARS-CoV-2 , Idoso , Anticorpos Antivirais , Humanos , Hospedeiro Imunocomprometido , Masculino , Glicoproteína da Espícula de Coronavírus
10.
J Infect Public Health ; 13(7): 1047-1050, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32224109

RESUMO

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 (<3 months) non-traumatic, non-metabolic, non-vascular neurological injuries. Acute hepatitis E was defined as anti-HEV IgM-positive serum in immunocompetent patient, and as anti-HEV IgM-positive serum or HEV RNA-positive serum in immunocompromised patients. RESULTS: One hundred fifty-nine patients were included. Anti-HEV IgM seroprevalence in our cohort of non-traumatic, non-metabolic, non-vascular neurological injuries was 6.9% (eleven patients, including 4 Parsonage-Turner syndrome (PTS) and 2 Guillain-Barré syndrome (GBS)). Elevated transaminases were observed in only 64% of hepatitis E patients and cholestasis in 64%. CONCLUSION: In this study, 6·9% of patients with acute non-traumatic, non-metabolic, non-vascular neurological injuries had a probable recent HEV infection. HEV serology should be systematically performed in this population, even in patients with normal transaminase level.


Assuntos
Anticorpos Anti-Hepatite/sangue , Vírus da Hepatite E/imunologia , Hepatite E/epidemiologia , Doenças do Sistema Nervoso/epidemiologia , Doença Aguda/epidemiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Neurite do Plexo Braquial/epidemiologia , Feminino , França/epidemiologia , Síndrome de Guillain-Barré/epidemiologia , Hepatite E/sangue , Hepatite E/diagnóstico , Hepatite E/imunologia , Humanos , Imunoglobulina M/sangue , Masculino , Pessoa de Meia-Idade , Doenças do Sistema Nervoso/imunologia , Estudos Prospectivos , RNA Viral/sangue , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Estudos Soroepidemiológicos , Transaminases/sangue , Adulto Jovem
11.
PLoS One ; 14(5): e0216308, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31063477

RESUMO

STUDY OBJECTIVE: To investigate the performance of a rapid RT-PCR assay to detect influenza A/B at emergency department admission. METHODS: This 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. RESULTS: A 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. CONCLUSION: Rapid RT-PCR assay performed by nurses at triage to detect influenza A/B is feasible and highly accurate.


Assuntos
Vírus da Influenza A/genética , Vírus da Influenza B/genética , Influenza Humana , Técnicas de Diagnóstico Molecular , Sistemas Automatizados de Assistência Junto ao Leito , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Adulto , Idoso , Idoso de 80 Anos ou mais , Serviço Hospitalar de Emergência , Feminino , França/epidemiologia , Humanos , Influenza Humana/diagnóstico , Influenza Humana/epidemiologia , Influenza Humana/genética , Influenza Humana/virologia , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Estações do Ano , Manejo de Espécimes
12.
Clin Res Hepatol Gastroenterol ; 42(4): 313-318, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29551607

RESUMO

INTRODUCTION: Presence of anti-E1E2 antibodies was previously associated with spontaneous cure of hepatitis C virus (HCV) and predictive before treatment of a sustained virological response (SVR) to bi- or tri-therapy in naïve or experienced patients, regardless of HCV genotype. We investigated the impact of anti-E1E2 seroprevalence at baseline on treatment response in patients receiving direct-acting antiviral (DAA) therapy. MATERIAL AND METHODS: We screened anti-E1E2 antibodies by ELISA in serum samples collected at treatment initiation for two groups of patients: 59 with SVR at the end of DAA treatment and 44 relapsers after DAA treatment. Nineteen patients received a combination of ribavirin (RBV) or PEG-interferon/ribavirin with sofosbuvir or daclatasvir and others received interferon-free treatment with DAA±RBV. HCV viral load was measured at different time points during treatment in a subgroup of patients. RESULTS: A significant association was observed between presence of anti-E1E2 and HCV viral load<6log10 prior treatment. Among patients with anti-E1E2 at baseline, 70% achieved SVR whereas among patients without anti-E1E2, only 45% achieved SVR. Conversely, 66% of patients experiencing DAA-failure were anti-E1E2 negative at baseline. In the multivariate analysis, presence of anti-E1E2 was significantly associated with SVR after adjustment on potential cofounders such as age, sex, fibrosis stage, prior HCV treatment and alanine aminotransferase (ALT) level. CONCLUSIONS: The presence of anti-E1E2 at treatment initiation is a predictive factor of SVR among patients treated with DAA and more likely among patients with low initial HCV viral load (<6log10). Absence of anti-E1E2 at baseline could predict DAA-treatment failure.


Assuntos
Anticorpos/sangue , Antivirais/uso terapêutico , Hepatite C Crônica/tratamento farmacológico , Peptídeos/imunologia , Idoso , Biomarcadores/sangue , Carbamatos , Quimioterapia Combinada , Ensaio de Imunoadsorção Enzimática , Feminino , Hepacivirus/fisiologia , Hepatite C Crônica/virologia , Humanos , Imidazóis/uso terapêutico , Interferon-alfa/uso terapêutico , Masculino , Pessoa de Meia-Idade , Polietilenoglicóis/uso terapêutico , Pirrolidinas , Proteínas Recombinantes/uso terapêutico , Estudos Retrospectivos , Ribavirina/uso terapêutico , Estudos Soroepidemiológicos , Sofosbuvir/uso terapêutico , Valina/análogos & derivados , Carga Viral/efeitos dos fármacos
13.
PLoS One ; 13(3): e0194396, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29566005

RESUMO

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 <200 IU/mL (p = 0.0061). The workflow allowed a non-specialized technician to obtain results in less than 4 hours whereas 2 to 3 days and experienced staff were required with the in-house assay. In conclusion, the Roche cobas® HCV GT kit is easy and rapid to use and provides reliable results. The high rate of uninterpretable results particularly for low viral load samples and less frequent genotypes, and the absence of subtyping for non-genotype 1 could require sending complex samples to a specialized laboratory.


