The Ultimate Parasite

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The millions of invaders are borne in by air or water. Judged by the ordinary signs of life--growth, motion, the need for food--they are dead. One by one, they find individual targets, which may be a thousand times bigger than their sub-microscopic selves. In such an unequal contest, they should have little chance. But these invaders are superlative saboteurs; they are virus particles, and their targets are cells in the human body.

Alongside the cell it is about to attack, the invader acts like an enemy agent skulking outside a big factory. Suddenly the virus slips into the complex living cell which, factory-like, has a definite schedule for receiving raw materials and processing them for the benefit of the whole organism. Either before or just after it slips in, the virus sheds its coat, a swatch of protein.

The coatless inner part of the virus is essentially a molecule of nucleic acid, but inside the factory it behaves like an immensely efficient agent taking over a captured industry. It seems to know where the production schedules and blueprints are--and it throws them away. In their place, it issues orders for the production of nothing but hundreds or thousands of copies of itself, plus an equal number of protein coats to fit. The cell-factory rushes to fill the massive order. It becomes strewn with waste materials. The strain tells. About the time the cell fills the invader's order and completes a myriad new molecules of nucleic acid, it falls into irreparable ruin. And just in time, the new molecules pick up their freshly tailored protein coats, leave the cell and go off in search of new cell-factories to conquer.

When a human victim of viral infection gets a stuffy head and a sore throat, or suffers a splitting headache and the feeling that his bones are breaking, or develops the blisters of cold sores or the rash of measles, his body is reacting to the biochemical disturbances that come from invasion by viruses. Viruses kill millions of people around the world every year, and give the miseries to hundreds of millions more.

Upstaging the Bacteria. Until only 20 years ago, the disease makers that dominated medicine's attention were the bacteria, hulking big microbes (by comparison with viruses) that generally attack by producing systemic poisons rather than by invading the body's cells. Antibiotics have wiped out or brought under control virtually all the major bacterial diseases: tuberculosis, some forms of pneumonia, diphtheria, scarlet fever, typhoid fever, gonorrhea, syphilis and most of the other illnesses that stir memories of Paul de Kruif's heroic Microbe Hunters.

But drugs cannot affect true viruses. The effect is to leave the virus diseases supreme, and virologists are multiplying with a speed reminiscent of the particles they study. Having only in the last dozen years dug out the mechanics of viral cell invasion, researchers now know of fascinating variations in the process. Occasionally and inexplicably; they have learned, viral nucleic-acid particles enter cells and "go underground," lying there dormant or masked for years. Or the nascent "pro-virus" molecules of new nucleic acid may play possum in this way. Virologists are having to coin words such as virion, capsid and capsomere to describe viral particles and their parts.

Virologists study a world of marvels, a world measured by the millimicron (a twenty-five millionth of an inch) and yet big enough to hold the mystery of life. How do a few hundred molecules from dead chemistry become a virus that can replicate itself, and mutate, and is thus most definitely alive?

In this short period of mushrooming knowledge, vaccines against one of the most feared of viral diseases, poliomyelitis, have been made and given to hundreds of millions of people. Influenza, which has staged two deadly worldwide onslaughts in this century, threatens a renewed attack this fall and winter, but virology has devised vaccines to fight it. There is solid hope that measles may soon be controlled, if not conquered outright, by a vaccine, no mean accomplishment. Long neglected and underrated, mainly because it is one of the supposedly unavoidable children's diseases, measles is a universal plague. In backward parts of the world it is a major killer. In the U.S. it kills an estimated 800 a year, and permanently disables 2,500. At an international symposium on measles in Washington last week, Nobel Prizewinner John F. Enders hailed the victory over measles. "Man," he said, "has been searching for an effective measles vaccine for over 200 years." Now, "by a fresh approach to an old objective." a measles vaccine has been developed that has proved effective in 96% of the children inoculated.

