91av

ONE IN TEN people in Britain is more afraid of mad cow disease than of AIDS, according to an opinion poll commissioned by The Observer newspaper. A huge 60 per cent of the people polled thought that the government is not doing enough to stop bovine spongiform encephalopathy (BSE) from spreading to humans.

Such mistrust of the Ministry of Agriculture’s efforts to safeguard human health is understandable, given its past record. The comparison of a disease that, as far as anyone can tell, has not claimed a single human life, with one that has killed hundreds of thousands is harder to fathom.

The problem, for both the ministry and the public, is that the ‘organism’ that causes BSE is a mysterious entity. No one has ever seen it. The disease belongs to a group of nervous diseases caused by what are known, for want of a better phrase, as ‘unconventional agents’. These are neither bacteria nor true viruses, but something unlike any other organism. They have eluded researchers for decades.

Various groups of researchers around the world have differing views on what type of life form these agents might be. The simplest idea, proposed by Heino Diringer of the Robert Koch Institute in Berlin, is that it is a tiny filamentous virus. A second theory, hotly debated for many years, invoked a ‘rogue protein’ that could replicate without having a nucleic acid. If this were true it would call into question the very nature of life. Stan Prusiner, of the University of California, at San Francisco, identified a protein that he thought fitted the bill. He called it a ‘prion’.

A third suggestion, put forward by Alan Dickinson, a former director of the Institute of Animal Health’s Neuropathogenesis Unit in Edinburgh, is that the agent is a tiny piece of nucleic acid that hijacks some of the host’s protein to form a protective coat around itself. This hypothetical agent is called a ‘virino’.

Whatever the agents are, they cause some devastating diseases: scrapie in sheep and goats, chronic wasting disease in captive deer, transmissible mink encephalopathy in ranched mink – and two human diseases, kuru and Creutzfeldt-Jakob disease. Years of research into these diseases have failed to reveal the nature of the agent, but have provided some clues for British scientists investigating BSE.

One of the first questions was: Where did BSE come from? BSE appeared very suddenly, out of the blue. A few cases in 1985 and 1986 have swelled into an epidemic that has claimed some 14 000 cows and is increasing by between 250 and 300 cases a week. Infected cows show no signs of disease for several years, but once symptoms appear the course of the disease is rapid.

Sick cows become edgy and irritable; they begin to lose coordination of their limbs and may eventually become very aggressive. The brains of infected animals are irreparably damaged, with holes in certain parts of the brain tissue, giving it a characteristic spongy look. Another telltale sign is the presence of clumps of protein fibrils, which show up in the electron microscope.

Scientists who have worked on scrapie say that what they see in the brains of mad cows is so similar to that in scrapie sheep that it is probably the same disease. Epidemiologists at the Central Veterinary Laboratory in Weybridge, Surrey, tend to agree. Their laborious researches in the early days of the epidemic pinpointed the source of infection to concentrated cattle feeds that included rendered down carcasses of other ruminant animals sold to feed manufacturers as meat and bone meal (91av, This Week, 7 January 1989).

Sheep infected with scrapie were entering the cows’ food supply, and the suggestion was that the scrapie agent crossed species from sheep to cattle. The government banned the feeding of ruminant remains to ruminant livestock in June 1988, in an attempt to cut off the source of infection.

The general view is that several factors operated to produce the sudden outbreak of BSE. The number of sheep in Britain increased in the late 1970s and early 1980s; and with it the incidence of scrapie. Around this time, farmers began to wean their dairy calves earlier, feeding the young animals on concentrated ‘weaner rations’ rich in protein. Older milk cows received similar rations to increase their yields.

In the best-quality feeds, the protein comes from soya or fish meal, which are more reliable sources of protein. Lower quality feeds include varying amounts of meat and bone meal. The composition of feedstuffs – and the amount of rendered down ruminant material – is determined by the cost of the various components at any particular time. When the price of soya and fish meal is high, the proportion of meat and bone meal increases. Feed is sold guaranteeing a certain proportion of protein, without saying where it comes from.

Also in the late 1970s, rendering plants in Britain changed the way they processed carcasses into meat and bone meal for animal feeds. Instead of processing batches of carcass material at high temperatures, they adopted a system of continuous processing at lower temperatures. All the firms except one also abandoned the use of solvents, such as benzene, hexane and trichloroethylene, to remove excess fat from the meat and bone meal. Perhaps more significant was the loss of a final heating stage to drive off the solvents. This final heating involved very high temperatures.

