The Coming Plague (105 page)
Authors: Laurie Garrett
DNA viruses also possessed special stretches of nucleotides that seemed to command polymerases to act with greater care, making accurate, often multiple copies of the stretch of genes located next along the “zipper.” These DNA sections were called “enhancers.” Studies showed that a key viral characteristic coded for by enhancers was the infectivity of the virus: the range of cell types it could invade and the ways in which the microbe could spread.
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Given such a vast range of mutational options for change, Harvard's Dr. Bernard Fields asked his colleagues, “Why haven't viruses wiped out all life on Earth?” It was, he felt, the crucial question.
And the answer, Fields said, lay in the difference between studying viruses in test tubes and studying them in the animals or humans that they infected. The venerable scientist, who had written the book on virology,
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chastised his colleagues for being “overly reductionist,” deriving too much from the fact there was one mutation in every 10,000 viral replicationsâin a test tube. In the real world those mutants still had to deliver their genetic payloads to the proper types of cells inside an animal or person in order to cause disease. And that necessary leap proved too great an obstacle for most mutant microbes, he said.
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chastised his colleagues for being “overly reductionist,” deriving too much from the fact there was one mutation in every 10,000 viral replicationsâin a test tube. In the real world those mutants still had to deliver their genetic payloads to the proper types of cells inside an animal or person in order to cause disease. And that necessary leap proved too great an obstacle for most mutant microbes, he said.
On the macrolevel, as Fields called it, little was well understood. There was no discipline of microbial ecology dedicated to studies of the behavior of microbes inside the human body.
“That's the big black box,” Fields said, “and it's the secret to evaluating all the conjectured risks of emergence of a new pathogenic virus.”
Some of the uncertainties in that black box included knowing how, exactly, viruses gained entry into the human body via alveoli in the lungs, M cells in the intestines, or lymphatic cells in the bloodstream; the roles various immune system chemicals played in either stifling or promoting viral activity; how viruses got past the thick membrane of cellular nuclei and past the chromatin mélange of proteins and carbohydrates to gain access to the host's DNA; which host chemical systems viruses exploited to their advantage; and how viruses got back out of hosts in order to be spread to other animals or humans.
Also in the black box were factors that seemed to make hosts more susceptible to viruses, Fields said, such as starvation, stress, and additional disease burdens. Though the catchall phrase “lowered immune response” was traditionally used to sidestep the mystery, little was known at the microbial level about how such factors influenced events. A starving child might make less protective mucus for his intestinal and stomach linings, for example, exposing more M-cell receptors to passing viruses. Was that a genuine phenomenon in nature, linking starvation and disease? Or was the mucosa depleted as a result of the infection?
“We know what many of the instruments are,” Fields said, “but we haven't a clue about the orchestration. The problem isn't a flute problem; it's an orchestra problem.”
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Virtually all the questions raised about viruses could also be directed toward bacteria and parasites, although the black box was somewhat smaller. There was plenty of evidence that bacterial and parasitic mutations were occurring in nature and that they often conferred new advantages on the microbes. Resistance to antibiotics and antimalarial drugs spoke volumes on the matter.
There, debate centered on the question originally raised in 1988 by John Cairns: was all bacterial mutation random, or were there directed changes that occurred in response to specific environmental pressures in the microbe's ecosphere?
The general dogma had it that such evolutionary events were random. New types of organisms emerged by chance mutations and haphazard genetic exchanges. If chance favored a certain type of organism when it surfaced, the microbe would thrive. In the meantime, the endless DNA dance of transposons, mutations, plasmids, and sexual conjugation went on, its pace essentially unaltered by environmental events. Genes shuffled and recombined, swapped and moved, whether or not the microbes were threatened.
“All DNA is recombinant DNA,” said the bible of biology,
Molecular Biology of the Gene
.
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“Genetic exchange works constantly to blend and rearrange chromosomes.” The emergence of, say, a penicillin-resistant
Streptococcus
was a “rare event, typically occurring in less than one per million cell divisions.” Of course,
Streptococcus
underwent more than a million individual cell divisions in twenty-four hours, starting with a single bacterium and expanding exponentially.
Molecular Biology of the Gene
.
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“Genetic exchange works constantly to blend and rearrange chromosomes.” The emergence of, say, a penicillin-resistant
Streptococcus
was a “rare event, typically occurring in less than one per million cell divisions.” Of course,
Streptococcus
underwent more than a million individual cell divisions in twenty-four hours, starting with a single bacterium and expanding exponentially.
Multiply resistant organisms that carried plasmids with dozens of advantageous genes were also considered the result of chance mixtures of DNA pieces that combined and recombined over microbial generations.
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If randomness was at the root of microbial evolution, humanity needn't fear unexpected changes in the rates of emergence of new mutants.
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If randomness was at the root of microbial evolution, humanity needn't fear unexpected changes in the rates of emergence of new mutants.
Well before scientists appreciated the extreme mobility of discrete pieces of DNA, seminal laboratory experiments were done with
Escherichia coli
proving that mutant abilities to withstand attacks from either viruses or antibiotics
preceded
the appearances of those threats in the bacteria's environment.
