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1.
Food Environ Virol ; 2019 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-31446609

RESUMO

This study aimed to optimize a method to identify human enteric viruses in sewage and stool samples using random primed next-generation sequencing. We tested three methods, two employed virus enrichment based on the binding properties of the viral capsid using pig-mucin capture or by selecting viral RNA prior to library preparation through a capture using the SureSelect target enrichment. The third method was based on a non-specific biophysical precipitation with polyethylene glycol. Full genomes of a number of common human enteric viruses including norovirus, rotavirus, husavirus, enterovirus and astrovirus were obtained. In stool samples full norovirus genome were detected as well as partial enterovirus genome. A variety of norovirus sequences was detected in sewage samples, with genogroup II being more prevalent. Interestingly, the pig-mucin capture enhanced not only the recovery of norovirus and rotavirus but also recovery of astrovirus, sapovirus and husavirus. Documenting sewage virome using these methods provides information for molecular epidemiology and may be useful in developing strategies to prevent further spread of viruses.

2.
Sci Rep ; 9(1): 10076, 2019 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-31296922

RESUMO

Infants (under 1-year-old) are at most risk of life threatening respiratory syncytial virus (RSV) disease. RSV epidemiological data alone has been insufficient in defining who acquires infection from whom (WAIFW) within households. We investigated RSV genomic variation within and between infected individuals and assessed its potential utility in tracking transmission in households. Over an entire single RSV season in coastal Kenya, nasal swabs were collected from members of 20 households every 3-4 days regardless of symptom status and screened for RSV nucleic acid. Next generation sequencing was used to generate >90% RSV full-length genomes for 51.1% of positive samples (191/374). Single nucleotide polymorphisms (SNPs) observed during household infection outbreaks ranged from 0-21 (median: 3) while SNPs observed during single-host infection episodes ranged from 0-17 (median: 1). Using the viral genomic data alone there was insufficient resolution to fully reconstruct within-household transmission chains. For households with clear index cases, the most likely source of infant infection was via a toddler (aged 1 to <3 years-old) or school-aged (aged 6 to <12 years-old) co-occupant. However, for best resolution of WAIFW within households, we suggest an integrated analysis of RSV genomic and epidemiological data.

3.
Viruses ; 11(5)2019 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-31035503

RESUMO

Advances in DNA sequencing technology are facilitating genomic analyses of unprecedented scope and scale, widening the gap between our abilities to generate and fully exploit biological sequence data. Comparable analytical challenges are encountered in other data-intensive fields involving sequential data, such as signal processing, in which dimensionality reduction (i.e., compression) methods are routinely used to lessen the computational burden of analyses. In this work, we explored the application of dimensionality reduction methods to numerically represent high-throughput sequence data for three important biological applications of virus sequence data: reference-based mapping, short sequence classification and de novo assembly. Leveraging highly compressed sequence transformations to accelerate sequence comparison, our approach yielded comparable accuracy to existing approaches, further demonstrating its suitability for sequences originating from diverse virus populations. We assessed the application of our methodology using both synthetic and real viral pathogen sequences. Our results show that the use of highly compressed sequence approximations can provide accurate results, with analytical performance retained and even enhanced through appropriate dimensionality reduction of sequence data.

4.
Euro Surveill ; 24(4)2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30696531

RESUMO

In November 2018, yellow fever was diagnosed in a Dutch traveller returning from a bicycle tour in the Gambia-Senegal region. A complete genome sequence of yellow fever virus (YFV) from the case was generated and clustered phylogenetically with YFV from the Gambia and Senegal, ruling out importation into the Netherlands from recent outbreaks in Brazil or Angola. We emphasise the need for increased public awareness of YFV vaccination before travelling to endemic countries.

