Chitinase Production from Locally Isolated Bacillus cereus GS02 from Chitinous Waste Enriched Soil
Garima Dukariya
School of Biotechnology, Devi Ahilya University, Khandwa Road, Indore, 452001, India.
Anil Kumar *
School of Biotechnology, Devi Ahilya University, Khandwa Road, Indore, 452001, India.
*Author to whom correspondence should be addressed.
Abstract
Background and Objective: Chitin is world’s second most abundant structural carbohydrate in nature and is found as a structural component in the cell wall of fungi and exoskeletons of invertebrates. During senescence, it is degraded by the enzyme, chitinase. In addition, chitinase has been exploited for various commercial applications such as control of insects and fungal pathogens in order to protect the crops, waste management, cosmetics and food industries. Chitinases have been found to be widely distributed in various organisms including viruses, animals, bacteria, fungi, higher plants and insects. In the present study, various soil samples enriched in chitinous waste were screened for the isolation of bacteria capable of producing chitinase.
Methodology: Chitinase producing bacteria were isolated using serial dilution plating technique onto different agar media. Primary and secondary screening were performed and the isolate producing maximum chitinase was selected for biochemical identification and 16s rRNA sequencing. The secretion of extracellular chitinase by this strain was optimized with respect to pH, temperature, incubation time, substrate concentration, carbon and nitrogen sources and inoculum size. All these components were optimized using OFAT (one factor at a time) approach.
Results: A total of 29 bacterial isolates were found exhibiting secretion of extracellular chitinase as determined using zones of clearance. Based on the area of zone of clearance, six isolates were selected for secondary screening and the most potent isolate was identified as Bacillus cereus. The maximum chitinase production by this strain was obtained at 37°C and pH 7.0 after 48 hours of incubation. The maximum chitinase secretion was observed on addition of 1% colloidal chitin and 0.05% yeast extract in the medium.
Keywords: Chitinases, chitin, isolation, Bacillus cereus, 16S rRNA, media optimization.