RESUMO
We used variant typing polymerase chain reaction to describe the evolution of severe acute respiratory syndrome coronavirus 2 Omicron sublineages between December 2021 and mid-March 2022. The selective advantage of the BA.2 variant over BA.1 is not due to greater nasopharyngeal viral loads.
Assuntos
COVID-19 , Humanos , Carga Viral , Reação em Cadeia da Polimerase , SARS-CoV-2/genética , Testes SorológicosRESUMO
In cervical cancer screening programs, the detection of high-risk human papillomavirus (HR-HPV) is now widely implemented on physician-collected samples and has expanded to include self-collected samples. The use of a cellularity control (CC) is needed to reduce false-negative HPV results. An external mRNA CC for the HPV APTIMA® assay was assessed for its analytical performance and the results were compared with both cervix cytobrush samples taken by physicians and self-collected vaginal samples from 148 women. The performance of the CC was adjusted to control for the presence of cellular mRNA in the ThinPrep® and Multitest® transport media. This CC is user-friendly but implies to perform two independent assays on PANTHER® automate. Self-collected vaginal sampling gives a lower median CC results (13.2 vs. 16.9 min) but a higher risk of negative CC results (3.3 vs. 0%). The usefulness of the CC for the HR-HPV assay may be optimized by the definition of a threshold for a minimum cell number to be tested to increase confidence in HPV-negative results. The systematic use of an RNA CC increases confidence for HPV RNA assays on self-collected vaginal samples.
Assuntos
Infecções por Papillomavirus , Displasia do Colo do Útero , Neoplasias do Colo do Útero , Feminino , Humanos , Neoplasias do Colo do Útero/diagnóstico , Infecções por Papillomavirus/diagnóstico , Sensibilidade e Especificidade , Esfregaço Vaginal/métodos , Detecção Precoce de Câncer/métodos , Papillomaviridae/genética , RNA Mensageiro/genética , Manejo de Espécimes/métodos , Papillomavirus HumanoRESUMO
Background: Minority resistant variants of human immunodeficiency virus type 1 (HIV-1) could influence the virological response to treatment based on nonnucleoside reverse transcriptase inhibitors (NNRTIs). Data on minority rilpivirine-resistant variants are scarce. This study used next-generation sequencing (NGS) to identify patients harboring minority resistant variants to nucleos(t)ide reverse transcriptase inhibitors and NNRTIs and to assess their influence on the virological response (VR). Methods: All the subjects, 541 HIV-1-infected patients started a first-line regimen containing rilpivirine. VR was defined as a HIV-1 RNA load <50 copies/mL at month 6 with continued suppression at month 12. NGS was performed at baseline (retrospectively) on the 454 GS-FLX platform (Roche). Results: NGS revealed resistance-associated mutations accounting for 1% to <5% of variants in 17.2% of samples, for 5%-20% in 5.7% of samples, and for >20% in 29% of samples. We identified 43 (8.8%) and 36 (7.4%) patients who harbored rilpivirine-resistant variants with a 1% sensitivity threshold according to the French National Agency for Research on AIDS and Viral Hepatitis and Stanford algorithms, respectively. The VR was 96.9% at month 12. Detection of minority rilpivirine resistant variants was not associated with virological failure (VF). Multivariate analysis indicated that VF at month 12 was associated with a CD4 count <250 cells/µL at baseline, a slower decrease in viral load at month 3, and rilpivirine resistance at baseline using the Stanford algorithm with a 20% threshold. Conclusions: Minority resistant variants had no impact on the VR of treatment-naive patients to a rilpivirine-based regimen.
