Characterizing clinically isolated Vibrio cholerae bacteriophages for the development of a phage prophylaxis cocktail
cholerae is the causative agent of cholera, a severe diarrheal disease that is a
substantial health burden on the developing world. It is a Gram-negative bacterium that
lives in the aquatic ecosystem and can colonize the human small intestine upon
ingestion. Virulent bacteriophages are hypothesized to play a crucial role in modulating
the dynamics of cholera epidemics and the... read moreprogression of disease. Studies to date,
however, have not demonstrated this in the context of human cholera infection.
Therefore, the goals of my thesis are to understand the impact of intra-patient phage
predation on V. cholerae population structure and how phages can be used to prevent
disease. In the first study, we investigated the heterogeneity of phage resistance in
Bangladeshi and Haitian stool samples containing V. cholerae and high titers of the
virulent phage ICP2. Using whole-genome sequencing, we determined that ICP2 uses the
major outer membrane protein OmpU as its receptor to initiate infection. The potential
fitness consequences of OmpU mutants were addressed using assays measuring in vitro
growth, bile sensitivity, and colonization in an infant mouse model. We were also able
to recapitulate the infection dynamics thought to occur in human cholera patients that
lead to the selection of OmpU mutants. Our results indicate phage pressure during
infection results in an altered V. cholerae population structure that exits the host. In
the second study, we determined that ICP2 is widespread in Haiti. We purified 34 ICP2
isolates from clinical samples collected from 2012 to 2014 and performed phylogenetic
analyses to determine evolutionary relationships. Although there was a high degree of
similarity, the relatedness does not seem to be associated with geographic location or
time of isolation. In the third study, we developed a prophylactic three-phage cocktail
for the prevention of cholera. Oral administration of the phages up to 24 hours before
V. cholerae challenge reduced colonization of the intestinal tract and prevented the
onset of disease symptoms in two animal models of cholera pathogenesis. For acute
infections, such as cholera, phage prophylaxis could provide a strategy to limit the
impact of bacterial disease on human health.
Thesis (Ph.D.)--Tufts University, 2016.
Submitted to the Dept. of Molecular Microbiology.
Advisors: Andrew Camilli, and Stephen Calderwood.
Committee: Ralph Isberg, Carol Kumamoto, and Wai-Leung Ng.
Keyword: Microbiology.read less