BSE-Safety of Milk

Roland Heynkes, 29.5.2001 (last update 12.12.2001)

Solely responsible for the English translation:
Ingrid Schütt-Abraham

Structure
		Summary
		Failed attempts to transmit BSE by intracerebral inoculation of milk
		Pattison and Millson 1961
		Hadlow, Kennedy, Race and Eklund 1980
		Hadlow, Kennedy and Race 1982
		Hourrigan (de Camp) 1990
		Fraser and Foster 1993
		Taylor (Edinburgh), Ferguson and Bostock (Compton), and Dawson (New Haw) 1995
		Failed attempts to transmit BSE by feeding milk and udder tissue to mice
		Introduction
		Barlow and Middleton 1990-1993 using CRH-mice
		Barlow and Middleton 1990-1993 using C57B1-mice
		Taylor, Ferguson, Bostock and Dawson 1995
		Offspring of BSE cows which had been fed for at least 1 month by their dams did not develop the disease.
		The Weybridge cohort study
		Is BSE itself hereditary or only the disposition for BSE infections?
		So far no child of a Kuru- or CJK-diseased mother seems to have contracted the disease.
		CJD-infectious colostrum?
		Scrapie-transmission by semen or embryos?
		New experiments
		Literature
		Acknowledgements

Summary

Since November 2000 BSE-infections have also been detected in German dairy cows. This raised concerns as to the BSE safety of German dairy produce, a question relevant for public health and agriculture politics. This report presents and evaluates in detail the currently available information. On this occasion it became apparent that due to various methodological inadequacies the experimental and epidemiological investigations which have so far been carried out actually do not guarantee the BSE safety of milk to a satisfactory extent. Safety declarations which have been given so far from various sides were obviously based on superficial examination of the data and are in the following refuted.

Nevertheless in the most sensitive experiment carried out so far to detect BSE infectivity in milk, the milk of scrapie infected goats proved to be at least 100.000 times less infectious than the most infectious areas of the brain [AEZR]. In a comparable experiment the brain of a BSE sick cow proved to be at least 100.000 times more infectious than its milk [ALNN]. According to this, milk needs not necessarily be substantially less infectious than liquor of scrapie sick goats [AEZR]. Though it is quite possible that milk of BSE-sick cows is totally free of infectivity, so far there have been no experiments which would even approximately demonstrate this. For obvious reasons the BSE infectivity for man cannot be established by experiments. To be on the safe side the SSC therefore recommended to take into account the worst case scenario of a non-existing species barrier, a linear dose-response-correlation down to the smallest dose and a cumulating effect of repeated infectivity uptake. However, even if for the want of better data one optimistically compares the infectivity of BSE for man to the infectivity of BSE for mice one would, based on the experiments in mice, only be able to state that the chance to survive the ingestion of half a liter of milk of a BSE-sick cow would exceed 50% [ALNN].

It can only be hoped that the new experiments mentioned farther down will bring more clarity.

Failed attempts to transmit BSE by intracerebral inoculation of milk

Pattison and Millson 1961

In the Compton Field Station located 59 km west of London near Newbury in Berkshire staff members of the Institute for Research on Animal Diseases (Agricultural Research Council) inoculated two 3-month-old goats intracerebrally with 1 ml each of milk sampled under sterile conditions from a scrapie-infected goat. Within an observation period of at least 29 months none of these two goats developed Scrapie. Two goats which had each been intracerebrally inoculated with 1 ml urine and two which had each been inoculated with 1 ml saliva of a scrapie-infected goat, stayed also healthy, though. Even with blood the authors did not succeed in transmitting the disease. [ANBI,ANBJ].

