A prominent feature of disease induced by Mycoplasma gallisepticum is a lymphoproliferative response in the respiratory tract. Although this is also seen in other mycoplasma infections, including Mycoplasma pneumoniae, the phenotype of the lymphocytes infiltrating the respiratory tract has not been determined. In this study, the numbers and distribution of lymphocytes in the tracheas of chickens infected with a virulent strain of M. gallisepticum were examined. Three groups of chickens were experimentally infected with M. gallisepticum and three unchallenged groups were used as controls. One infected and one control group were culled at 1, 2 and 3 weeks post infection. Tracheas were removed and examined for the presence and number of T cells carrying CD4 CD8, TCR gamma delta, TCR alpha beta 1 or TCR alpha beta 2 markers. There was no significant difference in the number of CD8(+) cells in the upper, middle and lower trachea. High numbers of both CD4(+) and CD8(+) cells were found with variable numbers of TCR alpha beta 1(+) and TCR alpha beta 2(+), but no TCR gamma delta(+), cells throughout the time course. The distribution of CD4 cells was dispersed, while the CD8(+) cells were clustered in follicular-like arrangements. No difference was detected in the distribution of TCR alpha beta 1(+) and TCR alpha beta 2(+) cells. The titre of mycoplasma genomes in the trachea decreased significantly from 1 to 2 weeks, while the mucosal thickness of the trachea increased significantly from 1 to 2 weeks then decreased from 2 to 3 weeks, indicating resolution of the lesions following control of infection. This study is the first to examine the phenotypes of T lymphocytes infiltrating the respiratory tract during mycoplasma infections. The findings suggest involvement of specific stimulation of CD8(+) cells, particularly in the acute phase of disease.