BETTE HILEMAN, C&EN WASHINGTON

The transmissible spongiform encephalopathies (TSEs)--a very unusual group of neurodegenerative diseases involving microscopic holes in the brain--affect both humans and animals. Cattle with bovine spongiform encephalopathy (BSE) or mad cow disease, lose weight, have difficulty standing, drool, wave their heads, and threaten other animals. A few months after symptoms appear, the cattle die.

CONTAINMENT Cattle throughout Europe are now being prepared for slaughter in an attempt to control the mad cow epidemic.

COURTESTY OF USDA

People with a similar disease known as new-variant Creutzfeldt-Jakob Disease (vCJD), which is almost certainly caused by the same infectious agent as BSE, become depressed and paranoid, lose the ability to walk and swallow, grow blind and comatose, and eventually die. Both diseases are incurable and, in most cases, can be diagnosed positively only after death. The latency period between exposure and disease can be as short as 10 years and may be as long as 60 years.

Since it first appeared in the U.K. in 1986, BSE has killed about 200,000 British and European cattle, and since 1996, 98 cases of vCJD have been suspected or confirmed in Britain, France, and Ireland. The human victims seem to have contracted the disease from eating beef contaminated with nervous system tissue, and they have had an unusual daisylike floral pattern in the brain in which an amyloid plaque core is surrounded by "petals" of spongelike tissue. The pathologic agent known as a prion that kills the cattle is nearly identical to the prion that claims victims of vCJD.

For years after BSE first appeared in Britain, authorities believed the disease would not spread beyond the U.K. They also believed it would not jump species to infect humans, reasoning that it would behave like scrapie, a similar prion disease, which has existed since about 1730 and apparently has been confined to sheep and goats.

They were wrong. Over the past year, BSE has cropped up in 12 European countries and as far afield as Canada and Oman. It is not known how many human victims vCJD will eventually claim.

According to the U.S. Food & Drug Administration, two factors led to the dissemination of BSE to countries outside the U.K.: Britain continued to export livestock after the epidemic began, and it rendered BSE-infected animals into meat and bone meal and sold it around the world as cattle feed. Britain stopped feeding rendered cattle to other cattle in 1988, but for eight years it continued to ship infected meat and bone meal to more than 80 countries. Asian nations bought nearly a million tons from 1988 to 1996. The U.S. bought 21 tons in 1989 until a new regulation banning feed imports from the U.K. went into effect. Compounding the problem, Britain exported 3.2 million live cattle to 36 countries on every continent between 1988 and 1996.

HEALY

In late January, the United Nations Food & Agriculture Organization (FAO) said BSE may be a global threat. It warned countries around the world to be concerned about BSE and vCJD. "All countries which have imported cattle or meat and bone meal from Western Europe, especially the U.K., can be considered at risk from the diseases," FAO said.

  SO FAR, BSE has not shown up in U.S. cattle. Some experts believe safeguards already in place will prevent it. The Council on Agricultural Science & Technology says the risk of BSE infecting U.S. cattle is "extremely low." Federal and state agencies "have taken aggressive action to reduce the risk that BSE could be introduced and spread in this country," FDA says.

Other researchers say current regulations are still not strong enough. They warn that the contaminated feed imported from Britain after the appearance of mad cow disease may yet infect U.S. cattle, or that people could contract vCJD from nutritional supplements made from BSE-infected bovine tissues.

Some scientists are also concerned that blood donations from people unknowingly infected with vCJD will contaminate part of the blood supply. And they worry that if BSE does show up in U.S. cattle, it may spread quickly because the U.S. still renders cattle into feed for pigs and poultry, and sometimes this feed is inadvertently given to cows. Most "downer" cattle--those who can't stand up and are at highest risk of having BSE--are taken directly to rendering plants without being tested for BSE.

