Metagenomic Diagnosis for a Culture-Negative Sample From a Patient With Severe Pneumonia by Nanopore and Next-Generation Sequencing

Abstract

Rapid and accurate etiologic diagnosis accelerates targeted antimicrobial therapy. Metagenomic analysis has played a critical role in pathogen identification. In this study, we leveraged the advantages of both the MinION and BGISEQ-500 platforms to make a bacteriologic diagnosis from a culture-negative lung tissue sample from an immunocompromised patient with severe pneumonia. Real-time nanopore sequencing rapidly identified Klebsiella pneumoniae by an 823 bp specific sequence within 1 min. Genomic analysis further identified bla_SHV-12, bla_KPC-2, bla_TEM-1, bla_CTX-M-65, and other resistance genes. The same sample was further sequenced on the BGISEQ-500 platform, which presented consistent results regarding the most top dominant pathogens and provided additional information of resistance genes. Revised antibiotic treatment was followed by the patient's clinical recovery. Though sample preparation and the interpretation of final results still need to be improved further, metagenomic sequencing contributes to the accurate diagnosis of culture-negative infections and facilitates the rational antibiotic therapy.

Keywords: clinical diagnosis; metagenomic next-generation sequencing (mNGS); nanopore sequencing; severe pneumonia; whole genome sequencing.

Figure 1

Timeline from hospitalization to patient discharge. The markers in black show the events from hospitalization to pre sequencing and the markers in blue show the events from sequencing to patient discharged. The red pentagram indicates the dates of taking samples and sequencing, and the red dot indicates the dates of culturing and sequencing results, respectively.

Figure 2

The species composition of mNGS data from MinION (A) and BGISEQ-500 (B). “Others” represents the sum of all species which had a proportion <1%.