Heliminthosporium oryzae is a necrotrophic fungal pathogen that effect rice crops grown on millions of hectares. We evaluated nine newly establishing rice lines and one local variety for resistance against H. oryzae. Significant (P ≤ 0.05) differences in response to pathogen attack were recorded in all rice lines. Maximum disease resistance was recorded in Kharamana under pathogen attack as compared to uninfected plants. A comparison of decline in shoot length revealed that Kharamana and Sakh experienced minimum lost (9.21%, 17.23%) in shoot length respectively against control while Binicol exhibited highest reduction (35.04%) in shoot length due to H. oryzae attack. Post-infection observations of shoot fresh weight revealed 63% decline in Binicol and declared it as the most susceptible rice line. Sakh, Kharamana and Gervex exhibited minimum fresh weight decrease (19.86%, 19.24% and 17.64% respectively) as compared to other lines under pathogen attack. Maximum chlorophyll-a contents were recorded in Kharamana under control and post pathogen attackconditions. Following the inoculation of H. oryzae, SOD was increased up to 35% and 23% in Kharamana and Sakh. However, minimum POD activity was recorded in Gervex followed by Swarnalata, Kaosen and C-13 in non-inoculated and pathogen-inoculated plants. Significant decrease in ascorbic acid contents (73.7% and 70.8%) was observed in Gervex and Binicol that later contributed in their susceptibility to H. oryzae attack. Pathogen attack caused Significant (P ≤ 0.05) changes in secondary metabolites in all rice lines but minimum total flavonoids, anthocyanin and lignin were observed in Binicol in uninfected plants and attested its susceptibility to pathogen. In post-pathogen attack conditions, Kharamana showed best resistance against pathogen by exhibiting a significantly high and maximum value of morpho-physiological, and biochemical attributes. Our findings suggest that tested resistant lines can be further explored for multiple traits including molecular regulation of defense responses to breed immunity in rice varieties.