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1.
Emerg Infect Dis ; 30(1): 141-145, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38147067

RESUMO

In a 2-year study in Leuven, Belgium, we investigated the use of wastewater sampling to assess community spread of respiratory viruses. Comparison with the number of positive clinical samples demonstrated that wastewater data reflected circulation levels of typical seasonal respiratory viruses, such as influenza, respiratory syncytial virus, and enterovirus D68.


Assuntos
Enterovirus Humano D , Influenza Humana , Vírus Sincicial Respiratório Humano , Humanos , Bélgica/epidemiologia , Águas Residuárias , Vírus Sincicial Respiratório Humano/genética
2.
Virol J ; 21(1): 40, 2024 02 10.
Artigo em Inglês | MEDLINE | ID: mdl-38341597

RESUMO

Since the onset of the coronavirus disease (COVID-19) pandemic in Belgium, UZ/KU Leuven has played a crucial role as the National Reference Centre (NRC) for respiratory pathogens, to be the first Belgian laboratory to develop and implement laboratory developed diagnostic assays for SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) and later to assess the quality of commercial kits. To meet the growing demand for decentralised testing, both clinical laboratories and government-supported high-throughput platforms were gradually deployed across Belgium. Consequently, the role of the NRC transitioned from a specialised testing laboratory to strengthening capacity and coordinating quality assurance. Here, we outline the measures taken by the NRC, the national public health institute Sciensano and the executing clinical laboratories to ensure effective quality management of molecular testing throughout the initial two years of the pandemic (March 2020 to March 2022).


Assuntos
COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , COVID-19/diagnóstico , COVID-19/epidemiologia , Bélgica/epidemiologia , Teste para COVID-19 , Pandemias , Técnicas de Laboratório Clínico , Técnicas de Diagnóstico Molecular
3.
J Med Virol ; 95(10): e29127, 2023 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-37772540

RESUMO

During the Belgian winter and spring season 2022-2023, we investigated the potential of used paper tissue (UPT) as a noninvasive sampling method for the diagnosis of acute respiratory infections. Screening for respiratory pathogens was done using an in-house developed respiratory panel for simultaneous detection of 22 respiratory viruses and seven nonviral pathogens. The method allowed the identification and typing of respiratory pathogens in symptomatic individuals, as well as in collective samples taken at a community level. Pathogens that were identified in nasal swabs could also be detected in concurrent UPT from the same patient. In all cases that tested positive on an antigen-detection rapid diagnostic test, the corresponding virus could be detected in UPT. The collection of UPT could be useful in epidemiological surveillance of severe acute respiratory syndrome coronavirus 2 and other coronaviruses, as well as other respiratory pathogens such as influenzavirus, respiratory syncytial virus, entero/rhinoviruses including EV-D68, parainfluenzaviruses, and Streptococcus pneumoniae. Multiple respiratory pathogens could be detected in UPTs of collectivities, confirming its applicability for community testing. This is especially interesting for screening in nursing homes, centers for the disabled, schools or other settings were taking nasal or nasopharyngeal samples is cumbersome.

