Background: Fusobacterium nucleatum is a genetically diverse species and a key organism in the human oral microbiome. It has been associated with several extra oral diseases, most notably, colorectal cancer. Its extensive genomic mosaicism complicates understanding of the clinical impact of genetic variation. We aimed to resolve strain level diversity and identify functional variation linked to pathogenic potential. Methods: Whole genome assemblies were generated using Oxford Nanopore sequencing for five type strains and sixteen cultured isolates. Comparative analyses included phylogeny, pangenome analysis and evaluation of the presence of core genes from Bacillus subtilis and Escherichia coli. TBLASTN hits were categorised using defined confidence thresholds to predict gene presence and divergence.  Results: Virulence gene carriage varied among isolates, including differential carriage of the fad family of cancer associated virulence genes and antibiotic resistance determinants. The apparent absence of ftsZ from one type strain confirms challenges in identifying homology in a highly divergent species and may uncover unusual evolution of cell division machinery. We also observed variation in the presence of key genes involved in antibiotic sensitivity including penicillin binding proteins and ß-lactamases. In vitro screening additionally revealed surprising vancomycin sensitivity, unexpected for this Gram-negative species.   Conclusion: Identifying strain or clade specific variation in this human pathobiont will support the development of targeted therapies and diagnostics. Our work highlights the necessity of culturing and whole genome sequencing to reveal the functional variation in this diverse, clinically relevant species that shotgun metagenomics and metataxonomics would not be able to deliver.