# Identification of Ameobicidal Products Against Pathogenic Free-Living Amoebae Produced by Pseudomonas aeruginosa

> **NIH NIH R21** · YALE UNIVERSITY · 2020 · $208,464

## Abstract

Project Summary
Acanthamoeba castellanii and Naegleria fowleri are free-living amoebae (FLA) that cause untreatable, fatal
infections of the central nervous system (CNS). FLA are bacterivorous, and one environmental bacterium they
prey upon is Pseudomonas aeruginosa. Because of the evolutionary pressure for P. aeruginosa to defend itself
against amoebal grazing and its well-known ability to produce a plethora of antimicrobial compounds, we
hypothesize that P. aeruginosa may produce one or more secreted amoebicidal compounds that can be
identified and serve as leads for new and much-needed drugs to treat amoeba infections in humans.
We have demonstrated that the cell-free supernatants of a laboratory strain of P. aeruginosa contain stable small
molecule(s) that are lethal to A. castellanii. These compound(s) will be isolated through a proven pipeline of
organic extraction followed by chromatography and bioassay-guided fractionation. They will then be
characterized using mass spectrometry and NMR, and the genes responsible for compound production will be
identified by utilizing biosynthetic gene cluster mining tools. The isolated compound(s) will be tested for toxicity
against FLA and human cells and for efficacy in an in vitro model of FLA infection of human neurons. Separately,
we will identify bacterial genes required to kill amoebae by screening an arrayed P. aeruginosa transposon
mutant library for amoebicidal activity. Filtered supernatants from each mutant will be incubated with A. castellanii
and amoeba viability will be quantified. We have optimized a colorimetric assay to measure amoeba viability in
a scalable manner using sulforhodamine B (SRB). It is our goal to identify leads for novel therapeutic agents to
treat devastating human infections by A. castellanii and N. fowleri.

## Key facts

- **NIH application ID:** 9885697
- **Project number:** 1R21AI149628-01
- **Recipient organization:** YALE UNIVERSITY
- **Principal Investigator:** BARBARA I KAZMIERCZAK
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $208,464
- **Award type:** 1
- **Project period:** 2020-01-10 → 2021-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9885697

## Citation

> US National Institutes of Health, RePORTER application 9885697, Identification of Ameobicidal Products Against Pathogenic Free-Living Amoebae Produced by Pseudomonas aeruginosa (1R21AI149628-01). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9885697. Licensed CC0.

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