4.
J Med Virol ; 95(2): e28587, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-36799251

RESUMO

Wastewater surveillance plays an important role in the management of the coronavirus disease 2019 (COVID-19) pandemic all over the world. Using different wastewater collection points in Leuven, we wanted to investigate the use of wastewater surveillance as an early warning system for an uprise of infections and as a tool to follow the circulation of specific variants of concern (VOCs) in particular geographic areas. Wastewater samples were collected from local neighborhood sewers and from a large regional wastewater treatment plant (WWTP) in the area of Leuven, Belgium. After virus concentration, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA was quantified by real-time quantitative polymerase chain reaction (RT-qPCR) and normalized with the human fecal indicator pepper mild mottle virus (PMMoV). A combination of multiplex RT-qPCR assays was used to detect signature mutations of circulating VOCs. Fecal virus shedding of SARS-CoV-2 variants was measured in feces samples of hospitalized patients. In two residential sampling sites, a rise in wastewater SARS-CoV-2 concentration preceded peaks in positive cases. In the WWTP, viral load peaks were seen concomitant with the consecutive waves of positive cases caused by the original Wuhan SARS-CoV-2 strain and subsequent VOCs. During the Omicron BA.1 wave, the wastewater viral load increased to a lesser degree, even after normalization of SARS-CoV-2 concentration using PMMoV. This might be attributable to a lower level of fecal excretion of this variant. Circulation of SARS-CoV-2 VOCs Alpha, Delta, Omicron BA1/BA.2, and BA.4/BA.5 could be detected based on the presence of specific key mutations. The shift in variants was noticeable in the wastewater, with key mutations of two different variants being present simultaneously during the transition period. Wastewater-based surveillance is a sensitive tool to monitor SARS-CoV-2 circulation levels and VOCs in larger regions. In times of reduced test capacity, this can prove to be highly valuable. Differences in excretion levels of various SARS-CoV-2 variants should however be taken into account when using wastewater surveillance to monitor SARS-CoV-2 circulation levels in the population.


Assuntos
COVID-19 , SARS-CoV-2 , Humanos , Bélgica , Águas Residuárias , Vigilância Epidemiológica Baseada em Águas Residuárias , RNA Viral
5.
Euro Surveill ; 28(9)2023 03.
Artigo em Inglês | MEDLINE | ID: mdl-36862099

RESUMO

BackgroundLateral flow antigen-detection rapid diagnostic tests (Ag-RDTs) for viral infections constitute a fast, cheap and reliable alternative to nucleic acid amplification tests (NAATs). Whereas leftover material from NAATs can be employed for genomic analysis of positive samples, there is a paucity of information on whether viral genetic characterisation can be achieved from archived Ag-RDTs.AimTo evaluate the possibility of retrieving leftover material of several viruses from a range of Ag-RDTs, for molecular genetic analysis.MethodsArchived Ag-RDTs which had been stored for up to 3 months at room temperature were used to extract viral nucleic acids for subsequent RT-qPCR, Sanger sequencing and Nanopore whole genome sequencing. The effects of brands of Ag-RDT and of various ways to prepare Ag-RDT material were evaluated.ResultsSARS-CoV-2 nucleic acids were successfully extracted and sequenced from nine different brands of Ag-RDTs for SARS-CoV-2, and for five of these, after storage for 3 months at room temperature. The approach also worked for Ag-RDTs for influenza virus (n = 3 brands), as well as for rotavirus and adenovirus 40/41 (n = 1 brand). The buffer of the Ag-RDT had an important influence on viral RNA yield from the test strip and the efficiency of subsequent sequencing.ConclusionOur finding that the test strip in Ag-RDTs is suited to preserve viral genomic material, even for several months at room temperature, and therefore can serve as source material for genetic characterisation could help improve global coverage of genomic surveillance for SARS-CoV-2 as well as for other viruses.


Assuntos
COVID-19 , Ácidos Nucleicos , Humanos , Bélgica , Testes de Diagnóstico Rápido , COVID-19/diagnóstico , SARS-CoV-2/genética , Genômica , Teste para COVID-19
6.
Emerg Infect Dis ; 27(6): 1-9, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-34013862

RESUMO

Human respiratory syncytial virus (HRSV) is the leading viral cause of serious pediatric respiratory disease, and lifelong reinfections are common. Its 2 major subgroups, A and B, exhibit some antigenic variability, enabling HRSV to circulate annually. Globally, research has increased the number of HRSV genomic sequences available. To ensure accurate molecular epidemiology analyses, we propose a uniform nomenclature for HRSV-positive samples and isolates, and HRSV sequences, namely: HRSV/subgroup identifier/geographic identifier/unique sequence identifier/year of sampling. We also propose a template for submitting associated metadata. Universal nomenclature would help researchers retrieve and analyze sequence data to better understand the evolution of this virus.


