A novel approach to fight a drug resistant fungal pathogen

A novel approach to fight a drug resistant fungal pathogen

By: Lance Fredericks    Email:  fred6557@vandals.uidaho.edu

Home Town: Coeur d' Alene, Idaho    High School: Coeur d' Alene Charter Academy 2016

Major: Microbiology, Molecular Biol & Biotechnology
Department: Biological Sciences
College: College of Science

The initial discovery

We are investigating the potential of protein toxins naturally produced by Saccharomyces cerevisiae (Brewer’s/Baker’s yeast) to inhibit the growth of the human pathogen Candida glabrata. These toxin producers are known as killer yeasts to those who study them. We have tested over 15,0000 interactions between our killer yeasts and 130 isolates of the pathogen. We were pleased to find that killer toxins are capable of inhibiting the growth of every single isolate of Candida glabrata we have tested. These results lead us to believe the toxins could one day be developed into therapeutics to treat C. glabrata in humans.

A plate assay showing the ability of concentrated spots of killer toxin producing yeasts to inhibit the lawn of C. glabrata. Dark blue zones and clear zones surrounding the spots show the effects of the toxins.

The problem of drug resistance

Our focus on C. glabrata comes from a dire need for new antifungal drugs to treat recurrent infections. This particular pathogen has a tendency to develop resistance to drugs very quickly which reduces treatment options at an alarming rate. We hope to add another arrow to the medical quiver that has an entirely new mechanism of action. Killer toxins are able to inhibit the growth of both drug resistant and drug sensitive C. glabrata equally. This conclusion comes from our tests of over 70 drug resistant C. glabrata isolates from the CDC and the Wayne State clinic in Detroit.

Generating and testing mutants

With the issue of drug resistance in mind we realize that it would be short sighted to ignore the inevitable resistance to killer toxins. We chose to generate and isolate mutants that are resistant to killer toxins in the lab and study them to gain some insight to how resistance may occur clinically. Our interest lies in the fitness tradeoffs that come with acquiring resistance. We test this by comparing mutants to the parent strain we used to generate them. The tests include growth rate, resistance to other toxins, stress response to chemical agents such as caffeine, full genome sequencing, and the ability to cause lethal infection in an animal model.

Resistant mutants are subjected to chemical stressors and observed for growth. The red spots are colonies growing on a plate containing a semi-toxic chemical.

Testing mutant pathogenicity

While testing the ability of mutants to survive in harsh environments is important, the best test of the pathogenicity (the ability to cause disease) is to actually cause disease in a living host. We chose to do this in the larvae of the Greater Wax Moth (Galleria mellonella). The test is a simple one in which we inject 20 larvae with a consistent amount of C. glabrata. We then compare the survival of the 20 larvae injected with one mutant to 20 larvae injected with the wild type parent strain. We have identified several mutant strains that cause noticeably less disease than the non-mutated parent strain which indicates some mechanisms of toxin resistance cause the C. glabrata to be less pathogenic.

A larva is immobilized in an injection chamber, held with a needle resistant glove, and injected with a dose of C. glabrata. (We wrote a methods paper on this. It is listed in my products produced–check it out!)

About Lance Fredericks

Lance FredericksI've been working as an undergraduate researcher in some capacity since October of 2016. I have been involved with The Rowley Lab since February of 2017. Since then I have developed a deep professional interest in the study of pathogens and infectious disease. After graduating in May of 2020 I will continue my research full time until 2021 when I will volunteer abroad. I plan to pursue an MD-PhD focusing on Infectious Diseases.

 


Products Produced:

Type: Title: Date Published/Presented: DOI:
Poster Exploring Antifungal Drugs Produced by Brewer's Yeasts June 26, 2019
Publication March 10, 2020 https://doi.org/10.1101/2020.03.10.985481
Poster

Additional Project Information:

Year in College Project Started:  Sophomore

Faculty Advisor:   Paul  Rowley

Faculty Advisor Email:   prowley@uidaho.edu

Faculty Advisor Website:   https://www.uidaho.edu/sci/biology/people/faculty/prowley

Funding Source:  Idaho INBRE

External Link to Project Information:   https://www.rowleylab.com/

Project Location:   Moscow, Idaho