Moreover, experts worry that a similar prion disease called chronic wasting disease (CWD), now infecting deer and elk in Colorado and Wyoming, could sicken humans or cattle. State notification systems for warning hunters not to eat deer or elk that appear diseased are weak. And elk meat from herds in which a few animals are sickened by CWD has occasionally entered the human food supply.

"The U.S. should expand its relatively small national surveillance center to better ascertain the presence of vCJD and BSE in suspected cases," says Bernadine Healy, president of the American Red Cross. "Now is the time to establish well-designed banks of tissue from routine tonsillectomies, appendectomies, and lymph node dissections to define and follow the prevalence of prions in the U.S. population," she continues. Healy was director of the National Institutes of Health from 1991 to 1993.

In animals, the major TSEs are BSE, scrapie in sheep and goats, CWD in deer and elk, transmissible mink encephalopathy, and feline spongiform encephalopathy. Some of the human spongiform encephalopathies are vCJD, sporadic or classic CJD, and kuru. On a worldwide basis, sporadic CJD occurs spontaneously in about one person per million each year, usually in victims over 55. An estimated 300 Americans die from the disease annually.

Prions, the causative agents in all TSEs, are unusual pathogens. They are not viruses, bacteria, protozoa, or fungi. They are a unique naked protein that has no nucleic acid--that is, no DNA or RNA--and resists digestion. The pathologic prions isolated from BSE-infected cattle and human cases of vCJD have identical biological features, but the numbers of amino acids differ somewhat, says Paul W. Brown, chairman of FDA's TSE Advisory Committee.

Unlike microbes, prions are not completely destroyed by sterilization, autoclaving, disinfectants, radiation, or cooking, and they remain intact for years after an infected animal is buried. Some of them also survive the heat applied (about 270 ºF) when cattle remains are rendered into meat and bone meal, and some remain intact even at 600 ºF. They are totally degraded only with incineration at temperatures greater than 1,000 ºF or treatment with strong sodium hydroxide solutions. Therefore, most cattle exposed or infected with BSE are now incinerated.

According to the most widely accepted theory, prions have a unique method of replication. First, these uniquely twisted molecules, which are merely a misfolded form of normal prion protein, enter the host through BSE-infected beef or some other means. Then, they contact normal prion proteins (PrP^c) on the membranes of nerve cells and induce the benign molecules to change shape. The originally benign protein molecules, now misshapen and abnormal, continue the process, inducing more prion proteins to change shape. As the chain reaction continues, the infective prions build up in the brain and eventually cause microscopic holes. Because the infective prions are insoluble and resist enzyme digestion, they are called protease-resistant prions.

After a human is exposed to prions from BSE-infected beef or by some other route, 10 or 15 years may pass before symptoms appear. Since prions are not microorganisms, they do not evoke the production of antibodies by the host animal, and prion diseases cannot be diagnosed by the routine blood and serum tests used to detect microbial infections.

Currently, the only definitive way to diagnose TSEs is through postmortem tests on brain samples. These tests are cheaper and faster than they were even a year ago (see page 38). But what would be most useful is a sensitive blood test that could be used on a live animal or human before symptoms even appear.

"A lot of groups are trying to develop a reliable blood test," Healy says. "Most of the work is being done in commercial labs and therefore tends to be proprietary. It is impossible to know how close we are," she explains. It is important for the government "to aggressively invest in finding a better, more sensitive test--a blood test or some other body-fluid test," she says.

When a human or animal is infected with one of the TSEs, prions move through the lymph system before they infect the brain. In an infected mammal during various stages of the disease, prions can be found in the spinal cord, spleen, tonsils, intestines, lymph nodes, eyes, and eventually the brain, but they have not been found in the muscle or skin. Until recently, there was very little solid evidence that prions would infect the blood.

However, in 1999, as a precautionary measure, FDA banned blood donations from people who had spent more than six months in the U.K. between 1980 and 1996. The agency wanted to guard against the possibility that people who contracted vCJD from eating BSE-infected meat in the U.K. would donate blood and transmit infective prions to others via pooled blood.