William Reilly, chairman of the British Veterinary Association’s committee on public health, believes that it may have been this final heating that killed the scrapie agent. The one firm of renderers that still uses solvent treatment is in Scotland, and Scotland has a disproportionately small number of cases of BSE (in mid-May 424 of the 13 648 cases in Britain). John Wilesmith, an epidemiologist at the Central Veterinary Laboratory, believes that solvent treatment is a factor, but only partially accounts for the geographical differences.

Following the recommendations in January of the Tyrrell committee, commissioned to assess the need for research on BSE, the government committed Pounds sterling 12 million over the next three years to studies of the disease. Some of the studies were already in train; others have just begun.

In 1987, Jim Hope and Laura Reekie, at the Neuropathogenesis Unit in Edinburgh, showed that an abnormal protein was present in the brains of infected cows. This was the bovine equivalent of a protein that clumps into fibrils in the brains of sheep with scrapie. Hope’s colleague, Hugh Fraser, immediately began experiments to try to transmit BSE to mice. He injected ground up brain tissue from cows that had died from BSE into the brains of the mice, which went on to develop the disease. This meant that, as with scrapie, researchers could use mice as a model for investigating many aspects of the disease.

At Weybridge, researchers showed that they could transmit BSE from sick cows to healthy cows by direct inoculation. Meanwhile, at the Royal Veterinary College in Hertfordshire, Richard Barlow and Deborah Middleton transmitted the disease by feeding mice large amounts of infected cow brain. This was the first sign that BSE could be contracted by eating infected tissue (91av, This Week, 3 February 1990). No other tissues from infected cows – spleen, spinal cord, lymph tissues, milk and udder tissue, or muscle (the beef we eat) – have produced any sign of disease in mice to date.

Transmission studies are a vital aspect of research into BSE. In Edinburgh and Weybridge and at the Clinical Research Centre in Harrow, experiments on a range of animals are under way. They are designed to show which species can contract the disease and by what route – through injection directly into the brain, into the body cavity or orally, in foodstuffs. Tests on pigs and chickens – which still receive meat and bone meal in their rations – have produced no results so far. Nor have tests on marmosets at the Clinical Research Centre.

The important point of these experiments is to find out more about the ‘species barrier’. When one of these agents crosses from its natural host into another species (as achieved in the laboratory), it usually meets some resistance, which manifests in the form of a longer than normal incubation period. Once it has crossed species, however, the agent meets less resistance in other individuals of that species and the incubation period, usually shorter, becomes fixed for the new host.

Transmission experiments should show how ‘high’ the barrier is in different species. This would be important in assessing the risk of a particular species succumbing to BSE.

Faced with a disease that is impossible to diagnose until after death, one of the greatest priorities for research is to find a test that can detect animals that are in the early stages of the disease. Diagnosis before slaughter would help to ensure that infected cattle do not enter the human food chain.

Researchers at the Institute of Animal Health’s unit in Edinburgh have made some progress on a diagnostic test. Working with scrapie in mice, they have devel oped polyclonal antibodies to the protein fibrils that form in the brain. The fibrils are aggregates of a cell protein that has been altered in some way by the scrapie agent. The antibody test identifies the abnormal protein.

‘This protein is present quite early in the course of the disease,’ says John Bourne, director of the institute. At the moment it can be detected in brain tissue, but only postmortem. But the abnormal protein is also present in spleen, and could be in other tissues too. Bourne hopes to identify the protein in blood cells, which would allow diagnosis in live animals. ‘It’s a long step from mice to sheep and cows, but it can be done,’ says Bourne. ‘The big step is from spleen to blood.’

A second type of test is needed to find which, if any, animals are genetically susceptible to the disease. Such a test would enable farmers to breed from resistant stock and produce herds that could be almost guaranteed healthy.

In Edinburgh, Hope and his colleague Nora Hunter have already found a way to distinguish sheep that are susceptible to scrapie from those that are ‘resistant’. In reality, the difference is that in one type the incubation period for the disease is short, and the animal succumbs; in the other, incubation takes longer than the lifespan of the animal. The two types have different forms (alleles) of a gene called sip (for scrapie incubation period). A simple blood test reveals within a few days which of the forms a sheep has (91av, Technology, 19 August 1989). The next step is to find out if cattle have any similar genetic susceptibility – and then to identify the gene responsible.

Such tests would reassure both consumers and farmers. Still greater reassurance might come from Wilesmith’s continued epid emiological studies. One of the most important questions that remains is whether cows are ‘dead end hosts’, in which the infection dies with the animal that contracted BSE from its food. Mink, which contract TME from contaminated food, do not pass on the infection to their offspring or to each other. Scrapie, by contrast, passes from ewe to lamb, either in the womb or during birth, by what is known as vertical transmission. Scrapie can also pass ‘laterally’, from sheep to sheep, most probably when they come into contact with pasture contaminated by placentas from ewes that have just given birth.

