Due to their broad-spectrum and bactericidal activity, antimicrobial proteins and peptides (AMPPs) are considered as future drug alternatives to combat the escalating problem of antimicrobial resistance in medicine. The community of competitive culturable and non-culturable bacteria in the rumen present a potential source for the discovery of novel bioactive compounds including AMPPs. Metagenomic and bioinformatics based techniques were used to prospect two rumen bacterial metagenomes for potentially novel antimicrobial genes (proteins) and peptide sequences. Novel short antimicrobial peptides (peptides 1-181) and eleven longer antimicrobial genes/miniproteins (Gene 6, 17A, 17B palG1 and palG2, H-G1, H-G2, H-G3, H-G4, H-G5) were identified. Eight of these (peptides 2, 3, 4, 5, 7, 8, 15 and palG1) were selected for further analysis. These antimicrobials displayed potent antimicrobial activity (minimum inhibitory concentrations ranging from 32 to 64 μg/ml) against both Gram positive bacteria strains (including Methicillin sensitive and resistant Staphylococcus aureus strains MSSA RN4220 and EMRSA-15, Enterococcus faecalis JH2-2 and Listeria monocytogenes NCTC 11994 (serovar 4b)), as well as Gram negative bacteria strains (Escherichia coli K12, Salmonella enterica serovar Typhimurium SL1344 and Pseudomonas aeruginosa (15692) PAO1 strain H103) in Mueller Hinton broth. No haemolytic activity against red blood cells was seen. Data obtained indicate that loss of cell viability is due to cytoplasmic leakage and there is some evidence of interference with the cell division mechanism. The rumen AMPPs identified in this study show great activity against clinically relevant human pathogens and to our knowledge are the first rumen AMPs identified using metagenomics. Overall, the data support the potential use of AMPs (2, 3, 7), AMPs (2, 3, 4, 5, 7, 8, 15) and the polypeptide palG1 in the treatment of S. aureus, E. coli and Ent. faecalis infections respectively in the future.
|Date of Award||01 Nov 2015|
|Supervisor||Sharon Huws (Supervisor) & Christopher Creevey (Supervisor)|
Prospecting Rumen Bacteria for Novel Antimicrobials
Oyama, L. B. (Author). 01 Nov 2015
Student thesis: Doctoral Thesis › Doctor of Philosophy