Assuntos
Infecções por Vírus Respiratório Sincicial , Vírus Sincicial Respiratório Humano , Criança , Variação Genética , Genótipo , Humanos , Epidemiologia Molecular , Filogenia , Vírus Sincicial Respiratório Humano/genética
7.
J Virol Methods ; 304: 114512, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35257682

RESUMO

SARS-CoV-2 has kept the world in suspense for almost 2 years now. The virus spread rapidly worldwide and several variants of concern have emerged: Alpha, Beta, Gamma, Delta and recently Omicron. A rapid method to detect key mutations is needed because these variants may jeopardize the effectiveness of immune protection following vaccination or past infection. This article describes an easy, cheap and fast method for the detection of mutations in the spike protein that are indicative for specific variants. This method can easily distinguish Omicron from other variants.


Assuntos
COVID-19 , SARS-CoV-2 , COVID-19/diagnóstico , Humanos , Mutação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , SARS-CoV-2/genética
8.
PLoS One ; 17(6): e0268532, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35666728

RESUMO

BACKGROUND: We aimed to provide regional data on clinical symptoms, medical resource utilization (MRU), and risk factors for increased MRU in hospitalized respiratory syncytial virus (RSV)-infected Belgian pediatric population. METHODS: This prospective, multicenter study enrolled RSV (+) hospitalized children (aged ≤5y) during the 2013-2015 RSV seasons. RSV was diagnosed within 24h of hospitalization. Disease severity of RSV (+) patients was assessed until discharge or up to maximum six days using a Physical Examination Score (PES) and a derived score based on ability to feed, dyspnea and respiratory effort (PES3). MRU (concomitant medications, length of hospitalization [LOH], and oxygen supplementation) was evaluated. Kaplan-Meier survival analysis was performed to compare MRU by age and presence of risk factors for severe disease. Association between baseline covariates and MRU was analyzed using Cox regression models. RESULTS: In total, 75 children were included, Median (range) age was 4 (0-41) months, risk factors were present in 18.7%, and early hospitalization (≤3 days of symptom onset) was observed in 57.3% of patients. Cough (100%), feeding problems (82.2%), nasal discharge (87.8%), and rales and rhonchi (82.2%) were frequently observed. Median (range) LOH and oxygen supplementation was 5 (2-7) and 3 (1-7) days. Oxygen supplementation, bronchodilators, and antibiotics were administered to 58.7%, 64.0%, and 41.3% of the patients, respectively. Age <3 months and baseline total PES3 score were associated with probability and the duration of receiving oxygen supplementation. LOH was not associated with any covariate. CONCLUSION: RSV is associated with high disease burden and MRU in hospitalized children. Oxygen supplementation but not length of hospitalization was associated with very young age and the PES3 score. These results warrant further assessment of the PES3 score as a predictor for the probability of receiving and length of oxygen supplementation in RSV hospitalized children. REGISTRATION: NCT02133092.


Assuntos
Infecções por Vírus Respiratório Sincicial , Vírus Sincicial Respiratório Humano , Bélgica/epidemiologia , Criança , Criança Hospitalizada , Hospitalização , Hospitais , Humanos , Lactente , Estudos Prospectivos , Infecções por Vírus Respiratório Sincicial/diagnóstico , Infecções por Vírus Respiratório Sincicial/epidemiologia , Infecções por Vírus Respiratório Sincicial/terapia , Fatores de Risco , Índice de Gravidade de Doença
9.
J Virol ; 83(17): 8759-70, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19553340

RESUMO

Papillomaviruses (PVs) are a large family of small DNA viruses infecting mammals, reptiles, and birds. PV infection induces cell proliferation that may lead to the formation of orogenital or skin tumors. PV-induced cell proliferation has been related mainly to the expression of two small oncoproteins, E6 and E7. In mammalian PVs, E6 contains two 70-residue zinc-binding repeats, whereas E7 consists of a natively unfolded N-terminal region followed by a zinc-binding domain which folds as an obligate homodimer. Here, we show that both the novel francolin bird PV Francolinus leucoscepus PV type 1 (FlPV-1) and the chaffinch bird PV Fringilla coelebs PV contain unusual E6 and E7 proteins. The avian E7 proteins contain an extended unfolded N terminus and a zinc-binding domain of reduced size, whereas the avian E6 proteins consist of a single zinc-binding domain. A comparable single-domain E6 protein may have existed in a common ancestor of mammalian and avian PVs. Mammalian E6 C-terminal domains are phylogenetically related to those of single-domain avian E6, whereas mammalian E6 N-terminal domains seem to have emerged by duplication and subsequently diverged from the original ancestral domain. In avian and mammalian cells, both FlPV-1 E6 and FlPV-1 E7 were evenly expressed in the cytoplasm and the nucleus. Finally, samples of full-length FlPV-1 E6 and the FlPV-1 E7 C-terminal zinc-binding domain were prepared for biophysical analysis. Both constructs were highly soluble and well folded, according to nuclear magnetic resonance spectroscopy measurements.