Virology's Thinker. Progress in virology is the work of many men, microbiologists, chemists and doctors of medicine, and in this task Harvard University's John Enders plays a unique part. He is, by virtue of temperament and academic qualifications, one of the deepest thinkers in virology. A philosopher of natural science, his contributions have been long-leap deductions and intuitions that guide other men's research, hypotheses that bypass a thousand experiments.

Enders' laboratory in the Jimmy Fund Building on Boston's Binney and Black-fan Streets, part of the Children's Hospital Medical Center, reflects the man and his methods. His corner office is lined with books, and contains a big table that offers plenty of room for papers and makes a good base for sandwich-luncheon conferences with his staff. Down one whole wall runs a long laboratory bench with a Bunsen burner that doubles for making instant coffee and two microscopes : a brass monocular antique that Enders keeps for sentiment's sake, and a modern binocular through which he checks virus damage to cells in tissue cultures. His group of laboratories, eight rooms, is small by modern, crash-program standards, and Enders runs the whole plant on a penurious $110,000 a year. As befits a man of some inherited wealth who cares little for what money buys, he lives comfortably but unostentatiously in a Brookline home with his second wife Carolyn.

"My Spirits Fail." He came late to the field he has made intuitively his own. John Franklin Enders was born in Hartford, Conn., in 1897, the son of a banker and grandson of a founder of Aetna Life Insurance Co. He has a childhood memory of Mark Twain, a friend of the family, coming to call in a characteristic white suit. From St. Paul's, where he rowed in the crew, played hockey and football. Enders went to Yale, only to have his education interrupted by World War I. He joined the Naval Reserve Flying Corps, became a flight instructor in a pontoon plane with a 50-h.p. engine.

After getting an A.B. at Yale, he briefly tried selling real estate (he flopped), went to Harvard to try for a Ph.D. in English. He started a thesis on the origin of genders, worked two years before he found that a student at Heidelberg had long since done the subject with unimprovable thoroughness. "I mouth the strange syllables of ten forgotten languages, letting my spirits fail, my youth pass," he youthfully wrote. Then a roommate, Australian Bacteriologist Hugh Ward, introduced John Enders to Hans Zinsser, Harvard University's professor of bacteriology and immunology, and one of the great fertilizing minds of his era (Rats, Lice and History for the layman. Infection and Resistance for the profession). Enders was then 30. "A man of superlative energy," Enders wrote of Zinsser after their first meeting at a hard-cider party: "A golden heart and sufficient intelligence."

Zinsser told Enders that if he would work up his chemistry and physics in the summer, he could have a laboratory assignment in the fall. Enders did not leap at the offer; he moves too thoughtfully and slowly for that. But he took it. In May 1928 he wrote to a friend: "This antipodal revolution of my studies has been of large value in helping me to obtain that Pisgah* sight of things and people that perhaps is the ultimate aim of my apparently inconsistent, faltering and obscure action." In 1930, at the age of 33, Enders got a Ph.D. in microbiology with a thesis on anaphylaxis (severe allergic reaction).

Through the Filter. Enders worked at first on tuberculosis and bacterial pneumonia. But in the early '30s there was growing awareness of the importance of smaller infectious particles, so small that, with negligible exceptions, they were invisible to medical researchers under even the strongest microscopes.

Edward Jenner had found, before 1800, an empirical method of protecting man against one dread disease now known to be caused by a virus: infection with cowpox ("vaccinia," hence the general term vaccination) would ward off later infection with the deadly and disfiguring smallpox (so called to distinguish it from syphilis, "the great pox"). Louis Pasteur achieved a similar triumph of empiricism. Unable to isolate the microbe of rabies, he simply assumed that it was too small to be seen and developed the Pasteur treatment for victims of bites by rabid animals.

Most of the bacteria studied by Pasteur and his early followers were big enough to be trapped in fine porcelain filters, devised by Pasteur's assistant Charles Chamberland, and to be seen under the 19th century light microscope. It was a temperamental Dutch botanist, Martinus Beijerinck (1851-1931), who found that whatever caused mosaic disease in tobacco plants could slip through the minute pores of these filters. In 1897 he concluded that this infectious, filter-passing fluid was a "filterable virus." The word virus had been loosely used for centuries to denote any "poison" that caused infectious disease.

