Isolation and molecular characterization of cellulase-producing bacteria from waste dump site

Abstract

Cellulases are collections of extracellular enzymes and a complex mixture of enzyme proteins with various specificities. Cellulases work together to hydrolyze glycosidic bonds and generate monomers of glucose for fermentation. This investigation aims to isolate and molecularly characterize Bacillus species with cellulolytic ability. Bacillus species were isolated from soil samples retrieved from a dump site and screened for cellulolytic ability on carboxyl methylcellulose (CMC) agar. The percentage hydrolysis efficiency of isolates was determined and cellulase produced was quantified using CMC assay method. Biochemical identification was by Analytical Profile Index (API) Kit 50CHB/20E and API web software while molecular characterization employed 16S rRNA gene sequencing and blast search analysis. Bacillus megaterium (FSP1) and Bacillus zanthoxyli (FSP4) exhibited their cellulolytic potentials by presenting zones of clearance of about 21 ± 2.08 and 7 ± 1.00 mm on CMCA with hydrolysis efficiency of 250 and 600 % respectively. Quantification of crude cellulase revealed cellulase activity of 85 and 74μmol/ml for both bacteria species. Biochemically, the cellulolytic bacteria were identified as Bacillus megaterium (FSP1) and Bacillus zanthoxyli (FSP4) while molecularly, they were identified as Bacillus megaterium 14581 (FSP1) and Bacillus zanthoxyli 1433 (FSP4) with Reference Sequence (RefSeq) accession numbers NR_116873 and NR_164882, and showing maximum sequence similarity of 99 and 96 % respectively. Results obtained from this investigation, suggests that both bacteria species characterised, possesses good cellulolytic ability and hence can be utilized for the production of the enzyme cellulase which has a wide range of industrial application.