Assuntos
Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , HIV-1/genética , Rilpivirina/uso terapêutico , Adulto , Farmacorresistência Viral , Feminino , Variação Genética , Humanos , Masculino , Mutação , Rilpivirina/administração & dosagem , Carga ViralRESUMO
Objectives: To evaluate the diagnostic performance of the Vela next-generation sequencing (NGS) system in conjunction with the Sentosa SQ HIV Genotyping Assay for genotyping HIV-1. Methods: Plasma RNA was extracted and templates prepared with the Sentosa SX instrument before sequencing the HIV-1 polymerase on the Sentosa SQ301 Sequencer (PGM IonTorrent). The Vela NGS System was compared with direct sequencing and the 454 GS-FLX (Roche) and MiSeq (Illumina) systems for genotypic resistance testing on clinical samples. Results: The Vela NGS system detected majority resistance mutations in subtype B and CRF02-AG samples at 500 copies/mL and minority variants with a sensitivity of 5% at 100â¯000 copies/mL. The Vela NGS system and direct sequencing identified resistance mutations with 97% concordance in 46 clinical samples. Vela identified 1/20 of the 1%-5% mutations identified by 454, 5/12 of the 5%-20% mutations and 60/61 of the >20% mutations. Vela identified 3/14 of the 1%-5% mutations identified by MiSeq, 0/2 of the 5%-20% mutations and 47/47 of the >20% mutations. The resistance mutation quantifications by Vela and 454 were concordant (bias: 2.31%), as were those by Vela and MiSeq (bias: 1.06%). Conclusions: The Vela NGS system provides automated nucleic acid extraction, PCR reagent distribution, library preparation and bioinformatics analysis. The analytical performance was very good when compared with direct sequencing, but was less sensitive than two other NGS platforms for detecting minority variants.
Assuntos
Automação Laboratorial/métodos , Farmacorresistência Viral , Técnicas de Genotipagem/métodos , Infecções por HIV/virologia , HIV-1/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Testes de Sensibilidade Microbiana/métodos , Biologia Computacional/métodos , Genes Virais , Genótipo , HIV-1/efeitos dos fármacos , HIV-1/isolamento & purificação , Mutação , RNA Viral/genética , Produtos do Gene pol do Vírus da Imunodeficiência Humana/genéticaRESUMO
The present study compares the performances of an in-house sequencing protocol developed on MiSeq, the Sanger method, and the 454 GS-FLX for detecting and quantifying drug-resistant mutations (DRMs) in the human immunodeficiency virus polymerase gene (reverse transcriptase [RT] and protease [PR]). MiSeq sequencing identified all the resistance mutations detected by bulk sequencing (n = 84). Both the MiSeq and 454 GS-FLX platforms identified 67 DRMs in the RT and PR regions, but a further 25 DRMs were identified by only one or other of them. Pearson's analysis showed good concordance between the percentage of drug-resistant variants determined by MiSeq and 454 GS-FLX sequencing (ρ = .77, P < .0001). The MiSeq platform is as accurate as the 454 GS-FLX Roche system for determining RT and PR DRMs and could be used for monitoring human immunodeficiency virus type 1 drug resistance.
Assuntos
Variação Genética , Infecções por HIV/virologia , HIV-1/classificação , HIV-1/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Mutação , Produtos do Gene pol do Vírus da Imunodeficiência Humana/genética , Farmacorresistência Viral , Protease de HIV/genética , Transcriptase Reversa do HIV/genética , HIV-1/isolamento & purificação , HumanosRESUMO
Direct-acting agents (DAAs) are highly efficient at treating hepatitis C virus (HCV) infections after kidney transplantation. Although drug agencies have recently warned of the risk of hepatitis B virus (HBV) reactivation after patients have received DAAs, reports have discrepant results in HBsAg-positive and HBsAg-negative patients. We report on 3 cases of HBV reactivation that were detected after achieving a DAA-associated sustained virological response in 3 kidney-transplant recipients initially HBsAg-negative. In the first case, retrospective virological analysis revealed that HBsAgs had become positive and HBV DNA was detectable before initiating DAA therapy. In the second and third cases, HBV reactivation occurred 2 months and more than 1 year after stopping anti-HCV therapy. These cases underline the discrepancies and highlight the need for comprehensive information before making definitive conclusions regarding the causal link between DAAs and HBV reactivation.
