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Rev. sci. tech. Off. int. Epiz., 1996, 15 (3), 827-852 A review of the epidemiology of scrapie in sheep L.J. HOINVILLE * Summary: The aim of this review is to summarise and evaluate the data available about the aetiology of scrapie in naturally affected sheep flocks, particularly data concerning the possible transmission of infection between related animals. The author examines data taken from various relevant studies carried out over the last thirty years. The main conclusions are that scrapie is an infectious disease with a genetic influence on the incubation period. The increased risk of disease in the offspring of affected animals is thought to be largely the result of increased genetic susceptibility, with a large proportion of the cases occurring in high-incidence flocks being the result of horizontal transmission of infection. KEYWORDS: Epidemiology - Genetics - Scrapie - Transmission. INTRODUCTION Although scrapie was first reported in sheep over two hundred years ago (85), the natural means of transmission within affected populations remains unclear. This is largely due to the long incubation period between infection and the development of clinical signs of disease, and the absence of an ante-mortem diagnostic test to detect infected animals during this period. Even histopathological examination of brain tissue may fail to give positive results in recently infected animals incubating the disease. In addition, the genetic influence on the occurrence of clinical disease complicates the interpretation of study results. These factors combine to make experimental studies of the disease difficult, but such factors also have more serious consequences for the interpretation of observational studies of the 'naturally occurring' disease. In these studies the length of time for which potentially exposed animals are monitored cannot be controlled and this may introduce problems of outcome misclassification. The aim of this review is to evaluate and summarise the relevant, available epidemiological data about the occurrence of scrapie in naturally affected sheep flocks, particularly the possible transmission of infection between related animals. Some data from experimental studies are presented, where such data provide information that may aid in the understanding of the aetiology in naturally affected flocks. The epidemiological data were taken from observational and experimental studies carried out over the last thirty years. As knowledge of the disease has developed, it has become clear that some of these studies did not take into account all confounding * Epidemiology Department, Central Veterinary Laboratory, New Haw, Addlestone, Surrey KT15 3NB, United Kingdom. 828 factors. Consequently, the results presented are more an indication of the likely effects of scrapie, rather than an accurate quantitative assessment. The occurrence of scrapie in different countries and within flocks is briefly described. The evidence for a genetic aetiology and for transmission of an infectious agent is discussed. Finally, some of the available information about the incidence of scrapie in the offspring of affected animals is summarised, and used to assess the likely roles of genetic susceptibility, maternal transmission and horizontal transmission of infection in the occurrence of the disease. OCCURRENCE OF DISEASE The world-wide distribution of scrapie is difficult to determine accurately. There is no pre-clinical diagnostic test; clinical signs are variable, and pathological examination of brain tissue or experimental transmission studies provide the only means of confirming infection. The stigma associated with the occurrence of disease means that some farmers may be reluctant to report cases. This means that the disease may remain undetected in some countries unless comprehensive surveillance systems are in operation. Scrapie is known to be endemic in many European countries, such as Iceland (69, 72), as it is in India and the United States of America (USA) (42, 96). The disease has been reported in many other countries as well (48), often, initially, in imported animals. However, in some regions, like Cyprus, the disease was first detected in native sheep (although imported animals may have been the original source of infection) (90). Australia and New Zealand did not, apparently, import the disease with their original European breeding stock. These countries have used vigorous importation, quarantine and culling policies to prevent the subsequent entry of scrapie, and to eliminate outbreaks in imported animals when necessary (4, 5), and have thus remained free of disease. Fluctuations in the incidence and regional distribution of scrapie have been reported within affected countries (72). The exact prevalence of affected flocks within affected countries is difficult to determine for the reasons given above. A team of researchers has conducted several self-administered questionnaire surveys