There is, however, a report on a successful transmission of Creutzfeldt-Jakob disease by urine to mice [ALLU]. Another experiment failed to detect infectivity in urine but nevertheless demonstrated the presence of protease-resistant prionprotein [AKSU]. In saliva the detection of scrapie-infectivity has not yet been successful, but infectivity has more than once been found in tonsils [AEZS,AEZR,AEZP], nasal mucosa [AEZS,AEZR,AEZP] and saliva glands [ANBJ,AEZS] of sheep [AEZP] and goats [ANBJ,AEZS,AEZR]. It seems therefore highly improbable that saliva should actually be free of infectivity. There have been many reports on successful transmissions of Creutzfeldt-Jakob disease und rodent scrapie by blood, but in September 2000 also the successful transmission of BSE-induced scrapie from sheep to sheep by this means has been reported [AFPS]. Examples like urine, saliva and blood underline the assumption that the transmission experiment of Pattison and Millson was simply not sensitive enough to detect low infectivity levels even in the absence of a species barrier. An obvious reason for this might be the extremely small size of the sample consisting of only two recipients.

Hadlow, Kennedy, Race and Eklund 1980

30 µl of the 1/10th diluted milk of three naturally scrapie infected goats from an experimental enclosure in Mission (Texas, USA) were intracerebrally inoculated into 10 Swiss mice per goat in the Rocky Mountain Laboratories in Hamilton (Montana, USA), a branch of the National Institute of Allergy and Infectious Diseases (National Institutes of Health). Moreover, 10 mice per goat were intracerebrally inoculated with 30 µl of the homogenized and 1/10 diluted udder tissue of these 3 goats. During the two-year observation period none of these 60 mice developed Scrapie. On the other hand in the same experiment infectivities were detected between 16.706 mouse-i.c.-LD50/mg in the mid-brain and 0,17 mouse-i.c.-LD50-units per mg lung. Because the milk was less infectious than the lung, and because the infectivity of the lung was already in the range of the detection limit the milk must have been at least more than 100.000 times less infectious than the most infectious areas of the brain, or at least 3.600 times less infectious than the upper spinal cord. According to the results of this experiment, however, the milk need not necessarily be significantly less infectious than the spinal liquor of the scrapie diseased goats. [AEZR]

Hadlow, Kennedy and Race 1982

A flock of Suffolk sheep was kept in an experimental enclosure in Mission (Texas, USA) in which Scrapie spread naturally. There colostrum was sampled from a ewe which later developed Scrapie, and in the Rocky Mountain Laboratories in Hamilton (Montana, USA), a branch of the National Institute of Allergy and Infectious Diseases (National Institutes of Health) 30 µl each of this milk were directly (intracerebrally) inoculated into the brains of 10 female 21- to 23-day-old Swiss mice. The mice were observed for 24 months, but none developed the disease during this observation period. [AEZP] This experiment, too, was very small, but the main problem would have been the species barrier between sheep and mouse.

Hourrigan (de Camp) 1990

In transmission experiments carried out in Swiss White mice allegedly directly in Mission (Texas, USA) by Dr. Marguerite de Camp 6 Swiss-White-mice are reported to have been intracerebrally inoculated with milk and 3 Swiss-White mice with colostrum of scrapie diseased sheep, without developing the disease. Unfortunately the authors do not supply methodological details, but the transmission experiment was obviously not very sensitive as only 4 of 14 mice inoculated with ovary tissue and 4 of 13 mice inoculated with uterus tissue developed the disease. [AFPP]

The much too superficial description of this experiment does neither allow to distinguish it from the one published by Hadlow et al. nor to establish their identity. [AEZR,AEZP].

Fraser and Foster 1993

In the Neuropathogenesis Unit of the Institute for Animal Health in Edinburgh a 1/10 diluted udder homogenate from a BSE sick cow was intracerebrally (20 µl) and additionally intraperitoneally (100 µl) injected into 24 RIII-mice, and during an observation period of at least 650 days none of the recipient mice developed BSE [ANCG]. However, in the same experiment infectivity was neither found in the ileum nor the bone marrow [ANCG], although at least the ileum had definitely been infectious in the mouse bioassay [AMJJ,AMJG]. Not quite as clear, but nevertheless in a mouse bioassay being part of the not yet completed pathogenesis study even bone marrow of an orally BSE infected bovine had emerged as infectious [AMJF] This shows that this experiment of Fraser and Foster had simply not been sufficiently sensitive to detect potential infectivity in the udder tissue. And actually the detection limit of the experiment carried out by Fraser and Foster with combined intracerebral and intraperitoneal inoculation is said to have been calculated as 10^1.4 LD50/g [AMJF].