Assuntos
Antivirais/uso terapêutico , Coinfecção/virologia , Vírus da Hepatite B/fisiologia , Transplante de Rim/efeitos adversos , Ativação Viral/efeitos dos fármacos , Antivirais/administração & dosagem , Coinfecção/tratamento farmacológico , DNA Viral/sangue , Feminino , Hepacivirus/efeitos dos fármacos , Hepatite B/tratamento farmacológico , Hepatite B/virologia , Antígenos de Superfície da Hepatite B/análise , Vírus da Hepatite B/efeitos dos fármacos , Hepatite C/tratamento farmacológico , Hepatite C/virologia , Hepatite C Crônica/tratamento farmacológico , Hepatite C Crônica/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Resposta Viral Sustentada , TransplantadosRESUMO
The analytical performance of the Veris HIV-1 assay for use on the new, fully automated Beckman Coulter DxN Veris molecular diagnostics system was evaluated at 10 European virology laboratories. The precision, analytical sensitivity, performance with negative samples, linearity, and performance with HIV-1 groups/subtypes were evaluated. The precision for the 1-ml assay showed a standard deviation (SD) of 0.14 log10 copies/ml or less and a coefficient of variation (CV) of ≤6.1% for each level tested. The 0.175-ml assay showed an SD of 0.17 log10 copies/ml or less and a CV of ≤5.2% for each level tested. The analytical sensitivities determined by probit analysis were 19.3 copies/ml for the 1-ml assay and 126 copies/ml for the 0.175-ml assay. The performance with 1,357 negative samples demonstrated 99.2% with not detected results. Linearity using patient samples was shown from 1.54 to 6.93 log10 copies/ml. The assay performed well, detecting and showing linearity with all HIV-1 genotypes tested. The Veris HIV-1 assay demonstrated analytical performance comparable to that of currently marketed HIV-1 assays. (DxN Veris products are Conformité Européenne [CE]-marked in vitro diagnostic products. The DxN Veris product line has not been submitted to the U.S. FDA and is not available in the U.S. market. The DxN Veris molecular diagnostics system is also known as the Veris MDx molecular diagnostics system and the Veris MDx system.).
Assuntos
Automação Laboratorial/métodos , Infecções por HIV/diagnóstico , HIV-1/isolamento & purificação , Técnicas de Diagnóstico Molecular/métodos , Europa (Continente) , Humanos , Sensibilidade e EspecificidadeRESUMO
The analytical performance of the Veris HCV Assay for use on the new and fully automated Beckman Coulter DxN Veris Molecular Diagnostics System (DxN Veris System) was evaluated at 10 European virology laboratories. Precision, analytical sensitivity, specificity, and performance with negative samples, linearity, and performance with hepatitis C virus (HCV) genotypes were evaluated. Precision for all sites showed a standard deviation (SD) of 0.22 log10 IU/ml or lower for each level tested. Analytical sensitivity determined by probit analysis was between 6.2 and 9.0 IU/ml. Specificity on 94 unique patient samples was 100%, and performance with 1,089 negative samples demonstrated 100% not-detected results. Linearity using patient samples was shown from 1.34 to 6.94 log10 IU/ml. The assay demonstrated linearity upon dilution with all HCV genotypes. The Veris HCV Assay demonstrated an analytical performance comparable to that of currently marketed HCV assays when tested across multiple European sites.
Assuntos
Automação Laboratorial/métodos , Hepacivirus/isolamento & purificação , Hepatite C Crônica/virologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Carga Viral/métodos , Europa (Continente) , Humanos , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: Viral reservoirs represent an important barrier to HIV cure. Accurate markers of HIV reservoirs are needed to develop multicenter studies. The aim of this multicenter quality control (QC) was to evaluate the inter-laboratory reproducibility of total HIV-1-DNA quantification. METHODS: Ten laboratories of the ANRS-AC11 working group participated by quantifying HIV-DNA with a real-time qPCR assay (Biocentric) in four samples (QCMD). RESULTS: Good reproducibility was found between laboratories (standard deviation ≤ 0.2 log10 copies/106 PBMC) for the three positive QC that were correctly classified by each laboratory (QC1Assuntos
Técnicas de Laboratório Clínico/normas
, DNA Viral/análise
, HIV-1/isolamento & purificação
, HIV-1/fisiologia
, Controle de Qualidade
, Reação em Cadeia da Polimerase em Tempo Real/normas
, Carga Viral/normas
, Técnicas de Laboratório Clínico/métodos
, Reservatórios de Doenças/virologia
, HIV-1/genética
, Humanos
, Colaboração Intersetorial
, RNA Viral/sangue
, Reação em Cadeia da Polimerase em Tempo Real/métodos
, Reprodutibilidade dos Testes
RESUMO
UNLABELLED: Most cases of hepatitis E viral (HEV) infection in developed countries are autochthonous. Nevertheless, the reported seroprevalence of HEV varies greatly depending on the geographical area and the performance of the immunoassay used. We used validated assays to determine the prevalence of anti-HEV immunoglobulin G (IgG) and IgM among 10,569 French blood donors living in mainland France and three overseas areas. Epidemiological information was collected using a specific questionnaire. We found an overall IgG seroprevalence of 22.4% (8%-86.4%) depending on the geographical area (P < 0.001). The presence of anti-HEV IgG was associated with increasing age (P < 0.001) and eating pork meat (P = 0.03), pork liver sausages (P < 0.001), game meat (P < 0.01), offal (P < 0.001), and oysters (P = 0.02). Conversely, drinking bottled water was associated with a lower rate of anti-HEV IgG (P = 0.02). Overall IgM seroprevalence was 1% (0%-4.6%). The frequency of anti-HEV IgM was higher in donors living in a high anti-HEV IgG seroprevalence area (1.9% versus 0.7%, P < 0.001) and in those eating pork liver sausage (1.4% versus 0.7%, P < 0.01), pâté (1% versus 0.4, P = 0.04), and wild boar (1.3% versus 0.7%, P < 0.01). CONCLUSION: HEV is endemic in France and hyperendemic in some areas; eating habits alone cannot totally explain the exposure to HEV, and contaminated water could contribute to the epidemiology of HEV infection in France.