in Britain (67), and found that between 17% and 34% of sheep farmers reported having seen at least one case of scrapie in their flock at some time. A similar survey in the Netherlands found that 6% of farmers reported ever having seen a case in their flock (80). These studies relied on the opinions of the farmers as to whether the animals were affected with scrapie, and may also suffer from selection bias as response rates for postal questionnaires were poor and, in Britain, a non-random sample was used. Although the exact figures may be inaccurate they do show that a considerable number of flocks are affected. The annual within-flock incidence found in these two studies was between 0.3% and 1.8%. The within-flock incidence has been reported as 2 % in Britain (95), 3 % to 5% in Iceland (69) and 1% to 10% in India (56). A within-flock incidence of 2 0 % to 30% has been reported in some flocks in Iceland (82), as well as in animals born in certain years in some British flocks (71). Scrapie occurs in both sexes and in the majority of sheep breeds, although the incidence may be higher in some breeds than in others (13, 95). This may reflect a variation in the genetic susceptibility between breeds. 829 AGE OF AFFECTED ANIMALS The majority of clinical cases occur in sheep between two and five years of age (15). However, cases have been reported in animals as young as twelve months (56), ten months (55), and seven months (82) and in animals as old as eleven years (70). Although only 10% to 15% of cases reported in Britain were found in sheep of more than four-and-a-half years old (70, 71), it is possible that many older animals in the pre-clinical stage of infection were culled before the disease was detected (16). Histopathological changes have been reported in lambs as young as eleven months (17), and the infectious agent is thought to have been present in the brains of eight-month-old lambs which were used to prepare a vaccine for louping-ill (ovine encephalomyelitis). These lambs were the offspring of scrapie-infected ewes, and the use of the vaccine resulted in the transmission of scrapie to vaccinated animals in flocks with no history of clinical disease (34, 41). AETIOLOGICAL CONTROVERSY Early observations on the occurrence of scrapie in flocks that had previously been free of disease, after the introduction of affected animals, suggested that scrapie could be a contagious disease (64). A contagious disease is defined as a condition caused by an infectious agent, which can be transmitted from infected to uninfected animals. This theory was supported by the experimental transmission of infection to sheep in 1936 (10) and to goats in 1939 (11). In the 1950s it was shown that scrapie was caused by a filterable, self-replicating agent (86, 93). In the early 1960s the disease was transmitted to various experimental animals (6, 97). But the study of scrapie has been complicated by the fact that an infectious agent has never been isolated, by the long incubation period between infection and the onset of clinical signs of disease, and by the problem of detecting animals which are pre-clinically infected. As a result the aetiology of scrapie has been a subject of controversy for many years. Despite the experimental evidence that scrapie was an infectious condition, some of those working with the disease in the 1960s believed that scrapie was an inherited condition. The variation in the incidence of scrapie between different breeds of sheep, and especially between different families, suggested a genetic influence on the occurence of the disease. The natural disease appeared to favour flocks of certain breeds (95), although experimental studies to investigate this variation may have overestimated the difference in susceptibility between breeds (15). It was later shown that there is considerable variation in the incidence of scrapie between different flocks of the same breed (68). There was also, however, experimental evidence that some breeds were more susceptible to inoculation with sheep scrapie brain pool (SSBP/1) (37). Stronger evidence for a genetic influence was provided by the variation in the incidence of disease among different families within affected flocks (71). This was further supported by experimental evidence for a variation in the incidence of scrapie among families of sheep after inoculation with SSBP/1 (39). 