Assuntos
Doenças das Aves/virologia , Proteínas Oncogênicas Virais/química , Proteínas Oncogênicas Virais/genética , Papillomaviridae/genética , Infecções por Papillomavirus/veterinária , Sequência de Aminoácidos , Animais , Aves , Núcleo Celular/química , Análise por Conglomerados , Citoplasma/química , DNA Viral/química , DNA Viral/genética , Evolução Molecular , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Dados de Sequência Molecular , Proteínas Oncogênicas Virais/análise , Papillomaviridae/isolamento & purificação , Filogenia , Dobramento de Proteína , Estrutura Terciária de Proteína , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência
10.
Virus Evol ; 6(2): veaa052, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33072402

RESUMO

Since the first human respiratory syncytial virus (HRSV) genotype classification in 1998, inconsistent conclusions have been drawn regarding the criteria that define HRSV genotypes and their nomenclature, challenging data comparisons between research groups. In this study, we aim to unify the field of HRSV genotype classification by reviewing the different methods that have been used in the past to define HRSV genotypes and by proposing a new classification procedure, based on well-established phylogenetic methods. All available complete HRSV genomes (>12,000 bp) were downloaded from GenBank and divided into the two subgroups: HRSV-A and HRSV-B. From whole-genome alignments, the regions that correspond to the open reading frame of the glycoprotein G and the second hypervariable region (HVR2) of the ectodomain were extracted. In the resulting partial alignments, the phylogenetic signal within each fragment was assessed. Maximum likelihood phylogenetic trees were reconstructed using the complete genome alignments. Patristic distances were calculated between all pairs of tips in the phylogenetic tree and summarized as a density plot in order to determine a cutoff value at the lowest point following the major distance peak. Our data show that neither the HVR2 fragment nor the G gene contains sufficient phylogenetic signal to perform reliable phylogenetic reconstruction. Therefore, whole-genome alignments were used to determine HRSV genotypes. We define a genotype using the following criteria: a bootstrap support of ≥ 70 per cent for the respective clade and a maximum patristic distance between all members of the clade of ≤0.018 substitutions per site for HRSV-A or ≤0.026 substitutions per site for HRSV-B. By applying this definition, we distinguish twenty-three genotypes within subtype HRSV-A and six genotypes within subtype HRSV-B. Applying the genotype criteria on subsampled data sets confirmed the robustness of the method.

11.
Vet Microbiol ; 133(1-2): 43-53, 2009 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-18676105

RESUMO

Papillomaviruses (PVs) have been shown as being the etiologic agents of various benign and malignant tumours in many vertebrate species. In dolphins and porpoises, a high prevalence of orogenital tumours has recently been documented with at least four distinct novel species-specific PV types detected in such lesions. Therefore, we generated the immunological reagents to establish a serological screening test to determine the prevalence of PV infection in Atlantic bottlenose dolphins [(Tursiops truncatus (Tt)]. Using the baculovirus expression system, virus-like particles (VLPs) derived from the L1 proteins of two TtPV types, TtPV1 and TtPV2, were generated. Polyclonal antibodies against TtPV VLPs were produced in rabbits and their specificity for the VLPs was confirmed. Electron microscopy and enzyme-linked immunosorbent assay (ELISA) studies revealed that the generated VLPs self-assembled into particles presenting conformational immunodominant epitopes. As such, these particles are potential antigen candidates for a TtPV vaccine. Subsequently, the VLPs served as antigens in initial ELISA tests using sera from six bottlenose dolphins to investigate PV antibody presence. Three of these sera were derived from dolphins with genital tumour history and showed positive PV ELISA reactivity, while the remaining sera from lesion-free dolphins were PV antibody-negative. The results suggest that the developed screening test may serve as a potential tool for determining PV prevalence and thus for observing transmission rates in dolphin populations as the significance of PV infection in cetaceans starts to unfold.


