# Rapid AST through Metabolic Imaging at Single Cell Level

> **NIH NIH R01** · BOSTON UNIVERSITY (CHARLES RIVER CAMPUS) · 2024 · $621,858

## Abstract

Project Summary
In order to combat infections and reduce antimicrobial resistance, it is essential to detect and characterize
bacterial/fungal susceptibility to antimicrobials in the early stages of infections to reduce the inappropriate use of
antimicrobials and the rate of death. Under R01AI141439 (2018 to 2022), we have been addressing this urgent
need by developing a rapid AST through high-speed stimulated Raman scattering (SRS) imaging of single-cell
metabolism. Through SRS imaging of glucose-d7 metabolism, we determined the antimicrobial susceptibility
profile of bacteria and fungi within 1 cell cycle. More recently, through SRS imaging of D2O incorporation, we
obtained a single-cell metabolism inactivation concentration (SC-MIC) in less than 2.5 h from culturing a colony
to obtaining results. Despite this initial success, there are remaining gaps in translating this technology into clinic.
First, the single-color SRS signal depends on the bacterial size and thus is not quantitative in predicting single-
cell metabolic activity. Second, the laser used in our past studies is too bulky to be used in clinic. Third, SRS
imaging alone does not tell the identity of the pathogens. Fourth, the single-well imaging chamber does not allow
high-throughput measurement of samples separately treated by a panel of antibiotics at various concentrations.
In this competitive renewal of R01AI141439, we propose to overcome these barriers through three technical
innovations. To address the first and second gaps, we will build a fast-tuning SRS microscope based on a
compact fiber-OPO laser. We have demonstrated the feasibility of this approach in multi-window SRS imaging
of single cell metabolism. By fast tuning between C-D and C-H vibration, we will be able to use CD/(CD+CH)
SRS signal ratio to quantify the metabolic activity of a single cell. To address the third gap, we will build multi-
color fluorescence in situ hybridization (FISH) into the compact SRS microscope to enable rapid identification
and fast AST on a single microscope platform. We have demonstrated the feasibility of FISH-SRS for imaging
metabolic activity of bacteria in a gut microbiome. To address the fourth gap, we will demonstrate high-throughput
AST through robotic handling of liquid specimens and a multi-well cartridge design. A team with complementary
expertise will carry out the proposed research. While we use urinary tract infection as a focused testbed in this
proposal, our technology is broadly applicable to rapid determination of susceptibility for other important
infections (e.g., bloodstream infections, meningitis, pneumonia), thus having a broad, positive impact on global
public health.

## Key facts

- **NIH application ID:** 10884403
- **Project number:** 5R01AI141439-06
- **Recipient organization:** BOSTON UNIVERSITY (CHARLES RIVER CAMPUS)
- **Principal Investigator:** Ji-Xin Cheng
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $621,858
- **Award type:** 5
- **Project period:** 2018-12-01 → 2027-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10884403

## Citation

> US National Institutes of Health, RePORTER application 10884403, Rapid AST through Metabolic Imaging at Single Cell Level (5R01AI141439-06). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10884403. Licensed CC0.

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