If cows are dead-end hosts, the disease should die out when all infected animals have been slaughtered. By 1992, which will be four years after the Ministry of Agriculture banned the use of ruminant offal in cattle feed, the number of cases should begin to fall off.

If BSE is transmitted to the calves of infected animals, the number will continue to increase for several more years as these animals succumb and are destroyed. Four calves with nervous symptoms have been slaughtered. At postmortem, three showed no sign of BSE. Results on the fourth are due shortly. In the worst scenario, with transmission from cow to cow, the epidemic will ease off slightly as the first batch of infected animals is slaughtered, and will then soar again as the disease begins to affect their contacts.

The Ministry of Agriculture has begun a seven-year study to monitor the health of 300 calves born to infected cows.

Another area of research involves trying to kill off the agent. This is extremely difficult when the only way to check that the agent has been deactivated is to inoculate experimental animals with the treated material and wait to see if they develop the disease.

All of the ‘unconventional agents’ that cause these diseases are extremely hardy. Few of the chemical or physical treatments that kill viruses have much effect on them; they withstand high temperatures, irradiation and doses of ultraviolet light that are normally used in sterilisation. Only autoclaving at very high temperatures or high concentrations of sodium hypo chlorite seem to have any effect. Work has begun in Edinburgh to test various combi nations of treatments – permutations of time and temperature, steam treatment and chemicals.

Despite the evidence from pathology and molecular biology, some people are not convinced that BSE is simply scrapie that has crossed into cows. There are some close similarities between BSE and scrapie, but also some differences. BSE could be a natural encephalopathy of cows which used to be so rare that no one had recognised it.

‘Whether it is scrapie in the feed or BSE is the question,’ said Francis Anthony, chairman of the British Veterinary Association’s farm animals committee. ‘In 25 years of general practice, I and several colleagues reckon we’ve seen cases of BSE before – odd, sporadic cases. Others say they’ve never seen it and it is a brand new disease. But I’m not totally convinced that it’s scrapie.’

If the disease has always been around, cropping up just occasionally, the picture of a ‘rogue scrapie’ that can cross from one species to another, no longer holds. If BSE is a natural disease, then it might be present in other countries, too. The outbreak in Britain has prompted researchers elsewhere to look for signs of mad cows.

In 1986, Dick Marsh, of the University of Madison-Wisconsin, pointed out that most of the outbreaks of transmissible mink encephalopathy involved mink that were fed meat from ‘downer’ cows – those that were dead or paralysed before they reached the knackers. Marsh hinted that the cows might have been suffering from a bovine form of scrapie.

‘We will be looking at brains from cows with undiagnosed neurological disorders, and if we find any lesions that look like BSE we will send the samples to Britain for confirmation,’ said Gary Colgrove of the US Department of Agriculture’s Animal Health Inspection Unit. ‘There is no evidence that we have BSE but we need to carry out this surveillance to be sure. We hope to look at 200 to 300 brains in the next 6 months.’

Another country taking a keen interest in BSE is France, which could be the next country to face an epidemic of the disease. France has scrapie in its sheep population; it also includes meat and bone meal in its cattle feed. Last year, it doubled its imports of meat and bone meal from Britain because the price had fallen so low.

Some vets suggest that in both France and the US, a few of the cattle that are destroyed because they are thought to have rabies might conceivably have had BSE instead. The early symptoms of rabies include weakness and general disability followed by paralysis in the hindquarters, causing very obvious staggering, as do a number of other diseases of cattle.

Evelyne Maillot, head of the office of epidemiology and rabies at the French Ministry of Agriculture in Paris, said ‘We have begun to look for signs of BSE in cattle in France but so far we have not found any. It is possible that the symptoms can be hidden by rabies but there are important differences between the two diseases. Nevertheless, we plan to look closely at this possibility, and will carry out a detailed study of cattle this summer.’

There are still other views about what BSE is or isn’t: the agent may be a form of scrapie that has mutated – either in the sheep, or in cows after they have eaten the contaminated food. A changed form of agent is probably the most worrying prospect. If BSE has always existed, then it is likely to remain a disease of cattle. If it is scrapie that has crossed into cows, then it has already made one jump over the species barrier. The recent discovery that several species of antelopes in zoos and two pet cats suffered from diseases very similar to BSE has renewed this fear.

‘The real point,’ says Richard Lacey, professor of clinical microbiology at the University of Leeds, ‘is that after infective agents have been through a new host, they are likely to change their host range. The fact that it has turned up in cats and zoo animals is worrying.’