Assuntos
Anticorpos Antivirais/sangue , Antígenos Virais/imunologia , Golfinho Nariz-de-Garrafa/virologia , Papillomaviridae/imunologia , Infecções por Papillomavirus/veterinária , Animais , Especificidade de Anticorpos , Golfinho Nariz-de-Garrafa/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Ensaio de Imunoadsorção Enzimática/veterinária , Programas de Rastreamento/métodos , Programas de Rastreamento/veterinária , Fases de Leitura Aberta , Infecções por Papillomavirus/diagnóstico , Infecções por Papillomavirus/epidemiologia , Infecções por Papillomavirus/transmissão , Prevalência , Coelhos , Especificidade da Espécie , Vacinas Virais
12.
Diagn Microbiol Infect Dis ; 94(1): 33-37, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30638655

RESUMO

The objective of this study was to compare the performance of the Idylla™ Respiratory (IFV-RSV) panel to the GeneXpert Xpert® Flu/RSV assay and establish the performance of a midturbinate swab compared to nasopharyngeal sampling. Considering GeneXpert® assay as imperfect reference standard, a positive percentage agreement between both assays of 98-100% for influenza A and 96-99% for influenza B could be calculated when 354 nasopharyngeal and 325 midturbinate swabs were retrospectively analyzed. Comparing midturbinate samples to nasopharyngeal specimens of 321 subjects, positive percentage agreement varied from 42% to 94% depending on both target virus and assay used. Negative percentage agreements ranged from 98% to 100% for both methods and sample type comparison. The Idylla™ assay showed excellent performance compared to the GeneXpert® assay for the detection of influenza virus. The study also showed a slightly better performance for nasopharyngeal sampling compared to the use of a midturbinate swab.


Assuntos
Influenza Humana/diagnóstico , Técnicas de Diagnóstico Molecular/métodos , Cavidade Nasal/virologia , Nasofaringe/virologia , Orthomyxoviridae/isolamento & purificação , Infecções por Vírus Respiratório Sincicial/diagnóstico , Vírus Sincicial Respiratório Humano/isolamento & purificação , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Manejo de Espécimes/métodos , Adulto Jovem
13.
J Virol ; 81(24): 13280-90, 2007 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17898069

RESUMO

Conservation efforts to prevent the extinction of the endangered western barred bandicoot (Perameles bougainville) are currently hindered by a progressively debilitating cutaneous and mucocutaneous papillomatosis and carcinomatosis syndrome observed in captive and wild populations. In this study, we detected a novel virus, designated the bandicoot papillomatosis carcinomatosis virus type 1 (BPCV1), in lesional tissue from affected western barred bandicoots using multiply primed rolling-circle amplification and PCR with the cutaneotropic papillomavirus primer pairs FAP59/FAP64 and AR-L1F8/AR-L1R9. Sequencing of the BPCV1 genome revealed a novel prototype virus exhibiting genomic properties of both the Papillomaviridae and the Polyomaviridae. Papillomaviral properties included a large genome size ( approximately 7.3 kb) and the presence of open reading frames (ORFs) encoding canonical L1 and L2 structural proteins. The genomic organization in which structural and nonstructural proteins were encoded on different strands of the double-stranded genome and the presence of ORFs encoding the nonstructural proteins large T and small t antigens were, on the other hand, typical polyomaviral features. BPCV1 may represent the first member of a novel virus family, descended from a common ancestor of the papillomaviruses and polyomaviruses recognized today. Alternatively, it may represent the product of ancient recombination between members of these two virus families. The discovery of this virus could have implications for the current taxonomic classification of Papillomaviridae and Polyomaviridae and can provide further insight into the evolution of these ancient virus families.


