BACKGROUND: Our understanding of adaptive immune responses in COVID-19 patients is rapidly evolving, but information on the innate immune responses by natural killer (NK) cells is still insufficient. OBJECTIVE: We aimed to examine the phenotypic and functional status of NK cells and their changes during the course of mild and severe COVID-19. METHODS: We performed RNA sequencing (RNA-seq) and flow cytometric analysis of NK cells from patients with mild and severe COVID-19 at multiple time points in the course of the disease using cryopreserved peripheral blood mononuclear cells (PBMCs). RESULTS: In RNA-seq analysis, the NK cells exhibited distinctive features compared to healthy donors, with significant enrichment of pro-inflammatory cytokine-mediated signaling pathways. Intriguingly, we found that the unconventional CD56(dim)CD16(neg) (uCD56(dim)) NK cell population expanded in cryopreserved PBMCs from COVID-19 patients regardless of disease severity, accompanied by decreased NK cell cytotoxicity. The NK cell population was rapidly normalized alongside the disappearance of uCD56(dim) NK cells and the recovery of NK cell cytotoxicity in mild COVID-19 patients, but this occurred slowly in severe COVID-19 patients. CONCLUSION: The current longitudinal study provides a deep understanding of the NK cell biology in COVID-19.