LAST SEPTEMBER, scientists presented strong, though not conclusive, evidence that BSE can be transmitted through blood transfusions, at least in sheep. Researchers at the Institute of Animal Health in Edinburgh, Scotland, infected a sheep with BSE via the oral route with brain from an infected cow. Before symptoms appeared, blood was transferred from this sheep into a healthy sheep. The healthy sheep then developed symptoms typical of scrapie. "This report suggests that blood donated by symptom-free vCJD-infected human beings may represent a risk of spread of vCJD infection among the human population," Chris Bostock wrote in the Lancet [356, 999 (2000)].

In January, FDA's TSE Advisory Committee recommended that the restriction on blood donations from people who have lived in the U.K. for six months or more be extended to people who have spent 10 years or more in Ireland, France, and Portugal since 1980. The American Red Cross, however, goes even further. It will not accept blood from potential donors who have lived anywhere in Western Europe for six months or more since 1980. "There is enormous medical uncertainty about whether vCJD can be transmitted with blood. But the research on sheep and other research in hamsters indicate a low-level potential for transfusion-related infection," Healy says.

Demented Cows
U.K. has vast majority of cases
COUNTRY	CASES
U.K.	180,501
Ireland	587
Portugal	509
Switzerland	366
France	218
Germany	31
Belgium	22
Spain	12
Netherlands	9
Denmark	3
Liechtenstein	2
Italy	2
Omana	2
Luxembourg	1
Canadaa	1
Falkland Islandsa	1
NOTE: Data as of Feb. 6. a Bovine spongiform encephalopathy in cattle imported from the U.K.
SOURCE: Office of International Epizootics

Another way humans might conceivably be infected with mad cow disease is via vaccines made with bovine materials, such as blood and fetal calf serum. FDA's Center for Biologics Evaluation & Research has been concerned about eliminating any potential for contamination of biological products with BSE prions. To minimize that possibility, FDA in 1993 and again in 1996 recommended that vaccine manufacturers not use materials derived from cattle from countries where BSE is known to exist. In 1998, this list of countries was expanded to include not only those where BSE exists, but those where BSE surveillance is inadequate.

However, some companies did not follow FDA's recommendations and continued to use bovine-derived materials from the prohibited list of countries for some of their vaccines. These, according to FDA, include Aventis Pasteur, North American Vaccine Inc. (now a unit of Baxter International), Lederle Laboratories, and GlaxoSmithKline. In some cases, the companies have no records indicating where the bovine materials came from. FDA has requested that these companies replace all the vaccines made from prohibited bovine materials or unknown source materials with vaccines made from bovine materials from approved countries.

The companies have now agreed to stop using suspect materials, FDA says. But the existing stock of vaccines will still be used until the firms reformulate the products, a process that will be completed by 2003.

A statement on FDA's website recommends that all children and adults continue to be immunized according to accepted immunization schedules, even if the raw materials for the vaccines being administered are from an unapproved country. The agency's recommendations were made to "reduce even the remote potential risk of vCJD," FDA says. There is not enough risk from the vaccine to justify a recall, FDA says.

CWD in captive and wild deer and elk may be the most mysterious and intractable of the prion diseases. In the long run, it may stay confined to deer and elk. On the other hand, it may eventually cross species barriers into livestock or into humans who eat venison. Some experts consider CWD the most likely way by which a mad-cow-like disease could infect people in the U.S. "I think the best available science says there is a risk of transmission of CWD to humans. No human or animal should eat any product from an animal with any prion disease," says Thomas Pringle, a biochemist who heads the Sperling Biomedical Foundation, Eugene, Ore.

CWD is characterized by listlessness, blank facial expression, repetitive walking in set patterns, excessive drinking and urination, and a long irreversible weight loss that eventually kills the victim. It first appeared in captive deer in Colorado in 1967, and over a 10-year period, it showed up in deer and elk in wildlife research facilities in both Colorado and Wyoming.

