Summary: | The escalating transmission of hospital-acquired infections, especially those due to antimicrobial-resistant bacteria, is a major health challenge worldwide. In this study, a culturomic analysis of bacterial community in a tertiary care hospital in the western region of Saudi Arabia is performed using environmental samples. The genome sequencing of four <i>Acinetobacter baumannii</i> was performed on isolates recovered from an intensive care unit (ICU) environment and clinical samples. A total of 361 bacterial isolates from surface and air samples were identified by MALDI-TOF technique or 16S rRNA gene sequencing. The isolates were classified into 70 distinct species, including ESKAPE pathogens. Resistance in Gram-positive isolates was mainly found to be against benzylpenicillin, azithromycin, ampicillin, and trimethoprim/sulfamethoxazole. Carbapenem- and multidrug-resistant isolates of <i>A. baumannii</i> and <i>Klebsiella pneumonia</i> were found on the ICU surfaces. Genome sequencing revealed that the carbapenem-resistant <i>A. baumannii</i> isolate from ICU environment was linked with those of clinical origin. The isolate Ab133-HEnv was classified as a novel sequence type (ST2528) based on a new allele of Oxf_gdhB-286. Three beta-lactam-antibiotic-resistance genes, <i>bla</i><sub>ADC-25</sub>, <i>bla</i><sub>OXA-23</sub>, and <i>bla</i><sub>OXA-66</sub>, were found in most of the analyzed genomes. Collectively, the results of this study highlight the spread of antimicrobial-resistant nosocomial pathogens in a health care facility in Saudi Arabia.
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