The recent renaissance in bacteriophage therapy has led to a number of successful cases in human patients. Various studies in mice and humans have shown that bacteriophages are rapidly cleared, meaning the bacteriophages cannot be observed in patient blood after the treatment. Thus, one of the main challenges facing the expanded use of bacteriophage therapy is the creation of bacteriophages that are not cleared rapidly from the patient. The argument for this is clear -- the longer infectious bacteriophages can stay in the body, the greater their chance of finding their pathogenic bacteria host. The longevity of circulating bacteriophages has been investigated several times. These studies showed that longer circulating bacteriophages can be isolated. Unfortunately, often the genomes of these bacteriophages were not sequenced nor were the bacteriophages characterized, leaving a gap in our understanding of how these bacteriophages were altered to survive in the body for extended times. The objective of this proposal is to understand the mechanisms leading to longer bacteriophage survival in the circulatory system. Our work is designed to provide important translational insights for making synthetic bacteriophages able to circulate longer inside patients and thereby increasing the efficacy of bacteriophage therapy. Our central hypothesis is that enhanced circulation times are due to amino acid substitution(s) in the major capsid protein that either stabilize the bacteriophage or allow the bacteriophage to evade capture by immune cells. A classic study of longer-circulating lambda bacteriophages revealed that a single mutation in the capsid protein was responsible for the phenotype, but the mechanism for this phenotype was not investigated. We propose to focus on bacteriophages P22 and Sf6 to understand the mechanism for in vivo stabilization in these two different but tractable and well-characterized bacteriophages, with the following specific aims:
Aim 1) Hypothesis: the circulation time is increased by amino acid substitutions in the major capsid protein.
Aim1 a) Isolate and characterize bacteriophages with enhanced circulation times.
Aim1 b) Determine if the mutated residues resulting in increased circulation time is a generalizable feature.
Aim 2) Hypothesis: the long circulating mutant bacteriophages are not degraded by macrophages.
The longer infectious bacteriophages can stay in the body during bacteriophage therapy treatment, the greater their chance of finding their pathogenic bacteria host. This project aims to isolate long-circulating bacteriophages using a mouse model with well-characterized model bacteriophages, and then determine why they exhibit extended circulation times by biophysical and structural characterization.