Although state officials stopped the movement of animals from these facilities soon after CWD was diagnosed as a TSE, the disease spread to free-ranging deer and elk populations in three states--a total of about 14,600 sq miles. It has also spread to elk farms in five mountain and midwestern states, and the Canadian provinces of Alberta and Saskatchewan.

  "AT PRESENT, there appear to be two distinct epidemics," says Michael Miller, a veterinarian at the Colorado Division of Wildlife. "One epidemic focuses on free-ranging cervids [deer and elk] in southeastern Wyoming, northeastern Colorado, and extreme southwestern Nebraska," he explains. The other has occurred on at least 18 elk farms scattered across the U.S. and Canada, he says.

"CWD has not been identified in deer from the farmed deer industry yet, but we really need to be doing surveillance to have assurance that it is not there," says Lynn H. Creekmore of USDA's National Animal Health Program.

In Larimer County, Colorado, the rate of CWD infection is now up to 15% in deer and 1% in elk, Miller says. Although the disease is spreading very slowly, it is gradually claiming a larger portion of the deer and elk populations. "It stands to reason that since the disease started in one location and spread out from there that it is still spreading," says Tom Thorne, a wildlife veterinarian with the Wyoming Game & Fish Department.

It is difficult, if not impossible, to eradicate CWD. After several deer and elk in a research facility near Wheatland, Wyo., came down with CWD, all the animals were killed and buried, and the pens remained empty for at least six months. Then, healthy animals were brought into the pens. After five years, some of them showed symptoms of CWD.

At the research facility in Fort Collins, Colo., personnel tried even harder to eliminate whatever it was that was sickening their captive deer and elk. They killed all the animals in the facility, plowed up the soil in the pens, and sprayed disinfectant on structures and pastures. Twelve healthy wild elk were then housed in the pens. After five years, two died from CWD.

  THE PRIONS causing CWD are probably spread through feces that comes in contact with the animals' food, says Elizabeth S. Williams, professor of veterinary medicine at the University of Wyoming. Sometimes, maternal deer seem to transmit CWD to their offspring. But in these cases, transmission may also occur through feces, she points out.

It would be "pointless" to kill the 80,000 deer and elk in the regions affected by CWD, Williams says, because healthy animals that move into the areas would pick it up from the environment. However, the Colorado Division of Wildlife hopes to slow the spread of the disease by reducing the populations of deer and elk in areas where CWD is most prevalent, spokesman Todd Malmsbury says.

Currently, there is no conclusive evidence that people have contracted vCJD from eating deer or elk meat. Between 1997 and 2000, the Centers for Disease Control & Prevention (CDC) investigated three suspicious cases of CJD, two in hunters and one in a daughter of a hunter. All three contracted CJD when they were less than 30 years old, and all were reported to have eaten venison, but not from areas where CWD is known to be endemic.

According to CDC, it is extremely unusual for people under the age of 30 to contract CJD unless they have been infected with a contaminated surgical instrument or human growth hormone from cadavers. Sporadic CJD in people under 30 occurs with an annual incidence of about five cases per billion people worldwide.

MAJESTIC THREAT A prion disease, chronic wasting disease, now infecting deer and elk in some parts of Colorado and Wyoming, may also sicken humans.

COURTESTY OF USDA

A prepared statement from Ermias D. Belay, a CDC medical epidemiologist who headed the investigation of CJD among hunters, says, "Although the occurrence of these unusually young CJD cases suggested a possible relationship of their illness with CWD, the follow-up investigation found no causal link between CWD and CJD in the three patients." CDC scientists refuse to discuss these cases with the press. Nor will they explain how they can diagnose this disease as sporadic CJD when they have never observed vCJD symptoms caused by exposure to CWD prions.

Other lines of evidence are somewhat less reassuring. Using test tube experiments, Byron Caughey and his colleagues at NIH's Rocky Mountain Laboratories in Hamilton, Mont., have found that prions from infected deer and elk are able to convert normal human prion protein to the protease-resistant form [EMBO J., 19, 4425 (2000)]. The efficiency of the process is low, but it is about the same as that with which BSE prions convert normal human prion protein to the protease-resistant form.

