Background Identifying the causes of community-acquired pneumonia (CAP) is challenging due to the disease’s complex etiology and the limitations of traditional microbiological diagnostic methods. Recent advances in next generation sequencing (NGS)-based metagenomics allow pan-pathogen detection in a single assay, and may have significant advantages over culture-based techniques. Results We conducted a cohort study of 159 CAP patients to assess the diagnostic performance of a clinical metagenomics assay and its impact on clinical management and patient outcomes. When compared to other techniques, clinical metagenomics detected more pathogens in more CAP cases, and identified a substantial number of polymicrobial infections. Moreover, metagenomics results led to changes in or confirmation of clinical management in 35 of 59 cases; these 35 cases also had significantly improved patient outcomes. Conclusions Clinical metagenomics could be a valuable tool for the diagnosis and treatment of CAP. Trial registration Trial registration number with the Chinese Clinical Trial Registry: ChiCTR2100043628.

Fast and accurate identification of microbial pathogens is critical for the proper treatment of infections. Traditional culture-based diagnosis in clinics is increasingly supplemented by metagenomic next-generation-sequencing (mNGS). Here, RNA/cDNA-targeted sequencing (meta-transcriptomics using NGS (mtNGS)) is established to reduce the host nucleotide percentage in clinic samples and by combining with Oxford Nanopore Technology (ONT) platforms (meta-transcriptomics using third-generation sequencing, mtTGS) to improve the sequencing time. It shows that mtNGS improves the ratio of microbial reads, facilitates bacterial identification using multiple-strategies, and discovers fungi, viruses, and antibiotic resistance genes, and displaying agreement with clinical findings. Furthermore, longer reads in mtTGS lead to additional improvement in pathogen identification and also accelerate the clinical diagnosis. Additionally, primary tests utilizing direct-RNA sequencing and targeted sequencing of ONT show that ONT displays important potential but must be further developed. This study presents the potential of RNA-targeted pathogen identification in clinical samples, especially when combined with the newest developments in ONT.

BackgroundTo evaluate the value of metagenomic next-generation sequencing (mNGS) for the early diagnosis of psittacosis, and to investigate its epidemiology by whole-genome capture.MethodsTwenty-one bronchoalveolar lavage fluid (BALF) and blood samples of 16 psittacosis patients from multiple centers during August 2019 to September 2021 were analyzed retrospectively. mNGS with normal datasets (10 M 75-bp single-end reads after sequencing) and larger datasets (30 M 150-bp paired-end reads after sequencing) as well as quantitative real-time polymerase chain reaction (qPCR) were used to detect the pathogen. Also, whole-genome capture of Chlamydophila psittaci was applied to draw the phylogenetic tree.ResultsmNGS successfully detected the pathogen in all 16 cases (100%), while qPCR was positive only in 5 out of 10 cases (50%), indicating a significantly higher sensitivity of mNGS than qPCR (p < 0.01). BALF-mNGS performed better than blood-mNGS (16/16 versus 3/5, p < 0.05). In addition, larger datasets (the read counts have tripled, and the base number was 12-fold larger compared to clinical mNGS with a normal dataset) of mNGS showed significantly increased contents of human DNA (p < 0.05) and decreased reads per million of the pathogen, suggesting no improvement. Whole-genome capture results of five samples (>60% coverage and >1 depth) were used to construct the phylogenetic tree.ConclusionSignificant advantages of mNGS with normal datasets were demonstrated in early diagnosing psittacosis. It is the first study to use whole-genome capture to analyze C. psittaci epidemiological information.

ObjectiveCritical illnesses in the intensive care unit (ICU) have been a global burden. We aimed to determine the correlation between the lung and gut in critically ill patients to find novel evidence of the lung-gut axis, which may be a new treatment for patients with critical illness in the ICU.MethodsWe collected bronchoalveolar lavage specimens and fecal samples of 31 patients with critical illness within 24 h after admission. Metagenomics was used to detect lung and intestinal samples. Immune cells were detected by flow cytometry.ResultsThere are 86 common species in both lung and gut. The abundance of Enterococcus faecium is high in both the lung and gut of patients with critical illness in the respiratory intensive care unit (RICU). Corynebacterium striatum in the lung and gut is correlated with different immune cells. In addition, C. striatum in the lung and gut might share the same source, supporting the concept of a gut-lung axis in humans.ConclusionsThe microbiome in the lung and gut showed a correlation to some extent, and C. striatum in the lung and gut might share the same source. In addition, the microbiome showed a correlation with immunity, indicating a potential therapeutic target in patients with critical illness. The lung-gut axis might play an important role in patients with critical illness in the RICU.