830 These findings led some of those investigating the disease in the 1960s to believe that scrapie was an autosomally recessive inherited condition, which could be transmitted experimentally but which could not be transmitted from an affected animal to an unaffected animal, except by inoculation (71). This belief was based on the following three pieces of evidence. First, the pattern of disease occurrence in naturally affected flocks and in experimentally inoculated animals was shown to be similar to that of an autosomally recessive condition (39, 7 1 , 72). Secondly, it was shown that epidemics of a genetically transmitted disease could, theoretically, occur after the introduction of infected animals (25). Finally, some of the early attempts to introduce the disease to unaffected flocks, by transferring animals from affected flocks or exposing healthy animals to infected stock, gave negative results (71, 74, 86). However, Parry demonstrated the autosomal recessive pattern of inheritance by investigating the occurrence of scrapie in commercial flocks (72). It has been suggested that there may have been considerable misclassification of animals in these flocks, due to the culling of infected animals before the onset of clinical signs of disease. It was also suggested that the allocation of a presumed genotype to animals on the basis of the presence or absence of scrapie in their progeny gave rise to a circular argument. Dickinson et al. presented further evidence on the occurrence of scrapie in affected flocks which was not consistent with either recessive or dominant genetic transmission (17). Both Parry and Draper assumed that animals carrying the scrapie-susceptible allele were likely to be of superior conformation and so would be given preference in the selection of flock replacements (25, 72). They suggested that this would account for the deviance from the expected pattern for an autosomal recessive condition, but this assumption has been questioned (95). In the experimental work of Gordon, the autosomal recessive pattern of susceptibility to inoculation with SSBP/1 did not occur in all breeds. He suggested that this may be due to errors in the recording of relationships or incomplete penetration or dominance of the genetic effect (39). The transmission of infection from affected to unaffected animals provides the strongest evidence that scrapie is a contagious disease and not an inherited condition. The studies referred to earlier, which failed to demonstrate the spread of disease, did not take into account the need for long observation periods. The studies that have shown that scrapie can be transmitted from affected sheep to unaffected sheep, with no inoculation of infected tissue, are listed in Table I. The studies conducted by Brotherston et al. in 1968 (1), Dickinson et al. in 1974 (21) and Hourrigan et al. in 1979 (48) are the most reliable, as these researchers ensured that exposed animals were very unlikely to be incubating the disease before exposure, and examined all exposed animals pathologically to confirm the occurrence of scrapie. For example, the study of Dickinson et al. exposed sheep from a large isolated flock that had been free of scrapie for at least twelve years (21). The remaining studies listed in Table I provide some evidence for the transmission of infection but could be criticised, as it is possible that the animals involved were pre-clinically infected before exposure to infected animals. The occurrence of scrapie in a goat born into a flock of sheep with a high prevalence of scrapie in the 1940s provided strong evidence for the spread of the disease, as scrapie is rarely reported in goats (8). There have been several subsequent reports of the occurrence of the disease in goats kept in contact with affected sheep ( 1 , 48, 84, 85, 87, 91). I ally animals were followed (months) t ced bs 3.2 13.5 3.2 18.8 3.0 17.4 10.7 35.7 28.0 42.8 35.0 16.7 22.0 3.3 57.1 25.0 37.5 25.0 18.5 13.0 25.0 gg 5 S 4 S ? ä ^ ^' ^ P c£ S ^ ^ ^ ^ ^ I IIIIIIII~ ~~ ~~ ~~ ~ IIIIIIIIIIIIllIIIIlIl M» ti * Animals not in direct contact with infected animals were grazed on pasture previously used by infected animals ? Information unknown 3 months From birth 6 months From birth 8 months From birth 7-9 months From birth From birth 1-3 days Girnrncr Adult Birth ? From birth Adult Weaning From birth ? Birth only 2.5 years infected Time for itact Oi 1 l! 31 111 31 101 33 23 28 28 75 11 20 6 9 30 14 6 8 20 27 23 4 P contact with : aturally or w contacts s Rambouillet Rambouillet Targhee Targhee Hampshire Hampshire Suffolk Suffolk Blackface Blackface Halfbred Cheviot Halfbred x Che\not Cheviot Suffolk Suffolk Suffolk Suffolk Suffolk Suffolk Suffolk Age when first exposed Í Number TABLE I Transmission of infection from infected sheep to scrapie-free is) it Reference 831 ^ ^' á 832 Most experts now recognise scrapie as an infectious condition which can be transmitted from affected to unaffected animals. DEMONSTRATION OF A DOSE-RESPONSE RELATIONSHIP The evidence for the transmission of infection from affected to unaffected animals would be strengthened if a dose-response relationship could be demonstrated. If animals which were exposed to a greater dose of infection had either a higher incidence of disease, or were younger when clinical signs of disease developed, this would indicate the existence of a dose-response relationship. In experimental transmission studies the incidence of disease is lower and the incubation period longer when the inoculum is diluted (38, 58). Variations in the incidence of disease, or the age at the onset of