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Roughly one out of every 10 million
E. coli
in a petri dish might randomly mutate to be resistant to, say, penicillin. Then, if the drug were poured into the petri dish, 9,999,999 bacteria would die, but that one resistant
E. coli
would survive, and divide and multiply, passing its genes for resistance on to its progeny.
Escherichia coli
proving that mutant abilities to withstand attacks from either viruses or antibiotics
preceded
the appearances of those threats in the bacteria's environment.
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Roughly one out of every 10 million
E. coli
in a petri dish might randomly mutate to be resistant to, say, penicillin. Then, if the drug were poured into the petri dish, 9,999,999 bacteria would die, but that one resistant
E. coli
would survive, and divide and multiply, passing its genes for resistance on to its progeny.
In 1988, however, John Cairns of the Harvard School of Public Health challenged that central dogma of biology.
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Using recombinant DNA techniques, his laboratory made a set of specific
E. coli
mutants that had unusual nutritional needs. They then altered the bacteria's environments, making them deficient in chemicals the mutants couldn't manufacture on their own. And they showed that the
E. coli
would specifically change two separate sets of genes to adapt to the situation and survive, doing so in far less time than random mutation would permit.
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Using recombinant DNA techniques, his laboratory made a set of specific
E. coli
mutants that had unusual nutritional needs. They then altered the bacteria's environments, making them deficient in chemicals the mutants couldn't manufacture on their own. And they showed that the
E. coli
would specifically change two separate sets of genes to adapt to the situation and survive, doing so in far less time than random mutation would permit.
“That such events ever occur seems almost unbelievable,” Cairns wrote, “but we have also to realize that what we are seeing probably gives us only a minimum estimate of the efficiency of the process, since in these cases the stimulus for change must fairly quickly disappear once a few mutant clones have been formed ⦠. It is difficult to imagine how bacteria are able to solve complex problems like theseâand do so without, at the same time, accumulating a large number of neutral and deleterious mutationsâunless they have access to some reversible process of trial and error.”
Cairns used computer metaphors to describe what he believed was going on in the microbial world. The essential genetic material that made an
E
.
coli
an
E. coli
was the organism's hard disk. The bacteria had an almost endless number of ways to scan that basic disk, turning off and on various genetic programs and data bases. Plasmids and transposons were “drifting floppy disks,” carrying additional bits of genetic data and programming.
E
.
coli
an
E. coli
was the organism's hard disk. The bacteria had an almost endless number of ways to scan that basic disk, turning off and on various genetic programs and data bases. Plasmids and transposons were “drifting floppy disks,” carrying additional bits of genetic data and programming.
There was a limit, Cairns argued, to how large any given organism's hard disk could be. Furthermore, energy needs placed restrictions on how many genes could be expressed, or turned on, at any given time. Some genetic programs would remain silent most of the time, stored against emergencies in the bacteria's data bank. Among those, he felt, were programs that actually ordered mutations, or changes, in elements of the basic hard disk. Since the bacteria couldn't afford to contain enough DNA to carry programs in anticipation of every possible crisis, a direct mutation command was, Cairns argued, the next-best alternative.
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The British-born biologist was convinced that such mechanisms were at play in some cases of drug resistance, as well as microbial evasion of the human immune system. In laboratory experiments it was possible to induce lysisâor ruptureâof bacterial cells and see the microbes' DNA “hard disk” flood into the fluid petri dish. There, other healthy bacteria would absorb the roaming DNA. And if antibodies were added to the mixture the scavenger bacteria would use the newly absorbed DNA to make new proteins to coat their membranes. In this way, the bacteria would disguise themselves from the antibodies, successfully evading immune system attack.
Even the scavenging activity was less random than it seemed. Studies by Rockefeller University's Alexander Tomasz of
Neisseria gonorrhoeae
and
Hemophilus influenzae
showed that these organisms had special proteins on the outer surface of their cell walls. The proteins scanned passing DNA, looking for useful genetic sequences. When something good drifted past, the protein grabbed it and pulled the DNA into the bacterium. And the pneumococci, which absorbed any “promiscuous DNA,” as Tomasz called it, had a special internal enzyme system that scanned the scavenged genetic material and rejected useless chunks of DNA.
Neisseria gonorrhoeae
and
Hemophilus influenzae
showed that these organisms had special proteins on the outer surface of their cell walls. The proteins scanned passing DNA, looking for useful genetic sequences. When something good drifted past, the protein grabbed it and pulled the DNA into the bacterium. And the pneumococci, which absorbed any “promiscuous DNA,” as Tomasz called it, had a special internal enzyme system that scanned the scavenged genetic material and rejected useless chunks of DNA.
There were, by 1992, several identified “mutator alleles” along the
E. coli
genomeâsites in the hard disk that ordered neighboring programs to alter themselves. And under experimental conditions it was possible to see a sort of “trial and error” mechanism in play, in which the microbe rejected useless or harmful mutations, but placed beneficial mutations in its permanent bacterial hard disk.
