A Phage Foundry Framework to Systematically Develop Viral Countermeasures to Combat Antibiotic-Resistant Bacterial Pathogens

Mutalik, Vivek K.; Arkin, Adam P.

doi:10.1016/j.isci.2022.104121

Cited by 21 publications

(13 citation statements)

References 269 publications

(358 reference statements)

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“…59 Key to this will be the creation of phage biofoundries that aim to fill the knowledge and technological gaps in phage therapeutics. 60 By combining such coordinated efforts with cutting-edge synthetic biology, rational therapeutic design, and automation, generally applicable tools for rapid phage engineering can be built to enable the next generation of phage therapies.…”

Section: Resultsmentioning

confidence: 99%

“…Finally, the cost-effective and rapid production of future phage therapies will require centralized infrastructure pertaining to the academic understanding of phages, their clinical use, and methods for their engineering into effective therapies . Key to this will be the creation of phage biofoundries that aim to fill the knowledge and technological gaps in phage therapeutics . By combining such coordinated efforts with cutting-edge synthetic biology, rational therapeutic design, and automation, generally applicable tools for rapid phage engineering can be built to enable the next generation of phage therapies.…”

Section: Resultsmentioning

confidence: 99%

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SpyPhage: A Cell-Free TXTL Platform for Rapid Engineering of Targeted Phage Therapies

et al. 2022

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The past decade has seen the emergence of multidrug resistant pathogens as a leading cause of death worldwide, reigniting interest in the field of phage therapy. Modern advances in the genetic engineering of bacteriophages have enabled several useful results including host range alterations, constitutive lytic growth, and control over phage replication. However, the slow licensing process of genetically modified organisms clearly inhibits the rapid therapeutic application of novel engineered variants necessary to fight mutant pathogens that emerge throughout the course of a pandemic. As a solution to this problem, we propose the SpyPhage system where a “scaffold” bacteriophage is engineered to incorporate a SpyTag moiety on its capsid head to enable rapid postsynthetic modification of their surfaces with SpyCatcher-fused therapeutic proteins. As a proof of concept, through CRISPR/Cas-facilitated phage engineering and whole genome assembly, we targeted a SpyTag capsid fusion to K1F, a phage targeting the pathogenic strain Escherichia coli K1. We demonstrate for the first time the cell-free assembly and decoration of the phage surface with two alternative fusion proteins, SpyCatcher-mCherry-EGF and SpyCatcher-mCherry-Rck, both of which facilitate the endocytotic uptake of the phages by a urinary bladder epithelial cell line. Overall, our work presents a cell-free phage production pipeline for the generation of multiple phenotypically distinct phages with a single underlying “scaffold” genotype. These phages could become the basis of next-generation phage therapies where the knowledge-based engineering of numerous phage variants would be quickly achievable without the use of live bacteria or the need to repeatedly license novel genetic alterations.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

SpyPhage: A Cell-Free TXTL Platform for Rapid Engineering of Targeted Phage Therapies

et al. 2022

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“…Phage vB_EcoM_SHAK9454 is an interesting case, as it is very similar to phage T7 (Figure S4) but can efficiently lyse a wide range of UPEC strains (Table 2) across K1, K2, and K5 capsule types, although it seems that R1 LPS strains may be non-permissive (Table 2). This makes vB_EcoM_SHAK9454 a potentially useful chassis for engineering UPEC phage (Ando et al 2015; Weynberg and Jaschke 2019; Mutalik and Arkin 2022) as the extensive knowledge of phage T7 and its tail fibers, which play important roles in host recognition and DNA delivery in the Autographiviridae (Casjens and Molineux 2012; Garcia-Doval and van Raaij 2012), can be used to guide its modification (Molineux 2006; Ando et al 2015; Huss et al 2021; Klumpp et al 2023),.…”

Section: Discussionmentioning

confidence: 99%

Discovery and characterisation of new phage targeting uropathogenicEscherichia coli

Kangachar,

Logel,

Trofimova

et al. 2024

Preprint

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Antimicrobial resistance (AMR) is increasing at an escalating rate with few new therapeutic options in the pipeline. Urinary tract infections (UTIs) are one of the most prevalent bacterial infections globally and are particularly prone to becoming recurrent and antibiotic resistant. The aim of this study was to discover and characterise new bacterial viruses (phage) against uropathogenic Escherichia coli (UPEC), which is the leading cause of UTIs. Six phages from the Autographiviridae family and Guernseyvirinae sub-family were isolated from wastewater and sequenced. The length of the isolated phage genomes was between 39,471 bp and 45,233 bp, with a GC content between 45.0% and 51.0%, and 57 to 84 predicted coding sequences (CDS) per genome. These phages were found to infect between 25 - 75% of the twelve UPEC strains tested. Using sequence comparison and predicted structural alignments, we show a similarity between the C-terminal domain of the tail fiber proteins of two phage that correlates with their host range. In vitro characterisation of phage cocktails against a single bacterial strain did not perform better than the best-performing phage, but did show synergistic improvement against a mixed UPEC strain population. Lastly, we measured the effectiveness of treatment with phage with different lytic kinetics in a sequential treatment and found it was slightly improved over single phage treatment.

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“…The most obvious use of phages in biomedicine is in the phage therapy of bacterial disease. That topic has been discussed extensively (recent reviews [ 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 ]). Here, I make the point that the following biomedicine-wide lesson is, in my opinion, worth learning from the low productivity [ 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 ] of recent efforts in phage therapy.…”

Section: Phages and Phage Dna Packaging In Relation To Biomedicine: A...mentioning

confidence: 99%

“…That topic has been discussed extensively (recent reviews [ 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 ]). Here, I make the point that the following biomedicine-wide lesson is, in my opinion, worth learning from the low productivity [ 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 ] of recent efforts in phage therapy. As noted for DNA packaging above, strategy should be focused on identifying and solving the key problem that inhibits progress.…”

Section: Phages and Phage Dna Packaging In Relation To Biomedicine: A...mentioning

confidence: 99%

A Perspective on Studies of Phage DNA Packaging Dynamics

Serwer

2022

IJMS

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The Special Issue “DNA Packaging Dynamics of Bacteriophages” is focused on an event that is among the physically simplest known events with biological character. Thus, phage DNA (and RNA) packaging is used as a relatively accessible model for physical analysis of all biological events. A similar perspective motivated early phage-directed work, which was a major contributor to early molecular biology. However, analysis of DNA packaging encounters the limitation that phages vary in difficulty of observing various aspects of their packaging. If a difficult-to-access aspect arises while using a well-studied phage, a counterstrategy is to (1) look for and use phages that provide a better access “window” and (2) integrate multi-phage-accessed information with the help of chemistry and physics. The assumption is that all phages are characterized by the same evolution-derived themes, although with variations. Universal principles will emerge from the themes. A spin-off of using this strategy is the isolation and characterization of the diverse phages needed for biomedicine. Below, I give examples in the areas of infectious disease, cancer, and neurodegenerative disease.

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A Phage Foundry Framework to Systematically Develop Viral Countermeasures to Combat Antibiotic-Resistant Bacterial Pathogens

Cited by 21 publications

References 269 publications

SpyPhage: A Cell-Free TXTL Platform for Rapid Engineering of Targeted Phage Therapies

SpyPhage: A Cell-Free TXTL Platform for Rapid Engineering of Targeted Phage Therapies

Discovery and characterisation of new phage targeting uropathogenicEscherichia coli

A Perspective on Studies of Phage DNA Packaging Dynamics

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