The management of melon fly Zeugodacus cucurbitae (Coquilett) having wide host range is most challenging. Bacillus thuringiensis remains a potential pest management candidate of biological origin. In the present study, nine indigenous Bt isolates were characterised and evaluated against neonate (~12 hrs old) maggots. The isolates showed a diversified nature of colony and crystal morphology. Concentration-response toxicity assay revealed that three isolates viz., T166, T60 and T184 were highly toxic with LC50 values of 0.38, 0.41 and 0.40 µg/ µl, respectively on par with the reference strain Bti 4Q2 with 0.37 µg/ µl. LC95 of Bt cultures ranged from 1.15 to 2.09 µg/ µl. Gene profiling revealed the occurrence of cry4Aa in T166 and T184, cry4Ba in T60, cry11Aa in T166 and cyt1 in T166 with protein profiling showing proteins of ~134, ~128, ~72 and ~27kDa, respectively.