Phage attacks shown in new light

Date:
March 6, 2023
Source:
University of Pittsburgh
Summary:
New methodology and tools provide an opportunity to watch in
unprecedented detail as a phage attacks a bacterium.


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FULL STORY ==========================================================================
As antibacterial resistance continues to render obsolete the use of some antibiotics, some have turned to bacteria-killing viruses to treat acute infections as well as some chronic illnesses.


========================================================================== Graham Hatfull, the Eberly Family Professor of Biotechnology in the
Kenneth P.

Dietrich School of Arts and Sciences at Pitt, has pioneered the use of
these viruses -- bacteriophages, phages for short -- to treat infections
in chronic diseases such as cystic fibrosis. Although the importance of resistance may have eluded the early discovers of antibiotics, Hatfull
is intent on understanding how bacteria become resistant to phages.

His lab has just discovered how a specific mutation in a bacterium results
in phage resistance. The results were published Feb. 23, in the journal
Nature Microbiology.

The new methodology and tools his team developed also gave them the
opportunity to watch in unprecedented detail as a phage attacks a
bacterium. As the use of phage therapy expands, these tools can help
others better understand how different mutations protect bacteria against invasion by their phages.

For this study, the team started with Mycobacterium smegmatis, a harmless relative of the bacteria responsible for tuberculosis, leprosy and other
hard- to-treat, chronic diseases. They then isolated a mutant form of the bacterium that is resistant to infection by a phage called Fionnbharth.

To understand how the specific mutation in the lsr2 gene helps these
resistant bacteria fight off a phage, the team first needed to understand
how phages killed a bacteria without the relevant mutation.

Carlos Guerrero-Bustamante, a fourth-year graduate student in Hatfull's
lab, genetically engineered two special kinds of phages for this
study. Some produced red fluorescence when they entered a bacterial
cell. Others had segments of DNA that would stick to fluorescent molecules
so phage DNA would light up in an infected cell.

Following the fluorescent beacons, "We could see where the phage DNA
entered the cell," Guerrero-Bustamante said. The imaging methods they used
were designed by Charles Dulberger, a collaborator and co-first author
of the paper who was then at Harvard T.H. Chan School of Public Health.

"We saw for the first time how the phages take that first step of binding
to cells and injecting their DNA into the bacteria," said Hatfull, who
is also a Howard Hughes Medical Institute Professor. "Then we applied
those insights to ask, 'So, how's it different if we get rid of the
Lsr2 protein?'" The link between Lsr2 and phage resistance has not been previously known, but with their new methods and tools, the team clearly
saw the critical role it played.

Typically, Lsr2 helps bacteria replicate its own DNA. When a phage
attacks, however, the virus co-opts the protein, using it to replicate
phage DNA and overwhelm the bacteria. When the lsr2 gene is missing or defective -- as in the phage-resistant Mycobacterium smegmatis -- the
bacteria doesn't make the protein and phages don't replicate enough to
take over the bacterial cell.

This was a surprise.

"We didn't know Lsr2 had anything to do with bacteriophages," Hatfull
said.

These new tools can be used to uncover all manner of surprises written
in the genes of phage-resistant bacteria. It may also help today's
researchers and tomorrow's clinicians to better understand and take
advantage of phages' abilities while avoiding the missteps that led to antibiotic resistance.

"This paper focuses on just one bacterial protein," and its resistance
to just one phage, Hatfull said, but its implications are wide. "There
are lots of different phages and lots of other proteins."
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========================================================================== Story Source: Materials provided by University_of_Pittsburgh. Original
written by Brandie Jefferson. Note: Content may be edited for style
and length.


========================================================================== Journal Reference:
1. Charles L. Dulberger, Carlos A. Guerrero-Bustamante, Sia^n V. Owen,
Sean
Wilson, Michael G. Wuo, Rebecca A. Garlena, Lexi A. Serpa, Daniel A.

Russell, Junhao Zhu, Ben J. Braunecker, Georgia R. Squyres,
Michael Baym, Laura L. Kiessling, Ethan C. Garner, Eric J. Rubin,
Graham F. Hatfull.

Mycobacterial nucleoid-associated protein Lsr2 is required for
productive mycobacteriophage infection. Nature Microbiology, 2023;
DOI: 10.1038/ s41564-023-01333-x ==========================================================================

Link to news story: https://www.sciencedaily.com/releases/2023/03/230306143446.htm

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