5.
Wellcome Open Res ; 3: 21, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30483597

RESUMO

Background: High-throughput whole genome sequencing facilitates investigation of minority virus sub-populations from virus positive samples. Minority variants are useful in understanding within and between host diversity, population dynamics and can potentially assist in elucidating person-person transmission pathways. Several minority variant callers have been developed to describe low frequency sub-populations from whole genome sequence data. These callers differ based on bioinformatics and statistical methods used to discriminate sequencing errors from low-frequency variants. Methods: We evaluated the diagnostic performance and concordance between published minority variant callers used in identifying minority variants from whole-genome sequence data from virus samples. We used the ART-Illumina read simulation tool to generate three artificial short-read datasets of varying coverage and error profiles from an RSV reference genome. The datasets were spiked with nucleotide variants at predetermined positions and frequencies. Variants were called using FreeBayes, LoFreq, Vardict, and VarScan2. The variant callers' agreement in identifying known variants was quantified using two measures; concordance accuracy and the inter-caller concordance. Results: The variant callers reported differences in identifying minority variants from the datasets. Concordance accuracy and inter-caller concordance were positively correlated with sample coverage. FreeBayes identified the majority of variants although it was characterised by variable sensitivity and precision in addition to a high false positive rate relative to the other minority variant callers and which varied with sample coverage. LoFreq was the most conservative caller. Conclusions: We conducted a performance and concordance evaluation of four minority variant calling tools used to identify and quantify low frequency variants. Inconsistency in the quality of sequenced samples impacts on sensitivity and accuracy of minority variant callers. Our study suggests that combining at least three tools when identifying minority variants is useful in filtering errors when calling low frequency variants.

6.
Arch Virol ; 2018 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-30460488

RESUMO

Lemurs are highly endangered mammals inhabiting the forests of Madagascar. In this study, we performed virus discovery on serum samples collected from 84 wild lemurs and identified viral sequence fragments from 4 novel viruses within the family Flaviviridae, including members of the genera Hepacivirus and Pegivirus. The sifaka hepacivirus (SifHV, two genotypes) and pegivirus (SifPgV, two genotypes) were discovered in the diademed sifaka (Propithecus diadema), while other pegiviral fragments were detected in samples from the indri (Indri indri, IndPgV) and the weasel sportive lemur (Lepilemur mustelinus, LepPgV). Although data are preliminary, each viral species appeared host species-specific and frequent infection was detected (18 of 84 individuals were positive for at least one virus). The complete coding sequence and partial 5' and 3' untranslated regions (UTRs) were obtained for SifHV and its genomic organization was consistent with that of other hepaciviruses, with one unique polyprotein and highly structured UTRs. Phylogenetic analyses showed the SifHV belonged to a clade that includes several viral species identified in rodents from Asia and North America, while SifPgV and IndPgV were more closely related to pegiviral species A and C, that include viruses found in humans as well as New- and Old-World monkeys. Our results support the current proposed model of virus-host co-divergence with frequent occurrence of cross-species transmission for these genera and highlight how the discovery of more members of the Flaviviridae can help clarify the ecology and evolutionary history of these viruses. Furthermore, this knowledge is important for conservation and captive management of lemurs.

7.
J Infect Dis ; 217(11): 1728-1739, 2018 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-29741740

RESUMO

Background: Human coronavirus NL63 (HCoV-NL63) is a globally endemic pathogen causing mild and severe respiratory tract infections with reinfections occurring repeatedly throughout a lifetime. Methods: Nasal samples were collected in coastal Kenya through community-based and hospital-based surveillance. HCoV-NL63 was detected with multiplex real-time reverse transcription PCR, and positive samples were targeted for nucleotide sequencing of the spike (S) protein. Additionally, paired samples from 25 individuals with evidence of repeat HCoV-NL63 infection were selected for whole-genome virus sequencing. Results: HCoV-NL63 was detected in 1.3% (75/5573) of child pneumonia admissions. Two HCoV-NL63 genotypes circulated in Kilifi between 2008 and 2014. Full genome sequences formed a monophyletic clade closely related to contemporary HCoV-NL63 from other global locations. An unexpected pattern of repeat infections was observed with some individuals showing higher viral titers during their second infection. Similar patterns for 2 other endemic coronaviruses, HCoV-229E and HCoV-OC43, were observed. Repeat infections by HCoV-NL63 were not accompanied by detectable genotype switching. Conclusions: In this coastal Kenya setting, HCoV-NL63 exhibited low prevalence in hospital pediatric pneumonia admissions. Clade persistence with low genetic diversity suggest limited immune selection, and absence of detectable clade switching in reinfections indicates initial exposure was insufficient to elicit a protective immune response.