Assuntos
Carcinoma/veterinária , Infecções por Vírus de DNA/veterinária , Genoma Viral , Marsupiais/virologia , Papiloma/veterinária , Papillomaviridae/classificação , Polyomaviridae/classificação , Animais , Antígenos Virais de Tumores/genética , Carcinoma/patologia , Carcinoma/virologia , Conservação dos Recursos Naturais , Infecções por Vírus de DNA/patologia , Infecções por Vírus de DNA/virologia , Vírus de DNA/classificação , Vírus de DNA/genética , Vírus de DNA/isolamento & purificação , DNA Viral/análise , DNA Viral/isolamento & purificação , Evolução Molecular , Dados de Sequência Molecular , Fases de Leitura Aberta/genética , Papiloma/patologia , Papiloma/virologia , Papillomaviridae/genética , Reação em Cadeia da Polimerase , Polyomaviridae/genética , Recombinação Genética , Análise de Sequência de DNA , Proteínas Virais/genética
14.
Vet Microbiol ; 129(1-2): 108-16, 2008 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-18215475

RESUMO

Papillomatosis has been documented in several carnivores, and papillomavirus (PV) types have been characterized from lesions in a number of carnivore species: the canine oral PV (COPV), the Felis domesticus PV type 1 (FdPV-1) isolated from a Persian cat, the Procyon lotor PV type 1 (PlPV-1) isolated from a raccoon, the canine PV type 2 (CPV-2) from a dog's foot pad lesion and the canine PV type 3 (CPV-3) associated with a canine epidermodysplasia verruciformis - like disease. A tissue sample was taken from a papillomatous lesion on the oral mucosa of a polar bear (Ursus maritimus). Extracted DNA was used as a template for multiply primed rolling-circle amplification (RCA), and restriction enzyme analysis of the RCA product indicated the presence of papillomaviral DNA. The genome of this PV was cloned and the complete genomic sequence was determined. The Ursus maritimus PV type 1 (UmPV-1) genome counts 7582 basepairs and is smaller than that of other papillomaviruses from carnivore species. UmPV-1 contains the typical noncoding region NCR1, but unlike the carnivore PVs of the Lambda genus, UmPV-1 does not possess a second noncoding region NCR2. Phylogenetic analysis based on a nucleotide sequence alignment of the L1 ORF of UmPV-1 and 51 other PV types indicates that UmPV-1 does not cluster with any of the other carnivore PVs, but branches off near the root of the common branch of the genus Alphapapillomavirus.


Assuntos
Mucosa Bucal/virologia , Papillomaviridae/genética , Infecções por Papillomavirus/veterinária , Ursidae/virologia , Animais , Animais de Zoológico , Masculino , Infecções por Papillomavirus/virologia , Filogenia , Língua/patologia
15.
Vector Borne Zoonotic Dis ; 18(8): 440-444, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29893639

RESUMO

Cases of wart-like lesions in humans and dromedary camels occurred in eastern Sudan in 2015 were described. Involvement of papillomavirus (PV) in causing these cases was affirmed by PCR and immunoperoxidase test. Mostly, the lesions were observed on the skin of the chest and forearms in addition to lips and mandible. Sequence analysis revealed Camelus dromedarius PV types 1 and 2 genotypes as the causative genotypes. We also observed cases of wart-like lesions on hands and legs of two herders attending the infected camel herd. Partial genome sequencing revealed human PV type 2 in one of the two human samples providing no indications for interspecies transmission of camel PVs, yet provides, for the first time evidence of active circulation of this virus in eastern Sudan.