Furthermore, the researchers found that prions from CWD-infected deer and elk convert normal prion protein in cattle to a protease-resistant prion, but at a rate one-fifth to one-twelfth the rate of cervid-to-cervid prion conversion.

"These experiments suggest that CWD prions from deer and elk could infect humans as readily as BSE--mad cow--prions from cattle sicken humans," says Michael Hansen, a research associate at Consumers Policy Institute, the research arm of Consumers Union.

  CAUGHEY EXPRESSES a similar concern: "Since humans have apparently been infected by BSE, it would seem prudent to take reasonable measures to limit exposure of humans--as well as sheep and cattle--to CWD infectivity, as has been recommended for other animal TSEs."

Colorado and Wyoming officials have taken some precautions to protect deer and elk hunters from CWD. They warn hunters not to shoot or eat a sickly animal. They also tell hunters to minimize contact with the animals' brain and spinal cord and to remove and discard the bones when butchering the meat. The warnings are widely known and strong enough to protect hunters, Williams says.

After killing a deer or elk in Colorado or Wyoming, about 60% of hunters submit the heads to wildlife officials for testing, which takes several weeks. If it turns out that the animal has CWD, the hunter is notified. However, hunters often eat the game meat before the results are known. "People up there aren't concerned about it," Malmsbury says.

Hunters should be prohibited from eating venison from endemic areas, Hansen says, because deer and elk can harbor the infection long before they show any symptoms. "If you want to avoid disaster, you have to take a precautionary approach," he says.

Whether or not CWD can infect humans who eat venison is a "giant question mark that needs to be tracked very carefully and looked at," Healy says.

Deer and elk are increasingly being raised in captivity for commercial purposes. The USDA is in the process of writing a proposal for a herd certification program to better control CWD on elk farms, Thorne says. Currently, elk farms--which collectively have annual sales of $1 billion, mostly from sales of velvet antler (uncalcified antler tissue for use in nutritional supplements)--are regulated by the states, but some states have no rules governing their management.

If an elk is infected with CWD, there are no federal laws requiring that the herd be quarantined. And there are as yet no tests to determine whether a healthy-looking animal is in fact infected with CWD. Sometimes meat from a herd with a case of CWD is sold to consumers, says Peter G. Lurie, medical researcher at Public Citizens Health Research Group.

  CRUCIAL FEATURES of vCJD are still unknown, including the minimum exposure to prions that causes the disease, the number of people exposed, and the incubation period. Consequently, scientists say it is impossible to predict with any accuracy the number of anticipated vCJD cases that will occur in the U.K. Last August, British researchers in the CJD Surveillance Unit in Edinburgh reported that the number of U.K. residents who have come down with the disease rose 23% annually between 1994 and 2000 [Lancet, 356, 1224 (2000)]. Extrapolating from this rate of increase, Roy Anderson of Oxford University put an upper bound of 136,000 on the eventual number of infections in the U.K., but says the toll could be much lower.

Peter Bacchetti, adjunct professor in the department of epidemiology and biostatistics at the University of California, San Francisco, says trying to extrapolate the ultimate shape of the vCJD disease curve is nearly impossible at this point. "We don't know how many people ate how many prions. We don't know if the dose affects the incubation time," he says.

In some species subject to prion diseases, susceptibility seems to depend on the amino acids coded for at a particular site--codon 129--on the prion protein gene. The most susceptible humans--40% of the population--are those with methionine-methionine encoded at the site. So far, all vCJD victims fit this genetic profile. If people who do not have "met-met" are also susceptible to vCJD, but with a longer incubation period, the disease toll will likely be higher.

So what should be done in the U.S. to learn more about vCJD, BSE, and CWD? One obvious step, Healy says, is to greatly expand the federal budget for prion research. "Prion research in general has been very skimpily funded, amazingly so in the sense that we have already won two Nobel Prizes in this country for prion work," she says.

