Direct multiplexed whole genome sequencing of respiratory tract samples reveals full viral genomic information

Zoll, Jan; Rahamat‐Langendoen, Janette; Ahout, Inge M. L.; Jonge, Marien I. de; Jans, Jop; Huijnen, Martijn A.; Ferwerda, Gerben; Warris, Adilia; Melchers, Willem J. G.

doi:10.1016/j.jcv.2015.02.010

Cited by 31 publications

(25 citation statements)

References 23 publications

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“…Another strategy is to perform shotgun sequencing of nucleic acids isolated from the entire, unseparated clinical sample (i.e., metagenomics). This method has been previously applied to total nucleic acids (RNA and DNA) extracted from clinical samples for viral detection [10][11][12][13]. Although metagenomic sequencing of total nucleic acids can detect viruses, its feasibility is highly dependent on the level of virus present in the sample and the amount of host cell nucleic acid present, which can in turn demand high levels of sequence depth for virus detection.…”

Section: Introductionmentioning

confidence: 99%

Dual RNA-seq reveals viral infections in asthmatic children without respiratory illness which are associated with changes in the airway transcriptome

et al. 2017

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BackgroundRespiratory illness caused by viral infection is associated with the development and exacerbation of childhood asthma. Little is known about the effects of respiratory viral infections in the absence of illness. Using quantitative PCR (qPCR) for common respiratory viruses and for two genes known to be highly upregulated in viral infections (CCL8/CXCL11), we screened 92 asthmatic and 69 healthy children without illness for respiratory virus infections.ResultsWe found 21 viral qPCR-positive and 2 suspected virus-infected subjects with high expression of CCL8/CXCL11. We applied a dual RNA-seq workflow to these subjects, together with 25 viral qPCR-negative subjects, to compare qPCR with sequencing-based virus detection and to generate the airway transcriptome for analysis. RNA-seq virus detection achieved 86% sensitivity when compared to qPCR-based screening. We detected additional respiratory viruses in the two CCL8/CXCL11-high subjects and in two of the qPCR-negative subjects. Viral read counts varied widely and were used to stratify subjects into Virus-High and Virus-Low groups. Examination of the host airway transcriptome found that the Virus-High group was characterized by immune cell airway infiltration, downregulation of cilia genes, and dampening of type 2 inflammation. Even the Virus-Low group was differentiated from the No-Virus group by 100 genes, some involved in eIF2 signaling.ConclusionsRespiratory virus infection without illness is not innocuous but may determine the airway function of these subjects by driving immune cell airway infiltration, cellular remodeling, and alteration of asthmogenic gene expression.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-016-1140-8) contains supplementary material, which is available to authorized users.

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Section: Introductionmentioning

confidence: 99%

Dual RNA-seq reveals viral infections in asthmatic children without respiratory illness which are associated with changes in the airway transcriptome

et al. 2017

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“…A third study demonstrated that HTS could be used to detect a pathogen (rhinovirus C) in a sample from a child with respiratory symptoms in which no virus had been detected by PCR. 26 Taken together, these data show that in some cases HTS could be advantageous compared with PCR assays, but sensitivity of sequencing can be a limitation.…”

Section: High-throughput Sequencing In the Clinic Sequencing Assays Cmentioning

confidence: 84%

The Virome of the Human Respiratory Tract

Wylie

2017

Clinics in Chest Medicine

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The human respiratory tract virome is defined here as the viruses present in the human respiratory tract that can infect human cells. Sensitive, culture-independent molecular assays (polymerase chain reaction and high-throughput sequencing) reveal that in addition to common viruses that cause acute, symptomatic infections the virome also includes viruses that do not cause clinical symptoms, have unknown pathogenic effect, or cause symptoms but are not among the most common viral respiratory tract pathogens. These molecular tools provide means for better defining the virome and studying the effects of viral infections on the dynamics of chronic lung diseases.

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“…Additionally, the predominant genera identified included HRSV, influenza virus A and HRV, whereas influenza A and HRV represented a small proportion of viral reads in the present analysis. Zoll et al [22] reported that HRV, TTV and TTMV were frequently detected in children and that viral populations were less complicated than those found in children from Beijing. The only viral metagenomic research on respiratory samples was reported by Yang et al [17] in China, in which only 0.05% of the sequences were virally derived.…”

Section: Discussionmentioning

confidence: 99%

Metagenomic analysis of viral genetic diversity in respiratory samples from children with severe acute respiratory infection in China

Wang

Zhu

et al. 2016

Clinical Microbiology and Infection

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Severe acute respiratory infection (SARI) in children is thought to be mainly caused by infection with various viruses, some of which have been well characterized; however, analyses of respiratory tract viromes among children with SARI versus those without are limited. In this study, nasopharyngeal swabs from children with and without SARI (135 versus 15) were collected in China between 2008 and 2010 and subjected to multiplex metagenomic analyses using a next-generation sequencing platform. The results show that members of the Paramyxoviridae, Coronaviridae, Parvoviridae, Orthomyxoviridae, Picornaviridae, Anelloviridae and Adenoviridae families represented the most abundant species identified (>50% genome coverage) in the respiratory tracts of children with SARI. The viral population found in the respiratory tracts of children without SARI was less diverse and mainly dominated by the Anelloviridae family with only a small proportion of common epidemic respiratory viruses. Several almost complete viral genomes were assembled, and the genetic diversity was determined among several samples based on next-generation sequencing. This research provides comprehensive mapping of the viromes of children with SARI and indicates high heterogeneity of known viruses present in the childhood respiratory tract, which may benefit the detection and prevention of respiratory disease.

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Direct multiplexed whole genome sequencing of respiratory tract samples reveals full viral genomic information

Cited by 31 publications

References 23 publications

Dual RNA-seq reveals viral infections in asthmatic children without respiratory illness which are associated with changes in the airway transcriptome

Dual RNA-seq reveals viral infections in asthmatic children without respiratory illness which are associated with changes in the airway transcriptome

The Virome of the Human Respiratory Tract

Metagenomic analysis of viral genetic diversity in respiratory samples from children with severe acute respiratory infection in China

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