Assuntos
Genótipo , Técnicas de Genotipagem/métodos , Hepacivirus/genética , Hepatite C Crônica/diagnóstico , Testes Sorológicos/métodos , Antivirais/farmacologia , Antivirais/uso terapêutico , Infecções Assintomáticas , Hepacivirus/efeitos dos fármacos , Hepacivirus/isolamento & purificação , Hepatite C Crônica/sangue , Hepatite C Crônica/tratamento farmacológico , Hepatite C Crônica/virologia , Humanos , RNA Viral/genética , RNA Viral/isolamento & purificação , Kit de Reagentes para Diagnóstico , Reação em Cadeia da Polimerase em Tempo Real , Estudos Retrospectivos , Análise de Sequência de RNA , Carga Viral/efeitos dos fármacos
14.
J Med Virol ; 90(5): 994-997, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29350410

RESUMO

The use of new Direct Acting Antivirals, specific of HCV, has greatly improved the HCV treatment. Most of the DAA are specific of HCV genotypes. Genotyping methods may target different regions of the HCV genome, though only the whole genome sequencing could confirm the correct genotype. The present study describes the virological investigation of a treatment failure due to the false identification of an unusual 2k/1b recombinant HCV form. It describes the sequencing methods, and the similarity analysis of the sequences to different genotype query sequences, to identify the recombination breakpoint.


Assuntos
Antivirais/administração & dosagem , Erros de Diagnóstico , Genótipo , Hepacivirus/classificação , Hepacivirus/genética , Hepatite C Crônica/tratamento farmacológico , Adulto , Técnicas de Genotipagem , Hepacivirus/isolamento & purificação , Hepatite C Crônica/virologia , Humanos , Masculino , Recombinação Genética , Análise de Sequência de DNA , Falha de Tratamento
16.
Antivir Ther ; 23(4): 307-314, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-28730994

RESUMO

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.


Assuntos
Antivirais/farmacologia , Farmacorresistência Viral , Hepacivirus/efeitos dos fármacos , Hepacivirus/genética , Hepatite C/virologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Proteínas não Estruturais Virais/genética , Antivirais/uso terapêutico , Análise Mutacional de DNA , Quimioterapia Combinada , Feminino , Genótipo , Hepatite C/tratamento farmacológico , Humanos , Masculino , Falha de Tratamento , Resultado do Tratamento , Carga Viral
17.
J Clin Virol ; 95: 13-19, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28830014

RESUMO

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.


Assuntos
Fármacos Anti-HIV/uso terapêutico , Farmacorresistência Viral/genética , Variação Genética , Infecções por HIV/tratamento farmacológico , HIV-1/genética , Mutação , Adulto , Fármacos Anti-HIV/farmacologia , Terapia Antirretroviral de Alta Atividade , Contagem de Linfócito CD4 , Feminino , Genótipo , Infecções por HIV/imunologia , Infecções por HIV/virologia , Transcriptase Reversa do HIV/genética , HIV-1/classificação , HIV-1/efeitos dos fármacos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Masculino , Pessoa de Meia-Idade , Plasma/virologia , RNA Viral/genética , Inibidores da Transcriptase Reversa/uso terapêutico , Análise de Sequência de DNA , Carga Viral
18.
Virol J ; 14(1): 161, 2017 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-28830495

RESUMO

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.


Assuntos
Aedes/virologia , Hepacivirus/genética , Insetos Vetores/virologia , Replicação Viral/fisiologia , Animais , Linhagem Celular , Genótipo , Hepacivirus/isolamento & purificação , Hepatite C/sangue , Hepatócitos/virologia , Humanos , Mutação , Peptídeos/metabolismo , Filogenia , Reação em Cadeia da Polimerase/métodos , Polimorfismo Genético , RNA Viral , Análise de Sequência , Proteínas do Envelope Viral/metabolismo
19.
Emerg Infect Dis ; 23(8): 1237-1245, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28726611

RESUMO

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.


Assuntos
Tipagem de Sequências Multilocus , Pneumocystis carinii/classificação , Pneumocystis carinii/genética , Pneumonia por Pneumocystis/epidemiologia , Pneumonia por Pneumocystis/microbiologia , Adolescente , Adulto , Idoso , Estudos de Casos e Controles , Criança , Pré-Escolar , Biologia Computacional/métodos , Surtos de Doenças , Feminino , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Filogenia , Pneumonia por Pneumocystis/transmissão , Polimorfismo Genético , Sensibilidade e Especificidade , Adulto Jovem
20.
PLoS One ; 12(4): e0174651, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28394908

RESUMO

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.


Assuntos
Evolução Molecular , Duplicação Gênica , Hepacivirus/genética , Recombinação Genética , Proteínas não Estruturais Virais/genética , Teorema de Bayes , Carcinoma Hepatocelular/virologia , Estudos de Coortes , Loci Gênicos , Hepatite C/virologia , Interações Hospedeiro-Patógeno , Humanos/virologia , Neoplasias Hepáticas/virologia , Mutação , Filogenia , Polimorfismo Genético , Proteínas do Envelope Viral/genética
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