The respiratory virome and exacerbations in patients with chronic obstructive pulmonary disease

Abstract

Introduction: Exacerbations are major contributors to morbidity and mortality in patients with chronic obstructive pulmonary disease (COPD), and respiratory bacterial and viral infections are an important trigger for the occurrence of such exacerbations. However, using conventional diagnostic techniques, a causative agent is not always found. Metagenomic next-generation sequencing (mNGS) allows analysis of the complete virome, but has not yet been applied in COPD exacerbations. Objectives: To study the respiratory virome in nasopharyngeal samples during COPD exacerbations using mNGS. Study design: 88 nasopharyngeal swabs from 63 patients from the Bergen COPD Exacerbation Study (2006-2010) were analysed by mNGS and in-house qPCR for respiratory viruses. Both DNA and RNA were sequenced simultaneously using an Illumina library preparation protocol with in-house adaptations. Results: By mNGS, 23/88 samples tested positive. Sensitivity and specificity were both 96% for diagnostic targets (23/24 and 1067/1120, respectively). Viral pathogens only detected by mNGS were herpes simplex virus type 1 and coronavirus OC43. A positive correlation was found between Cq value and mNGS viral species reads (p=0.008). Patients with viral pathogens had lower percentages of bacteriophages (p<0.000). No correlation was found between viral reads (species and genus level) and clinical markers. Conclusions: The mNGS protocol used was highly sensitive and specific for semi-quantitative detection of respiratory viruses. Excellent negative predictive value implicates the power of mNGS to exclude any infectious cause in one test, with consequences for clinical decision making. Reduced abundance of bacteriophages in COPD patients with viral pathogens implicates skewing of the virome, and speculatively the bacterial population, during infection.

An Optimized Metagenomic Approach for Virome Detection of Clinical Pharyngeal Samples With Respiratory Infection

“…Results presented in pharyngeal clinical samples showed that the greatest amounts of viral sequences from mammalian viruses were present in the families Coronaviridae , Anelloviridae , and Paramyxoviridae , Coronaviridae and Anelloviridae comprising the major part of the composition. Both Coronaviridae and Anelloviridae were detected in the same sample in the present research ( Figure 4B ), which was inconsistent with previous studies where Anelloviridae were not detected in the Coronaviridae -positive samples and Coronaviridae were not detected in the Anelloviridae -positive samples ( van Rijn et al, 2019 ). We also observed that the M3 did not detecte Herpesviridae , which were thought to have possibly integrated into the human genome.…”

“…Our results for M3 proved to be robust for viral verification but also to be accurate for viral genome analysis and evolutionary study. We further verified OC43 viruses in three original samples using RT-PCR with pan-coronavirus primers ( Woo et al, 2005 ), indicating that it might have been one of the causes of acute COPD exacerbation ( Wylie, 2017 ; van Rijn et al, 2019 ). Furthermore, the other nine Anelloviridae family viruses were also characterized ( Supplementary Table S2 ).…”

“…Among all 12 subsamples, the majority of the viral reads (average: ∼57%) aligned to a crucial respiratory virus ( Coronaviridae , HCov-OC43) ( Wylie, 2017 ; van Rijn et al, 2019 ). To explore the effects of the different methods on the efficiency of genome recovery, we compared the corresponding HCov-OC43 reference (MG197719) genome site sequencing depth profiles (see Figure 4 ).…”

“…Common human respiratory tract viruses have been summarized for the period from 2009 to 2016, including Rhinoviruses, Paramyxoviruses, Orthomyxoviruses, Coronaviruses, Adenoviruses, Parvoviruses, Herpesviruses, Anelloviruses, Papillomaviruses , and Polyomaviruses ( Wylie, 2017 ). To date, only one study characterized respiratory virome in the areas of COPD in 63 patients with acute COPD exacerbation from Norway, in which an obtained median of 11 million sequence reads per sample contained ∼93% of human reads, 3% of bacterial, 0.1% of viral, and 3% of unknown reads, as identified using NGS without any viral enrichment steps ( van Rijn et al, 2019 ). In this study, we aimed to optimize the pretreatment method to enrich viral particles for NGS sequencing to investigate the virome in pharyngeal clinical samples from hospitalized patients with acute COPD exacerbation.…”

Coronavirus discovery by metagenomic sequencing: a tool for pandemic preparedness

“…Library preparation and sequencing were performed using a previously validated protocol [ 15 , 16 ]. Briefly, 200 μl of patient samples were spiked with equine arteritis virus (EAV) and phocid herpesvirus-1 (PhHV-1) prior to NA extraction using the Magnapure 96 DNA and Viral NA Small volume extraction kit on the MagnaPure 96 system (Roche, Basel, Switzerland) resulting in 100 μL nucleic acid-containing eluate.…”