For 40 years, the one clear mark of the virus was this ability to slip invisibly through porcelain filters. In those four decades, without waiting to see what a virus looked like, brilliant men did brilliant things about viruses and viral diseases. At Manhattan's Rockefeller Institute, Dr. Peyton Rous in 1910 proved that a filterable virus is the cause of sarcoma (a kind of cancer) in chickens. At Harvard and then at the Rockefeller Foundation, South Africa-born Max Theiler performed the delicate and dangerous feat of getting yellow-fever virus to grow in the brains of mice. With infinite patience, Theiler in 1936 grew 176 generations of virus in tissue cultures of chick embryo cells,* weakening the virus with each "pass" and seeking a generation that would be too feeble to induce the disease, yet strong enough in a vaccine to spur the system to create antibodies. The vaccine that he achieved won Max Theiler a Nobel Prize.

A Ton of Tobacco. But the study of viruses in the 1930s was still a toddler among the sciences; no U.S. university even had a chair in virology. Medical texts of the period were studded with such notations as: "The cause of this disease is believed to be a filterable virus, but has not been isolated," Virology needed new foundations to build on.

One appeared in 1938: the electron microscope, in which beams of electrons are focused sharply enough to take photographs of objects less than a millionth of an inch across. This made many virus particles visualizable--and another Rockefeller fellow had something to visualize. Indiana-born Wendell Stanley went back to Beijerinck's favorite, the tobacco mosaic virus, or TMV, and spent years in a Princeton laboratory cooking down a ton of sickly tobacco leaves, filtering and re-filtering, dissolving and redissolving, until he had isolated the cause of this economically costly disease. What he had to show for years of imaginative perseverance was about a teaspoonful of white crystals that looked no more impressive than powdered sugar. It was pure TMV, and the feat won Stanley a Nobel Prize in chemistry.

Stanley thus gave a crystal-clear answer to the question: What is TMV? Electron micrographs showed thin rod-shaped crystals, little more than a hundred-thousandth of an inch long. This answer raised an intriguing new question. Is a virus animate or inanimate, living or dead, animal or mineral? Dr. Stanley's way out of the dilemma is to broaden the definition of "living'' to include any particles that are capable of reproducing or replicating themselves. That covers viruses.

"If You're a Tulip." Thanks largely to chemists like Stanley (who now runs the University of California's Virus Laboratory) and the electron microscopists, a virus can now be defined as an infectious particle that has no metabolism of its own and reproduces itself only by taking over the metabolic processes of the living cell it invades. Viruses are the ultimate parasites. They parasitize everything in nature from bacteria and flowering plants up through invertebrates such as mosquitoes, and the vertebrates from fish, amphibians, reptiles, birds and mammals to man.

There is no such thing as a beneficial virus, though some do no harm. Whenever a virus has a detectable effect, it is bad. All healthy tulips are solid-colored. The tulip-streak (or "breaking") virus creates variegated color patterns of great beauty in the eye of the human beholder (and of great cash value to Dutch growers). But, says Stanley, "if you have the tulip-streak virus and you happen to be a tulip, you're sick.'' This lack of evident purpose in viruses leaves teleological philosophers at a loss. Yet viruses must have influenced evolution through natural selection. In deed, the close resemblance between the virus' core of nucleic acid and the gene, or "unit of heredity;" suggests that virus particles are lost genes in search of evolution.

No man knows how many viruses there are or how to classify them. John Enders and six other internationally famous virologists* have just made a stab at classification in Virology, conceding that they are making "some dogmatic statements and sweeping suggestions based on grossly inadequate knowledge." They recognize 400 viruses as infecting vertebrates, rate 50 of them, including rabies, as unclassifiable, and put the rest in six groups (see chart). They leave out the large "mantle viruses'' of parrot fever and trachoma, which are vulnerable to antibiotics and other drugs.

