Isolation, Screening and Identification of Cellulolytic Bacteria from Soil

Background: Cellulose is the most abundant carbohydrate on earth and is considered as a good candidate for production of second generation biofuel (ethanol) and many other products of routine use. For degradation, cellulases are used which are mostly secreted by microbes such as fungi. Cellulases also play an important role in senescence of plants and in host-parasite relationship for invading the plant cell wall. However, comparatively lesser studies have been carried out on cellulase producing bacteria. Therefore, present study was aimed to isolate cellulase (Endo-β-1,4-D-glucanase; EC. 3.2.1.4.) from bacterial sources.

Methodology: To isolate thermophilic/ mesophilic cellulase producing bacteria, soil samples were collected from wood furnishing area and agricultural farm around Indore. Besides, soil sample was also collected from the vicinity of Amlai Paper Mill in Budhar district, Madhya Pradesh. These soil samples after suitable dilutions were streaked on different nutrients agar petri-dishes having carboxymethyl cellulose (CMC) as an inducer. After screening, four colonies were isolated capable of producing good amount of cellulase. Screening was done using Congo red staining and confirmation was done after growth of the bacteria in liquid nutrient medium having CMC. These colonies individually were grown in suitable nutrient media having CMC as an inducer and enzyme activity was determined in the nutrient media after harvesting bacterial cells by centrifugation.

Results: The highest enzyme producing bacteria were identified as Bacillus lichenoformis and Ochrobactrum anthropi after biochemical analyses, 16S rRNA sequencing and subsequently phylogenetic tree analysis.