loading
loading
NIH grant 1R01AI192998-01A1: Impact of bacterial phenotypic variation on phage resistance
Institute: {'code': 'AI', 'abbreviation': 'NIAID', 'name': 'National Institute of Allergy and Infectious Diseases', 'admin_org_id': '12500003', 'admin_funding_url': ''}. PI: Anna Kuchina. Award: $874085.
Start: 2026-09-25T00:00:00.
Project Summary
Bacteroides fragilis is a pathobiont found in the human intestinal tract, causing abscesses and bacteremia with
high mortality rates. In addition, B. fragilis is associated with inflammatory bowel disease and colorectal cancer.
As multidrug-resistant isolates of pathogenic B. fragilis are emerging, effective phage therapies are needed.
However, this organism has a potent capacity to evade bacteriophages by using phenotypic plasticity. Multiple
recombinases acting on a wide array of invertible genomic regions cause variation of expression in single B.
fragilis cells. This genome-wide phase variation creates extensive phenotypic diversity across the clonal bacterial
population, leading to co-existence of cells with distinct types of capsular polysaccharides, fimbria, phage
immunity systems, and other diverse phenotypic features in a seemingly isogenic population. We hypothesize
that variable expression of bacterial surface components and antiphage defense systems in single cells
collectively influence B. fragilis susceptibility to bacteriophage infection and potentiate the emergence of
phenotypic phage resistance. In this proposal, we aim to unravel the impact of heterogeneous expression of
multiple genes, including antiphage defense systems, on the susceptibility of B. fragilis to phages. Importantly,
the combinatorial impacts of these systems will be studied in the native host with the intact regulation
mechanisms. For a high-throughput, transcriptome-wide characterization of individual B. fragilis cells infected
with a human gut phage, we will use the technology we recently developed for bacterial single-cell RNA
sequencing, microSPLiT (microbial split-pool ligation transcriptomics). Since multiple pathogens use
heterogeneous gene expression to enhance their survival, we extend this framework to study how phenotypic
plasticity in expression of capsule components and antiphage defenses affects phage infection dynamics in a
high priority pathogen Acinetobacter baumannii. In Aim 1, we will identify the B. fragilis heterogeneous
phenotypic states and quantify their effect on phage resistance in the native host using microSPLiT in conjunction
with CRISPR-interference and phage infection assays. In Aim 2, we will use microSPLiT to interrogate how the
sensitivity of phenotypic subpopulations of B. fragilis to phage changes upon exposure to human gut metabolites
such as bile acids, as well as the mouse gut environment. Finally, in Aim 3, we will isolate a collection of human
gut bacteriophages against B. fragilis targeting different phenotypic subpopulations of the host. Further, we will
use microSPLiT to investigate how variability in gene expression enables distinct phage infection dynamics of
clinical A. baumannii strains and recently isolated phages. Our findings will directly inform the rational design of
therapeutic phage cocktails against these pathogens. Overall, our investigation of how genome-wide phenotypic
plasticity enables bacteria to evade phages will provide a broadly applicable framework for studying strategies
of phage resistance across bacterial species.document fetched 2026-10-04 02:43