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United States Department of Agriculture

Agricultural Research Service

Research Project: MOLECULAR MICROBIOLOGY AND CONTROL OF ENTERIC PATHOGENS THAT CONTAMINATE FRESH PRODUCE Title: An Engineered R-Type Pyocin Is a Highly Specific and Sensitive Bactericidal Agent for the Food-Borne Pathogen Escherichia coli O157:H7

Authors
item Scholl, D -
item Cooley, Michael
item Williams, S -
item Gebhart, D -
item Martin, D -
item Bates, Anne
item Mandrell, Robert

Submitted to: Antimicrobial Agents and Chemotherapy
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 27, 2009
Publication Date: July 1, 2009
Citation: Scholl, D., Cooley, M.B., Williams, S.R., Gebhart, D., Martin, D., Bates, A.H., Mandrell, R.E. 2009. An Engineered R-Type Pyocin Is a Highly Specific and Sensitive Bactericidal Agent for the Food-Borne Pathogen Escherichia coli O157:H7. Antimicrobial Agents and Chemotherapy. 53(7):3074-3080.

Interpretive Summary: Some strains of Pseudomonas aeruginosa produce R-type pyocins, which are high molecular weight phage tail-like protein complexes that have bactericidal activity against other Pseudomonas strains. These particles recognize and bind to bacterial surface structures via tail fibers; their primary spectrum determinant. R-type pyocins kill the cell by contracting a sheath-like structure and inserting their hollow core through the cell envelope, resulting in dissipation of the cellular membrane potential. We have re-targeted an R-type pyocin to E. coli O157:H7 by fusing a "tail spike" protein from an O157-specific phage, phiV10, to the pyocin tail fiber. The phiV10 tail spike protein recognizes and degrades the O157 lipopolysaccharide. This engineered pyocin, termed AVR2-V10, is sensitive and specific, killing 100% of diverse E. coli O157:H7 isolates but no other serotypes tested. AVR2-V10 can kill E. coli O157:H7 on beef surfaces, making it a candidate agent for eliminating this pathogen from food products. All rare AVR2-V10-resistant mutants isolated and examined have lost the ability to produce O157 antigen and are expected to have compromised virulence. In addition, E. coli O157:H7 exposed to and killed by AVR2-V10 do not release shiga toxin, as is often the case with many antibiotics, suggesting potential therapeutic applications. The demonstration that a novel R-type pyocin can be created in the laboratory by fusing a catalytic Podoviridae tail spike to a Myoviridae tail fiber is evidence that the plasticity observed among bacteriophage tail genes can, with modern molecular techniques, be exploited to produce non-natural, targeted antimicrobial agents.

Technical Abstract: Some strains of Pseudomonas aeruginosa produce R-type pyocins, which are high molecular weight phage tail-like protein complexes that have bactericidal activity against other Pseudomonas strains. These particles recognize and bind to bacterial surface structures via tail fibers; their primary spectrum determinant. R-type pyocins kill the cell by contracting a sheath-like structure and inserting their hollow core through the cell envelope, resulting in dissipation of the cellular membrane potential. We have re-targeted an R-type pyocin to E. coli O157:H7 by fusing a "tail spike" protein from an O157-specific phage, phiV10, to the pyocin tail fiber. The phiV10 tail spike protein recognizes and degrades the O157 lipopolysaccharide. This engineered pyocin, termed AVR2-V10, is sensitive and specific, killing 100% of diverse E. coli O157:H7 isolates but no other serotypes tested. AVR2-V10 can kill E. coli O157:H7 on beef surfaces, making it a candidate agent for eliminating this pathogen from food products. All rare AVR2-V10-resistant mutants isolated and examined have lost the ability to produce O157 antigen and are expected to have compromised virulence. In addition, E. coli O157:H7 exposed to and killed by AVR2-V10 do not release shiga toxin, as is often the case with many antibiotics, suggesting potential therapeutic applications. The demonstration that a novel R-type pyocin can be created in the laboratory by fusing a catalytic Podoviridae tail spike to a Myoviridae tail fiber is evidence that the plasticity observed among bacteriophage tail genes can, with modern molecular techniques, be exploited to produce non-natural, targeted antimicrobial agents.

Last Modified: 10/21/2014
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