A Closer Look. Thus viruses got defined and classified. But just how the virus core gets into a cell remained a mystery, even after Dr. Robley C. Williams, a member of Stanley's California team, devised the method of plating the particles with gold or uranium to get clearer electron micrographs. Then, two years ago at Cambridge University's Cavendish Laboratory, Drs. Sydney Brenner and Robert W. Home made an illuminating refinement on electron micrography, revealing far more intimate details of virus structures and differences, and clues to how viruses work.

Influenza viruses were found to have horns or spikes. On some of these is an enzyme that can dissolve part of a cell's outer coating. Presumably, this is what the flu virus uses to open a hole in the cell-factory wall for its nucleic-acid core to slip through. A virus known as T2 bacteriophage (it attacks bacteria) was found to have a tadpole shape; the "tail" is like a coiled spring around a tiny hypodermic needle that stabs the cell wall, and through this the nucleic-acid core is injected. Micrographs show whether viruses are basically cubic or helical in structure. They also reveal that viruses may have an exquisitely complex symmetry around as many as five axes, and contain hundreds of submolecules, each of which may have a hollow hexagonal structure. Chemical tests show whether viruses have cores of ribonucleic acid (RNA) or deoxyribonucleic acid (DNA), and whether they have enzymes or fats in their coats.

The Vaccination Mechanism. The most practical results so far of virological research are vaccines, and vaccines depend on the basic concept of viral structure as a nucleic-acid core with a protein overcoat. The coat is a foreign substance to the body it invades, and in the higher animals, including man, the system fights back by making antibodies that gang up on a virus particle, surround it and neutralize it. Unhappily, it takes days or weeks for the body to mobilize its antibody police, so the first viral invasion is likely to succeed and make the invaded victim sick, or may even kill him. But if the body survives such an invasion, it learns to remobilize its defenses quickly, like emergency police, whenever it recognizes an old viral enemy or one wearing a similar protein overcoat.

Vaccination imitates the natural process for creating antibodies, using similar but less harmful viruses (cowpox instead of smallpox, for example) or weakened viruses, or even killed viruses. But in some individuals, and against some viruses more than others, the antibody memory is short --hence revaccination. If a virus mutates, as often happens with flu, a new vaccine containing the mutant must be prepared.

When John Enders got interested in measles in the 1930s, it was not clear even to him that he was sliding over from bacteriology to virology. "There was still conjecture as to whether measles was caused by a virus.'' he recalls. "Measles intrigued me as a problem that had been elusive for so long." Dr. Enders, working with Dr. William McD. Hammon, promptly ran into frustration of his own. The only animals that would catch measles were monkeys, and only a few of these. The researchers thought that they had got measles virus to infect a cat, only to discover that the animal had a different virus disease: cat distemper. This led to the production of a valuable veterinary vaccine, but not what Enders was looking for.

Enders tried again. Drs. Alice Woodruff and Ernest Goodpasture of Vanderbilt University had recently given virology (and vaccination) a big boost with the discovery that some viruses grow well in incubating eggs. Enders put fluid from a measles patient into eggs, but had no luck. Searching for a better medium, he turned his attention to embryonic tissue culture, sensing that growing viruses in live cells--the technique that Harrison pioneered--held unrealized possibilities.

Dr. Enders pondered while he puttered with cowpox virus in tissue cultures of chick embryo cells. Through an ordinary binocular microscope, he could see that the cells were damaged and began to fall apart as the virus multiplied. Others had seen this phenomenon; to the thoughtful Dr. Enders its significance eventually became clear and astonishingly simple: the nature and amount of cell damage were indexes to the nature and amount of viral activity. "It seems incredibly obvious now," he says.

