vanB-Gene-Dominated Resistance in Enterococcus spp. and Silent vanA-Gene Carriage in Phenotypically Susceptible Isolates: Genomic Epidemiology in Two Hospitals in Latvia

Background/Objectives: Vancomycin-resistant (VRE) and vancomycin-variable (VVE) Enterococcus spp. represent an increasing clinical challenge due to limited treatment options and the potential for undetected dissemination of such resistance genes. Data on Enterococci genomic epidemiology in healthcare settings remain rather limited. Our study aimed to investigate vancomycin resistance determinants in Enterococcus spp., clonal structure, and occurrence of VVE using whole-genome sequencing (WGS) in Latvia. Methods: Clinical isolates collected from hospitalised patients in two tertiary-level hospitals in Latvia (2021–2024) were analysed using WGS following routine laboratory identification. Vancomycin resistance determinants were identified in silico, along with MLST and cgMLST genotyping. Results: Of 532 sequenced isolates, 482 met the quality and inclusion criteria. E. faecalis (56.64%) and E. faecium (40.25%) predominated. Among 125 isolates carrying vancomycin resistance genes, vanB (54.40%) was the most frequent, followed by vanA (38.20%) and vanC (6.40%); vanC was restricted to E. gallinarum and E. casseliflavus. Vancomycin resistance was more prevalent in E. faecium (51.03%) than in E. faecalis (6.59%). cgMLST identified outbreak clusters among E. faecium ST80 and ST78 with complex type-specific resistance patterns and hospital specificity. E. faecalis showed polyclonal endemicity with the vanB gene present in different clades. Three (0.62%) vancomycin-variable E. faecium (VVE) isolates were identified in one hospital, harbouring vanA-type gene clusters comprising vanHAX but lacking the sensory gene vanS and the regulatory gene vanR. Conclusions: The VanB gene predominated in both hospitals, driven by clonal expansion of hospital-adapted E. faecium ST80/ST78, contrasting with earlier vanA predominance in Europe but aligning with recent regional vanB trends. The detection of VVE highlights clinically relevant genotype–phenotype discordance, underscoring the importance of integrating genomic surveillance with routine phenotypic testing to detect cryptic resistance and guide effective antimicrobial therapy.