Phage-delivered CRISPR-Cas9 for strain-specific depletion and genomic deletions in the gut microbiome
Kathy N. Lam,
Peter Spanogiannopoulos,
Paola Soto-Perez,
Margaret Alexander,
Matthew J. Nalley,
Jordan E. Bisanz,
Renuka R. Nayak,
Allison M. Weakley,
Feiqiao B. Yu,
Peter J. Turnbaugh
Affiliations
Kathy N. Lam
Department of Microbiology & Immunology, University of California, San Francisco, San Francisco, CA 94143, USA
Peter Spanogiannopoulos
Department of Microbiology & Immunology, University of California, San Francisco, San Francisco, CA 94143, USA
Paola Soto-Perez
Department of Microbiology & Immunology, University of California, San Francisco, San Francisco, CA 94143, USA
Margaret Alexander
Department of Microbiology & Immunology, University of California, San Francisco, San Francisco, CA 94143, USA
Matthew J. Nalley
Department of Microbiology & Immunology, University of California, San Francisco, San Francisco, CA 94143, USA
Jordan E. Bisanz
Department of Microbiology & Immunology, University of California, San Francisco, San Francisco, CA 94143, USA
Renuka R. Nayak
Department of Microbiology & Immunology, University of California, San Francisco, San Francisco, CA 94143, USA
Allison M. Weakley
Chan-Zuckerberg Biohub, San Francisco, CA 94158, USA; Stanford ChEM-H: Chemistry, Engineering and Medicine for Human Health, Stanford University, Stanford, CA 94305, USA
Feiqiao B. Yu
Chan-Zuckerberg Biohub, San Francisco, CA 94158, USA
Peter J. Turnbaugh
Department of Microbiology & Immunology, University of California, San Francisco, San Francisco, CA 94143, USA; Chan-Zuckerberg Biohub, San Francisco, CA 94158, USA; Corresponding author
Summary: Mechanistic insights into the role of the human microbiome in the predisposition to and treatment of disease are limited by the lack of methods to precisely add or remove microbial strains or genes from complex communities. Here, we demonstrate that engineered bacteriophage M13 can be used to deliver DNA to Escherichia coli within the mouse gastrointestinal (GI) tract. Delivery of a programmable exogenous CRISPR-Cas9 system enables the strain-specific depletion of fluorescently marked isogenic strains during competitive colonization and genomic deletions that encompass the target gene in mice colonized with a single strain. Multiple mechanisms allow E. coli to escape targeting, including loss of the CRISPR array or even the entire CRISPR-Cas9 system. These results provide a robust and experimentally tractable platform for microbiome editing, a foundation for the refinement of this approach to increase targeting efficiency, and a proof of concept for the extension to other phage-bacterial pairs of interest.
