Genomic analysis of antimicrobial resistance and virulence among gram-negative bloodstream isolates from Lebanon

The increasing prevalence of multidrug-resistant (MDR) gram-negative bacteria has reduced therapeutic options for bloodstream infections (BSIs) and complicates clinical management. In Lebanon, limited genomic surveillance has restricted insight into the molecular determinants underlying resistance and virulence in invasive pathogens. In this study, whole-genome sequencing was used to characterize antimicrobial resistance, mobilome composition, and virulence gene content among 24 gram-negative isolates recovered from BSIs. Six species were identified, including Escherichia coli , Klebsiella pneumoniae , Citrobacter farmeri , Citrobacter portucalensis , Proteus mirabilis , and Morganella morganii . Phenotypic testing showed widespread resistance to β-lactam and non-β-lactam antimicrobial classes. Extended-spectrum β-lactamase production was detected in 62.5% of isolates, and 37.5% were classified as carbapenem-resistant Enterobacterales. Genomic analyses revealed that resistance in E. coli and K. pneumoniae was largely driven by plasmid-mediated ESBL and carbapenemase genes associated with IncF replicons and insertion sequences, whereas chromosomal mechanisms predominated in Citrobacter species. A chromosomally encoded bla OXA-244 variant was identified in an E. coli ST69 isolate, and a high-risk K. pneumoniae ST147 isolate carried multiple β-lactamase genes across distinct plasmids. Colistin resistance in one K. pneumoniae isolate was associated with a frameshift mutation in mgrB in the absence of plasmid-mediated mcr genes. Virulence profiling showed heterogeneous repertoires across species, with expanded adhesion, iron acquisition, and serum resistance determinants in E. coli . This study provides the first genome-level characterization of C. farmeri and C. portucalensis BSIs in Lebanon and supports the integration of genomic surveillance into routine monitoring of invasive infections. IMPORTANCE This study presents a pilot whole-genome sequencing analysis of 24 gram-negative bloodstream isolates from a tertiary care hospital in Lebanon. By integrating phenotypic susceptibility testing with genomic, mobilome, and virulence analyses, it reveals marked species- and isolate-level diversity. Resistance and virulence determinants were found within specific genetic contexts rather than being uniformly linked to species or sequence types, underscoring the value of genome-based surveillance to inform antimicrobial stewardship and infection control strategies.