8.
Genome Announc ; 6(13)2018 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-29599155

RESUMO

Genetic characterization of wild-type measles virus (MV) strains is a critical component of measles surveillance and molecular epidemiology. We have obtained complete genome sequences of six MV strains belonging to different genotypes, using random-primed next generation sequencing.

9.
AIDS ; 32(6): 709-714, 2018 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-29369160

RESUMO

OBJECTIVE: HIV-1 frequently adapts in response to immune pressure from cytotoxic T-lymphocytes (CTL). Many HIV-2 infected individuals have robust capsid-specific CTL responses associated with viral control. Despite this CTL pressure, adaptive changes in this key immunogenic HIV-2 protein have not previously been described. We sought to compare selective pressure on HIV-1 and HIV-2 capsids and identify HLA-associated viral polymorphisms in HIV-2. DESIGN AND METHODS: Bioinformatic algorithms to identify sites under positive and negative selective pressure and a statistical model of evolution to identify HLA-associated polymorphisms in HIV-2 was applied to sequences from a community cohort in Guinea-Bissau. IFN-γ ELISpots were used to compare T-cell responses to wild-type and variant epitopes. RESULTS: We identified greater purifying selection and less sites under positive selective pressure in HIV-2 compared with HIV-1. Five HIV-2 codons with HLA-associated polymorphisms were detected all within or around known or predicted CTL epitopes. One site was within the HLA-B58 SuperType (ST)-restricted epitope (TSTVEEQIQW), the HIV-2 equivalent of the HIV-1 TW10 epitope. In contrast to HIV-1, where a T→N mutation at position 3 is associated with resulting loss of CTL control, an E→D mutation at position 5 was observed in HIV-2. Robust CTL responses to the variant HIV-2 epitope were seen, suggesting that HIV-2 adaptation may be at the level of T-cell receptor recognition. CONCLUSION: Greater constraints on evolution may exist in HIV-2, resulting in more purifying selection and different immune adaptation pathways in HIV-1 and HIV-2 capsids. This may allow CTL responses to persist in HIV-2.

10.
J Infect Dis ; 216(12): 1513-1524, 2017 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-29029115

RESUMO

Background: The genus Norovirus comprises large genetic diversity, and new GII.4 variants emerge every 2-3 years. It is unknown in which host these new variants originate. Here we study whether prolonged shedders within the immunocompromised population could be a reservoir for newly emerging strains. Methods: Sixty-five fecal samples from 16 immunocompromised patients were retrospectively selected. Isolated viral RNA was enriched by hybridization with a custom norovirus whole-genome RNA bait set and deep sequenced on the Illumina MiSeq platform. Results: Patients shed virus for average 352 days (range, 76-716 days). Phylogenetic analysis showed distinct GII.4 variants in 3 of 13 patients (23%). The viral mutation rates were variable between patients but did not differ between various immune status groups. All within-host GII.4 viral populations showed amino acid changes at blocking epitopes over time, and the majority of VP1 amino acid mutations were located at the capsid surface. Conclusions: This study found viruses in immunocompromised hosts that are genetically distinct from viruses circulating in the general population, and these patients therefore may contain a reservoir for newly emerging strains. Future studies need to determine whether these new strains are of risk to other immunocompromised patients and the general population.

