How bacteria fight antibiotics
Two mechanisms of antibiotic tolerance are demonstrated in separate studies
in Science
By Cathy Holding

Researchers report in two separate papers in Science this week on novel
methods used by bacteria to avoid being killed by antibiotics. In one study,
scientists at the Rockefeller University report that the bacterial cells
known as persisters, which tolerate but do not become resistant to
antibiotics, preexist in a population and that their random switching
between normal and slow-growing persister states enables them to escape
antibiotic killing.

And in an accompanying paper, Stanford University researchers demonstrate
that certain antibiotics trigger the SOS response in bacteria, resulting in
shutdown of DNA replication and transient dormancy, enabling survival of the
antibiotic sensitive bacteria. The SOS response prevents damaged DNA from
being copied at cell division, said Christine Miller, lead author of the
Stanford study. "It's common throughout the whole plant, animal, bacterial
world," she said.

Miller and colleagues were investigating conditions that affected the dpiAB
gene operon-overexpression of which results in overcoming of the replication
proteins DnaA and DnaB and induction of the SOS response-while it was
connected in a plasmid to a lacZ reporter gene. They found that exposure to
antibiotics triggered reporter gene expression, but only the beta-lactam or
penicillin group of antibiotics-which work by binding cell wall components
and causing cell lysis-had this effect.

"What they discovered is an interconnection among regulatory pathways that
people did not suspect talked to each other-that is, the cell wall, DNA
replication, and SOS," Kim Lewis, professor of biology at Northeastern
University, told The Scientist.

But Lewis, who was not involved in the study, said there was a disconnect
between the observations and the model the authors propose. "The cell can do
considerably simpler and more elegant things-and it doesn't, which means we
don't understand something," Lewis said. "We're missing something."

Nathalie Balaban's group at the Rockefeller cultured single bacterial cells
and monitored the effect on them and their progeny in the presence and
absence of antibiotics. "It's a technique that's based on Steve Quake's
microfluidic devices, and we just adapted it for this experiment," Balaban
said. They found that the slow-growing persisters flip into normal growth
mode and back again in a stochastic fashion and therefore escape antibiotic
killing.

The authors write that even before antibiotic treatment, persisters could be
clearly distinguished from the normal cells by their reduced growth rate.
The group mathematically modeled the switching from a normal to a persister
state and vice versa. "We have described it is as stochastic, but we don't
know a specific mechanism [to account for the switch]," Balaban said.

"This is interesting, but for somebody who doesn't know why this is
important, this will just stay cute and insignificant," said Lewis, whose
group has already described how persisters are largely responsible for the
complete tolerance of biofilms to killing by antibiotics. "That is what
makes persisters so really important-because biofilms are responsible for
something like 65% of all infections in the West. It's an enormous,
intractable problem."

And Lewis felt the use of the term "phenotypic switch" was unfortunate and
misleading. "A switch in microbiology is a very particular thing," he said.
"Phenotypic switches always in all cases that have been described so far
affect the genome."

Still, Denis A. Mitchison, emeritus professor at the Department of Cell and
Molecular Sciences at St. George's Hospital Medical School, London, welcomed
Balaban et al.'s findings. Mitchison, who was not involved in the study,
said that when he began working on the concept of persistence 50 years ago,
in the area of tuberculosis, he had grant proposals turned down because the
referees thought these bacterial populations do not preexist and are the
result of interaction with drugs. "This is very nice because it's providing
evidence that that's not so," said Mitchison.

Links for this article
N.Q. Balaban et al., "Bacterial persistence as a phenotypic switch,"
Science, DOI:10.1126/science.1099390, August 12, 2004.
http://www.sciencemag.org/

C. Miller et al., "SOS response induction by b-lactams and bacterial defense
against antibiotic lethality," Science, DOI:10.1126/science.1101630, August
12, 2004.
http://www.sciencemag.org/

Stanley N. Cohen Laboratory at Stanford University
http://sncohenlab.stanford.edu/

Kim Lewis
http://www.biology.neu.edu/faculty03/lewis03.html

Rockefeller University Laboratory of Living Matetr
http://www.rockefeller.edu/research/abstract.php?id=88

C. Holding, "Lab on a chip," The Scientist, March 15, 2004.
http://www.biomedcentral.com/news/20040315/02/

I. Keren et al., "Persister cells and tolerance to antimicrobials," FEMS
Microbiol Lett, 230:13-18, January 15, 2004.
[PubMed Abstract] [Publisher Full Text]

N. Johnston, "Debaffling biofilms," The Scientist, 18:34, August 2, 2004.
http://www.the-scientist.com/yr2004/aug/hot_040802.html

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