Genome reconstruction and characterisation of extensively drug-resistant bacterial pathogens through direct metagenomic sequencing of human faeces

Whole-genome sequencing of microbial pathogens is revolutionising modern approaches to outbreaks of infectious diseases and is reliant upon organism culture. Culture-independent methods have shown promise in identifying pathogens, but high level reconstruction of microbial genomes from microbiologically complex samples for more in-depth analyses remains a challenge. Here, using metagenomic sequencing of a human faecal sample and analysis by tetranucleotide frequency profiling projected onto emergent self-organising maps, we were able to reconstruct the underlying populations of two extensively-drug resistant pathogens, Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae and vancomycin-resistant Enterococcus faecium. From these genomes, we were able to ascertain molecular typing results, such as MLST, and identify highly discriminatory mutations in the metagenome to distinguish closely related strains. These proof-of-principle results demonstrate the utility of clinical sample metagenomics to recover sequences of important drug-resistant bacteria and application of the approach in outbreak investigations, independent of the need to culture the organisms.