11.
Virus Evol ; 3(2): vex022, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28948041

RESUMO

Porcine stool-associated RNA virus (posavirus), and Human stool-associated RNA virus (husavirus) are viruses in the order Picornavirales recently described in porcine and human fecal samples. The tentative group (Posa and Posa-like viruses: PPLVs) also includes fish stool-associated RNA virus (fisavirus) as well as members detected in insects (Drosophila subobscura and Anopheles sinensis) and parasites (Ascaris suum). As part of an agnostic deep sequencing survey of animal and human viruses in Vietnam, we detected three husaviruses in human fecal samples, two of which share 97-98% amino acid identity to Dutch husavirus strains and one highly divergent husavirus with only 25% amino acid identity to known husaviruses. In addition, the current study found forty-seven complete posavirus genomes from pigs, ten novel rat stool-associated RNA virus genomes (tentatively named rasavirus), and sixteen novel bat stool-associated RNA virus genomes (tentatively named basavirus). The five expected Picornavirales protein domains (helicase, 3C-protease, RNA-dependent RNA polymerase, and two Picornavirus capsid domain) were found to be encoded by all PPLV genomes. In addition, a nucleotide composition analysis revealed that the PPLVs shared compositional properties with arthropod viruses and predicted non-mammalian hosts for all PPLV lineages. The study adds seventy-six genomes to the twenty-nine PPLV genomes currently available and greatly extends our sequence knowledge of this group of viruses within the Picornavirales order.

12.
Virus Evol ; 3(1): vex006, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28458916

RESUMO

Detailed information on the source, spread and evolution of respiratory syncytial virus (RSV) during seasonal community outbreaks remains sparse. Molecular analyses of attachment (G) gene sequences from hospitalized cases suggest that multiple genotypes and variants co-circulate during epidemics and that RSV persistence over successive seasons is characterized by replacement and multiple new introductions of variants. No studies have defined the patterns of introduction, spread and evolution of RSV at the local community and household level. We present a whole genome sequence analysis of 131 RSV group A viruses collected during 6-month household-based RSV infection surveillance in Coastal Kenya, 2010 within an area of 12 km2. RSV infections were identified by regular symptom-independent screening of all household members twice weekly. Phylogenetic analysis revealed that the RSV A viruses in nine households were closely related to genotype GA2 and fell within a single branch of the global phylogeny. Genomic analysis allowed the detection of household-specific variation in seven households. For comparison, using only G gene analysis, household-specific variation was found only in one of the nine households. Nucleotide changes were observed both intra-host (viruses identified from same individual in follow-up sampling) and inter-host (viruses identified from different household members) and these coupled with sampling dates enabled a partial reconstruction of the within household transmission chains. The genomic evolutionary rate for the household dataset was estimated as 2.307 × 10 - 3 (95% highest posterior density: 0.935-4.165× 10 - 3) substitutions/site/year. We conclude that (i) at the household level, most RSV infections arise from the introduction of a single virus variant followed by accumulation of household specific variation and (ii) analysis of complete virus genomes is crucial to better understand viral transmission in the community. A key question arising is whether prevention of RSV introduction or spread within the household by vaccinating key transmitting household members would lead to a reduced onward community-wide transmission.

13.
Nature ; 544(7650): 309-315, 2017 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-28405027

RESUMO

The 2013-2016 West African epidemic caused by the Ebola virus was of unprecedented magnitude, duration and impact. Here we reconstruct the dispersal, proliferation and decline of Ebola virus throughout the region by analysing 1,610 Ebola virus genomes, which represent over 5% of the known cases. We test the association of geography, climate and demography with viral movement among administrative regions, inferring a classic 'gravity' model, with intense dispersal between larger and closer populations. Despite attenuation of international dispersal after border closures, cross-border transmission had already sown the seeds for an international epidemic, rendering these measures ineffective at curbing the epidemic. We address why the epidemic did not spread into neighbouring countries, showing that these countries were susceptible to substantial outbreaks but at lower risk of introductions. Finally, we reveal that this large epidemic was a heterogeneous and spatially dissociated collection of transmission clusters of varying size, duration and connectivity. These insights will help to inform interventions in future epidemics.


