Six subspecies of Bacillus thuringiensis Berliner (Bt) were grown in minimal medium with chitin as the sole carbon source for 6 days to obtain Bt cell-free fermented broths, which were then evaluated for chitinolytic activity and tested against third-instar Aedes aegypti (L.) (Diptera: Culicidae) larvae. Bt Pakistani showed the highest chitinolytic activity (approximately >2700 mU/mL), Bt kurstaki showed the lowest activity (approximately <2000 mU/mL), and Bt thompsoni, Bt aizawai, Bt israelensis, and Bt alesti showed intermediate activities (approximately 2100–2400 mU/mL). Bt aizawai and Bt thompsoni broths showed the highest toxicity (LC50) against third-instar A. aegypti larvae (approximately <290 mU/mL). Bt kurstaki broth showed the lowest toxicity (approximately 420 mU/mL), while Bt pakistani, Bt israelensis, and Bt alesti broths showed intermediate toxicities (approximately 360–460 mU/mL). A purified and biochemically characterized Bt aizawai chitinase and commercial chitinases (from Serratia marcescens Bizio and Streptomyces griseus Waksman and Henrici) were evaluated and compared for synergistic effects on Bt israelensis spore-toxin complex against third-instar A. aegypti larvae. The synergism factor value of Streptomyces griseus and Bt aizawai chitinases were >2 and approximately 1.4; synergism was not evident for the Serratia marcescens chitinase (synergism factor value approximately 0.9).