Taylor (Edinburgh), Ferguson and Bostock (Compton), also Dawson (New Haw) 1995

With milk of 6 clinically BSE sick pregnant cows in different stages of lactation young RIII/FaDk mice were inoculated (each with 20 µl intracerebrally and 100 µl intraperitoneally). Of these mice 25 stayed healthy for a limited observation period of only 300-450 days, 43 for after all 451-600 days and 88 even for 601-702 days. Symptoms typical for TSE were not observed. [ALNN]

The injection directly into the brain is an admittedly less realistic mode of transmission than feeding, however, much better suited to detect small amounts of infectivity. A titration experiment with RIII-mice and highly infectious brain homogenate from a BSE-sick cow showed, however, that an intracerebral inoculation of 20 µl already at a dilution by the factor 1000 will lead to incubation periods of about 450 days [AEEK]. Therefore one could not expect the 25 RIII mice observed for only 300-450 days after intracerebral inoculation of milk to develop the disease. Accordingly there remain 131 RIII mice intracerebrally inoculated with 20 µl each, which lived for long enough to develop the disease despite the infectivity of the injected material being lower by 3 orders of magnitude.

But apart from the incubation period a lower infectivity affects predominantly the share of mice which develop the disease. In the said titration experiment on each dilution level 12 RIII mice were used and from a 10.000-fold dilution only single animals developed the disease [AEEK]. With 131 mice the milk could have carried 1/100.000 of the infectivity of the brain of BSE-sick bovines without this having become apparent in the experiment.

The titration experiment which could not detect BSE infectivity at a dilution factor of 1/100.000 in RIII mice established for the inoculated brain tissue an original infectivity of at least 10^5,1 i.c.-mouse-LD50 per gram cattle brain. Projected to a kilogram this would be 10^8,1 or 1,259 x 10⁸ i.c.-mouse-LD50-Units per kilogram cattle brain. If the infectivity of milk were only 1/100.000 of this value, 1 liter of milk would nevertheless contain 1,259 x 10³ or 1259 i.c.-mouse-LD50-Units. Even if one takes into account that the infectivity with oral ingestion by RIII mice would be lower by approximately a factor of 700 [ALMQ], one would still get 1259/700=1,799 i.c.-mouse-LD50-Units per liter or 1 i.c.-mouse-LD50-Unit in 0,556 liters with oral ingestion. This so far most sensitive of all experiments to investigate the BSE safety of milk thus only demonstrates that 10 liters of milk of a BSE-infected cow would kill not more than about (10/0,556)/2 = 9 mice max., if one fed this milk to a sufficiently large number of mice. How many mice would one need to lick up 10 liters of milk? To arrive at a satisfactory safety range for man the number of experimental animals would have to be higher by several orders of magnitude.

This simple calculation clearly demonstrates three facts:

1. The so far experimentally established BSE safety of milk is totally insufficient. Even if brain were 1 million times more infectious than milk a probability to survive the ingestion of 5.6 liters of milk of a BSE-sick cow of 50% would not be really reassuring.
2. Gigantically dimensioned mouse bioassays were required to even approximately represent the human consumption of milk. With relevance to humans the BSE safety of milk thus cannot be established in feeding experiments with mice.
3. In spite of the sensitivity being higher by about the factor 700 another experiment with intracerebral inoculation of milk into the brains of mice would have to use recipient mice in numbers which are by orders of magnitude higher than have so far been used in all past experiments.

Failed attempts to transmit BSE by feeding milk and udder tissue to mice

Introduction

There have been two experiments to detect BSE-infectivity in milk by feeding it to mice. Both have in common a relatively low sensitivity due to the species barrier between cattle and mice. A similar species barrier exists between cattle and humans, however, the number of recipient mice as well as the applicated amounts of milk are naturally a long way from the amounts of milk consumed by humans in the EU.