Assuntos
Ebolavirus/genética , Ebolavirus/fisiologia , Genoma Viral/genética , Doença pelo Vírus Ebola/transmissão , Doença pelo Vírus Ebola/virologia , Clima , Surtos de Doenças/estatística & dados numéricos , Ebolavirus/isolamento & purificação , Geografia , Doença pelo Vírus Ebola/epidemiologia , Humanos , Internacionalidade , Modelos Lineares , Epidemiologia Molecular , Filogenia , Viagem/legislação & jurisprudência , Viagem/estatística & dados numéricos
14.
Genome Res ; 27(2): 300-309, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27986821

RESUMO

We are rapidly approaching the point where we have sequenced millions of human genomes. There is a pressing need for new data structures to store raw sequencing data and efficient algorithms for population scale analysis. Current reference-based data formats do not fully exploit the redundancy in population sequencing nor take advantage of shared genetic variation. In recent years, the Burrows-Wheeler transform (BWT) and FM-index have been widely employed as a full-text searchable index for read alignment and de novo assembly. We introduce the concept of a population BWT and use it to store and index the sequencing reads of 2705 samples from the 1000 Genomes Project. A key feature is that, as more genomes are added, identical read sequences are increasingly observed, and compression becomes more efficient. We assess the support in the 1000 Genomes read data for every base position of two human reference assembly versions, identifying that 3.2 Mbp with population support was lost in the transition from GRCh37 with 13.7 Mbp added to GRCh38. We show that the vast majority of variant alleles can be uniquely described by overlapping 31-mers and show how rapid and accurate SNP and indel genotyping can be carried out across the genomes in the population BWT. We use the population BWT to carry out nonreference queries to search for the presence of all known viral genomes and discover human T-lymphotropic virus 1 integrations in six samples in a recognized epidemiological distribution.


Assuntos
Genoma Humano/genética , Genômica , Alinhamento de Sequência/métodos , Sequenciamento Completo do Genoma/métodos , Alelos , Compressão de Dados , Genótipo , Humanos , Mutação INDEL/genética , Análise de Sequência de DNA , Software
15.
Euro Surveill ; 21(48)2016 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-27934581

RESUMO

In October 2016, a severe infection with swine influenza A(H1N1) virus of the Eurasian avian lineage occurred in a child with a previous history of eczema in the Netherlands, following contact to pigs. The patient's condition deteriorated rapidly and required life support through extracorporeal membrane oxygenation. After start of oseltamivir treatment and removal of mucus plugs, the patient fully recovered. Monitoring of more than 80 close unprotected contacts revealed no secondary cases.


Assuntos
Oxigenação por Membrana Extracorpórea , Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Influenza Humana/diagnóstico , Infecções Respiratórias/virologia , Síndrome Respiratória Aguda Grave/terapia , Animais , Antivirais/uso terapêutico , Humanos , Influenza Humana/tratamento farmacológico , Influenza Humana/virologia , Unidades de Terapia Intensiva Pediátrica , Países Baixos , Infecções por Orthomyxoviridae/transmissão , Infecções por Orthomyxoviridae/veterinária , Infecções por Orthomyxoviridae/virologia , Oseltamivir/uso terapêutico , Reação em Cadeia da Polimerase em Tempo Real , Infecções Respiratórias/diagnóstico , Infecções Respiratórias/tratamento farmacológico , Síndrome Respiratória Aguda Grave/complicações , Suínos , Doenças dos Suínos/transmissão , Doenças dos Suínos/virologia , Resultado do Tratamento
16.
Genome Announc ; 4(6)2016 Dec 22.
Artigo em Inglês | MEDLINE | ID: mdl-28007850

RESUMO

To document the viral zoonotic risks in Vietnam, fecal samples were systematically collected from a number of mammals in southern Vietnam and subjected to agnostic deep sequencing. We describe here novel Vietnamese bunyavirus sequences detected in bat feces. The complete L and S segments from 14 viruses were determined.

