Genomic Surveillance and Resistance Profiling of Multidrug-Resistant Acinetobacter baumannii Clinical Isolates: Clonal Diversity and Virulence Insights

<i>Acinetobacter baumannii</i> is a multidrug-resistant opportunistic pathogen that poses critical challenges in hospital settings due to its environmental resilience and high resistance to antibiotics. Genomic surveillance has become essential for identifying transmission patterns, guiding antimicrobial stewardship, and informing infection control policies. We conducted whole-genome sequencing on 44 <i>A. baumannii</i> isolates collected between 2022 and 2023 from diverse wards in an Italian hospital. Illumina-based sequencing was followed by a comprehensive bioinformatics pipeline, including genome assembly, taxonomic validation, MLST, SNP-based phylogeny, pan-genome analysis, antimicrobial resistance (AMR) gene profiling, and virulence factor prediction. Most isolates were classified as ST2; SAMPLE-34 was ST1 and genetically distinct. Phylogenetic analysis revealed four clonal clusters with cluster-specific AMR and accessory gene content. The pan-genome included 5050 genes, with notable variation linked to hospital ward origin. ICU and internal medicine strains carried higher loads of AMR genes, especially against aminoglycosides, β-lactams, and quinolones. Virulence profiling highlighted widespread immune evasion mechanisms; "Acenovactin" was predominant, while some isolates lacked key adhesion or toxin factors. Our findings underscore the clinical relevance of integrating genomic epidemiology into routine hospital surveillance. Identifying clonal clusters and resistance signatures supports real-time outbreak detection, risk stratification, and targeted infection prevention strategies.