The process represented a passage made in heaven for microbes: all blood flow was unobstructed, the local human immune system was shut down, and the destination—the insect midgut—was a very comfortable microbial ecology.77
Once in the insect’s midgut, microbes could swiftly multiply, make their way back up to the salivary gland, and be injected into an unwitting host. Or they might remain in the insect, exerting unusual pressures on the creature. For example, only female insects fed on blood: to ensure a plentiful supply of females, some microbes made their way to the insect’s ovaries, where they genetically manipulated the male chromosomes, ensuring that offspring would be female. The organisms would then be passed on to the adult insect’s female offspring, which would be born already infected.
Evolution was a very dynamic and active process for microbes housed in an insect’s midgut. Some viruses changed very slowly over time, probably because they possessed extremely accurate mechanisms for replication and repair of their genetic material. But there were insect-borne viruses that were capable of sorting and resorting their chromosomes, shuffling RNA about seemingly at random. And under conditions of co-infection of an insect by more than one species of microbe, exchanges might occur. The end result could be new mutant organisms.
Barry Beaty of Colorado State University in Fort Collins pointed out that it was a simple matter to get a single mosquito infected with two different strains of bluetongue virus. The virus, which produced disease in ruminant livestock, was comprised of ten RNA segments, or chromosomes, each of which had to be properly duplicated and assorted each time the virus reproduced itself. Beaty’s group showed that insects that were co-infected with two strains of bluetongue virus rarely injected back into animals a viral strain that was identical to either of the original strains: rather, it was a mélange of the two.78
Beaty noted that the usually mild snowshoe hare virus was in the 1980s producing serious human disease in northern Russia. It was due, Beaty said, to such a recombination event, only the exchange resulted, not in a wider vector range, but in greater virulence. On the basis of genetic analysis of the pertinent viruses, Beaty believed that the new Russian epidemic—which by 1992 was causing encephalitis in more than 100,000 people a year—was the result of a gene swap between the Inkoo and Tahyna viruses. The two parent viruses produced little more than mild flu-like symptoms in human beings, but their recombination proved potentially deadly.
How commonly other viruses, or bacteria, exchanged genetic material while inside vectors wasn’t known. Nor was it clear how significant a role such mutations might play in the emergence of new diseases.79
Many insect-borne viruses were thought to have originally been plant microbes that, thousands of millions of years ago, infected insects as they fed on plant nectar. In the 1990s, amid evidence of rising rates of genetic change in many plant microbes, concern was expressed about the possible emergence of new species that might be absorbed by insects. In such a scenario, a microbe that was genuinely new, to which humans had no natural immunity, might quite suddenly emerge. Genetic change in plant microbes was accelerating due to agricultural practices that exerted strong selection pressures on the microbes; to changing geography of plant growth due to international trading of plant seeds and breeding practices; and to the deliberate release of laboratory genetically altered plant viruses that were intended to offer agricultural crops protection against pests.80
To minimize use of toxic pesticides, and to prevent incurable viral diseases in plants, scientists in the 1990s were developing ingenious genetic means to protect plants. Using crippled viruses to carry genes that would help vital food crops fend off dangerous pathogens, researchers were breeding plants that could withstand a range of types of infections. There was a catch, however. Studies showed that, in nature, plants such as corn, wheat, and tomatoes were commonly co-infected with up to five different viruses, and those viruses could exchange genetic material.81 A review of 125 plant strains produced through such laboratory manipulation showed that 3 percent of the time the crippled virus that was used to carry such genes into plant cells could swap genes with other viruses in the plant, producing active, pathogenic—new—viral species.82
“Microbes are masters at genetic engineering,†wrote Canadian microbiologist Julian Davies.83 He was referring to mechanisms bacteria use to become resistant to antibiotics, but Davies’s comment could just as well apply to viruses in an insect’s midgut, malarial parasites responding to chloroquine, or influenza cyclically reinventing itself. That recognition prompted many virologists in the late 1980s to ask, “What is the likelihood that a truly new virus capable of causing human disease will emerge?â€
One approach to answering that question was to use molecular techniques to sequence the DNA or RNA of a group of viruses and try to trace their family trees, searching for evidence of such recombination events: if they had occurred in the past, it seemed logical that dangerous gene swappings might in the future occur again. Researchers in San Diego concluded that several human and animal retroviruses shared sequences of RNA, which could have been the result of crossover RNA recombination events. However, those events had to have occurred hundreds or thousands of years ago, because most of the retrovirus species were as different from one another as were humans from fungus.84
Nobel laureate Howard Temin took a different approach to answering the question, which, at the outset, he said was “inherently unpredictable.†Temin tried to calculate rates of mutations or incremental RNA changes in the human immunodeficiency virus type 1. Overall, he estimated that the HIV viruses made a significant mutational change in seven out of every 100,000 viral replications. Considering that an ailing person might have millions of HIVs in his or her body at any given moment, all of which were undergoing constant replication, Temin’s figure was far from comforting. But it was also not a worst-case scenario. Scientists discovered so-called hypermutation sites on HIV that were particularly prone to change: there, the RNA would mutate significantly in one out of every 1,000 viral replications.85
Temin doubted that such mutations would result in a more dangerous form of HIV—he felt the virus was already perfectly adapted by virtue of having combined certain lethality with a decade-long period of invisible infection during which the microbe would be passed on to other human beings. Still, he felt the essentially labile nature of the virus made its future incalculable.86
At the 1989 “Emerging Viruses†conference convened in Washington, D.C., by Rockefeller University and the National Institutes for Allergy and Infectious Diseases, another Nobel laureate, Joshua Lederberg, questioned Temin’s confidence that HIV’s rapid mutation rate probably wouldn’t result in greater viral virulence.
