Next-generation sequencing to investigate antimicrobial resistance: a one-health perspective

Antimicrobial resistance (AMR) is a leading cause of death around the world and, given the occurrence and dissemination of antimicrobial-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) in different ecological niches, such as human population, wild and farm animals and environment, it is necessary to look at it from a One-Health perspective. Next-generation sequencing (NGS) allows to determine the whole genome of a single pathogen (culture-dependent) but also to detect microorganisms and ARGs composition by sequencing the DNA/RNA content of a given sample without previous isolation (culture-independent). This thesis aimed to use NGS technologies for the surveillance of ARB and to track the dissemination of ARGs in human healthcare and animal farms. We showed the potential of applying a culture-independent approach, shotgun metagenomics (SMg), to monitor the bacterial community and resistome profiles of tap-water samples collected from high-risk hospital units and oral fluid samples collected from pig farms. In addition, we use a culture-dependent approach to investigate the presence of ARB resistant to last-resort antibiotics, such as Carbapenem-resistant (CRE) and Colistin-Resistant Enterobacterales (ColRE). Moreover, we demonstrated the possibility of combining short- and long-read whole genome sequencing (WGS) to resolve mobile genetic elements carrying ARGs, to characterize environmental strains and investigate bacterial molecular epidemiology in the nosocomial environment.