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The chain of reactions to an epidemic

IN the spring of 1985, a cow on a farm in Kent became the first victim of BSE. It was more than a year before veterinary pathologists identified the disease as a spongiform encephalopathy, something akin to scrapie in sheep.

By the beginning of 1988, and several hundred cases later, the Ministry of Agriculture commissioned an independent working party, under the chairmanship of Sir Richard Southwood of the University of Oxford, to advise on what steps it should take to halt the epidemic and safeguard human health.

Following Southwood’s advice, the ministry made the disease notifiable in June 1988. Farmers were then legally bound to report all cases. In July, the government banned the practice of feeding cattle with rations made from the remains of sheep and cows.

By August, all cattle suspected of having BSE had to be slaughtered and their brains sent for investigation to the Central Veterinary Laboratory in Weybridge. Three months later, farmers were ordered to destroy milk from suspect cattle.

In March 1989 the government provided guidelines to manufacturers of medicines that use bovine materials. More radically, in June a total ban on the human consumption of certain cattle offal (from healthy animals) came into force, following hard on the heels of a ban on their inclusion in baby foods. Most pet food manufacturers operate a voluntary ban on these materials.

Following further advice from Southwood, the ministry set up an expert committee under the chairmanship of a virologist, David Tyrrell, to advise on research into BSE and related diseases. Publication of the Tyrrell report in January this year coincided with the government’s announcement of Pounds sterling 12 million to pay for most of the research the committee recommended.

Despite these actions a number of countries banned the import of British cattle. The EEC has banned the import of British cattle born before the introduction of the feed ban. Australia, Austria, Israel, the US, Finland, Sweden and the Soviet Union also operate bans. West Germany and France have banned all imports of beef, ignoring demands from the EEC to lift their boycotts.

Following the announcement that two pet cats had contracted a disease resembling BSE, hundreds of schools throughout Britain refused to serve beef for school meals.

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The human connection: where does Creutzfeldt-Jakob disease fit in?

CREUTZFELDT-JAKOB disease is devastating. Like the other spongiform encephalopathies it usually takes many years to incubate. When symptoms finally appear, usually in the sixth decade of life, the course of the disease is catastrophic. It begins with loss of memory and perhaps uncharacteristic behaviour, progressing to rapid mental deterioration, dementia, incoordination and involuntary movements of the limbs. CJD is invariably fatal and most people die within a few months.

Although CJD was first described in 1919, few scientists took much notice of the disease until the late 1960s when experiments showed that the disease could be transmitted to chimpanzees. No one knows how it is transmitted in nature. The disease occurs worldwide, and affects about one person in 2 million each year. Most cases are sporadic, seeming to crop up from nowhere. A small proportion are familial, but whether because of a genetic susceptibility or through contact is not clear. A few cases are the unfortunate result of infection during brain surgery or corneal transplants, or by past treatment for dwarfism with extracts of pituitary hormone from corpses.

Those few instances apart, most cases are a mystery. The question many people are now asking is: Is CJD scrapie that has crossed from sheep to humans? And might BSE do the same? Scrapie has been present in British flocks for more than 250 years and is an obvious suspect as the source of infection. But epidemiological studies of CJD show overwhelmingly that there is no link between the incidence of scrapie and that of CJD.

There are as many cases of CJD in Australia and New Zealand, where there is no scrapie, as there are in Britain and France, where scrapie is endemic. Icelanders probably eat more sheep meat (and sheep heads) than any other people. Thirty per cent of their sheep have scrapie, yet the incidence of CJD is no higher in Iceland than elsewhere.

Likewise, Japan has a similar number of cases, yet Japanese people eat little sheep meat and what they do eat comes from New Zealand.

Although all the evidence suggests that scrapie cannot infect people, the Department of Health last month agreed to provide Pounds sterling 350 000 for a new long-term project to monitor cases of CJD in Britain. The project will be run by Robert Will, a neurologist at the Western General Hospital in Edinburgh, in collaboration with the Neuropathogenesis Unit in Edinburgh. Will and his colleagues will collect data on every case – how they are distributed across the country, what jobs the patients did and what sort of diet they had. Eventually, such information should show if there has been an increase since the outbreak of BSE.

A similar study carried out at the University of Oxford between 1970 and 1984 provides an ideal comparison. ‘It may be 10 to 15 years or longer before it can be determined with any certainty that BSE does not represent a risk to the human population,’ says Will.

Further reading ‘At the limits of infection’, 91av, 15 May 1986. ‘Mad cows and ministers lose their heads’, 91av, 11 August 1988. Novel Infectious Agents and the Central Nervous System, Ciba Foundation Symposium 135, John Wiley & Sons 1988.