Metagenomic sequencing with spiked-in internal control to monitor cellularity and diagnosis of pneumonia

Abstract

We read with interest the study by Peng and colleges showing the performance of metagenomic Next-Generation Sequencing (mNGS) in detecting pulmonary pathogens was not superior to conventional microbiological tests (CMT) in a cohort of 101 immunocompromised adults. 1 Peng J.M. Du B. Qin H.Y. Wang Q. Shi Y. Metagenomic next-generation sequencing for the diagnosis of suspected pneumoni in immunocompromised patients. J Infect. 2021; 82 (AprPubMed PMID: 33609588): 22-27 Abstract Full Text Full Text PDF PubMed Scopus (17) Google Scholar Indeed, although mNGS enables untargeted “pan-pathogen” detection that covers a broad array of microorganisms with known genomic sequences, clinical application of this test has encountered challenges. For instance, the diagnostic sensitivity is affected by the quantity of host DNA, which varies considerably from sample to sample. 2 Schlaberg R. Chiu C.Y. Miller S. Procop G.W. Weinstock G. Professional practice CValidation of metagenomic next-generation sequencing tests for universal pathogen detection. Arch Pathol Lab Med. 2017; 141 (JunPubMed PMID: 28169558): 776-786 Crossref PubMed Scopus (216) Google Scholar Host cell depletion techniques have been used to improve the sensitivity of mNGS but may lead to unspecific removal of pathogens. 3 Charalampous T. Kay G.L. Richardson H. Aydin A. Baldan R. Jeanes C. et al. Nanopore metagenomics enables rapid clinical diagnosis of bacterial lower respiratory infection. Nat Biotechnol. 2019; 37 (JulPubMed PMID: 31235920): 783-792 Crossref PubMed Scopus (186) Google Scholar In our independent study, we developed a spike-in internal control to assess the abundance of host and microbial DNA in bronchoalveolar lavage fluid (BALF) and evaluated the analytical and diagnostic performance of mNGS with and without host depletion in a cohort of 205 patients suspected of lower respiratory tract infections (Supplementary Table 1).