Thermophilin 109 is a naturally produced broad spectrum bacteriocin encoded within the blp gene cluster of Streptococcus thermophilus
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To demonstrate that S. thermophilus ST109 produces an antimicrobial peptide encoded within the bacteriocin-like peptide (blp) gene cluster, and to determine its broad spectrum activity against potential human pathogens.
Analysis of the cell free supernatant (CFS) revealed that antimicrobial activity was associated with the presence of a heat-stable peptide of approximately 5–6 kDa; and activity was lost after protease treatment or exposure to α-amylase. Deletion of blpC, which encodes a quorum sensing induction peptide, resulted in a loss of antimicrobial activity showing that thermophilin 109 was encoded within the blp gene cluster of ST109. Sequencing of the ST109 blp gene cluster showed 90% and 99% identity to clusters previously characterized in S. thermophilus strains LMD-9 and ST106, both of which are unable to naturally produce their bacteriocins. Real-time qPCR showed that blpC and blpD were expressed approximately 24 and 100-fold higher in ST109 as compared to strain LMD-9; and broad spectrum activity was demonstrated against lactobacilli, enterococci and the human pathogen Streptococcus pyogenes.
The high level of similarity observed between the ST109 and ST106 blp gene clusters at the nucleic acid level suggests bacteriocin expression may be regulated by factors encoded elsewhere on the chromosome. Activity against E. faecalis and S. pyogenes suggest S. thermophilus ST109 could be used for food safety and probiotic applications.
KeywordsStreptococcus thermophilus Bacteriocin Antimicrobial activity Lactic acid bacteria Quorum sensing
We would like to thank J. Hernandez, Temple University, for technical assistance in determining the antimicrobial spectra of thermophilin 109; and D. Needleman, USDA-ARS, for technical assistance for sequencing the ST109 blp gene cluster.
Supplementary Fig. 1—Genotype confirmation of the S. thermophilus ST109 blpC knock out mutant (ST109CKO). (A) PCR confirming the presence of both the erythromycin (erm; lane 1) and kanamycin (kan; lane 2) resistance genes on plasmid pSTKOC. (B) PCR analysis of strain ST109CKO, where both the erm (lane 1) and blpC (lane 3) were absent; and the kan gene was present (lane 2). blpC was successfully amplified from the parent ST109 culture (lane 4) to confirm proper function of the primer set. M: 1kb ladder.
Supplementary Table 1—Real-time PCR of blp components in S. thermophilus strains ST109, ST109CKO and LMD-9.
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