Postdoctoral Fellowship: PRFB: Quantifying single-cell response to phage-induced stresses and high hydrostatic pressure in bacteria.

This action funds an NSF Postdoctoral Research Fellowship in Biology for FY 2024, Integrative Research Investigating the Rules of Life Governing Interactions Between Genomes, Environment, and Phenotypes. The fellowship supports the research and training of the fellow who will contribute to the area of Rules of Life in innovative ways. Bacteria, as simple forms of life, can adapt to various stresses and thrive in extreme conditions on Earth.

This research will explore the adaptive bacterial response to two primary stressors: bacteriophage infection and the immense hydrostatic pressure faced by organisms in the deep ocean. Novel methods will be developed that will benefit the fields of fluorescence microscopy and extremophile biology. By observing wild-type and mutant bacterial cells over many generations, this project will increase understanding of life’s adaptations to phage infection and extreme environments, which can have an impact on biofuel engineering, biotechnology applications, and even in the search for extraterrestrial life.

This fellowship will also support the development of a program for local high school students to gain research experience in biophysics, with a focus on engaging underrepresented groups in STEM.

This project will test the hypothesis that cell membranes play essential roles in the cellular response to stresses from phage infection and high pressure. Genetic manipulation of model organisms Escherichia coli and Pseudomonas aeruginosa, and deep-sea organism Shewanella piezotolerans will be used to perturb cellular pathways hypothesized to be important in phage production and pressure adaptation.

Cellular fitness will be quantified through time-resolved fluorescence microscopy of bacteria within microfluidic devices where cells are maintained in the exponential growth phase. Fluorescence reporters coupled to target mutations will quantify growth rate, cell size, and morphological integrity as a function of phage infection and exposure to high-hydrostatic pressure (HHP) up to 100 MPa.

The development of this technique at HHP will contribute to the training of fellows who are passionate about the implementation of novel techniques. To characterize the mechanism of stress during phage infection, phage-infected cells will be compared with cells that produce only particular phage proteins, which are hypothesized to affect growth as they crowd the membrane space and affect biosynthetic flux across the membrane.

For HHP experiments, wild-type cells will be compared with mutants of hydrostatic stress response components to determine the dominant response pathway. Concurrent development of CMU-PREP (Pittsburgh area Research Experience in Physics at Carnegie Mellon University) will be taking place, bringing select local high schoolers onto campus each summer to partake in research projects in Physics.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.