A World War II project diverted Dr. Enders to mumps, which was feared as a menace to troops. He produced a moderately satisfactory killed-virus vaccine that helped him form his now firm opinion that a vaccine made from a live but attenuated virus is better than any made with killed virus: "I don't think you can do better than nature itself does when it gives you the disease." Back at tissue culture after the war, Enders was joined by two research fellows. Drs. Frederick C. Robbins and Thomas Weller. The work was bedeviled by bacteria contaminating the cultures, making them useless before a slow-starting virus multiplied. Virologists began adding antibiotics to their cultures to keep out bacteria; Enders hit upon a particularly successful combination of penicillin and streptomycin. Yet even in uncontaminated cultures, Enders failed to isolate and grow the obstreperous measles virus.

Culture in Human Embryo. Then came a lucky break. The lab happened to have some poliovirus tucked away. This had hitherto refused to grow except in brain cells, which are unsafe as a culture for a human vaccine because nerve-cell proteins can kill the vaccinated person. Enders suggested growing it in cultures of muscle and skin from human embryos recovered in therapeutic abortions. It worked. Watching the cell-damage effect, the Harvard researchers could see that the virus was multiplying. The virus could still cause paralytic polio. But when serum from a recent polio patient was mixed with the virus in tissue culture, the cells were protected. The antibodies were at work. In guarded, highly technical language in Science, meaningful only to other virologists, the three researchers reported their success.

It meant that poliovirus could at last be grown in a way to make a safe vaccine, and the discovery led the University of Pittsburgh's Dr. Jonas E. Salk to the next step, developing a formaldehyde-killed vaccine. It also meant a 1954 Nobel Prize, which Enders insisted that Robbins and Weller share with him equally.

Success in Measles. An ever more insistent backer of live-virus vaccines, Enders was a bit dismayed that the U.S. took up killed-virus polio vaccine with such zest. He experimented for a while attenuating poliovirus, sent a sample to the University of Cincinnati's Dr. Albert Sabin (who went on to make workable live-virus polio vaccines), then turned back to basic research. In 1954 another of his research fellows, Thomas Peebles, fulfilled Enders' longstanding dream of growing measles virus (obtained from a prep school student named David Edmonston) in tissue culture. This time, aiming for a safe and effective live-virus product, Enders decided to keep control of the vaccine project in his own lab.

There Peebles and other research fellows worked with infinite patience for two years, domesticating the virus. After 72 generations, the measles virus was adjudged sufficiently attenuated for safe trial in children. Dr. Samuel Katz, now No. 2 man in Enders' lab, gave some of the first shots to his own youngsters.

Last week's symposium reports made it clear that Enders' measles vaccine is effective in stimulating production of antibodies that should give permanent immunity. But in many children the vaccine causes something like a mild case of measles. A few youngsters get the rash, but more get a fever that may run as high as 104DEG or 106DEG. Strangely, this fever does not make the youngsters ill, and few of them need even to stay in bed. Doctors generally agree with Virologist Enders: if the facts about the dangers of natural measles are fully explained, parents will accept these side effects of vaccination without too much fuss. To be on the safe side, some physicians are giving, along with the vaccine, a shot of gamma globulin, hoping that it contains enough measles antibodies to cut down the rash and fever. But this may also curtail the immunity. Some manufacturers are preparing a killed-virus measles vaccine.

But usable vaccines are hardly the whole measure of the Enders team's technical breakthrough in tissue-culture methods, now 13 years old. When the method was devised, only 13 viruses that cause disease in man had been identified. Now at least 58 others have been cultured, plus about 300 that infect animals, most of them grown by Enders' methods. They range from a score or more of common cold viruses to more severe respiratory diseases (including some types of viral pneumonia). They include respiratory-intestinal infections, and apparently even infectious hepatitis.

The Cancer Challenge. The big prize in the pursuit of viruses is cancer. When Peyton Rous, 50 years ago, reported his chicken-sarcoma virus, he was scoffed at because it implied that a cancer (at least in fowl) was infectious, and every medical scientist of a half-century ago "knew" that cancer was not an infectious disease. Today, though none of them believe that cancer is infectious in the same way as measles, polio or flu, thousands of researchers are working feverishly in response to the challenge thrown down in 1956 by Wendell Stanley: if animal cancers are caused in some way by viruses, why not human cancers? Prove it or disprove it!