disease, could be the result of differences in the level of exposure to scrapie, the duration of exposure or the age when first exposed. The variations in the incidence of scrapie among animals that are naturally exposed to different levels of infection provide evidence of a dose-response relationship. These variations are demonstrated by the high within-flock incidence seen in some Icelandic flocks. This high incidence has been attributed to the high level of exposure to infected animals, and possibly an infected environment, during the winter housing period, with a higher incidence of scrapie in animals born during a long winter housing period (69, 82). An increase in the incidence of the disease and a decline in the age at which the disease begins have been reported within affected flocks, and been attributed to increasing levels of environmental contamination (7, 30, 57, 82). The variation in the incidence of disease in sheep which are exposed to an infected environment for different lengths of time also provides evidence of a dose-response relationship. This is demonstrated by studies in which lambs born into an affected flock are removed from exposure at various ages (Table II). The final observation that provides evidence for a relationship between the dose of exposure and the occurrence of disease is the variation in the incidence of disease with the age at first exposure. The data in Table I show that there was a higher incidence of disease in sheep which were first exposed to scrapie at birth, than in sheep which were first exposed when between three and nine months of age (48). Sheep which were first exposed at birth were also affected at younger ages. This may be partly explained by the confounding effect of environmental contamination. Assuming that the environmental contamination at the field study site increased over time, then those animals born at the study site were exposed to a greater level of environmental contamination than sheep that had been introduced to the site at the start of the study. A short incubation period in sheep exposed to the disease at an early age is also suggested in experimental studies by the occurrence of scrapie in the offspring of inoculated ewes at between seven and nine months of age (18, 37). TRANSMISSION FROM A CONTAMINATED ENVIRONMENT Evidence for the transmission of infection from a contaminated environment was provided when scrapie-free sheep developed the disease after grazing pasture used by clinically affected sheep approximately twice a week over a three-year period with no 833 TABLE II The relationship between the incidence of scrapie and the duration of exposure Exposure Infected flock Infected flock Offspring of naturally affected ewes Offspring of inoculated ewes Age when removed from exposure (months) Number exposed Incidence (%) Reference 0 63 10 (48) 4 32 16 9 17 29 20 32 41 0 23 13 - 20 25 0 7 8 29 75 (18) 109 115 4 15 (41) -0 - (17) direct contact between the scrapie-free and affected sheep (40). However, the possibility that the introduced animals may already have been infected could not be ruled out in this study. Stronger evidence of environmental contamination was provided by the attempts to eradicate maedi-visna and sheep pulmonary adenomatosis in Iceland, by slaughtering affected flocks and re-stocking with lambs from scrapie-free areas. Scrapie recurred on some sites that had previously had a high incidence of scrapie, but not on sites where scrapie had not previously occurred. This recurrence only on heavily affected sites makes contamination of the environment a more likely explanation than pre-clinical infection of the introduced stock (69, 83). The subsequent, more vigorous control policy, which included thorough disinfection of affected premises and a longer rest period, followed by re-stocking with a more reliable source of scrapie-free stock, has been more successful (82). The resistance of the scrapie agent to various disinfection procedures and its survival in soil for three years make contamination of the environment biologically plausible (2, 89). However, the survival of the scrapie agent in soil was demonstrated in an experimental study using a high titre of infectious agent, and the titre in the material that was recovered at the end of three years was much reduced. TRANSMISSION OF INFECTION FROM INOCULATED ANIMALS It has been observed that animals that are inoculated with the scrapie agent are less able to transmit the infection to goats or sheep than naturally infected animals (1, 73, 76, 77). However, scrapie has occurred in animals that have been in contact with inoculated sheep (35), as well as in the non-inoculated offspring of inoculated animals (18, 37). 