In this aspect one has to take into account that 300 ml milk ingested by mice are comparable to 300 ml milk ingested by man and must not be projected comparing the body weights of man and mouse. Taylor and colleagues themselves point out that the infectious dose is probably not dependent on the mass of the consumer [ALNN]. This is obviously correct, for 300 mg of BSE brain homogenate killed 12 of 15 RIII mice [ALMQ], while feeding a comparable amount of 500 mg each of a brain homogenate obtained from the brains of 4 BSE cows killed 1 out of 6 sheep after an incubation period of 734 days and two out of three goats after an incubation period of 941 resp. 1501 days [AEDA]. The weight of the sheep or the goats which is by orders of magnitude higher than that of mice obviously had no significant effect on the level of the infectious dose and therefore man's size alone does not render him more resistant to BSE. This was, however, not to be expected differently, as the infectious agent amplifies in the organism.

Apart from this the composition of bovine milk especially with regard to its cell content is by no means constant, and accordingly the milk of BSE-infected cows need not be equally infectious on different days. Samples taken only on a few days of lactation from few cows cannot really be considered representative. Therefore the results of these experiments do not exclude that the milk of BSE-sick cows might at times be much more infectious than the milk which has so far been tested.

If these experiments are not only superficially and emotionally viewed but evaluated with regard to a quantitative assessment the results can by no means be considered reassuring.

Barlow and Middleton 1990-1993 using CRH-mice

Mixed with feed resp. drinking water on average 144 g udder tissue plus 20,2 ml milk of a BSE sick cow were applied to 9 CRH mice. Within the observation period of not more than 689 days, however, none of these mice contracted the disease. In this aspect they did not differ, though, from 8 CRH mice which had on average been fed 12,9 g lymph nodes, or 8 CRH mice which had on average been fed 13,4 g placenta. As even 8 CRH mice which had on average been fed 6,25 g brain homogenate plus 4,52 ml cerebrospinal fluid did not develop the disease within 678 days the CRH mice were obviously much too BSE-resistant for the detection of BSE infectivity in orally ingested material. [ANCB,ANCC,AIIK]

Barlow and Middleton 1990-1993 using C57B1 mice

Mixed with feed or drinking water 10 C57B1 mice each received on average 128,9 g udder tissue plus 14,6 ml milk of a BSE sick cow. Within the observation period of not more than 571 days, however, none of these mice developed the disease. In this aspect they did not differ from 10 C57B1 mice fed on average 29,7 g lymph nodes from the udder region, 20 mice fed on average 42,6 g spleen, 10 mice fed on average 13,7 g lymph node, or 10 mice fed on average 26 g placenta. Only 7 out of 9 C57B1 mice, however, proved to be infected, each of which had been fed on average 9,1 g brain homogenate plus 4,5 ml cerebrospinal fluid. [ANCB,ANCC,AIIK]

The Bioassay system with C57B1 mice was much better than the one using CRH mice but nevertheless still so insensitive that not even all mice which had been fed BSE brains contracted the disease. Therefore it is no wonder that none of the only 10 C57B1 mice which had been fed udder tissue and cow milk developed the disease. Ultimately, from this experiment one can only draw the not really reassuring conclusion that 128,9 g udder tissue plus 14,6 ml milk of the BSE cows were at least one order of magnitude less infectious than 9,1 g brain homogenate plus 4,5 ml cerebrospinal fluid of these animals. If milk and udder tissue were about equally infectious their BSE infectivity would have been at least 2 orders of magnitude lower than the infectivity of the brain tissue of the examined 4 cows. Of course this safety margin is insufficient to declare milk of BSE infected cows to be safe. [ANCC,AIIK]

Taylor, Ferguson, Bostock and Dawson 1995

Young RIII/FaDk mice were each fed on average 300 ml milk from 6 clinically BSE sick pregnant cows in different stages of lactation. Of these mice 22 stayed healthy for the observation period of only 300-450 days, 25 after all for 451-600 days and 72 even for 601-702 days. [ALNN]

