Background & Aim: Vector-borne diseases are one of the major public health problems in developing countries. Culextritaeniorhynchus (Diptera: Culicidae) could be a vector of filariasis and West Nile encephalitis. The use ofentomopathogenic bacteria like Bacillus thuringiensis (Bt) as larvicides is a viable alternative for the biological controlmosquitoes with a low environmental impact. Hence, an attempt was made to explore the abundance of Bt in Mizoramsoils from different habitats, testing their toxicity against C. tritaeniorhynchus and characterization by following acetateselection, mortality, dissimilarity analysis and RAPD-PCR assay methods. Method: In this present study, nine soilisolates (Mzubt 2,4,5,6,11,23,25,26 and 29) were identified, characterized and evaluated against C. tritaeniorhynchus.The new isolates were compared with two reference strains - Bt israelensis 4Q1 and Bt aizawai 4J3 obtained fromBacillus Genetic Stock Center, USA. Results: The mzubt 2 and mzubt 6 isolates showed higher larvicidal activity thanother isolates, against C. tritaeniorhynchus. RAPD-PCR assay has been optimized that discriminate Bacillusthuringiensis (Bt) isolates from nine different habitats soil covering five districts of Mizoram. All the studied isolatesshowed a diverse RAPD patterns and were different from each other in relation to habitats, toxicity and type of cry genepresent. High polymorphism was observed between Bt isolates which was authenticated through high PIC, RP, EMR andMI values. Further, three major clusters were identified through dissimilarity analysis. Conclusion: No relationshipfound between the type of cry gene in Bt and their toxicity against mosquitoes. The importance of the isolation of nativestrains of Bt in the mosquito management program in Mizoram is highlighted based on the findings.