NIH, which does the lion's share of the work, spent less than $14 million in fiscal 2000 on prion research. That funding is "very, very small in the context of an enormously growing NIH budget, which is over $20 billion," Healy says. It is also very small compared with the $100 million that all of Europe is investing in prion research. "We should be doing at least that," she says. The situation today may be analogous to the early days of the AIDS epidemic when NIH was doing very little research on retrovirology, she says. "When it turned out that HIV was a retrovirus, we looked around and saw we didn't have the retrovirus investment that we should have had," she explains.

Another needed step, Healy says, is to greatly expand surveillance for BSE and vCJD. Surveillance needs to be increased for both humans and animals who die from unclear neurological problems, she says. According to FDA, 60 diagnostic labs are examining hundreds of cattle brains each year. In contrast, France is testing 40,000 cows each week with a new test developed by Prionics AG. Germany didn't realize it had BSE in its cattle until a private producer started testing thousands of animals.

In addition, Healy says, the tissue banks that she advocates starting in the U.S. can be used to "see if we can detect prion disease in a cross-section of otherwise healthy people." Britain and Switzerland have done this for 30,000 people. "It's not that expensive. You can go back and use that tissue as a reference if something occurs here five or 10 years from now," she explains.

NIH's Brown, however, says creating such a tissue bank now would be too expensive. "I suspect that would be spending an enormous amount of money for almost no yield," he says. "We do not have a problem with BSE. vCJD is not going to creep up on us as an epidemic without first having some signal from animals," he explains. "I think a much greater payoff would come from the examination of 10 times as many cattle."

In the face of all the scientific uncertainty about prion diseases and whether BSE and vCJD outbreaks in the U.S. would involve only a handful of people or thousands, "we are in the uncomfortable position of having to make decisions that have a high likelihood of being wrong," Healy says. Her advice to every decisionmaker is "make sure you can live with being wrong."

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ELUSIVE
Prion Science: Still Plagued With Uncertainty

Scores of research groups are ardently pursuing the elusive prion (protein infectious particle), and numerous advances in prion chemistry and biology have been reported over the past few years. But the prion has always been a fairly tricky research subject, and knowledge of the mechanisms by which it forms and presumably causes disease continue to be difficult to nail down. Somehow, the true nature of the mysterious prion always seems to lie just beyond reach.

Creutzfeldt-Jakob disease (CJD) and related neurodegenerative conditions, like the sheep disease scrapie, resemble viral diseases in some ways. But the infectious agent in these transmissible spongiform encephalopathies (TSEs) withstands radiation, extreme heat, and chemical treatments that would normally kill a virus. Furthermore, no disease-associated viruses have been found in humans and animals suffering from these conditions.

In 1982, neurologist Stanley B. Prusiner and coworkers at the University of California, San Francisco, first isolated and characterized a possible infectious agent from scrapie-infected tissue and called it prion protein. Based on several lines of evidence, they proposed the protein-only hypothesis--that TSEs are caused not by infectious viruses, but by infectious prion protein alone.

The proposal was roundly attacked by the scientific community as heretical, since proteins alone had never before been found to replicate, infect animals and people, and cause disease. However, over the years, the idea became more generally accepted, and its critics eventually piped down--especially after the Nobel Foundation gave its imprimatur to the hypothesis by awarding Prusiner the 1998 Nobel Prize in Physiology or Medicine.

Nevertheless, due to the prion's rather ornery nature, the hypothesis has not yet been proven definitively either. A key test of the protein-only hypothesis--demonstrating that prion protein scrapie (PrP^Sc) newly created in a cell-free (and thus virus-free) system is actually infectious--has never been carried out successfully.

In 1985, researchers isolated, characterized, and cloned the gene for prion protein. Studies of the gene's expression revealed that prion protein is not found only in TSE-infected animals. Instead, it is an endogenous glycoprotein found on brain cell membranes and in other tissues of healthy animals and people.

