A new lignolytic micromycete fungus Myrothecium roridum LG7 was isolated and selected for biological delignification of agro residue-paddy straw and herbaceous weed Parthenium sp. Physical and chemical modifications in the biomass following pretreatment with M. roridum LG7 for 7days in term of structural modification and lignin removal, changes in lignin skeleton, and alteration of cellulose crystallinity was observed through SEM-EDXA, FTIR and XRD analysis, respectively. Colonization of the fungus led to high amount of lignin removal (5.8–6.98mg/gds) from pretreated biomass which could be recovered as a value added product. Enzymatic hydrolysis of M. roridum LG7 pretreated biomass released significantly higher amount of reducing sugars (455.81–509.65mg/gds) as compared to respective raw biomass within 24h. This study illustrates the promise of M. roridum LG7 for biological pretreatment through structural and chemical alteration of biomass beside creation of alkaline environment which prevent the growth of other contaminants.