Assessment of Microbiome-Based Pathogen Detection Using Illumina Short-Read and Nanopore Long-Read Sequencing in 144 Patients Undergoing Bronchoalveolar Lavage in a University Hospital in Germany

Lower respiratory tract infections (LRTIs) represent a significant global health concern, and the accurate identification of pathogens is crucial for patient care. Culture-based methods are the gold standard, but their detection abilities are limited. Next-generation sequencing (NGS) offers a promising method for comprehensive microbial detection, providing valuable information for clinical practice. In this study, 144 bronchoalveolar lavage fluid samples were collected, culture-based diagnostics were performed, and bacterial microbiome profiles were generated by short-read sequencing of the V4 region of the 16S rRNA gene using Illumina technologies and long-read sequencing with Oxford Nanopore Technologies (ONT) to determine the full-length 16S rRNA gene. The most common genera detected by NGS included Streptococcus, <i>Staphylococcus</i>, <i>Veillonella</i>, <i>Prevotella</i>, <i>Rothia</i>, <i>Enterococcus</i>, and <i>Haemophilus</i>. Short-read sequencing detected cultured bacteria at the genus level in ~85% of cases, while long-read sequencing demonstrated agreement with cultured species in ~62% of cases. In three cases, long-read sequencing identified the uncommon potential lung pathogen <i>Tropheryma whipplei</i> not detected with traditional culturing techniques. The NGS results showed a partial overlap with culture as the current diagnostic gold standard in LRTI. Additionally, NGS detected a broader spectrum of bacteria, revealed fastidious potential pathogens, and offered deeper insights into the complex microbial ecosystem of the lungs.