SCIENCE OBSERVER
Free Upgrades, Unfortunately
Elsa Youngsteadt
Contrary to infectious-disease dogma, the mutations that enable
bacteria to resist antibiotics do not always result in weaker
strains, according to a study published in the June 30 issue of the
journal Science. This is bad news for public-health
efforts, especially because the germ in question is the
tuberculosis-causing Mycobacterium tuberculosis, once the
leading cause of death in the United States.
Classic laboratory experiments once suggested that bacteria pay a
price for antibiotic resistance—that resistant bacteria are
weaker than their susceptible counterparts and should not spread
through the human population when forced to compete with hardier strains.
But the new study, headed by scientists at Stanford University, has
undermined this comforting conventional wisdom. It shows that in
real human patients, tuberculosis bacteria can evolve resistance to
antibiotics and still be just as aggressive as their susceptible ancestors.
"It's generally bad news for the world that some tuberculosis
strains can get something for nothing," said Sebastien Gagneux,
one of the lead authors of the study and a research associate at the
Institute of Systems Biology. "Even though many drug-resistant
strains are less hardy than susceptible strains, others evolve over
the course of treatment and remain virulent."
M. tuberculosis has plagued humans throughout history.
Improved public health brought the epidemic under control, and the
advent of antibiotics in the 1940s seemed to ensure successful
treatment. But the bacteria have adapted to nearly every drug used
against them. With its recent resurgence, the disease now kills a
person every 15 seconds, worldwide. Public-heath experts say the
antibiotic-resistant strains may be around to stay. "Evolution
is a powerful engine," says Peter Small, senior program officer
of the Tuberculosis Global Health Program and contributing author of
the Sciencepaper.
Gagneux and coauthor Clara Davis Long of Stanford suspected that the
lab studies underpinning the epidemiological credo might be missing
the big picture. A human being is, after all, a very different
environment than a culture plate. So Gagneux, Davis Long and their
collaborators took a more thorough look at the cost of antibiotic
resistance in tuberculosis bacteria, using not only the usual
lab-generated strains, but also those sampled directly from human
tuberculosis patients.
The investigators looked at the evolution of resistance to the drug
rifampin, one of the preferred first-line treatments for the
disease. Rifampin binds to the molecule that makes, or polymerizes,
bacterial RNA. The drug disables the so-called polymerase molecule
and prevents the crucial flow of information from DNA to RNA.
Without RNA, bacteria can't make the proteins they need to survive.
But simple mutations in the gene that encodes RNA polymerase can
change its structure. The different shape decreases the drug's
ability to bind and allows M. tuberculosis to persist in
the face of antibiotic onslaught. Classic studies suggested that
such a change would carry some cost, such as decreased efficiency of
the polymerase molecule. Such a penalty would cause the mutants to
grow more slowly than unmodified strains, thereby retarding the
spread of the antibiotic-resistant mutation in the population.
Not so, according to the new study, which found that some resistant
bugs are every bit as robust as unmodified strains. The Stanford
group collected tuberculosis bacteria from the sputum of patients,
first at the beginning of their infections, and a second time after
some of those patients developed rifampin-resistant infections. The
investigators then pitted the resistant strains against their
susceptible counterparts in antibiotic-free competition assays.
These tests force the two strains to compete for limited resources
in a common culture flask, so the hardier bug should take over as
the weaker one gets crowded out. Contrary to expectations, five of
the ten resistant strains held their own in these tests, and one
actually dominated its antibiotic-susceptible ancestor.
These results suggested that the evolutionary changes that lead to
drug resistance occur differently in human patients and culture
dishes. To confirm this conclusion, the scientists generated several
rifampin-resistant strains in the lab, simply by exposing M.
tuberculosis to growth media that contained a small amount of
the drug. Each resistant strain that arose had a single mutation in
the RNA-polymerase gene, and some of the changes were identical to
those found in strains isolated from human patients. However, unlike
the bacteria from the clinic, all the mutated laboratory strains
were weaker than their susceptible progenitor. This part of the
study parallels the methodology underpinning the conventional
wisdom, and it highlights the difference between what goes on in the
lab and what goes on in a human patient.
"Bacteria exist in heterogeneous populations of different
strains. If you just look at lab-adapted strains, you won't get the
right answer," Gagneux said. "By looking at clinical
isolates, you find that some kind of selection is going on in
patients that is different from what happens in a culture plate."
Although the laboratory mutants were less fit than their clinical
counterparts, the cost of the lab-generated mutations was still
related to their prevalence in clinical samples. The specific
mutations that were least costly in the lab bacteria were the same
ones that carried no cost for clinical strains and occurred most
often in patients. The costliest mutations generated in the lab
never turned up in a patient. In other words, "you don't see
wimpy bugs in the real world," Small said.
Despite its grim conclusion that our old enemy is more versatile
than we believed, the study also yielded some insights that may
prove useful in the fight. "The good news is that some very
simple in vitro competitive assays reflect the behavior of
these bacteria in the real world," Small added. "Now we
have a link between a pathogen's in vitro ecology and
its epidemiology, and that link can be incorporated into predictive
models. Laboratory data can tell us in advance whether a particular
bug is a concern. We don't have to wait and see."
Just how clinical isolates outperform lab-generated strains with
identical mutations remains a mystery. Some changes simply have a
low cost to begin with, and, the study also found, the cost depends
on the strain in which the mutation occurs. However, the real key is
likely to be compensatory mutation—one or more additional
changes, in the same RNA polymerase gene or in related genes, that
make up for the diminished function caused by the original mutation.
Gagneux is planning future studies to find these putative
compensatory mutations and learn how they restore the performance of
strains that acquire antibiotic-resistance mutations.
The authors don't want their study to fuel an alarmist panic.
Indeed, the data don't call for it, says Bruce Levin, an expert on
the evolution of antibiotic resistance at Emory University who was
not involved with the study. Levin points out that "the spread
of tuberculosis does not depend solely on the efficacy or lack of
efficacy of antibiotics." He cites public-health practices and
better nutrition as bulwarks against 19th-century-style epidemics.
However, the paper does highlight a sobering trend in human
epidemiology. Drug-resistant bacteria are here to stay, even if society
stopped abusing antibiotics right now. Furthermore, evolution doesn't
just work on bacteria. Levin explains, "the drug-resistant
mutations that aren't costly are the ones that will take over, not only
in the bacteria responsible for tuberculosis but also in [organisms]
responsible for other diseases."
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