A Syntenic Pangenome for Gardnerella Reveals Taxonomic Boundaries and Stratification of Metabolic and Virulence Potential across Species

Bacterial vaginosis (BV) is a prevalent condition associated with an imbalance in the vaginal microbiota, often involving species of Gardnerella . The taxonomic complexity and inconsistent nomenclature of Gardnerella have impeded progress in understanding the role of specific species in health and disease. In this study, we conducted a comprehensive genomic and pangenomic analysis to resolve taxonomic ambiguities and elucidate metabolic and virulence potential across Gardnerella species. We obtained complete, closed genomes for 42 Gardnerella isolates from women with BV and curated publicly available genome sequences (n = 291). Average nucleotide identity (ANI) analysis, digital DNA-DNA hybridization (dDDH), and the cpn60 gene sequences identified nine species and eleven subspecies within Gardnerella , for which we refined species and subspecies boundaries and proposed updated nomenclature. Pangenome analysis revealed species-specific gene clusters linked to metabolic pathways, virulence factors, and niche adaptations, distinguishing species specialized for mucin degradation in the vaginal environment from those potentially adapted to urinary tract colonization. Notably, we identified lineage-specific evolutionary divergence in gene clusters associated with biofilm formation, carbohydrate metabolism, and antimicrobial resistance. We further discovered the first cryptic plasmids naturally present within the Gardnerella genus. Our findings provide a unified framework for Gardnerella taxonomy and nomenclature, and enhance our understanding of species-specific functional capabilities, with implications for Gardnerella research, diagnostics, and targeted therapeutics in BV.