Keragaman dan Potensi Selulolitik Bakteri dan Jamur dari Saluran Pencernaan Oryctes rhinoceros L.

Abstract

The gut of insects is the natural habitat of various microbes that have the ability to degrade cellulose, which is the main component of plant walls. Oryctes rhinoceros L., is one of the pests in oil palm and is thought to have potential cellulolytic bacteria and fungi in the digestive tract that have not been widely explored. This study aims to uncover the diversity and potential of cellulolytic bacteria and fungi from the gut of O. rhinoceros L. as a new source of enzymes. Isolation of bacteria and fungi is conducted by the spread plate method using the medium of Nutrient Agar (NA) for bacteria and Potato Dextrose Agar (PDA) for fungi. Screening of cellulolytic activity was conducted using the selective medium of Carboxymethyl Cellulose (CMC) 1%. Identification of isolates could potentially be conducted based on morphological characters as well as molecular analysis of the 16S rRNA gene for bacteria and ITSrDNA for fungi. The cellulase enzyme was purified using gel filtration chromatography and ion exchange chromatography using DEAE-cellulose. Enzyme characterization includes molecular weight analysis with SDS-PAGE, measurement of specific activity, pH, optimal temperature, and the influence of metal ions on enzyme activity. A total of 42 bacterial and fungal isolates were successfully isolated from the digestive tract of instar III larvae and O. rhinoceros L. beetles, consisting of 19 bacterial isolates and 23 fungal isolates. Based on the Shannon and Simpson index, it shows that the diversity of bacteria and fungi in the gut of O. rhinoceros L. is high with a Shannon index value of 1.768-1.979 and a Simpson index of 0.826-0.847. Four isolates showing the highest cellulolytic activity index were identified as Bacillus tequilensis B01L (1.28) and Fusarium oxysporum F05L (0.90) isolated from the gastrointestinal tract of instar III larvae. Bacillus cereus B19L (2.50) and Aspergillus niger KOr15J (1.93) from the digestive tract of beetles. Pure cellulase yields a molecular weight of 52.23 kDa for bacterial enzymes and 72.61 kDa for fungal enzymes obtained from ion exchange chromatography. The highest specific activity was shown by the cellulase enzyme of B. cereus B19L at 9.221 U/mg, followed by B. tequilensis B01L (8.529 U/mg), F. oxysporum F05L (8.077 U/mg), and A. niger KOr15J (4.242 U/mg). The cellulase activity of the bacteria shows optimum at pH 6– 7 and pH 5–6 for fungi, an optimum temperature of 30°C to 60°C. Ca²⁺ ions increase the activity of cellulase enzymes from bacteria, while Fe²⁺ ions increase cellulase activity from fungi. This study shows that bacteria and fungi from the digestive tract of O. rhinoceros L. have high diversity and great potential in the stable and efficient production of cellulase enzymes. This potential also shows the alpha ability to degrade filterpaper, making it very prospective for the bioconversion of cellulose waste.