Construction of a genetically modified T7Select phage system to express the antimicrobial peptide 1018

David J. Lemon; Matthew K. Kay; James K. Titus; April A. Ford; Wen Chen; LCDR Nicholas J. Hamlin; Yoon Y. Hwang

doi:10.1007/s12275-019-8686-6

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Construction of a genetically modified T7Select phage system to express the antimicrobial peptide 1018: David J. Lemon , Matthew K. Kay , James K. Titus , April A. Ford , Wen Chen , LCDR Nicholas J. Hamlin , Yoon Y. Hwang; Journal of Microbiology 2019;57(6):532-538.
DOI: https://doi.org/10.1007/s12275-019-8686-6
Published online: May 27, 2019

Craniofacial Health and Restorative Medicine, Naval Medical Research Unit San Antonio, Fort Sam Houston, TX, 78234, USA

Corresponding author: Yoon Y. Hwang , Tel: +1-210-539-6263,

Received: 12 December 2018 • Revised: 10 January 2019 • Accepted: 29 January 2019

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Abstract

Bacteriophage therapy was an ascendant technology for combating bacterial infections before the golden age of antibiotics, but the therapeutic potential of phages was largely ignored after the discovery of penicillin. Recently, with antibioticresistant infections on the rise, these phages are receiving renewed attention to combat problematic bacterial infections. Our approach is to enhance bacteriophages with antimicrobial peptides, short peptides with broad-spectrum antibiotic or antibiofilm effects. We inserted coding sequences for 1018, an antimicrobial peptide previously shown to be an effective broad-spectrum antimicrobial and antibiofilm agent, or the fluorescent marker mCherry, into the T7Select phage genome. Transcription and production of 1018 or mCherry began rapidly after E. coli cultures were infected with genetically modified phages. mCherry fluorescence, which requires a 90 min initial maturation period, was observed in infected cultures after 2 h of infection. Finally, we tested phages expressing 1018 (1018 T7) against bacterial planktonic cultures and biofilms, and found the 1018 T7 phage was more effective than the unmodified T7Select phage at both killing planktonic cells and eradicating established biofilms, validating our phage-driven antimicrobial peptide expression system. The combination of narrow-spectrum phages delivering relatively high local doses of broad-spectrum antimicrobials could be a powerful
method
to combat resistant infections. The experiments we describe prove this combination is feasible in vitro, but further testing and optimization are required before genetically modified phages are ready for use in vivo.

Keywords: T7 phage;genetic modification;antimicrobial peptide;antibiotic-resistant infection;peptide 1018;mCherry

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Construction of a genetically modified T7Select phage system to express the antimicrobial peptide 1018

J. Microbiol. 2019;57(6):532-538. Published online May 27, 2019

DOI: https://doi.org/10.1007/s12275-019-8686-6

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