Phage Foundry: Establishing Capabilities for High-Throughput Phage-Host Interaction Characterization and Prediction
Authors:
Denish Piya1* ([email protected]), Mohamad Alayouni1, Archana Anand4, Adam P. Arkin2, Hans Carlson1, Brady Cress2, Adam Deutschbauer1, Darian Doakes2, Michael Hajkowski4, Jamie Inman1, Alexey Kazakov1, Britt Koskella2, Petr Leiman5, Catherine Mageeney6, Ryan Melnyk1, Mark Mimee3, Harshini Mukundan1, Avery Noonan1, Ella Rotman3, Hemaa Selvakumar1, Antoine Snijders1, Jessica Trinh6, Simon Roux1, Vivek K. Mutalik1 (PI)
Institutions:
1Lawrence Berkeley National Laboratory; 2University of California–Berkeley; 3University of Chicago; 4San Francisco State University; 5University of Texas Medical Branch; 6Sandia National Laboratory
Abstract
Increase in the incidence of antimicrobial-resistant (AMR) bacterial pathogens currently poses an immense threat to normal world order. In addition to the tragic impact on human health, AMR is estimated to have a worldwide economic cost running into trillions of U.S. dollars by severely debilitating agriculture, dairy, aquaculture, livestock, and poultry industries. Bacteriophages (phages) have been proposed as an alternative to antibiotics due to a dearth of new antimicrobial molecules in the discovery pipeline. There have already been some successes in treating AMR pathogenesis by using phages under compassionate use protocols; however, biological tools for broad-scale mechanistic characterization of phage-host interactions in clinically and agriculturally relevant bacteria are still limited, hampering development and application of phages as robust antimicrobial agents.
In this project, researchers are developing tools for high-throughput characterization of phage-host interactions on highest priority ESKAPE human pathogens: Klebsiella pneumoniae, Acinetobacter baumannii and Pseudomonas aeruginosa, as well as important crop pathogen Pseudomonas syringae. By using panels of clinically relevant and genomically diverse strains, the team is building a highly diverse collection of phages (“phage banks”) and conducting large-scale phage-susceptibility assays for genome-wide association studies (GWAS)like analyses to identify bacterial and phage genes that drive host range and specificity. To guide phage isolation efforts and build a baseline understanding of the population diversity, mobile genetic elements, and phages associated with pathogens, the team is using a multiomics approach on a large wastewater system. To generate systematic genotype-phenotype mapping, powerful high-throughput genetic technologies, such as genome-wide loss-of-function randomly barcoded transposon sequencing, CRISPR interference and overexpression dual barcoded shotgun expression library sequencing, are being applied to a set of select strains.
This systematic effort in phenotyping and high-throughput genetics will provide fitness landscapes in presence of phages, antibiotics, metals, and other stressors, as well as enable researchers to map the cross-resistance and collateral sensitivities between phages and antibiotics. The characterization workflows, resources and datasets generated at the Biopreparedness Research Virtual Environment Phage Foundry will provide crucial foundational knowledge necessary to develop machine-learning models and facilitate quick and effective prediction of therapeutic formulations for countering any emerging recalcitrant infections.
Funding Information
This material by Biopreparedness Research Virtual Environment (BRaVE) Phage Foundry at Lawrence Berkeley National Laboratory is based upon work supported by the DOE, Office of Science, BER program under contract number DE-AC02-05CH11231.