Assuntos
Doadores de Sangue , Transmissão de Doença Infecciosa/estatística & dados numéricos , Anticorpos Anti-Hepatite/sangue , Hepatite E/epidemiologia , Imunoglobulina G/imunologia , Adulto , Distribuição de Qui-Quadrado , Estudos Transversais , Ensaio de Imunoadsorção Enzimática , Feminino , França/epidemiologia , Hepatite E/etiologia , Vírus da Hepatite E/isolamento & purificação , Humanos , Imunoglobulina G/sangue , Masculino , Pessoa de Meia-Idade , Prevalência , Medição de Risco , Estudos Soroepidemiológicos , Estatísticas não Paramétricas , Inquéritos e QuestionáriosRESUMO
BACKGROUND: Malignancies and lymphoma are common complications after kidney transplantation. However, no link has been made between the incidence of malignancies and hepatitis C virus (HCV) infection in this setting. This case-controlled study compared the incidence of malignancies, including lymphoma, between kidney transplant (KT) patients with or without HCV replication. PATIENTS AND METHODS: A total of 99 HCV-positive RNA-positive KT patients were matched with 198 (1:2) anti-HCV-negative patients according to age, gender, and date of transplantation, and were followed for 145.8±78.4 months. RESULTS: During the follow-up period, 28 HCV-positive (28%) cases developed at least one cancer, and 64 (32%) patients developed cancer in the HCV-negative group (P=not significant [ns]). Survival without a cancer was similar between both groups. Thirteen HCV-positive patients (13%) developed at least one solid cancer vs 29 (15%) HCV-negative patients (P=ns). Survival without a solid cancer was similar between both groups. Three patients from the HCV-positive and 4 from the HCV-negative group developed a lymphoma. Only 2 patients from the HCV group died from hepatocellular carcinoma. Survival without a skin cancer was similar between both groups. Patient and death-censored graft survival rates were significantly lower in the HCV group. CONCLUSION: The incidences and types of malignancies were similar in the HCV-positive and HCV-negative KT patients.
Assuntos
Carcinoma Hepatocelular/complicações , Hepacivirus/isolamento & purificação , Hepatite C/epidemiologia , Transplante de Rim/efeitos adversos , Linfoma/epidemiologia , Neoplasias/epidemiologia , Adulto , Carcinoma Hepatocelular/epidemiologia , Carcinoma Hepatocelular/mortalidade , Carcinoma Hepatocelular/virologia , Estudos de Casos e Controles , Feminino , Sobrevivência de Enxerto , Hepatite C/complicações , Hepatite C/mortalidade , Hepatite C/virologia , Humanos , Incidência , Linfoma/complicações , Linfoma/mortalidade , Linfoma/virologia , Masculino , Pessoa de Meia-Idade , Neoplasias/complicações , Neoplasias/mortalidade , Neoplasias/virologia , TransplantadosRESUMO
Hepatitis E virus (HEV) can lead to chronic infection in solid-organ transplant patients. Ribavirin is efficient for treatment of chronically infected patients. Recently, the1634R mutation in the HEV polymerase has been associated with treatment failure. However, it is unclear if this mutation can be used as a prognostic marker of treatment outcome. We studied the prevalence of the 1634R mutation in the HEV polymerase of patients starting ribavirin therapy, the influence of the 1634R variants on the viral response, the frequency of the 1634R mutation in patients whose treatment failed, and its impact on ribavirin retreatment. We analyzed pretreatment samples from 63 solid-organ transplant patients with chronic hepatitis E using deep sequencing; 42 patients had a sustained virologic response (SVR), and 21 were non-SVR patients. We detected the 1634R variant by deep sequencing in 36.5% (23/63) of the patients (proportions, 1.3 to 100%). The 1634R variant was detected in 31.0% (13/42) of baseline plasma samples from patients with SVR and in 47.6% (10/21) in the other patients (P = 0.2). The presence of this mutation did not influence the initial decrease in viral RNA. Lastly, a second prolonged ribavirin treatment led to SVR in 70% of the patients who initially did not have SVR, despite the presence of the 1634R variant. We conclude that the presence of the 1634R variant at ribavirin initiation does not lead to absolute ribavirin resistance. Although its proportion increased in patients whose treatment failed, the presence of the 1634R variant did not compromise the response to a second ribavirin treatment.
