Bacillus subtilis chitinase identified by matrix-assisted laser desorption/ionization time-of flight/time of flight mass spectrometry has insecticidal activity against Spodoptera litura Fab

Abstract

An extracellular chitinase was identified and purified (CS1 and CS2) from Bacillus subtilis. The 16S rRNA sequencing was submitted in GenBank (accession numbers KC336487 and KC412256). The purified crude enzymes were identified through matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF MS) analysis. The peptide sequences were matched with chitinase sequences. The peak m/z with 1297. 592 and 3094.570 mascot search resulted sequence was blasted with NCBI protein sequences and confirmed that it is a chitinase enzyme. The effects of chitinase on gut enzymes lactate dehydrogenase, acid phosphatase, alkaline phosphatase and adenosine triphosphatase of the tobacco cutworm Spodoptera litura larvae were investigated. At all concentrations tested, chitinase decreased the activities of these gut enzymes relative to the control. When chitinase treated leaves were fed to larvae in bioassays, gut tissue and gut enzymes were affected. The histological study clearly shows the chitinase treated larval gut, peritrophic membrane and epithelial cells were affected significantly. Chitinase isolated from B. subtilis has effectively reduced the gut enzyme activity and growth of S. litura. The chitin based bioformulation may serve as an effective biocide against the polyphagous pest like S. litura.

Keywords: Chitinase; Gut enzymes; Histology; MALDI-TOF MS; Spodoptera litura.