Faizal bin Abdul Hamid2024-10-092024-10-092008https://studentrepo.iium.edu.my/handle/123456789/11376Sea cucumbers have been shown to produce antibacterial agents. This research tested the hypothesis that the antibacterial agents were produced by bacteria indigenous to the sea cucumber, as part of a symbiotic relationship. Research was done using 2 specimens of Holothuria tubulosa and a total of 23 bacterial strains were isolated from 5 zones of the sea cucumber and also from the sea-water as a control. All strains were grown using brain-heart agar and broth prepared with sterile sea-water. From the 23 strains, seven strains were selected for further analysis. These 7 were identified as J1:2 = K. pneumoniae, J2:2 = B. subtillis, J2:3 = B. licheniformis, J2:6 = B. licheniformis, J2:7 = K. pneumoniae, J2:10 = B. subtillis and J1:12 = K. pneumoniae, based on PCR 16S rRNA gene specific forward and reverse primers, and subsequent sequencing of the approximately 700bp products. Selection was based on antibacterial well diffusion testing results, with the 7 selected strains having higher and more consistent antibacterial activity in comparison to the other isolated strains from H. tubulosa. In turbidometric analysis, all 7 strains’ extracts delayed the exponential growth phase of test pathogens K. pneumoniae and S. typhimurium compared to the negative control (negative control = approx. 120 minutes, test samples = approx. 200 minutes). Size exclusion chromatography using Sephadex G-25 (1-5kDa separation) was carried out on all 7 strains. A hundred 1ml fractions were collected for each strain and absorbance readings were taken at 600nm. Two major peaks were seen in the chromatograms for all 7 strains, the first between fractions 20 to 40 (5kDa range) and the second at fractions 50 to 70 (1kDa range). SDS-PAGE analysis on fractions 27-33 of bacterial strain J2:2 (B. subtilis strain) showed 3 peptide bands, X (MW = 20413), Y (MW = 18742) and Z (MW = 5792). The results from this research showed that is possible that antibacterial agents found in sea cucumbers are made by microflora indigenous to the sea cucumber.enCopyright International Islamic University MalaysiaBacteriology -- Cultures and culture media -- MalaysiaAntibacterial agents -- MalaysiaHolothurians -- MalaysiaMaster Thesishttps://lib.iium.edu.my/mom/services/mom/document/getFile/hHQqkqBBxG716Qdf7IdNnSy1RH17dcAT20090518095235000