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Title: Isolation and Purification of Enteroicin E-760 with a Broad Antimicrobial Activity Against Gram-positive and Gram-negative Bacteria (AAC01569-06 Version 3)

Author
item Line, John
item SVETOCH, E - STATE DEPT RUSSIA
item ERUSLANOV, B - STATE DEPT RUSSIA
item PERELYGIN, V - STATE DEPT RUSSIA
item MITSEVICH, E - STATE DEPT RUSSIA
item MITSEVICH, I - STATE DEPT RUSSIA
item POKHILENKO, V - STATE DEPT RUSSIA
item LEVCHUK, V - STATE DEPT RUSSIA
item SVETOCH, O - STATE DEPT RUSSIA
item Seal, Bruce
item Siragusa, Gregory
item Stern, Norman

Submitted to: Antimicrobial Agents and Chemotherapy
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/9/2007
Publication Date: 3/1/2008
Citation: Line, J.E., Svetoch, E.A., Eruslanov, B.V., Perelygin, V.V., Mitsevich, E.V., Mitsevich, I.P., Pokhilenko, V.D., Levchuk, V.P., Svetoch, O.E., Seal, B.S., Siragusa, G.R., Stern, N.J. 2008. Isolation and Purification of Enteroicin E-760 with a Broad Antimicrobial Activity Against Gram-positive and Gram-negative Bacteria (AAC01569-06 Version 3). Antimicrobial Agents and Chemotherapy. 52:1094-1100.

Interpretive Summary: Campylobacter jejuni is a Gram-negative human food-borne pathogen of primary importance. Poultry are frequently contaminated with C. jejuni during production with the majority of commercial U.S. flocks positive for the organism by the time the birds reach market age at about 6 weeks. There are currently few effective on-farm interventions for reducing colonization of poultry with C. jejuni. There is a need for effective intervention that may be practically applied in the poultry industry to reduce colonization of poultry with C. jejuni and subsequently reduce consumer exposure to this pathogen. In this paper we report the isolation and purification of a 5,362 Da bacteriocin produced by an Entercoccus species isolated from chicken ceca with broad spectrum activity against both Gram-positive and Gram-negative bacteria and demonstrate the ability of the purified bacteriocin to reduce colonization of poultry by Campylobacter spp. Administration of bacteriocin E-760 treated feed significantly (P< 0.05) reduced colonization of young broiler chicks experimentally challenged with two strains of C. jejuni by more than 8 log10 CFU. Bacteriocin E-760 also significantly (P<0.05) reduced colonization of naturally acquired Campylobacter spp. in market age broilers when administered in treated feed for four days prior to analysis. This information will be useful to researchers in government, academia and the poultry industry. Bacteriocin E-760 could be developed as a practical on-farm intervention for reducing consumer exposure to pathogenic campylobacter.

Technical Abstract: Strain NRRL B-30745, isolated from chicken ceca, and identified as an Enterococcus spp. of the durans/faecium/hirae group, produces a 5362 Da bacteriocin that inhibits the growth of Salmonella enteritidis, S. choleraesuis, S. typhimurium, S. gallinarum, Escherichia coli O157:H7, Yersinia enterocolitica, Citrobacter freundii, Klebsiella pneumoniae, Shigella dysenteriae, Pseudomonas aeruginosa, Proteus mirabilis, Morganella morganii, Staphylococcus aureus, S. epidermidis, Listeria monocytogenes, Campylobacter jejuni and 20 other Campylobacter spp. isolates. The bacteriocin, E-760, was isolated and purified by cation exchange and hydrophobic interaction chromatography. The proteinaceous nature of purified bacteriocin E-760 was demonstrated upon treatment with various enzymes. The antimicrobial peptide was found to be sensitive to Beta-chymotrypsin, proteinase K and papain, and resistant to lysozyme and lipase. The bacteriocin demonstrated thermostability by retaining activity after 15 min at 100oC. It was stable at pH values between 5.0 and 8.7, but activity was lost at pH 3.0 and above pH 9.5. Administration of bacteriocin E-760 treated feed significantly (P< 0.05) reduced colonization of young broiler chicks experimentally challenged with two strains of C. jejuni by more than 8 log10 CFU. Bacteriocin E-760 also significantly (P<0.05) reduced colonization of naturally acquired Campylobacter spp. in market age broilers when administered in treated feed for four days prior to analysis.