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Abstract Enterococci, listed as priority pathogens by the WHO, are frequently encountered in hospital-acquired infections. Additionally, owing to their capability to thrive in diverse environments and hosts, they can lead to various community-acquired infections. Enterococci are responsible for a broad spectrum of infectious diseases, including urinary tract infections, bloodstream infections, endocarditis, intraabdominal infections, skin and soft tissue infections, and surgical site infections. The genus Enterococcus includes several species. The most common species incriminated in human infections are E. faecalis and E. faecium. E. faecalis is known to be more virulent while E. faecium is known to be more resistant to antibiotics. Treating enterococcal infections poses significant challenges due to their inherent resistance to multiple classes of antibiotics and their remarkable ability to acquire resistance to almost all antibiotic groups. Due to the global spread of antimicrobial resistance in enterococci to most of the available antibiotics, the use of antimicrobials of the last resort is increasing. Linezolid, a critical last-resort antibiotic, demonstrates efficacy against various clinically significant Gram-positive cocci, including S. aureus, S. epidermidis, Enterococcus spp., and streptococci. Its mechanism of action involves binding to the V domain of the 23S rRNA component within the 50S ribosomal subunit. This binding impedes the formation of a functional 70S initiation complex, disrupting bacterial protein synthesis. Although still relatively uncommon, global rates of linezolid resistance are increasing. Resistance to linezolid can result from alterations at the target site, primarily through mutations in the 23S rRNA. Additionally, transferable linezolid resistance genes have been identified in resistant strains, with notable genes like Cfr, Cfr(B), Cfr(D), OptrA, and PoxtA contributing to this resistance mechanism. The objectives of this study were to isolate Enterococcus spp., specifically focusing on E. faecalis and E. faecium, from patients at Menoufia University Hospitals who suffered from either hospital or communityacquired infections. Additionally, the study aimed to determine the prevalence of these bacteria in different specimens and hospital departments, assess their antimicrobial susceptibility patterns, identify risk factors associated with infection by linezolid-resistant enterococci, and investigate the genetic basis of linezolid resistance. This study was carried out between October 2021 and April 2023 at the Medical Microbiology and Immunology Department of Menoufia University and the National Liver Institute. A total of 660 clinical samples were gathered and processed using the standard microbiological procedures. The most commonly collected specimens were urine (31.5%), followed by blood (21%), sputum (15.9%), wound discharge (10.6%), and ascitic fluid (9.7%). Among these samples, 102 Enterococcus spp. were isolated, comprising 15.7% of all culture-positive specimens. Species identification of enterococcal isolates was done by Vitek -2 automated system. E. faecalis represented 54.9% of all enterococcal isolates while E. faecium represented 43.1% and other species accounted for approximately 2%. Focusing specifically on E. faecalis and E. faecium isolates (totaling 100), 60% were from patients with healthcare-associated infections (HAIs) and 40% from patients with community-acquired infections (CAIs). HAIs were more prevalent among enterococcal infections. The mean age of infected patients was 46.25 ±16.72 years. Analysis revealed that 68% of infected patients had chronic medical conditions, 11% had malignant diseases, and 89% had a history of antibiotic administration. Additionally, 50% of patients utilized medical devices, and 29% underwent invasive procedures. |