Isolation, screening and characterization of cellulase producing bacteria from rumen fluids of ox and sheep samples

Abstract

Ruminant animals lack enzymes to break down fibrous feeds but they harbor different microorganisms capable of degrading their feeds. Rumen bacteria contribute a considerable part in the breakdown of cellulose biomass. The purpose of this study was to isolate, screen and characterize cellulase producing bacteria from rumen fluids of domestic ruminants and optimize their growth condition for cellulase production. Samples were collected from five fistulated oxen from Holetta agricultural research center and another sample were collected from three sheep and two goats from slaughter house at Holetta. A total of 85 pure isolates were collected and screened for cellulase activity by using Congo red stain on Carboxymethylcellulose (CMC) agar. Isolates which have shown maximum clear zone around the colony were characterized based on morphological and biochemical characteristics. These isolates were selected for enzyme production and optimization of the fermentation medium for maximum cellulase production. Dinitrosalisyclicacid (DNSA) method was used for measuring cellulase activity at 540 nm. Among all screened isolates, 26 bacterial isolates were found to hydrolyze CMC. From 26 cellulolytic bacterial isolates, 7 isolates have shown maximum clear zone around the colony. In this study, the optimum incubation time, carbon source, pH, temperature, and nitrogen source were found to be 72 hr., wheat straw, 7.0, 39ºC and Ammonium sulphate respectively. These cellulolytic bacterial isolates determine great potential for the study of enzymes in cellulose degradation and for improving the bioconversion of lignocellulose biomass. The current study showed that the 7 isolates were promising for cellulase production at their respective optimum parameters. From cheap lignocellulosic substrates, Wheat straw was found to be best digested substrate by cellulose degrading bacteria compare to Barley and Teff straw. Further characterization of the seven isolates at a molecular level and purification and detailed characterization of cellulase enzyme are recommended for effective utilization in feed industry.