Characterising the microbial and antimicrobial resistance signatures of hospital-acquired pneumonia using nanopore metagenomic sequencing

Hospital-acquired pneumonia (HAP) is a significant burden in nosocomial settings, yet its microbial underpinnings remain poorly understood. Here, we leverage shotgun nanopore sequencing to characterise the respiratory microbiomes of 250 HAP patients in a UK multi-site cohort, validating these using paired PCR and culture assays. Sequencing identified the dominant microbes implicated in HAP, including detection of probable pathogens in 49 PCR- and culture-negative cases. We found a high prevalence of fungi in 81/239 (34%) in HAP patients, of whom 26/81 (32%) were PCR/culture-negative, suggesting that fungi may represent an under-investigated component of HAP, whether as colonists or pathogens. Although HAP is clinically sub-categorised based on the use and duration of ventilation before disease onset, we found that the microbial profiles of these sub-groups were indistinguishable. We also found a concerningly high proportion of multi-drug-resistant microbes in HAP patients, with 21% of assembled bacterial genomes harbouring acquired antimicrobial resistance (AMR) genes that confer resistance to at least three classes of antimicrobials. This included high AMR gene carriage associated to Staphylococcus epidermidis , which may be an important reservoir of AMR, though typically viewed as a commensal. Our work provides extensive metagenomic characterisation of HAP, underscores the value of metagenomics in describing its complex aetiology, and further prompts its potential role for pathogen detection, resistance profiling and treatment.