Metagenomic investigation of VIM-type metallo-β-lactamase-producing multidrug-resistant Pseudomonas aeruginosa associated with a hospital outbreak across multiple hospital units in Saudi Arabia

BACKGROUND: Healthcare-associated infections (HAIs) caused by multidrug-resistant (MDR) Pseudomonas aeruginosa (P. aeruginosa) represent a public health challenge, particularly when associated with VIM-type metallo-β-lactamases (MBLs), which limit therapeutic options. Conventional microbiological methods may underestimate resistance determinants and transmission dynamics. Long-read metagenomic sequencing is a promising approach for Pathogen detection, resistome characterization, and genomic surveillance. OBJECTIVES: The study's objectives were to characterize the resistome, including detection of the blaVIM gene, assess genomic relatedness and potential transmission dynamics, and evaluate the diagnostic value of metagenomics compared with conventional microbiological approaches. METHODS: This retrospective infection control investigation included seven hospitalized patients from multiple hospital units. Clinical specimens included blood, respiratory specimens, surgical tissue, and device-associated material. Conventional microbiological investigations included bacterial culture, identification, and antimicrobial susceptibility testing (AST) using the VITEK 2 automated system. Carbapenemase genes were detected using Xpert Carba-R. Long-read metagenomic sequencing was conducted using Oxford Nanopore Technologies (ONT) on the GridION platform. Bioinformatic analysis was performed using the CosmosID platform for taxonomic profiling, antimicrobial resistance gene detection, and genomic relatedness assessment. RESULTS: Conventional microbiological methods identified carbapenem-resistant Pseudomonas aeruginosa (CRPA) in five cases, whereas ONT sequencing detected the blaVIM gene in all seven samples, demonstrating superior diagnostic sensitivity. A highly conserved resistome profile was identified across all investigated cases, including multiple β-lactamase and aminoglycoside, fluoroquinolone, and polymyxin-associated resistance determinants. Genomic relatedness analysis demonstrated close clustering patterns with minimal genomic variability, suggesting possible circulation of closely related MDR strains. CONCLUSION: These findings highlight the added value of ONT sequencing in identifying concealed resistance determinants and improving transmission tracking compared with conventional diagnostic approaches. Future investigations involving larger sample sizes and environmental surveillance are needed to further clarify transmission dynamics and potential reservoirs of VIM-producing P. aeruginosa.