Half a decade later than the above described experiment Taylor fed in a comparable experiment 300 mg of a BSE-infectious brain homogenate [ALMQ] to each of 15 R III mice. Subsequently 12 of the 15 mice developed the disease and the longest incubation period was 547 days [ALMQ]. Unfortunately the authors of this later publication do not give any information as to the incubation periods of the other mice which developed the disease [ALMQ]. But in the otherwise comparable feeding experiment of Barlow and Middleton 5 of 7 C57B1 mice - each fed on average 9,1 g brain homogenate plus 4,5 ml cerebrospinal fluid - developed the disease 435-504 days after the feeding commenced [ANCB,ANCC]. Because - at least after intracerebral inoculation - both strains of mice differ only with respect to the incubation period but not with respect to the susceptibility and the variability of the incubation periods [AEEK] the incubation periods of the 12 RIII mice fed 300 mg each of BSE-infectious brain homogenate [ALMQ] can be assumed to have also varied between 460 and 547 days. In the 22 RIII mice which had been observed after oral inoculation for maximally 450 days the disease would probably not have developed even if they had been fed 300 ml brain homogenate. Therefore these 22 animals should by no means be included when estimating the BSE safety of milk.

One could, however, even in the remaining 97 orally inoculated RIII mice not expect any or at least only few cases of disease as even 300 mg of a 10% brain homogenate of a BSE cow infected only 12 of 15 RIII mice [ALMQ]. Projected on the 97 mice this would refer to 78 infected mice, and already an infectivity less than 2 orders of magnitude lower would be below the detection limit. The entire experimental section in which 300 ml milk were fed per mouse does only demonstrate undiluted brain of BSE sick cows to be at least 1000 times more infectious than the milk of such animals.

Offspring of BSE cows which had been fed for at least 1 month by their dams did not develop the disease

With the aid of the British epidemiological BSE data base until August 1996 only 132 offspring of BSE sick cows could be found in beef suckler herds with known lineage (natural or foster dams), which had been fed by their dam for at least one month and had reached at least 20 months of age [AMMT]. The following table presents the age these animals reached without recognizable BSE disease and how many months had passed between their birth and the outbreak of BSE in their mothers.

Naturally no reasonable human would claim that a 20-month-old bovine which had been slaughtered without recognizable BSE-symptoms had obviously not been infected with BSE and that therefore its mother's milk could not have been infectious. For each animal one has thus to calculate on the base of the statistical data on British herds with endogenous BSE cases (animals which had not been added other than by birth) the probability by which it would have developed BSE at its age. For this I use the following factors derived from the British BSE statistic for 1996 being the year of death: 0,0000 (2 years), 0,0166 (3 years), 0,1278 (4 years), 0,2765 (5 years), 0,4649 (6 years), 0,6259 (7 years), 0,7725 (8 years), 0,8700 (9 years), 0,9292 (10 years), 0,9610 (11 years) [AMMT]. This statistic is comparable to the hypothetical cases of maternal transmission because most BSE cattle was obviously infected shortly after birth [ANDI]. This can be derived from the fact that firstly BSE occurs notably more often in 4-6 year-old cattle than in younger or older animals und that secondly the effect of the first British meat-and-bone-meal feed ban is reflected in the statistic only by those animals which have been born shortly before or after the ban.

If one multiplies the case numbers of the first table by the factors for the age-dependent probability of developing the disease one gets the following table:

The sum of the sums of the last line of the above table is 43,57. If all cattle in the study had been infected during their first weeks of life one would have to expect roughly 43 BSE cases in the 132 observed bovines. We know, however, that there is no maternal transmission of BSE in this magnitude of 100%. Wilesmith et al. interpreted the results of the Weybridge cohort study as hinting to a maternal transmission in the magnitude of 10% [AMMU], but in reality the data do not speak for a maternal transmission of BSE [AJZJ], but for hereditary differences in the susceptibility for BSE infections. The animals observed in the Weybridge cohort study, however, had not been fed by their dams[AMMU] and therefore this study does neither contradict a maternal transmission of BSE by milk.

