Diversity and functional annotation of chitinolytic Bacillus and associated chitinases from north western Indian Himalayas

Abstract

Chitinolytic Bacillus and associated chitinases have prospective implication in both industry and biological control of agricultural pests. A total of 83 chitinolytic Bacillus strains were isolated from native soils of Uttarakhand state, north western Indian Himalayas. A multiphasic characterization of the collection showed a vast diversity in bacterial kinetics (growth and enzyme production), pH stability in ten isolates, thermal-stability in 23 isolates and psychrotolerance in three isolates. The collection was characterized by chief existence of 148 kDa (65 isolates) and 152 kDa (7 isolates) chitinases, the high molecular weight enzymes. Partial sequencing of 16S rRNA and chi genes showed Paenibacillus species are the major chitinase producers of the region. In addition, licheniformis and circulans group chitinases were also found to be associated with the collection. The molecular variation in sequences revealed existence of different Paenibacillus species and especially a special evolutionary status of chitinases in phylogeny which is also evidenced by associated high molecular weight chitinases. The bioactivity of isolates against insect pests, Pieris brassicae and Helicoverpa armigera showed that the isolates were not lethal except at very high concentrations (108 cfu/ml) that to in only 9 isolates. However, they are primarily involved in growth reduction which led to identification of three prominent isolates (UKCH27, UKCH29 and UKCH77) that caused a significant larval weight reduction at lowest concentration tested (102 cfu/ml). Further testing of synergism between Cry toxins of B. thuringiensis strain HD 1 and these isolates showed near cent percent mortality of test insects at LC30 concentrations