The in vitro proliferation of peripheral blood lymphocytes (PBLs) without any mitogenic stimulation is one of the hallmarks of human T lymphotropic virus type I (HTLV-I) infection. Recent evidence suggests a difference in the degree of the phenomenon between HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and asymptomatic HTLV-I carriers (AC). In this article, we demonstrated several alterations in the features of the in vitro transformed lymphocytes between patients with HAM/TSP (n = 16) and AC (n = 8). The percentages of total CD8+ and CD8+CD28+ cells were significantly increased in the in vitro proliferating T lymphocytes derived from the patients with HAM/TSP when compared to those from AC. HAM/TSP was segregated from AC by the high degree of the proliferation of CD8+CD28+ cells. The expression of HTLV-I-specific antigens on the cultured PBLs was detected only in the subjects which showed low CD8+CD28+/CD4+ ratio of the in vitro proliferating lymphocytes. These findings suggest that this phenomenon distinguishes HAM/TSP from AC, not only in quantity but also in quality.