Assuntos
Antivirais/uso terapêutico , Vírus da Hepatite E/efeitos dos fármacos , Vírus da Hepatite E/genética , Hepatite E/tratamento farmacológico , RNA Polimerase Dependente de RNA/genética , Ribavirina/uso terapêutico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Sequência de Bases , Criança , Feminino , Marcadores Genéticos , Hepatite E/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Mutação/genética , RNA Viral/genética , Análise de Sequência de RNA , Resultado do Tratamento , Adulto JovemRESUMO
Little is known about virus adaptation in immunocompromised patients with chronic genotype 3 hepatitis E virus (HEV3) infections. Virus-host recombinant strains have been isolated recently from chronically infected patients. The nature and incidence of such recombinant events occurring during infections of solid-organ transplant (SOT) recipients are essentially unknown. The polyproline region (PPR) of strains isolated from SOT patients was sequenced during the acute-infection phase (n = 59) and during follow-up of patients whose infections became chronic (n = 27). These 27 HEV strains included 3 (11%) that showed recombinant events 12, 34, 48, or 88 months after infection. In one strain, parts of the PPR and the RNA-dependent RNA polymerase were concomitantly inserted. In the second, a fragment of a human tyrosine aminotransferase (TAT) gene was inserted first, followed by a fragment of PPR. A fragment of the human inter-α-trypsin inhibitor (ITI) gene was inserted in the third. All the inserted sequences were rich in aliphatic and basic amino acids. In vitro growth experiments suggest that the ITI insertion promoted more vigorous virus growth. In silico studies showed that the inserted sequences could provide potential acetylation, ubiquitination, and phosphorylation sites. We found that recombinant events had occurred in the HEV PPR in approximately 11% of the strains isolated from chronically infected transplant patients followed up in Toulouse University Hospital. These inserted fragments came from the HEV genome or a human gene and could enhance virus replication. Importance: Hepatitis E virus (HEV) can cause chronic infections in immunocompromised patients, including solid-organ transplant (SOT) recipients. Two strains that had undergone recombination with human ribosomal genes were described recently. The strains with inserted sequences replicated better in vitro. Little is known about the frequency of such recombinant events or how such an insertion enhances replication. We therefore investigated 59 SOT patients infected with HEV and found 3 strains with 4 recombinant events in 27 of these patients whose infection became chronic. The 4 inserted sequences were of different origins (human gene or HEV genome), but all were enriched in aliphatic and basic amino acids and provided potential regulation sites. Our data indicate that recombinant events occur in approximately 11% of strains isolated from chronically infected patients. The structures of the inserted sequences provide new clues as to how the inserted sequences could foster virus replication.
Assuntos
Vírus da Hepatite E/química , Hospedeiro Imunocomprometido , Transplante de Órgãos , Peptídeos/química , Sequência de Aminoácidos , Sequência de Bases , Primers do DNA , Genoma Viral , Vírus da Hepatite E/genética , Vírus da Hepatite E/patogenicidade , Humanos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Homologia de Sequência de AminoácidosRESUMO
Hepatitis E virus (HEV) can chronically infect immunocompromised patients. The polyproline region (PPR) and the macro domain of ORF1 protein may modulate virus production and/or the host immune response. We investigated the association between the genetic heterogeneity of HEV quasispecies in ORF1 and the outcome of infection in solid-organ transplant patients. Both sequence entropy and genetic distances during the hepatitis E acute phase were higher in patients whose infection became chronic than in those who cleared the virus. Hence, great quasispecies heterogeneity in the regions encoding the PPR and the macro domain may facilitate HEV persistence.