“My concern is not what we know, but what we don’t know,†Lederberg said. HIV was capable of infecting macrophage cells, he noted, asking, “Could the virus evolve the ability to infect macrophages in the lungs and, thus, become a respiratory disease?â€
At Caltech in southern California Jim and Ellen Strauss busied themselves with the task of mapping the evolution of all viruses whose genetic material was in the form of RNA. They concluded that all RNA viruses were descended from a single ancestor virus and that over the millennia the viruses had mutated a million times more rapidly than had their DNA-based hosts. Though rates of change varied from RNA virus to RNA virus, the Strausses were convinced that each and every one of the microbes had at some point come into existence through such a process.
“We now recognize that RNA viruses will continue to evolve rapidly as they have over the millennia,†the Strausses wrote. “As the recent epidemic of AIDS makes clear, new pathogens can and will arise.â€87
The Strausses felt that scientists, when referring to RNA viruses, shouldn’t really speak of species; rather, they should refer to “consensus sequences.�
� The rate of mutation was so high that RNA viral populations were actually pools of genetic concoctions, some particular form of which might dominate at any given time. It was a widely shared view. Many researchers spoke of “quasispecies†of viruses that moved about in “swarms.†John Holland, of the University of California at San Diego, felt that the high error rate of RNA polymerase, the enzyme responsible for making copies of RNA viruses, was the key to the extraordinary mutation rate. The polymerase was constantly “jumping†and “stuttering,†to use the official vernacular, to make different viruses.
“Natural selection among viruses isn’t about ending up with a specific genome that you call a species,†Holland said. “It’s about statistics.†More specifically, it was about the statistical odds that any specific genotype would dominate a “quasispecies swarm†at any given time.
RNA was nothing more than a long sequence of four different chemicals—base pairs, or nucleotides—the order of which comprised the genetic code. Microbiologist Peter Palese, of Mount Sinai School of Medicine, discovered in laboratory tests that if he examined a pool of 100 clones of flu viruses—clones being supposedly identical organisms—there were on average seven mutations for every 91.6 nucleotides. Similarly, in polio virus clones he found about one mutation in every 95.3 nucleotides.
The rate of mutation, of course, depended on the number of times the viruses reproduced themselves, as all the changes occurred during replication. That meant that mutations were most likely to occur among infectious microbes inside an extremely sick individual or when the rate of spread among people was very high.
“The greater the number of people infected,†Palese said, “the greater the rate of mutation.â€
As the Homo sapiens population swelled, greater opportunities would present themselves for both viral spread and mutation. It seemed perfectly reasonable therefore to assume that the evolution of microbes capable of infecting Homo sapiens would accelerate, perhaps dramatically.
Most mutations were, of course, deleterious to the individual microbes. But given such a high rate of change, it was inevitable that the microbes would occasionally hit on a mutation that increased their edge against the human immune system, gave them a wider range of cellular targets, allowed them to pass more efficiently from person to person, or rendered them in other ways more dangerous to Homo sapiens.
In laboratory studies these processes appeared to function by more stringent rules than pure happenstance. The polymerase and replicase enzymes that controlled replication of viral RNA and DNA seemed to jump about. The polymerase could be seen snaking its way down an RNA nucleotide chain like some molecular zipper slithering along the zipper track. But then the enzyme would jump tracks, taking the portion of new RNA it had already manufactured and joining it to another nucleotide stretch. The result would be a genetic hybrid. This had been seen with viruses as varied as polio and a microbe that infested tobacco plants. It sometimes occurred because there were “bumps†or “kinks†in the original RNA/DNA strand, which prompted the busy polymerase to jump tracks. These bumps themselves weren’t entirely random events, as they seemed to exist at significant points in the microbes’ genes.88
There were parts of microbial genomes that required constant mutation, particularly the genes that coded for proteins on their outer surfaces which were recognized by the human immune system. HIV, influenza, polio, schistosomes, Plasmodium falciparum, and staphylococci all had hypervariable mutation sites in the genetic regions that coded for such proteins. Change, at one pace or another, was essential to survival in the midst of human antibodies, T cells, and macrophages.89
Researchers at Louisiana State University felt they had evidence of “virus gumbos,†or mixtures of species of viruses which, combined, produced diseases that they seemed incapable of causing alone. In particular, they saw that some quasispecies of the feline leukemia virus were harmless to cats unless the cats were co-infected with other strains of the leukemia virus or the feline immunodeficiency virus (cat AIDS).90
In human beings similar “virus gumbos†were known to affect the AIDS disease process. Two herpes viruses, for example, were capable of directly activating replication genes inside HIV: herpes simplex Type 1 and HHV-6 (human herpes virus 6). There was even evidence that the polymerase zipper-jumping effect could occur inside patients, with strange hybrid viruses appearing: apparent mixtures of herpes and AIDS viruses.91
Cancer viruses were known to bring about their deleterious effects either by inserting themselves into specific oncogene sites in human or animal DNA or by manufacturing special proteins that switched on those cancer-causing genes. If the proper oncogenic signals were turned on (or switched off, in the case of cancer-suppression genes), a cascade effect might result in which other oncogenes and cellular signals were altered, ultimately transforming the cell into a cancerous entity. Most biologists believed that it was far from coincidental that such an intimate relationship between viruses and oncogenes had evolved over the millennia. And some went so far as to question whether genetic experiments with oncogenes and cancer cells might not result in infectious release of oncogene-carrying viruses or bacteria: perhaps altogether new species of viruses might result.92
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.93
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,94 chastised his colleagues for being “overly reductionist,†deriving too much from the fact there was one mutation in every 10,000 viral replications—in 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.â€
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.â€95
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.96 “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.
The Coming Plague Page 90