Enhanced production of chitinase from Bacillus cereus V5331 and evaluation of anti-candidal potential of enzyme: central composite design approach.

A chitinase-producing bacterium was isolated from a dumping ground soil sample and identified as Bacillus cereus V5331 based on 16S rRNA gene sequencing. The production of chitinase was enhanced through the application of solid-state fermentation (SSF). The effect of variables like pH, temperature, swollen chitin (SC) concentration, moisture content, inoculum size and incubation period were assessed by using a one-variable-at-a-time (OVAT) approach. A central composite design (CCD) using response surface methodology (RSM) was subsequently employed with the chitin concentration, moisture ratio, incubation period and inoculum size as found most effective independent variables. Optimization resulted in a 1.84 fold increase in chitinase production by meeting the parameters: 0.2% chitin, 1:1 moisture ratio, 1% inoculum and 120 h of incubation at 37 °C and pH 7 ± 0.2. The scanning electron microscopy (SEM) studies revealed that chitinase showed significant antifungal action against Candida albicans, causing cell wall disintegration, cell wall inhibition (1.5 cm) and substantial biofilm degradation (53.56%). Chitin breakdown was further confirmed by Fourier transform infrared spectroscopic (FTIR) analysis and p-DMAB assay of the enzyme-treated yeast, which showed the release of 211 µg mL-1 of N-acetylglucosamine (NAG). The current study demonstrates a newly isolated bacterium, which is non-hemolytic (gamma- hemolytic) and producing chitinase constitutively as well as in the presence of chitin in cost-effective locally available media. Further, its anti-fungal potential against C. albicans makes it a promising candidate for future studies to develop an antifungal topical therapeutic agent.