Virus particles have been found in some human cancers, but this does not prove that they caused the disease. Some tumor viruses invade an animal, yet they disappear for months or years, and then belatedly cause cancer. The hows and whys of this latent period are unknown. One partial explanation may lie in the ability of new "provirus" particles to remain undetected in cells, doing no evident damage until they are stimulated by chemicals or X rays. The important thing is that these nucleic-acid molecules can be infective by themselves, with no assist from the protein that normally accompanies them in the whole virus. Dr. Frank L. Horsfall, director of Manhattan's Sloan-Kettering Institute, the world's biggest private cancer research organization, sums it up: "We can now speak of infectious molecules in animal as well as plant diseases--something that was inconceivable only a decade ago."

It may be that in some unpredictable cases, a molecule of viral nucleic acid, without its protein overcoat, so closely resembles a gene that it can slip into the cell's chromosomal lineup, displacing a normal gene, and make the cell reproduce abnormally. Most of the resulting abnormal cells would probably die, but a few might retain the power to run wild and perpetuate themselves as cancer.

Where these infectious molecules might come from and what might trigger them into activity at unpredictable times are still mysteries. Perhaps they are inherited, and lie dormant for decades. This would go far to explain why some cancers, though not hereditary in the ordinary sense, tend to run in families. Or they may come from virus infections of the mother during pregnancy: if they cross the placental barrier, they could lodge in the fetus, which has little or no antibody-forming mechanism to reject them.

Triggering mechanisms are still more obscure. Stanford University's Radiologist Henry Kaplan has shown that if he gives a dose of X rays to seemingly virus-free mice, they develop cancers containing virus particles. The late Dr. Francisco Duran-Reynals argued that chemicals and viruses combine to cause cancer. Now many laboratories are confirming his basic thesis: mice painted with a low dose of a known carcinogen (cancer-causing chemical) get no tumors, and neither do those exposed only to viruses; but if mice get both the virus and minute amounts of the chemical, many of them soon develop cancer.

Interferon. From the chemists' laboratories there has come no drug that will selectively attack viruses while sparing the cells in which they seek sanctuary. But nature suggests that there is a way. If it takes days or weeks for protective antibodies to develop, why does not the pullulating virus overwhelm all the victim's susceptible cells in the meantime? London's Dr. Alick Isaacs last year found a partial answer. Virus-infected cells produce a substance that Isaacs calls interferon, which spreads to neighboring, uninfected cells. With their interferon guard up, these cells are unusually resistant to viral invasion.

Interferon is not a specific antiviral defense. Dr. Isaacs believes, but one of the body's general defenses. Just what it is or how it works, nobody knows. But the search is being pressed in many labs, including Enders'. Last week Enders told the Washington measles conference that he and a research fellow have just found evidence suggesting that interferon is important in changing a dangerous virus like that of wild measles to the tame Edmonston strain useful for vaccines. In such ways, the findings of many far-flung laboratories fit together, building largely on the Enders cultures that exposed so many viruses to isolation and attack. Once a starveling science that at any given time occupied only a few dozen men, virology has thus fanned out to become the task of thousands moving massively against man's diseases and toward life's most elusive secrets.

* The mountain from which Moses viewed the Promised Land in Deuteronomy 34:1.

* The feat of growing living tissues outside the body, in a bath of blood fractions or chemical nutrients, was first achieved by Dr. Ross G. Harrison (1870-1959) at Johns Hopkins University in 1907. Alexis Carrel later publicized it. Many scientists, among them Enders, believe that Harrison rated a Nobel Prize for his work.

* Including Britain's Sir Christopher Andrewes. Australia's Sir Macfarlane Burnet. Russia's Viktor M. Zhdanov.

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