THERMOSTABLE-CELLULASE PRODUCING ANOXYBACILLUS RUPIENSIS AND ROSEOMONAS SP. ISOLATED FROM SAWDUST

Abstract

Mesophilic enzymes are easily denatured by the heat usually generated in industrial processes, hence the need to search for thermophilic bacteria for production of this important enzyme. Sawdust samples were collected from sawmills in Ede, Osun State, Nigeria. Bacteria were isolated, identified and screened for cellulase production using standard microbiological procedures. Production of the enzyme was optimized using different carbon and nitrogen sources, temperature, pHs. Three Isolates with significantly high cellulase production were selected and identified by PCR amplification of 16SrDNA genes. Data were analysed using descriptive statistic and ANOVA at p= 0.05. Roseomonas sp. (A1), Anoxybacillus rupiensis E1 and Anoxybacillus rupiensis 5H were the best thermostable cellulase producers.Anoxybacillus rupiensis (5H) in medium with 1% carboxymethylcellulose produced 9.22 U/mL of cellulase while Anoxybacillus rupiensis (E1) produced 3.0 U/mL of cellulase in medium supplemented 1% (w/v) tryptone. Analysis of 16S rDNA indicated that the isolates E1 and 5H were genetically the same but different from A1. DNA sequencing of E1 and 5H have 96% and 100% similarities respectively with the sequences within the GenBank. The similarity match correspond to Anoxybacillus rupiensis AM 988776 strain for E1 and Anoxybacillus rupiensis AM 988775 for 5H. A1 has 100% similarity with Roseomonas alkaliterrae NR 134161. Optimum production of crude cellulase was at 50oC and pH of 7, while optimum activity of the enzyme was at 600C for A1 and pH of 9 for 5H. These isolates can be exploited for the production of thermostable cellulase of industrial importance.