Now the rates of the expected cases of the disease can easily be calculated for the 132 offspring of BSE cows. In the following table this has been done for 3 assumed scenarios. The milk of a BSE infected cow might:

1. more than 2 years prior to the outbreak of clinical disease be as infectious as at the end of the incubation period,
2. maximally 3 years prior to the outbreak of the disease be infectious,
3. or maximally 1 Jahr prior to the outbreak of the disease be infectious.

Even if the milk of BSE infected cows were more than 2 years before the outbreak of the disease as infectious as at the end of the incubation period one could expect no disease in this investigation with an assumed BSE transmission rate of 2%. Should, however, the milk of BSE infected cows be infectious not earlier than 2 years before the outbreak of the disease, then in this investigation one would not have to expect a clinical case even at a BSE transmission rate of 5%. If the milk were infectious only 1 year before the outbreak of the disease one could not expect any clinical case even with a BSE transmission rate of 10%.

That in this study not a single case of BSE was found in the 132 offspring which had been fed the milk of BSE infected cow for at least one month can therefore not really be considered convincing proof against the infectivity of milk.

The Weybridge cohort study

The Weybridge cohort study did not reveal proof for any maternal transmission of BSE. It points rather to hereditary differences in BSE susceptibility, because subsequent to the feed ban the frequency of BSE in the offspring of BSE infected cows decreased at the same rate as in the offspring of cows which did not develop BSE, and because the incubation periods in the offspring of BSE sick cows tended to be prolonged rather than shortened. Accordingly at least in this experiment calves could only to a statistically not measurable extent have become infected by the milk of BSE sick cows. This was not to be expected otherwise because most of the animals used in this study were born from dairy cows and had therefore received only colostrum for 1 - 2 days, but no milk. [AMMU]

Is BSE itself hereditary or only the diposition for BSE infections?

An analysis of the genealogy of 75 of the first known British BSE cases for up to 9 generations revealed BSE with the greatest possible probability to be no hereditary disease, while the data on the other hand do not exclude heredity of the disposition for BSE infections either by the father or the mother [ANDA].

So far no child of a Kuru or CJD diseased mother seems to have developed the disease.

Referring to a rather outmoded presentation by Michael Alpers dating back to the year 1987 [ANCH] Rosalind M. Ridley and Harry F. Baker report that no child born after 1959 to a Kuru sick mother had developed Kuru itself. [AJZJ]. However, the respective observation periods were less than 29 years and the number of offspring not stated but probably small.

The SSC claims to know that up to now still no child breast-fed by a Kuru- or CJD sick mother has developed the disease [ANCL]. Poof or references for this statement, however, are not given by the SSC. It is hard to believe, though, that the children of Kuru and CJD sick patients are in fact under constant observation. Probably the number of cases which could be observed over a sufficiently long period is so small that one cannot draw any conclusions from them. The SSC mentioned only a single case in which the child of a CJD patient reached at last the age of 30 years. But even this child could still die in 20 or 30 years from Creutzfeldt-Jakob disease.

CJD-infectious colostrum?

Colostrum gained 4 days after a cesarean section in the 30th week of pregnancy from a 38-year-old Japanese woman was injected directly into the brains of 20 BALB/c mice. 2 of these 20 mice became sick and showed according to the presentation given by the authors histopathologically recognizable spongiforme degeneration and astrogliosis. By injection of brain material from the diseased mice into the brains of other mice the authors demonstrated transmissibility of the disease. The child of this woman was not breast fed and has so far healthily reached at least its 6th year of life. [ALJP]

The disquietening result of this experiment was simply concealed in many opinions on the BSE safety of milk. Others question it without backing their doubts by facts which could be checked. Thus the Scientific Steering Committee of the EU in its Opinion on the safety of milk with regard to TSE refers to a letter from Professor K. Yamanouchi to Ray Bradley dated February 1997, which unfortunately was never published and is not even quoted word-for-word, according to which a Japanese authority by further histopathological resp. immunhistochemical examinations of fixated brain slices of mice which had been inoculated with colostrum and later became sick detected neither spongiforme degenerations nor protease resistant prion proteins [ANCL]. This is rather peculiar, as the authors of the study had clearly reported to have found spongiforme degenerations and astrogliosis. Why, then, did the Japanese authority not publish a clarifying report on the assumed fake or at least on the misinterpretation by Tamai et al., and why was their paper not officially retracted if the data were, in fact, incorrect?

