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Browsing by Author "Silver, Pamela A."

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    In situ reprogramming of gut bacteria by oral delivery
    Hsu, Bryan B.; Plant, Isaac N.; Lyon, Lorena; Anastassacos, Frances M.; Way, Jeffrey C.; Silver, Pamela A. (2020-10-06)
    Abundant links between the gut microbiota and human health indicate that modification of bacterial function could be a powerful therapeutic strategy. The inaccessibility of the gut and inter-connections between gut bacteria and the host make it difficult to precisely target bacterial functions without disrupting the microbiota and/or host physiology. Herein we describe a multidisciplinary approach to modulate the expression of a specific bacterial gene within the gut by oral administration. We demonstrate that an engineered temperate phage lambda expressing a programmable dCas9 represses a targeted E. coli gene in the mammalian gut. To facilitate phage administration while minimizing disruption to host processes, we develop an aqueous-based encapsulation formulation with a microbiota-based release mechanism and show that it facilitates oral delivery of phage in vivo. Finally we combine these technologies and show that bacterial gene expression in the mammalian gut can be precisely modified in situ with a single oral dose. It is difficult to precisely target bacterial populations in the mammalian gut. Here the authors use encapsulated phages to deliver dCas9 to E. coli in the mouse gut to modulate RFP expression.
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    Stable Neutralization of a Virulence Factor in Bacteria Using Temperate Phage in the Mammalian Gut
    Hsu, Bryan B.; Way, Jeffrey C.; Silver, Pamela A. (2020-01-28)
    Elimination or alteration of select members of the gut microbiota is key to therapeutic efficacy. However, the complexity of these microbial inhabitants makes it challenging to precisely target bacteria. Here, we deliver exogenous genes to specific bacteria by genomic integration of temperate phage for long-lasting modification. As a real-world therapeutic test, we engineered lambda phage to transcriptionally repress Shiga toxin by using genetic hybrids between lambda and other lambdoid phages to overcome resistance encoded by the virulence-expressing prophage. We show that a single dose of engineered phage propagates throughout the bacterial community and reduces Shiga toxin production in an enteric mouse model of infection without markedly affecting bacterial concentrations. Our work reveals a new framework for transferring functions to bacteria within their native environment. IMPORTANCE With the increasing frequency of antibiotic resistance, it is critical to explore new therapeutic strategies for treating bacterial infections. Here, we use a temperate phage, i.e., one that integrates itself into the bacterial genome, to neutralize the expression of a virulence factor by modifying bacterial function at the genetic level. We show that Shiga toxin production can be significantly reduced in vitro and in the mammalian gut. Alternative to traditional applications of phage therapy that rely on killing bacteria, our genetics-based antivirulence approach introduces a new framework for treating bacterial infections.