BACKGROUND: An alarming rise in reported Lassa fever cases continues in west Africa. Liberia has the largest reported per capita incidence of Lassa fever cases in the region, but genomic information on the circulating strains is scarce. The aim of this study was to substantially increase the available pool of data to help foster the generation of targeted diagnostics and therapeutics. METHODS: Clinical serum samples collected from 17 positive Lassa fever cases originating from Liberia (16 cases) and Guinea (one case) within the past decade were processed at the Liberian Institute for Biomedical Research using a targeted-enrichment sequencing approach, producing 17 near-complete genomes. An additional 17 Lassa virus sequences (two from Guinea, seven from Liberia, four from Nigeria, and four from Sierra Leone) were generated from viral stocks at the US Centers for Disease Control and Prevention (Atlanta, GA) from samples originating from the Mano River Union (Guinea, Liberia, and Sierra Leone) region and Nigeria. Sequences were compared with existing Lassa virus genomes and published Lassa virus assays. FINDINGS: The 23 new Liberian Lassa virus genomes grouped within two clades (IV.A and IV.B) and were genetically divergent from those circulating elsewhere in west Africa. A time-calibrated phylogeographic analysis incorporating the new genomes suggests Liberia was the entry point of Lassa virus into the Mano River Union region and estimates the introduction to have occurred between 300-350 years ago. A high level of diversity exists between the Liberian Lassa virus genomes. Nucleotide percent difference between Liberian Lassa virus genomes ranged up to 27% in the L segment and 18% in the S segment. The commonly used Lassa Josiah-MGB assay was up to 25% divergent across the target sites when aligned to the Liberian Lassa virus genomes. INTERPRETATION: The large amount of novel genomic diversity of Lassa virus observed in the Liberian cases emphasises the need to match deployed diagnostic capabilities with locally circulating strains and underscores the importance of evaluating cross-lineage protection in the development of vaccines and therapeutics. FUNDING: Defense Biological Product Assurance Office of the US Department of Defense and the Armed Forces Health Surveillance Branch and its Global Emerging Infections Surveillance and Response Section.