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E. coli
genomeâsites in the hard disk that ordered neighboring programs to alter themselves. And under experimental conditions it was possible to see a sort of “trial and error” mechanism in play, in which the microbe rejected useless or harmful mutations, but placed beneficial mutations in its permanent bacterial hard disk.
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A number of stress proteins were discovered in microbesâproteins (or the genes that coded for them) that were activated when the cell was challenged by a range of threats: heat, fevers, some human hormones, arachidonic acid (an immune system activator), and a variety of human disease states. When activated, these proteins acted rapidly to protect vital biochemical functions inside the microbe. Termed “molecular chaperones,” the proteins guided fragile compounds through their duties. The stress proteins could be turned on and off experimentally by inflicting definable changes upon their environments. There was no clearer example of a microbe's adaptation to its environmentâadaptation that required genetic as well as chemical change.
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Studies of vancomycin resistance in
Staphylococcus aureus
strains found in a handful of European clinical settings revealed that seven separate genes were required to render the bacteria invulnerable to the drug. The seven genes prompted one simple alteration in the chemistry of the microbe's cell wall, replacing an ester bond in a structural protein with an amide one. The ester bond was the target for vancomycin.
Staphylococcus aureus
strains found in a handful of European clinical settings revealed that seven separate genes were required to render the bacteria invulnerable to the drug. The seven genes prompted one simple alteration in the chemistry of the microbe's cell wall, replacing an ester bond in a structural protein with an amide one. The ester bond was the target for vancomycin.
Here was the amazing thing: those seven resistance genes were switched on only when vancomycin was in the bacteria's environment. How the bacteria knew of the threat's presence was an utter mystery.
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Researchers noted that many extremely divergent microbial species shared genetic signaling sites, called operons, that with very minor mutation conferred multiple antibiotic resistances on the organisms. For example, seven very different microbes (
E. coli, Salmonella, Shigella, Klebsiella, Citrobactr, Hofnia
, and
Enterobactr
) naturally shared an operon which, with a single point mutation, made the organisms resistant to tetracycline, chloramphenicol, norfloxacin, ampicillin, and quinolones.
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In Cairns's terms, this implied that all seven bacterial species shared a few bytes of hard disk space that was specifically designed to undergo a single data-bit alteration when necessary to respond to an antibiotic threat.
E. coli, Salmonella, Shigella, Klebsiella, Citrobactr, Hofnia
, and
Enterobactr
) naturally shared an operon which, with a single point mutation, made the organisms resistant to tetracycline, chloramphenicol, norfloxacin, ampicillin, and quinolones.
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In Cairns's terms, this implied that all seven bacterial species shared a few bytes of hard disk space that was specifically designed to undergo a single data-bit alteration when necessary to respond to an antibiotic threat.
Studies of various pathogenic
E. coli
strains showed that there was often a trade-off between genes for extreme virulence and those for antibiotic resistance. Rarely could the organisms carry enough genetic baggage to render them both highly lethal and resistant. Highly virulent strains didn't usually need resistance genes, however, because they could produce diseaseâand reproduce themselvesâso rapidly that
Homo sapiens
didn't have the opportunity to make antibodies before the bacteria had accomplished their essential tasks of invasion, reproduction, and spread.
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E. coli
strains showed that there was often a trade-off between genes for extreme virulence and those for antibiotic resistance. Rarely could the organisms carry enough genetic baggage to render them both highly lethal and resistant. Highly virulent strains didn't usually need resistance genes, however, because they could produce diseaseâand reproduce themselvesâso rapidly that
Homo sapiens
didn't have the opportunity to make antibodies before the bacteria had accomplished their essential tasks of invasion, reproduction, and spread.
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While infectious disease biologists debated questions of random versus directed mutations among the microbes, the overall evolutionary role of jumping genes was the subject of great debate among biologists of all stripes. Some scientists had, by the 1990s, come to believe that transposons and plasmids were a driving forceâperhaps
the
driving forceâof evolution, even in plants and animals. The grand biological soup of shifting genes, it was suggested, was constantly giving one creature the capabilities normally carried by another. Human beings, in fact, might be nothing more than four billion years of gene jumping.
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the
driving forceâof evolution, even in plants and animals. The grand biological soup of shifting genes, it was suggested, was constantly giving one creature the capabilities normally carried by another. Human beings, in fact, might be nothing more than four billion years of gene jumping.
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But pure random chaos in such a mutation soup seemed terrifying. How could any species survive if its cells absorbed any chunk of DNA that came their way, no matter how dangerous it might be? Most random mutations were lethal, or at least deleterious, to the altered organism.
A series of startling experiments performed in a variety of laboratories during the early 1990s significantly raised the stakes of that debate. Amber Beaudry and Gerald Joyce, of the Scripps Research Institute located in southern California, succeeded in forcing a particular protein, called a ribozyme, to evolve in a test tube. Normally the ribozyme's job was to make specific cuts and slices in the organism's RNA. But Beaudry and Joyce showed that after ten generations of reproduction the ribozyme could mutate, becoming capable of chopping DNA as well.
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