Genomic Epidemiology, Antimicrobial Resistance, and Virulence of Clinical Enterococcus Isolates from Hat Yai, Thailand: Genomic Relatedness and Lineage Persistence.

Background/Objectives: Enterococcus species are important causes of healthcare-associated infections, particularly multidrug-resistant (MDR) and vancomycin-resistant enterococci (VRE). This study characterized antimicrobial susceptibility, biofilm formation, genomic features, and molecular epidemiology of clinical Enterococcus isolates from southern Thailand. Methods: Forty clinical isolates, comprising E. faecalis (n = 22) and E. faecium (n = 18), were characterized by antimicrobial susceptibility testing, biofilm assays, and whole-genome sequencing. Results: All isolates were susceptible to linezolid and oritavancin, whereas non-susceptibility to ampicillin, fosfomycin, vancomycin, and daptomycin was 60.0%, 57.5%, 12.5%, and 5.0%, respectively. High-level gentamicin resistance and MDR occurred in 37.5% and 22.5%, respectively. Strong biofilm formation predominated in E. faecalis (86.4%), whereas 94.4% of E. faecium were weak producers. ST6 and ST17 predominated in E. faecalis and E. faecium, respectively. All five VRE isolates were E. faecium ST17 or ST262 and carried vanA. High-level ampicillin-resistant E. faecium harbored the PBP5 S466' insertion and multiple C-terminal substitutions. To our knowledge, this is the first report in Thailand of both daptomycin-resistant E. faecium and resistance-associated LiaS T120A and LiaR W73C substitutions. Closely related isolates within E. faecalis ST6 and E. faecium VRE ST17, ST117, and ST262 showed spatiotemporal patterns consistent with potential epidemiological linkage or lineage persistence. Three novel STs were identified, with E. faecium ST3096 exhibiting high-level ampicillin resistance, fosfomycin resistance, and reduced daptomycin susceptibility. Conclusions: These findings reveal emerging resistant lineages and spatiotemporal patterns of genomic relatedness, supporting continued genomic surveillance of clinical enterococci in Thailand.