Assuntos
Camelus/virologia , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/veterinária , Verrugas/veterinária , Adulto , Animais , Feminino , Humanos , Masculino , Infecções por Papillomavirus/epidemiologia , Infecções por Papillomavirus/virologia , Sudão/epidemiologia , Verrugas/epidemiologia , Verrugas/virologia , Zoonoses
16.
J Clin Virol ; 106: 34-40, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30031351

RESUMO

BACKGROUND: Virus-specific molecular assays such as real-time polymerase chain reaction (RT-PCR) are regularly used as the gold standard to diagnose viral respiratory tract infections, but simultaneous detection of multiple different pathogens is often challenging. A multiplex digital method of RNA quantification, nCounter (NanoString Technologies), can overcome this disadvantage and identify, in a single reaction, the presence of different respiratory viruses. OBJECTIVES: To evaluate the accuracy of nCounter to identify and quantify RSV-A and RSV-B in nasopharyngeal aspirates (NPA) of children (6-23-months-old) with acute respiratory infection. STUDY DESIGN: NPA was collected at enrolment in a prospective cross-sectional study conducted in Salvador, Brazil. A quantitative RT-PCR with a subgroup-specific primer and probeset for RSV-A and RSV-B was performed in parallel with a customized nCounter probeset containing viral targets in NPA. RESULTS: Of 559 NPA tested, RSV was detected by RT-PCR in 139 (24.9%), by nCounter in 122 (21.8%) and by any method in 158 (28.3%) cases. Compared to the gold standard of qRT-PCR, sensitivity of nCounter was 74.3% (95%CI:63.3%-82.9% RSV-A) and 77.6% (95%CI:66.3%-85.9% RSV-B); specificity was 98.4% (95%CI:96.8%-99.2% RSV-A) and 97.8% (95%CI:96.0%-98.8% RSV-B); positive predictive value was 87.3% (95%CI:76.9%-93.4% RSV-A) and 82.5% (95%CI:71.4%-90.0% RSV-B) and negative predictive value was 96.1% (95%CI:94.1%-97.5% RSV-A), and 96.9% (95%CI:95.1%-98.2% RSV-B). Accuracy was 95.2% (95%CI:93.1%-96.7%) for RSV-A and 95.3% (95%CI:93.3%-96.9%) for RSV-B, while both methods significantly correlated for RSV-A (r = 0.44, p = 8 × 10-5) and RSV-B (r = 0.73, p = 3 × 10-12) quantification. CONCLUSIONS: nCounter is highly accurate in detecting RSV-A/B in NPA. Robustness and high-throughput multiplexing indicate its use in large-scale epidemiological studies.


Assuntos
Técnicas de Diagnóstico Molecular/métodos , Reação em Cadeia da Polimerase Multiplex/métodos , RNA Viral/análise , Infecções Respiratórias/diagnóstico , Doença Aguda/epidemiologia , Brasil/epidemiologia , Estudos Transversais , Feminino , Humanos , Lactente , Masculino , Nasofaringe/virologia , Estudos Prospectivos , Reprodutibilidade dos Testes , Vírus Sincicial Respiratório Humano/genética , Vírus Sincicial Respiratório Humano/isolamento & purificação , Infecções Respiratórias/epidemiologia , Infecções Respiratórias/virologia , Sensibilidade e Especificidade
17.
Curr Protoc Microbiol ; 44: 1E.12.1-1E.12.18, 2017 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-28166385

RESUMO

Many methods for the discovery of novel viruses are based on amplification of the virus using consensus or degenerate PCR primers. A downside of this approach is that it requires prior knowledge of the viral nucleotide sequence to be applicable. Presented in this unit is a method for the sequence-independent amplification of circular viral genomes that is based on the rolling-circle mechanism used by certain viruses in their natural replication cycle. The amplification of the virus of interest is coupled to the isolation of the viral genome by gel extraction following a restriction digestion. Once isolated, the sequence of the viral genome can be determined by nanopore sequencing, a rapid and inexpensive next-generation sequencing technology that generates long reads in real time. The method described in this unit was originally developed for the discovery of papillomaviruses, but can be used for the identification of all types of circular DNA viruses. © 2017 by John Wiley & Sons, Inc.


Assuntos
Vírus de DNA/classificação , Vírus de DNA/isolamento & purificação , DNA Circular/genética , DNA Circular/isolamento & purificação , Técnicas de Amplificação de Ácido Nucleico/métodos , Virologia/métodos , Vírus de DNA/genética
18.
Open Vet J ; 7(2): 174-179, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28717601

RESUMO

We investigated two outbreaks of papillomatosis between 2013 and 2015 in Al Ahsa region of eastern Saudi Arabia involving fourteen dromedary camels. The disease affected both young and adult animals and occurred in coincidence with demodectic mange infestation. Diagnosis was made based on gross and histopathological characteristics of the wart lesion and was confirmed by PCR. Rolling circle amplification followed by degenerate primer PCR and sequencing of the amplicons revealed the presence of both Camelus dromedarius papillomavirus types 1 and 2, previously identified in infected dromedaries in Sudan.

