When the Last Line Fails: Characterization of Colistin-Resistant Acinetobacter baumannii Reveals High Virulence and Limited Clonal Dissemination in Greek Hospitals

<i>Acinetobacter baumannii</i> has emerged as a major pathogen responsible for healthcare-associated infections, particularly in intensive care units, contributing to significant morbidity and mortality due to its multidrug resistance and ability to persist in clinical environments. This study aimed to investigate the phenotypic and genomic characteristics of all multidrug-resistant <i>A. baumannii</i> isolates collected between January and June 2022 from two tertiary care hospitals in Thessaloniki, Greece. A total of 40 isolates were included. All isolates exhibited resistance to colistin; however, none harbored the <i>mcr</i>-1 to <i>mcr</i>-9 genes, as confirmed by polymerase chain reaction (PCR). PCR-based screening for virulence-associated genes revealed high prevalence rates of <i>basD</i> (100%), <i>pld</i> (95%), <i>csuE</i> (87.5%), and <i>bap</i> (77.5%). In contrast, <i>ompA</i> and <i>pglC</i> were not detected. Twitching motility ranged from 2 to 50 mm, with 25% of the isolates classified as non-motile and 20% as highly motile. Swarming motility was observed in all strains. Additionally, all isolates demonstrated positive α-hemolysis, suggesting a potential virulence mechanism involving tissue damage and iron acquisition. Pulsed-field gel electrophoresis (PFGE) revealed significant genomic diversity among the isolates, indicating a low likelihood of patient-to-patient or clonal transmission within the hospital setting. These findings highlight the complex relationship between antimicrobial resistance and virulence in clinical <i>A. baumannii</i> isolates and emphasize the urgent need for robust infection control strategies and continued microbiological surveillance.