Moreover the Japanese authority is reported to have actually detected spongiforme degenerations and protease resistant prion protein in brain slices of mice which had been inoculated with brain material from the mice which had been injected with colostrum [ANCL]. This raises the question how the allegedly not infected mice could have been infectious themselves.

Quite strange is, too, that the Japanese authority as well as the SSC doubt the results of Tamai et al. only because after the initial transmission crossing a species barrier one had not found the typical holes and amyloid plaques [ANCL]. Apart from the contradicting statements of Tamai et al. also Clark 1981, Fraser et al. 1992 and Lasmézas et al. 1997 had shown that likewise transmitting Scrapie to cattle [AFPP] or BSE to mice [AEEK,AHBH] not always holes and immunologically detectable protease resistant prion protein can be found. Similarly the Scrapie strain SSBP/1 in sheep partly causes only minimal degenerative changes, making a clear neuropatholocical diagnosis hardly possible [ADIO,AEDB].

Finally in all experiments on the transmission of CJD or BSE to mice it was observed that different strains of mice showed different susceptibilities to certain strains of the infectious agent [ANCB, ANCC, ALMQ]. Insuccessful attempts to repeat a transmission experiment therefore do not necessarily prove it to be untrustworthy.

Scrapie transmission by semen or embryos?

The embryo transfer study by Foster et al. [AEDB, AECX] intended to show whether scrapie infected sheep could infect their offspring already in the first days of gestation. The published first part of results in the year 1992 seemed to prove this [AEDB], but after an extension of the experiment by negative controls as published in 1996 this interpretation could no longer be upheld [AECX]. The biological offspring of not infected mothers developed the disease as often as the biological offspring of infected mothers. In view of all inadequacies of the experimental design this embryo transfer experiment by Foster and his colleagues shows that with Scrapie there must either occur transmission by clinically inapparent mothers or by ways of transmission other than the ingestion of infectious placenta. Moreover, this experiment demonstrates that a maternal transmission is highly unlikely up to the 6th day of gestation. Milk would have been only one of several possible ways of transmission in this experiment if there would actually exist scrapie resistant scrapie transmitters in sheep.

In another embryo transfer experiment not even all experimentally infected sheep developed the disease and only 2 of 20 offspring of Scrapie infected sheep intended as positive controls [AECB]. Of the offspring produced by embryo transfer none contracted the disease, thus even this experiment does not point at maternal transmission [AECB].

New Experiments

In an experiment which has presumably begun in the meantime it shall be attempted to detect protease resistant prion protein in the concentrated milk of experimentally infected cows by one of the validated BSE-Tests [SEA]. The SEAC states with respect to this that an intracerebral inoculation of milk of BSE sick cows directly into the brains of calves would be favourable and therefore recommended such bioassays [SEA]. Without a species barrier and circumventing the digestive tract such a transmission experiment promises a significantly higher sensitivity than all experiments carried out so far.

Also the SSC is aware of the limited significance of the studies carried out so far and of the existence of leukocytes in milk and made clear, that a precise estimation as to the BSE risk from milk and colostrum would not be possible without respective experiments using intracerebral inoculation in animals of the same species [ANCL]. Therefore the SSC advises against the consumption of colostrum, milk and milk products of BSE suspect animals and requires more significant transmission experiments [ANCL]. Until the end of March 2001 the SSC had no knowledge of plans for highly sensitive experiments in which milk of BSE sick cows was to be injected into the brains of calves [ANCL]. Commissioned by the Food Standards Agency only the milk of experimentally infected cows was to be examined for protease resistant prion protein [ANCL].

