Bacterial viruses, known as bacteriophages or phages, are among the most abundant biological entities on the
planet. These viruses vary broadly in terms of genome content, infection mechanisms, replication cycles, and
structure. With an increasing interest in using bacteriophages to combat antibiotic resistant bacteria, it is
necessary to understand mechanisms of how these viruses infect their hosts; replicate and regulate both their
genes and their hosts’ genes; and how they persist in various environments, including the human body and/or
storage conditions. However, our current model systems inherently only represent a fraction of bacteriophage
diversity. The long-term goal of this work is to develop an additional bacteriophage system—the Moogleviruses—
that appear to be ubiquitous in the environment and clinically useful qualities. They have, however, only recently
been isolated because they do not commonly infect the model species of bacteria Escherichia coli and
Salmonella enterica. Instead, these viruses are often isolated against the pathogenic bacteria Shigella flexneri
or opportunistic pathogens such as Citrobacter freundii. Because these viruses are obligately lytic and specific
to one of the most common etiological agents of diarrhea, S. flexneri, they could be used for controlling this
species of bacteria in food or water, or for treating antibiotic-resistant infections. At this time, however, we lack
sufficient understanding of their biological processes. Moogleviruses have distinct characteristics versus other
bacteriophages, including a semi-specific host range, a large number of tRNAs encoded in their genomes, and
uncommon capsid and genome sizes. The central hypothesis of this work is that these viruses use alternative
strategies to infect and persist compared to more thoroughly characterized model systems. The objectives of
this specific proposal are therefore to determine the mechanisms Shigella-infecting Moogleviruses use to identify
their hosts, modulate the expression of viral and host genes on a translational level, and assemble into new
particles that can persist in the environment. The expected outcomes of this proposal are: 1) the identification of
critical interacting regions between the phage receptor-binding proteins and both primary and secondary
receptors on the S. flexneri host, along with determining the kinetics of attachment and entry; 2) complete Ribo-
seq and RNA-seq datasets from a variety of environments and infection conditions, leading to a mechanistic
understanding of how phage infection alters translational efficiency of phage and host genes; and 3) a broadly
resolved assembly pathway of the Mooglevirus capsid, with an indication of proteins or protein domains
responsible for affecting capsid stability. This work will have impacts for both basic biology, expanding our
repertoire of known bacteriophage strategies and mechanisms for infection and persistence; and medical
application, informing how these phages themselves could be applied to combat S. flexneri infections in the
clinic, or how their properties could be used to engineer novel phages for medical or industrial applications.
