The evaluation of Antibacterial Activity of Fungal Endophyte Ceratobasidium ramicola IBRLCM127 Colonizing in Rhizomes of Medicinal Plant, Curcuma mangga Valeton & Zijp

Abstract

The current study was conducted to evaluate the antibacterial activity of fungal endophyte isolate, Ceratobasidium ramicola IBRLCM127 colonizing in rhizomes of a medicinal plant, Curcuma mangga Valeton & Zijp. Primary screening of its antibacterial potential was performed by employing agar plug assay, and the results revealed that the fungal isolate was capable to inhibit all the 11 test bacteria used in the study. All four Gram-positive bacteria (Staphylococcus aureus, Bacillus cereus, Bacillus subtilis and Methicillin‑resistant Staphylococcus aureus ATCC 33591) showed the highest susceptibility degree to the fungal isolate with the size of inhibition zones of ≥21 mm. As for Gram-negative bacteria, 6 out of 7 tested bacteria (Proteus mirabilis, Yersinia enterocolitica, Escherichia coli IBRL 0157, Salmonella typhimurium, Klebsiella pneumoniae ATCC 13883 and Acinetobacter antratus) were the most susceptible species with inhibition zone size of ≥21 mm, whilst only Pseudomonas aeruginosa ATCC 27844 less susceptible to fungal isolate with the size of inhibition zone of 11 to≤20 mm. Secondary screening using disc diffusion assay revealed that fungal ethyl acetate extract derived from fermentative broth (extracellular) demonstrated better potential of antibacterial activity as compared to the methanol extract derived from fungal biomass (intracellular). The results showed that 8 out of 11 test bacteria were susceptible to fungal ethyl acetate extract with the diameter of inhibition zone ranging from 8.0±0.0 mm to 13.0±0.0 mm and 8.3±0.6 mm to 11.7±0.6 mm in Gram-positive and Gram-negative bacteria, respectively. Contradictorily, methanol extract only capable to inhibit Staphylococcus aureus with inhibition zone of 8.3±0.6 mm in diameter. The minimal inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values of ethyl acetate extract towards Gram-positive bacteria were both in the range of 125.00 to 500.00 µg/mL, whilst for Gram-negative bacteria were in the range of 125.00 to 250.00 and 250.00 to 500.00 µg/mL, respectively. On the other hand, the MIC and and MBC values for methanol extract towards Gram-positive bacteria were 250.00 and 500.00 µg/mL, respectively. Both ethyl acetate and methanol extracts exerted bactericidal effects on test bacteria with the ratio of MBC/MIC ≤4. The detail of the ethyl acetate extract effects on the bacterial cells was observed using scanning electron microscopy (SEM), in which the micrographs obtained from SEM revealed that the severity of the cell damages caused by the extract were beyond repair, and the mode of action could possibly due to the disruption in the cell wall biosynthesis and cell membrane permeability.