EVALUATION OF CHITINOLYTIC GUT MICROBIOTA IN SOME CARPS AND OPTIMIZATION OF CULTURE CONDITIONS FOR CHITINASE PRODUCTION BY THE SELECTED BACTERIA

Abstract

Present study was aimed at isolation of autochthonous chitinase-producing bacteria from the gastrointestinal tracts of 3 Indian Major Carps (Labeo rohita, Catla catla, Cirrhinus mrigala) and 3 exotic carps (Hypophthalmichthys molitrix, Ctenopharyngodon idella, Cyprinus carpio). Altogether, 119 bacteria were isolated from both the proximal and distal intestine and screened for chitinolytic activity. On the basis of chitin hydrolysis zone, 63 isolates were primarily selected for chitinase production, from which 34 potent strains were further studied for quantitative enzyme assay. Amongst them, the strains HMH1 and CMF2 exhibited potent chitinolytic activity and were identified as Bacillus pumilus (KF454036) and Bacillus flexus (KF454035), respectively by 16S rRNA partial gene sequence analysis. Optimization of various fermentation parameters (e.g., temperature, pH, inoculums size, surfactant, colloidal chitin concentration, incubation time, carbon sources, organic and inorganic nitrogen sources) were carried out in chitinase production medium. Incubation for 72 h at 350C and initial pH 7.5 revealed optimum chitinase productions by B. pumilus HMH1 in the media supplemented with colloidal chitin 0.1% (w/v), maltose 2% (w/v), ammonium sulphate 1.0% (w/v) and Tween-80 0.2% (v/v). However, B. flexus CMF2 required 48 h incubation at 35°C and initial pH 8.0 with colloidal chitin 0.15% (w/v), sucrose 1% (w/v), yeast extract 2.0% (w/v) and Tween-20 0.2% (v/v) supplementation for optimum yield. The results indicate that there is ample scope for further research to appraise fish gut microorganisms for chitinase production or as probiotics to improve feed efficiency in fish.