A research cooperation including

• Prof. Erwin Märtlbauer, Chair of Hygiene and Technology of Milk at the Institute for Hygiene and Technology of Food of Animal Origin of the Veterinary Faculty of the Ludwig-Maximilians-University in Munich,
• Prof. Siegfried Scherer at the Institute for Microbiology of the Research Centre for Milk and Food in Weihenstephan of the Technical University of Munich,
• Prof. Manfred Gareis at the Institute for Microbiology and Toxicology of the Federal Institution for Meat Research in Kulmbach and
• Dr. Martin Groschup at the Institute for Immunology of the Federal Research Institution for Viral Diseases of Animals

strives to achieve a concentration of the agent by biochemical and/or immunochemical methods which allows for a 1000 fold more sensitive detection of PrPsc in body fluids like milk, blood and meat juice.

Moreover, according to an information given by Nora Hunter on the 14.3.2001 an experiment was to be carried out in which some scrapie susceptible lambs were to be hand-reared [ANCL].

Literature

ANCH . Alpers,M. - Epidemiological and clinical aspects of kuru - In: Prusiner,S.B. und McKinley,M.P. (Editoren) Prions: novel infectious pathogens causing scrapie and Creutzfeldt-Jakob disease. San Diego: Academic Press, 1987: 451-65

ANCC . Barlow,R.M.; Middleton,D.J. - Oral transmission of BSE to mice - Sub-Acute Spongiform Encephalopathies, edited by Ray Bradley, Marc Savey and Brian Marchant - Proceedings of a Seminar in the CEC Agricultural Research Programme, held in Brussels, 12-14 November 1990, 1991; 55: 33-9 in Current Topics in Veterinary Medicine and Animal Science

ANCB . Barlow,R.M.; Middleton,D.J. - Dietary transmission of bovine spongiform encephalopathy to mice - The Veterinary Record 1990 Feb 3; 126(5): 111-2

ADIO . Dickinson,A.G. - Scrapie in sheep and goats - Frontiers of Biology 1976; 44: 209-41

AECB . Foote,W.C.; Clark,W.; Maciulis,A.; Call,J.W.; Hourrigan,J.; Evans,R.C.; Marshall,M.R.; de Camp,M. - Prevention of scrapie transmission in sheep, using embryo transfer - American Journal of Veterinary Research 1993 Nov; 54(11): 1863-8

AECX . Foster,J.D.; Hunter,N.; Williams,A.; Mylne,M.J.A.; Mckelvey,W.A.C.; Hope,J.; Fraser,H.; Bostock,C. - Observations on the transmission of scrapie in experiments using embryo-transfer - The Veterinary Record 1996 Jun 8; 138(23): 559-62

AEDA . Foster,J.D.; Hope,J.; Fraser,H. - Transmission of bovine spongiform encephalopathy to sheep and goats - The Veterinary Record 1993 Oct 2; 133(14): 339-41

AEDB . Foster,J.D.; McKelvey,W.A.; Mylne,M.J.; Williams,A.; Hunter,N.; Hope,J.; Fraser,H. - Studies on maternal transmission of scrapie in sheep by embryo transfer - The Veterinary Record 1992 Apr 18; 130(16): 341-3

ANCG . Fraser,H.; Foster,J.D. - Transmission to mice, sheep and goats and bioassay of bovine tissues - Transmissible Spongiform Encephalopathies, R. Bradley and B. Marchant, eds.: Brussels: Working document for the European Commission Ref.F.II.3-JC/0003), pp. 145-159. - Proceedings of a Consultation on BSE with the Scientific Veterinary Committee of the Commission of the European Communities, 14-15 September 1993, (1994)

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AEZP Hadlow,W.J.; Kennedy,R.C.; Race,R.E. - Natural infection of Suffolk sheep with scrapie virus - Journal of Infectious Diseases 1982 Nov; 146(5): 657-64

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AMMU . Wilesmith,J.W.; Wells,G.A.; Ryan,J.B.; Gavier-Widen,D.; Simmons,M.M. - A cohort study to examine maternally-associated risk factors for bovine spongiform encephalopathy - Veterinary Record 1997 Sep 6; 141(10): 239-43

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Acknowledgements

I am thanking Mrs Dr Schütt-Abraham for a stimulating discussion of my work and the Agricultural Ministry in Nordrhein-Westfalen for a financial support.

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