19.
J Clin Virol ; 94: 72-78, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28772168

RESUMO

BACKGROUND: Acute Respiratory Infections (ARIs) are a major health problem, especially in young children and the elderly. OBJECTIVES: Insights into the seasonality of respiratory viruses can help us understand when the burden on society is highest and which age groups are most vulnerable. STUDY DESIGN: We monitored six respiratory viruses during five consecutive seasons (2011-2016) in Belgium. Patient specimens (n=22876), tested for one or more of the following respiratory viruses, were included in this analysis: Influenza viruses (IAV & IBV), Human respiratory syncytial virus (hRSV), Human metapneumovirus (hMPV), Adenovirus (ADV) and Human parainfluenza virus (hPIV). Data were analysed for four age categories: <6y, 6-17y, 18-64y and ≥65y. RESULTS: Children <6y had the highest infection rates (39% positive vs. 20% positive adults) and the highest frequency of co-infections. hRSV (28%) and IAV (32%) caused the most common respiratory viral infections and followed, like hMPV, a seasonal pattern with winter peaks. hRSV followed an annual pattern with two peaks: first in young children and ±7 weeks later in elderly. This phenomenon has not been described in literature so far. hPIV and ADV occurred throughout the year with higher rates in winter. CONCLUSIONS: Children <6y are most vulnerable for respiratory viral infections and have a higher risk for co-infections. hRSV and IAV are the most common respiratory infections with peaks during the winter season in Belgium.


Assuntos
Infecções por Vírus Respiratório Sincicial/epidemiologia , Infecções por Vírus Respiratório Sincicial/virologia , Vírus Sincicial Respiratório Humano/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Bélgica/epidemiologia , Criança , Pré-Escolar , Humanos , Lactente , Recém-Nascido , Pessoa de Meia-Idade , Prevalência , Estudos Retrospectivos , Estações do Ano , Adulto Jovem
20.
Vet Microbiol ; 201: 26-31, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28284619

RESUMO

Papillomaviridae form a large family of viruses that are known to infect a variety of vertebrates, including mammals, reptiles, birds and fish. Infections usually give rise to minor skin lesions but can in some cases lead to the development of malignant neoplasia. In this study, we identified a novel species of papillomavirus (PV), isolated from warts of four giraffes (Giraffa camelopardalis). The sequence of the L1 gene was determined and found to be identical for all isolates. Using nanopore sequencing, the full sequence of the PV genome could be determined. The coding region of the genome was found to contain seven open reading frames (ORF), encoding the early proteins E1, E2 and E5-E7 as well as the late proteins L1 and L2. In addition to these ORFs, a region located within the E2 gene is thought, based on sequence similarities to other papillomaviruses, to encode an E4 protein, although no start codon could be identified. Based on the sequence of the L1 gene, this novel PV was found to be most similar to Capreolus capreolus papillomavirus 1 (CcaPV1), with 67.96% nucleotide identity. We therefore suggest that the virus identified here is given the name Giraffa camelopardalis papillomavirus 1 (GcPV1) and is classified as a novel species within the genus Deltapapillomavirus, in line with the current guidelines for the nomenclature and classification of PVs.


Assuntos
Deltapapillomavirus/classificação , Genoma Viral/genética , Girafas/virologia , Infecções por Papillomavirus/veterinária , Animais , Deltapapillomavirus/genética , Deltapapillomavirus/isolamento & purificação , Masculino , Nanoporos , Fases de Leitura Aberta/genética , Infecções por Papillomavirus/patologia , Infecções por Papillomavirus/virologia , Filogenia , Análise de Sequência de DNA/veterinária , Pele/patologia , Pele/virologia
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