# Mechanisms of phenotypic antibiotic resistance in Gram-negative bacteria

> **NIH NIH R01** · EMORY UNIVERSITY · 2022 · $433,628

## Abstract

PROJECT SUMMARY/ABSTRACT
Antibiotic resistance is one of the most serious medical challenges of our time. This crisis puts patients at risk
of untreatable bacterial infections and threatens major advances of modern medicine that rely on antibiotics
(transplants, chemotherapy, etc). There are at least 2 million antibiotic resistant infections each year in the US,
leading to over 23,000 deaths [1]. It is estimated that without significant action, worldwide annual mortality due
to these infections will reach 10 million by 2050, surpassing the predicted mortality from cancer [2].
Understanding resistance mechanisms is critical to designing novel therapeutics to combat resistant bacteria.
Heteroresistance is an enigmatic form of antibiotic resistance in which a bacterial isolate harbors a resistant
subpopulation that can rapidly replicate in the presence of an antibiotic, while a genetically identical yet
susceptible subpopulation is killed [3, 4]. Not only do many species of bacteria exhibit this form of phenotypic
resistance, but it has been reported against different classes of antibiotics. Unfortunately, our understanding of
heteroresistance is extremely limited and its relevance during infection has been unclear. Using two clinical
isolates of the Gram-negative nosocomial pathogen Enterobacter cloacae, we recently showed that
heteroresistance to the last-line antibiotic colistin can cause treatment failure in an in vivo model [4].
Furthermore, one of the isolates harbored a very low frequency resistant subpopulation (<1 in 10,000 cells)
that was undetected by clinical diagnostic tests, leading to its incorrect classification as colistin susceptible [4].
Our unpublished national surveillance data reveal that colistin heteroresistance is present in 10% of
carbapenem-resistant Enterobacteriaceae (CRE), including 18% of carbapenem-resistant Enterobacter,
although the majority of these isolates are incorrectly classified as colistin susceptible. Such misclassification
could lead clinicians to prescribe colistin inappropriately, leading to treatment failures. Taken together, these
data highlight a largely unappreciated epidemic in which colistin heteroresistance is prevalent,
overwhelmingly undetected, and may cause unexplained antibiotic treatment failure in the clinic.
We will use a combination of genetics, biochemistry, single cell microscopy, flow cytometry and cell sorting to
make foundational insights into heteroresistance. Specifically, we will elucidate the dynamics of the resistant
subpopulation within colistin heteroresistant Enterobacter, as well as the molecular mechanism controlling
resistance. The knowledge gained from this work will form a paradigm with which to study heteroresistance in
other bacteria and against diverse antibiotics. Further, the impact of this research will likely extend to
eukaryotes as heteroresistance has been observed in fungi [5, 6] and human cancers [7]. Overall, this work will
significantly broaden our understan...

## Key facts

- **NIH application ID:** 10465291
- **Project number:** 5R01AI141883-05
- **Recipient organization:** EMORY UNIVERSITY
- **Principal Investigator:** DAVID S WEISS
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $433,628
- **Award type:** 5
- **Project period:** 2018-09-24 → 2023-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10465291

## Citation

> US National Institutes of Health, RePORTER application 10465291, Mechanisms of phenotypic antibiotic resistance in Gram-negative bacteria (5R01AI141883-05). Retrieved via AI Analytics 2026-06-11 from https://api.ai-analytics.org/grant/nih/10465291. Licensed CC0.

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