The protein has two forms--normal and infectious. The normal form, PrP^C (prion protein cellular), is soluble in water, sensitive to protease enzymes, and primarilya-helical in structure. The infectious form, PrP^Sc, is water-insoluble and protease-resistant, and it has a predominantlyb-sheet structure.

PrP^C and PrP^Sc have the same protein sequence but different shapes, or conformations. In the protein-only hypothesis, Prusiner proposed that when PrP^C comes in contact with PrP^Sc, it gets converted into the "bad" form. Infectious PrP^Sc molecules from the conversion then build up in the brain, and this is associated in some way with TSE-induced physiological changes, leading to neurodegeneration and death.

Researchers have tried to find evidence for the alternative hypothesis--that TSEs are caused by viruses. Some believed that the existence of different TSE strains--variations characterized by differences in incubation time and the type of neurological damage incurred--was strong evidence for a potential viral cause of TSEs because a simple protein was unlikely to exhibit sufficient variation to account for these strains.

However, "if a virus caused prion disease, then we could find the virus within sheep, and it would be the same virus as in the cow, but we can't find a virus," Prusiner explained last month at a symposium on detection of mad cow disease at the Pittsburgh Conference, in New Orleans. Researchers have spent many years looking for nucleic acids "without any success," he noted. Prusiner and others believe TSE strains can indeed be adequately accounted for by prion protein variations--such as nuances of protein shape or structural diversity arising from glycosylation.

The underlying mechanisms of prion formation and TSE causation remain uncertain. For example, there are two major alternative models for the molecular mechanism by which PrP^C converts into PrP^Sc. The template assistance mechanism--also called the heterodimer mechanism or the monomer-directed conversion model--was proposed by Prusiner. The idea is that PrP^C binds to PrP^Sc, forming a PrP^C-PrP^Sc heterodimer. PrP^C-to-PrP^Sc conversion occurs, turning the heterodimer into a PrP^Sc homodimer. The homodimer then dissociates, releasing PrP^Sc. Prusiner and coworkers also believe that a chaperone called Protein X may bind to PrP^C and facilitate its conversion to PrP^Sc.

The other major model is the nucleated polymerization mechanism, backed by Peter T. Lansbury Jr., associate professor of neurology at Harvard Medical School. It proposes that an oligomeric form of PrP^Sc recruits PrP^C monomer and mediates its refolding into PrP^Sc.

The normal function of endogenous prion protein in the brain likewise remains a mystery, although a signaling or transport role is suspected. And the way in which PrP^Sc formation causes disease is unknown. So in prion biochemistry much remains to be revealed.--STU BORMAN

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ORGANIC
Dietary Supplements: Possible Vector For Mad Cow Disease

Largely unregulated in the U.S., dietary supplements made of raw bovine organs may be one of the holes in the barriers erected to control mad cow disease, technically known as bovine spongiform encephalopathy (BSE).

A number of manufacturers make supplements from cattle organs and bones that would harbor BSE prions in an infected animal. For example, Atrium Inc., Hebron, Ill., sells a supplement called Brain 360, which contains 360 mg of raw bovine brain concentrate per dose. The company also produces a pill called Pituitary Whole, which contains 40 mg of raw bovine pituitary gland.

BITTER PILL A variety of supplements on the market today contain bovine bone, brain, or other glands.

PhytoPharmica markets a pill called Adrenal-Cortex Fractions, which contains in one dose raw bovine brain, lung, heart, and spleen. Except for heart, all of these organs are considered high risk for BSE. PhytoPharmica also sells a product called Brian Essentials, which contains 300 mg of freeze-dried bovine brain per dose. A number of companies grind up cattle bones and add them to sports nutrition products. While not glandular, the bones, and thus the products, could be contaminated with prions, if made from bones of BSE-infected cattle.

