gamma delta T cells represent an evolutionarily primitive T cell subset characterized by distinct T cell receptors (TCRs) and innate and adaptive immune functions. However, the presence of this T cell subset in ancient vertebrates remains unclear. In this study, gamma delta T cells from a zebrafish (Danio redo) model were subjected to molecular and cellular characterizations. The constant regions of zebrafish TCR-gamma (DrTRGC) and 6 (DrTRDC) were initially identified. Zebrafish gamma delta T cells accounted for 7.7-20.5% of the total lymphocytes in spleen, head kidney, peripheral blood, skin, gill, and intestine tissues. They possess typical morphological features of lymphocytes with a surface phenotype of gamma+delta+CD4(-)CD8(+). Zebrafish gamma delta T cells functionally showed a potent phagocytic ability to both soluble and particulate antigens. They can also act as an antigen-presenting cell to initiate antigen (KLH)-specific CD4(+) T-KLH cell activation and to induce B cell proliferation and IgM production. Particularly, zebrafish gamma delta T cells also play a critical role in antigen-specific IgZ production in intestinal mucus. These findings demonstrated that gamma delta T cells had been originated as early as teleost fish, which providing valuable insights into the evolutionary history of T cell subset. It is anticipated that this study would be used as a guide to develop a zebrafish model for the cross-species investigation of gamma delta T cell biology.