2,6-Dichloroisonicotinic acid (INA) seed treatment at different concentrations (0.5, 1, 2.5 and 5 mM) for 3 and 6 h was evaluated during the interaction of pearl millet with Sclerospora graminicola in association with chitinase activity. Susceptible pearl millet seeds treated with INA at 1 mM concentration for 3 h showed significant (P<0.05) downy mildew disease resistance compared to 3 h treatments. The present study revealed that INA treated @ 1 mM provided the maximum seed germination 95% and seedling vigor 1552 in comparison to control which offered 79% and 923 of seed germination and seedling vigor, respectively. The results on growth parameter studies revealed that INA treated plants showed enhanced growth parameters when compared to control under greenhouse conditions. Application of INA @ 1 mM reduced the severity of disease with a highest protection of 62% under greenhouse conditions at 6 h treatment and highest reduction found on third day of inoculation in time gap studies. The enhanced chitinase activity was observed at all test points in INA @ 1 mM treated seedlings as compared to the control and peak activity of 10.11 and 3.44 respectively, was found at 24 hai.