Improvement of Chitinase Production by Bacillus thuringiensis NM101-19 for Antifungal Biocontrol through Physical Mutation

Abstract

Chitinase is one of the most important enzymes due to its diversity and a variety of potential uses. This study is an attempt to enhance chitinase production for antifungal biocontrol by subjecting Bacillus thuringiensis NM101-19 strain grown on shrimp shell wastes to various doses of gamma irradiation. Six mutants (BM-4, BM-6, BM-8, BM-12, BM-15, and BM-17) obtained at gamma ray doses of 40, 60, 80, 100, 120 and 140 Gy, respectively produced higher levels of chitinolytic activities in comparision to the wild-type strain. The BM-15 mutant strain showed the highest chitinase production (65.41 U/mL) which was 2.60 times more than the wild type (25.11 U/mL). Biocontrol efficacy of the mutants was statistically superior to the wild-type strain against all tested phytopathogens. Random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) PCR techniques, with five primers for each, were used in order to detect the variation in DNA profile between the mutant and wild-type strains in response to gamma-radiation treatments. RAPD and ISSR analysis indicated the appearance and disappearance of DNA polymorphic bands at different gamma ray doses. The results confirmed that the mutagenesis technique is a potent strategy to enhance the chitinase activity for industrial and agricultural purposes.