Bioversity of fungal antagonist, Trichoderma from different locations of Nilgiri district of Tamil Nadu, India, were characterized through molecular methods. Thirty four isolates were tentatively identified as Trichoderma and confirmed upto species level through molecular tools. PCR amplification of the 18s-28s rRNA gene region revealed that all thirty four isolates produced amplicon size of 600bp and were further confirmed through sequencing. The sequences of various Trichoderma spp. were compared with both NCBI and TrichOKEY database to validate their molecular identity. Among 34 isolates, 31 isolates were identified as T. asperellum (KT462693, KU361372, KX533978 to KX533999, KX523262 to KX523264, KX555650, KX147092 to KX147094, KX5334000), 2 isolates as T. harzianum (KX533989, KX533990)  and 1 isolate as T. virens (KU666466) through NCBI data base. However, all the T. asperellum isolates identified through NCBI database were identified as T. koningiopsis using TricHOKEY data base. Identity of T. harzianum isolates (TRI 35 and TRI 36) and T. virens isolate (TRI 37) were same in both NCBI and TricHOKEY database. Antagonistic assay with diverse species of Trichoderma revealed that T. virens (TRI 37) was effective in inhibiting the radial growth of Pythium aphanidermatum (87.78%) followed by T. harzianum (TRI 35), (TRI 36) and T. asperellum (TRI 9) in vitro. The effective isolates T. virens (TRI 37), T. harzianum (TRI 35, TRI 36)  and T. asperellum  (TRI 9) were compatible with each other. Biopriming of cucumber seeds with talc based formulation of the consortia comprising of T. virens  isolate ( TRI 37), T. harzianum isolates (TRI 35 and TRI 36) and T. asperellum TRI 9 @ 10⁸ cfu/g and soil application suppressed damping off to an extent of 76.82%  over untreated control.