The fungal species complex of Fusarium oxysporum contains human and plant pathogenic strains. An F. oxysporum genome can be compartmentalized into highly conserved core chromosomes and strain-specific accessory chromosomes that are enriched in genes for host-specific pathogenicity and support the adaptability of this cross-kingdom pathogen. To better understand the adaptive evolution of F. oxysporum, my study focuses on a keratitis strain, MRL8996, which was responsible for the 2005–2006 major keratitis outbreak linked to contaminated contact lens solution using short-term experimental evolution (STEE). The ancestral isolate was passaged ten times in vivo in a mouse keratitis model to create independently evolved lineages. In vitro phenotyping characterization was carried out by measuring the growth rates of end populations of each evolved lineage in different abiotic stress conditions. Genotypically, we sequenced all end populations of each evolved lineage using whole genome shotgun sequencing. Compared to the genome of the ancestral isolate, we categorized all mutations, including SNPs, Indels, and transposon insertions. Most interestingly, we observed partial decrease in copy number or a complete depletion of chromosome 12, a core chromosome. Quantitative PCR (qPCR) of the ancestral strain, the end population and intermediate passages of the evolved lineages is conducted to track the dynamics of this deletion event. In addition, we are making efforts to correlate observed genotypic variation with interesting adapting phenotypes. These findings demonstrate that the keratitis-associated pathogen exhibits dynamic genome plasticity, including loss of core chromosomal material, facilitating adaptation to host-associated environments and resulting in altered responses to abiotic stress conditions.