Assuntos
Heterogeneidade Genética , Vírus da Hepatite E/imunologia , Hepatite E/imunologia , Hospedeiro Imunocomprometido , Proteínas Virais/imunologia , Adulto , Doença Crônica , Feminino , Hepatite E/virologia , Vírus da Hepatite E/genética , Humanos , Masculino , Pessoa de Meia-Idade , Transplante de Órgãos/efeitos adversos , Peptídeos/genética , Peptídeos/imunologia , Transplante , Proteínas Virais/genéticaRESUMO
We screened plasma samples (minipools of 96 samples, corresponding to 53,234 blood donations) from France that had been processed with solvent-detergent for hepatitis E virus RNA. The detection rate was 1 HEV-positive sample/2,218 blood donations. Most samples (22/24) from viremic donors were negative for IgG and IgM against HEV.
Assuntos
Doadores de Sangue , Vírus da Hepatite E/genética , Hepatite E/epidemiologia , Hepatite E/virologia , Adulto , Idoso , Feminino , França/epidemiologia , Genótipo , Geografia Médica , Hepatite E/transmissão , Vírus da Hepatite E/classificação , Humanos , Masculino , Pessoa de Meia-Idade , Razão de Chances , Prevalência , RNA Viral , Estudos Soroepidemiológicos , Carga ViralRESUMO
OBJECTIVES: To track changes in the V3 env region of HIV-1 quasispecies and determine virus coreceptor use in cell reservoirs of patients on long-term suppressive antiretroviral therapy (ART). PATIENTS AND METHODS: Ten patients whose plasma viraemia had been suppressed for a median of 5.5 years were followed for 5 years. The V3 env regions of viruses in peripheral blood mononuclear cells were analysed by ultra-deep sequencing (UDS). HIV-1 tropism was predicted using the geno2pheno 5.75 algorithm and a phenotypic assay. RESULTS: The UDS and phenotypic assay data were concordant for predicting HIV-1 tropism. CXCR4-using viruses detected by UDS accounted for 14.7%-76.5% of the virus populations in samples from five patients at enrolment. Five patients harboured pure R5 virus populations and no X4 viruses emerged during the 5 years. The selection pressures estimated by the dN/dS ratio were acting on the V3 region to produce diversification of the quasispecies in CXCR4-infected patients and purification of the quasispecies in R5-infected patients on effective ART. CONCLUSIONS: UDS showed that the virus quasispecies in cell reservoirs of patients on long-term suppressive ART continued to evolve. CXCR4-using variants became more diversified. Analysis of the selection pressures on the virus quasispecies could provide a clearer picture of virus persistence in patients on effective ART.
Assuntos
Antirretrovirais/uso terapêutico , Variação Genética , Infecções por HIV/virologia , HIV-1/classificação , HIV-1/fisiologia , Receptores de HIV/análise , Tropismo Viral , Adulto , DNA Viral/genética , Feminino , Genótipo , Técnicas de Genotipagem , Infecções por HIV/tratamento farmacológico , HIV-1/genética , HIV-1/isolamento & purificação , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Leucócitos Mononucleares/virologia , Masculino , Pessoa de Meia-Idade , Fenótipo , Cultura de Vírus , Produtos do Gene env do Vírus da Imunodeficiência Humana/genéticaRESUMO
OBJECTIVES: BK virus is a major cause of chronic renal allograft failure.Transplant ureteral stent use has been reported as a risk factorfor BK virus infection. Recently, the use of a new type of ureteral stent (Magnetic Black Star) was reported in kidney transplant recipients. The aim ofthis preliminary report was to compare BK virus viremia and viruria occurrence depending on the type of double-J stent (standard versus Magnetic Black Star). MATERIALS AND METHODS: We included all kidney transplants performed in our center from January to December 2022. Each case had double-J stent placement. Indwelling stents were either a 6- or 7-Fr standard double-J stent or a 6-Fr Magnetic Black Star double-J stent. The type of double-J stent was chosen according to the surgeon's preference. A standard BK virus screening protocol was followed during the study period, which consisted of routine polymerase chain reaction examination of plasma and urine samples during monthly follow-ups. RESULTS: We assessed 120 patients without missing data: 92 patients received standard double-J stents and 28 patients received Magnetic Black Star stents. Patients were mostly male in the standard group (70.7%) versus the Magnetic Black Star group (42.9%) (P = .01). ABO- and HLA-incompatible transplant rates were similar in both groups. BK viremia occurrence and BK viruria occurrence were similar between groups at 1 month, 3 months, and 6 months. CONCLUSIONS: This preliminary study showed no differences concerning BKvirus infection depending on the type of double-J stents used during kidney transplant.