"Half a gram of infected bovine brain is all you need to get vCJD"--one good dietary supplement pill--notes Paul Brown, chairman of the Food & Drug Administration's Transmissible Spongiform Encephalopathies Advisory Committee. Supplements made from BSE-infected cattle could become a wide vector for spreading new-variant Creutzfeldt-Jacob disease (vCJD) in humans, he says.

Supplements, unlike drugs, are not subject to FDA safety and efficacy requirements. Congress specifically exempted them. Nor are supplement manufacturers required to specify the country of origin for bovine parts on product labels.

"If brain and pituitary tissues used to make dietary supplements come from a BSE-infected country, then there is a real risk," says Brown, who directs the Laboratory of Central Nervous System Studies at the National Institutes of Health. "FDA needs the authority to require the producers to document the authenticity of what they are claiming--that is, that the organs they use do not originate from BSE-infected countries," he explains. He estimates that about 30% of nutritional and sports supplements contain bovine bone, brain, or other glands.

The Department of Agriculture has the authority to stop selected shipments of bovine parts or products from countries where BSE has been detected. But very few shipments are actually inspected.

FDA has all the authority it needs to keep contaminated products off store shelves and out of consumer hands, says David Seckman, executive director of National Nutritional Foods Association (NNFA), a trade group for the dietary supplements industry. But that authority hasn't been needed, he says, because no evidence has been found that any products made from bovine parts are contaminated.

In mid-March, responding to media reports that some dietary supplements could be contaminated with mad cow disease--reports Seckman calls "the worst kind of sensational journalism"--NNFA issued a guidance to its members encouraging them to eliminate all neurological bovine materials from their products. But NNFA did not ask them to eliminate other bovine tissues, such as bones and pituitary gland, that can also contain prions in a BSE-infected animal.

However, mad cow disease is showing up in more countries almost weekly. So a country that is BSE-free one day may not be so the next. Also, there is no way to test supplements for prions, Brown says.

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FEED TRAP
Major Barrier May Be Porous

Regulations governing the content of cattle feed are one of the major barriers erected to keep bovine spongiform encephalopathy (BSE) out of the U.S. In 1989, the Department of Agriculture prohibited the import of rendered animal protein, including meat and bone meal, from ruminant animals from the U.K. In 1997, the ban was extended to all of Europe and to other countries that present an undue risk of introducing BSE into the U.S.

And last December, USDA prohibited the import from Europe of all rendered animal protein products regardless of species. This latest ban was instituted because feed made from nonruminant animals was found to be contaminated with BSE prions.

The Food & Drug Administration in 1997 prohibited the practice of feeding most proteins derived from mammals--that is, meat and bone meal--to ruminant animals, such as cattle. It took this step to avoid spreading BSE through feed, in the event that the disease ever does show up in U.S. cattle.

Although FDA's feed rule sounds stringent, it is often not enforced. Between 1998 and 2000, FDA and state inspectors visited 9,184 rendering firms to increase awareness of the BSE regulation only to find that 1,688 of the firms "were not aware of the new regulation," Congress' General Accounting Office says in a report published last year. And 28% of the firms had failed to state on their packages of meat and bone meal derived from mammals that it should not be fed to cattle or other ruminants. In addition, 1,771 firms that make meat and bone meal for both ruminants and nonruminants did not have a system in place to prevent commingling of products, as required. FDA took only limited action against firms that were out of compliance.

FDA now says it is conducting additional inspections and reinspecting noncompliant facilities.

One exception to the ban on feeding cattle protein to cattle is that FDA allows calves to be given spray-dried blood plasma from cows as a milk replacer. If the blood from a BSE-infected cow does contain prions, this practice could spread BSE around the cattle population.

"Clinical experiments have not been able to show any infectivity in cattle blood, even from clinically infected cattle," says Paul W. Brown, chair of FDA's Transmissible Spongiform Encephalopathies Advisory Committee. "So it ought to be safe. But if you have a choice, you'd obviously use the blood of a healthy animal," he says.