17.
Virus Evol ; 2(1): vew005, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27774298

RESUMO

Several novel clades of astroviruses have recently been identified in human faecal samples. Here, we describe a novel astrovirus-like RNA virus detected in human stools, which we have tentatively named bastrovirus. The genome of this novel virus consists of 6,300 nucleotides organized in three open reading frames. Several sequence divergent strains were detected sharing 67-93 per cent nucleotide identity. Bastrovirus encodes a putative structural protein that is homologous to the capsid protein found in members of the Astroviridae family (45% amino acid identity). The virus also encodes a putative non-structural protein that is genetically distant from astroviruses but shares some homology to the non-structural protein encoded by members of the Hepeviridae family (28% amino acid identity). This novel bastrovirus is present in 8.7 per cent (35/400) of faecal samples collected from 300 HIV-1-positive and 100 HIV-1-negative individuals suggesting common occurrence of the virus. However, whether the source of the virus is infected human cells or other, for example, dietary, remains to be determined.

18.
BMC Med ; 14(1): 132, 2016 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-27604081

RESUMO

Media and World Health Organization (WHO) attention on Zika virus transmission at the 2016 Rio Olympic Games and the 2015 Ebola virus outbreak in West Africa diverted the attention of global public health authorities from other lethal infectious diseases with epidemic potential. Mass gatherings such as the annual Hajj pilgrimage hosted by Kingdom of Saudi Arabia attract huge crowds from all continents, creating high-risk conditions for the rapid global spread of infectious diseases. The highly lethal Middle Eastern respiratory syndrome coronavirus (MERS-CoV) remains in the WHO list of top emerging diseases likely to cause major epidemics. The 2015 MERS-CoV outbreak in South Korea, in which 184 MERS cases including 33 deaths occurred in 2 months, that was imported from the Middle East by a South Korean businessman was a wake-up call for the global community to refocus attention on MERS-CoV and other emerging and re-emerging infectious diseases with epidemic potential. The international donor community and Middle Eastern countries should make available resources for, and make a serious commitment to, taking forward a "One Health" global network for proactive surveillance, rapid detection, and prevention of MERS-CoV and other epidemic infectious diseases threats.


Assuntos
Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/prevenção & controle , Surtos de Doenças/prevenção & controle , Epidemias/prevenção & controle , Saúde Global , Coronavírus da Síndrome Respiratória do Oriente Médio , Atenção , Doenças Transmissíveis/epidemiologia , Doenças Transmissíveis/transmissão , Doenças Transmissíveis Emergentes/epidemiologia , Doenças Transmissíveis Emergentes/prevenção & controle , Doenças Transmissíveis Emergentes/transmissão , Infecções por Coronavirus/transmissão , Humanos , Coronavírus da Síndrome Respiratória do Oriente Médio/isolamento & purificação , Arábia Saudita/epidemiologia , Viagem
19.
Clin Infect Dis ; 63(10): 1353-1356, 2016 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-27585800

RESUMO

We report on an Ebola virus disease (EVD) survivor who showed Ebola virus in seminal fluid 531 days after onset of disease. The persisting virus was sexually transmitted in February 2016, about 470 days after onset of symptoms, and caused a new cluster of EVD in Guinea and Liberia.


Assuntos
Surtos de Doenças , Ebolavirus/genética , Doença pelo Vírus Ebola , Sêmen/virologia , Doenças Virais Sexualmente Transmissíveis , Ebolavirus/isolamento & purificação , Feminino , Guiné , Doença pelo Vírus Ebola/transmissão , Doença pelo Vírus Ebola/virologia , Humanos , Masculino , Reação em Cadeia da Polimerase , RNA Viral/análise , Doenças Virais Sexualmente Transmissíveis/transmissão , Doenças Virais Sexualmente Transmissíveis/virologia , Sobreviventes
20.
Genome Announc ; 4(4)2016 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-27469941

RESUMO

Primer-independent agnostic deep sequencing was used to generate three human rhinovirus (HRV) B genomes and one HRV C genome from samples collected in a household respiratory survey in rural coastal Kenya. The study provides the first rhinovirus genomes from Kenya and will help improve the sensitivity of local molecular diagnostics.

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