Assuntos
Vírus BK , Transplante de Rim , Infecções por Polyomavirus , Desenho de Prótese , Stents , Infecções Tumorais por Vírus , Viremia , Humanos , Transplante de Rim/efeitos adversos , Vírus BK/patogenicidade , Vírus BK/imunologia , Masculino , Viremia/diagnóstico , Viremia/virologia , Feminino , Pessoa de Meia-Idade , Infecções por Polyomavirus/virologia , Infecções por Polyomavirus/diagnóstico , Infecções por Polyomavirus/imunologia , Infecções por Polyomavirus/urina , Fatores de Risco , Resultado do Tratamento , Adulto , Infecções Tumorais por Vírus/virologia , Infecções Tumorais por Vírus/diagnóstico , Infecções Tumorais por Vírus/imunologia , Infecções Tumorais por Vírus/urina , Fatores de Tempo , Dados Preliminares , Estudos RetrospectivosRESUMO
BACKGROUND: HIV-1 resistance testing is recommended in clinical management and next-generation sequencing (NGS) methods are now available in many virology laboratories. OBJECTIVES: To evaluate the diagnostic performance of Long-Read Single Molecule Real-time (SMRT) sequencing (Sequel, PacBio) for HIV-1 polymerase genotyping. STUDY DESIGN: 111 prospective clinical samples (83 plasma and 28 leukocyte-enriched blood fraction) were analyzed for routine HIV-1 resistance genotyping using Sanger sequencing, Vela NGS, and SMRT sequencing. We developed a SMRT sequencing protocol and a bio-informatics pipeline to infer antiretroviral resistance on both haplotype and variant calling approaches. RESULTS: The polymerase was successfully sequenced by the three platforms in 98 % of plasma RNA samples for viral loads above 4 log copies/mL. The success rate decreased to 83 % using Sanger or Vela sequencing and to 67 % using SMRT sequencing for viral loads of 3 to 4 log copies/mL. Sensitivities of 50 %, 54 % and 61 % were obtained using SMRT, Vela, and Sanger sequencing, respectively, in cellular DNA from patients with prolonged undetectable plasma HIV-1 RNA. Ninety-eight percent of resistance-associated mutations (RAMs) identified with Sanger sequencing were detected using SMRT sequencing. Furthermore, 91 % of RAMs (> 5 % threshold) identified with Vela NGS were detected using SMRT sequencing. RAM quantification using Vela and SMRT sequencing was well correlated (Spearman correlation ρ = 0.82; P < 0.0001). CONCLUSIONS: SMRT sequencing of the full-length HIV-1 polymerase appeared performant for characterizing HIV-1 genotypic resistance on both RNA and DNA clinical samples. Long-read sequencing is a new tool for mutation haplotyping and resistance analysis.
RESUMO
INTRODUCTION: We evaluated the performance of the automated Altostar HEV RNA platform for detecting HEV RNA. METHODS AND RESULTS: Clinical performance was determined by testing 81 plasma samples and 10 fecal samples manually quantified previously with the Realstar RT-PCR assay using the Magnapure instrument for extraction. The assays were concordant for 79/81 plasma samples (97.5%) and 10/10 (100%) fecal samples. The two plasma samples that tested negative with the Altostar assay had a very low HEV RNA concentration (1.6 and 1.4â¯log10 IU/ml). Quantitative results obtained with the automated platform and the manual workflow were highly correlated (ρ= 0.98, p<0.01). The intra-run and inter-run standard deviation were 0.09 IU/ml and 0.13 IU/ml respectively. The assay was linear from 2 to 6â¯log IU/ml. The limit of detection determined by Probit analysis with the WHO HEV RNA standard was 7.6 [95% CI: 4.4-52.5] IU/ml. CONCLUSIONS: The Altostar platform enables highly accurate testing for the detection of HEV RNA in stool and the quantification of HEV RNA in plasma. This allowed us to shorten turnaround times and to save time for the technical staff.