title: Cloning of genes encoding colicin E2 in Lactococcus lactis subspecies lactis and evaluation of the colicin-producing transformants as inhibitors of Escherichia coli O157:H7 during milk fermentation.
abstract: Colicin E2 (ColE2) is a proteinaceous bacterial toxin produced by some strains of Escherichia coli and other members of the Enterobacteriaceae that exhibits inhibitory activity against some strains of E. coli O157:H7. A 2.0-kb DNA fragment, containing the ColE2 structural gene ceaB and immunity gene ceiB from E. coli NCTC 50133 (pColE2-P9), was cloned into the lactococcal plasmid vector pNZ2103. The lysis gene, celB, was not cloned. The plasmid, pLR-E2, encoding the cloned genes was transformed into E. coli DH5α and Lactococcus lactis ssp. lactis LM0230 and PN-1 using electroporation. The bacteriocin ColE2 was expressed in transformants of both E. coli and L. lactis ssp. lactis. Secretion of ColE2 into media was verified by spot-on-lawn assays and measurement of ColE2 activity in the growth medium of transformants. The level of ColE2 produced by transformants containing pLR-E2 was similar to that produced by the parental strain, E. coli NCTC 50133 (pColE2-P9). Evaluation of a ColE2-producing transformant of L. lactis ssp. lactis as a starter culture revealed that, although ColE2 was produced by transformants and could be detected in milk during fermentation, the inhibitory activity of ColE2 against E. coli O157:H7 was significantly decreased in milk compared with buffered growth medium.
authors.affiliation: Department of Food Science, The Pennsylvania State University, University Park, PA 16802, USA.
authors.forename: S M
authors.initials: SM
authors.lastname: Liu
authors.forename: D M
authors.initials: DM
authors.lastname: Miller
authors.forename: R F
authors.initials: RF
authors.lastname: Roberts
mesh_terms: D000818:Animals; D003086:Bacteriocin Plasmids / Q000235:genetics; D003001:Cloning, Molecular; D003087:Colicins / Q000235:genetics*; D019453:Escherichia coli O157 / Q000378:metabolism*; D005285:Fermentation; D013294:Lactococcus lactis / Q000235:genetics*; D008892:Milk / Q000382:microbiology
chemical_list: D003087:Colicins