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Title:	Isolation, characterisation and antimicrobial activity determination of bacteriocin from a marine bacillus isolate
Authors:	Fatemeh Shayesteh (P51222)
Supervisor:	Gires Usup, Prof. Dr.
Keywords:	Antimicrobial peptide Marine bacillus Bacteriocins
Issue Date:	15-Aug-2017
Description:	This study was carried out to isolate marine bacteria that produce antimicrobial peptide active against the biofilm-producing pathogen, Candida albicans ATCC 10231. One particular isolate that was obtained from the marine carpet clam Paphia textile was found to be a potent producer of bacteriocin with strong activity against C. albicans ATCC 10231 and several human and food borne pathogens. Based on biochemical tests, 16S rRNA gene sequencing, fatty acids analysis and carbon source utilisation kit, this isolate is proposed as a new species of Bacillus cereus group. This bacterium exhibited optimum bacteriocin production in minimal medium containing 2% trypton, 1% glucose, and 2% NaCl at pH 8.0, 30 ˚C and 200 rev/min aeration. Bacteriocin activity in the culture supernatant was detectable from late exponential phase, reached a peak at mid stationary phase and declined soon after. Purification of bacteriocin was achieved with ammonium sulfate precipitation followed by gel filtration chromatography. As revealed by SDS-PAGE, the active fraction showed a protein with an approximate molecular weight of 11 KDa. Profile analysis of this bacteriocin by electrospray mass spectrometry (LC/MS/MS) showed 43% protein sequence coverage with a putative uncharacterized protein from Bacillus cereus strain B4264. This bacteriocin was sensitive to trypsin but resistant to proteinase K, pepsin, lysozyme, lipase, catalase and α-amylase. It was heat-stable, surviving at 121 ˚C under pressure and active over a pH range of 2.0 - 12.0. It was also stable and active in the presence of different surfactants, solvents and heavy metals. The CC50 value of bacteriocin was to be 32 μg/mL. The bacteriocin was active against biofilm producing bacteria Enterobacter cloacae UCCa2, Escherichia coli UCCb1, Proteus mirabilis UCDe2, and E. coli UCAa4 isolated from urinary catheter. Additionally, the bacteriocin was able to inhibit the formation of biofilm and disrupt established biofilm of these isolates. Observation of biofilm by scanning electron microscopy showed that the bacteriocin decreased biofilm thickness as well as altering cell morphology. Bacteriocin displayed a bactericidal mode of action against C. albicans ATCC 10231 leading to loss of cell viability, efflux of UV absorbing materials, K+ ions, inorganic phosphate and ATP. Furthermore, scanning and transmission electron microscopy observations of treated cells indicated severe modification in cell morphology with a concomitant lysis.,Certification of Master's/Doctoral Thesis" is not available
Pages:	270
Call Number:	QR92.B3S533 2017 tesis
Publisher:	UKM, Bangi
Appears in Collections:	Faculty of Science and Technology / Fakulti Sains dan Teknologi

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