Next‐Generation Sequencing for Characterizing Respiratory Tract Virome and Improving Detection of Viral Pathogens in Children With Pneumonia

ABSTRACT Background Pneumonia is typically caused by a variety of pathogenic microorganisms. Traditional research often focuses on the infection of a few microorganisms, whereas metagenomic studies focus on the impact of the bacteriome and mycobiome on respiratory diseases. Reports on the virome characteristics of pediatric pneumonia remain relatively scarce. Methods We employed de novo assembly and combined homology‐ and feature‐based methods to characterize the respiratory virome in whole‐genome DNA sequencing samples from oropharynx (OP) swabs, nasopharynx (NP) swabs, and bronchoalveolar lavage fluids (BALF) of children with pneumonia. Results Significant differences were observed in the alpha and beta diversity indexes, as well as in the composition of the oropharyngeal virome, between pneumonia cases and controls. We identified 1137 viral operational taxonomic units (vOTUs) with significant differences, indicating a preference of pneumonia‐reduced vOTUs for infecting Prevotella , Neisseria , and Veillonella , whereas pneumonia‐enriched vOTUs included polyomavirus, human adenovirus, and phages targeting Staphylococcus , Streptococcus , Granulicatella , and Actinomyces . Comparative analysis revealed higher relative abundances and prevalence rates of pneumonia‐enriched OP vOTUs in NP and BALF samples compared to pneumonia‐reduced vOTUs. Additionally, virome analysis identified six pediatric patients with severe human adenovirus or polyomavirus infections, five of whom might have been undetected by targeted polymerase chain reaction (PCR)‐based testing. Conclusions This study offers insights into pediatric pneumonia respiratory viromes, highlighting frequent transmission of potentially pathogenic viruses and demonstrating virome analysis as a valuable adjunct for pathogen detection.