834 Pattison has suggested that experimentally inoculated animals were unable to transmit scrapie because they were usually males or non-breeding females and did not, therefore, excrete the agent in placental tissue (78). The placenta has been identified as one possible source of the infectious agent (see 'Source of infection'). TRANSMISSION FROM INFECTED ANIMALS IN THE PRECLINICAL PHASE Early reports from the United Kingdom suggested that scrapie occurred in unaffected flocks after the purchase of pre-clinically infected sheep (64, 88). This is now believed to be the usual means of introduction to unaffected flocks in Britain (73), the USA (48) and Iceland (82). The theory of transmission of infection from preclinically infected animals is supported by the pattern of disease occurrence in the offspring of infected ewes. There is no evidence that the incidence of scrapie in such offspring depends on whether or not the ewe had developed clinical signs of the disease when these offspring were born. The risk of scrapie developing in offspring that were born the year before the dam had developed clinical signs was the same as for offspring that were born in the year of the onset of the disease in the dam (15, 7 1 , 72). SOURCE OF INFECTION For infected animals to transmit infection to healthy animals, they must first excrete the agent, which must then enter the body of the healthy animal. The attempts to isolate the infectious agent of scrapie in various peripheral tissues and excretions taken from infected animals are summarised in Table III. The isolation of the agent from the placenta of infected sheep, and possibly from foetal cotyledon and amniotic fluid, suggests that the placenta is one source of infection. A study by Pattison of the transmission of infection using placental tissue (78, 79) has been criticised as no control animals were used (92), but the high incidence of disease in the exposed animals makes pre-clinical infection of those exposed animals an unlikely explanation of the results. Dickinson suggests that ewes may eat the placenta and then excrete the infectious agent over pastures with little or no reduction of infectivity after passage through the alimentary tract. This was based on evidence from unpublished experimental work in mice, which showed that the scrapie agent could pass through the alimentary tract with no loss of infectivity (15). In contrast, there has been no successful isolation of the scrapie agent from the faeces, urine or saliva of infected sheep or goats (Table III). Only a limited number of experiments using clinically infected donor sheep have been carried out and the results are crucially dependent on the sensitivity of the bioassay used to detect infection. The infectious agent may be present in these excretions at a lower titre and the assays used may have been unable to detect this level of infection. If these excretions were infected, they may represent a more important source of infection than the placenta. Even a low concentration of the agent continuously excreted in faeces or urine could result in a greater level of environmental contamination than a single excretion of a higher concentration of the agent in the placenta once a year. However, 835 TABLE I I I Isolation of scrapie agent from peripheral tissues and excretions Donor animals Tissue or excretion . . Natural or . . . Species Clinical rasps experimental infection c a s e s Number of donors in which infection was detected Assay used Reference R o u t e SDecies F o f inoculation Intestine Intestine Intestine Faeces Faeces Faeces Faeces Faeces Interna] parasite Haemonchus contortus Haemonchus contortus Retropharyngeal lymph node Retropharyngeal lymph node Retropharyngeal lymph node Nasal mucosa Nasal mucosa Salivary gland Salivary gland Salivary gland Sheep Sheep Goats Sheep Sheep Goats Goats Goats Sheep Sheep Yes No Yes No ? Yes Yes Yes ? Yes Natural Natural Natural Natural Natural Natural Experimental ? Natural Experimental '9/9 5/8 3/3 0/8 0/4 0/3 0/1 0/? 1/5 0/6 Mice Mice Mice Mice Mice Mice Goats Goats Mice Sheep Cerebral Cerebral Cerebral Cerebral Cerebral Cerebral Cerebral Oral Cerebral Cerebral (44) (44) (43) (44) (47) (43) (75) (73) (47) (26) Goats Yes Experimental 0/6 Goats Cerebral (26) Sheep Yes Natural 9/9 Mice Cerebral (44) Sheep No Natural 7/8 Mice Cerebral (44) Goats Yes Natural 3/3 Mice Cerebral (43) Sheep Goats Sheep Goats Goats Natural Natural Natural Natural Experimental 3/9 3/3 0/9 0/3 5/14 Mice Mice Mice Mice Goats Cerebral Cerebral Cerebral Cerebral Cerebral (44) (43) (44) (43) (77) Saliva Saliva Saliva Saliva Kidney Kidney Urine Urine Urine Urine Sheep Sheep Goats Goats Sheep Goats Sheep Goats Goats Goats Yes Yes Yes Yes Yes (3) No (11) Yes ? Yes Yes Yes Yes ? Yes Yes Yes Natural Natural Experimental ? Natural Natural Natural Natural Experimental Experimental 0/9 0/2 0/1 0/? 0/9 0/3 0/4 0/3 0/1 0/14 Mice Mice Goats Goats Mice Mice Mice Mice Goats Goats Cerebral Cerebral Cerebral Oral Cerebral Cerebral Cerebral Cerebral Cerebral Cerebral (44) (47) (75) (73) (44) (43) (47) (43) (75) (77) Mice Mice Sheep and goats Mice Mice Mice Cerebral Cerebral Cerebral (44) (47) (78, 79) Cerebral Cerebral Cerebral (47) (47) (47) M INO N Foetal cotyledon Foetal cotyledon Placenta Sheep Sheep Sheep Yes ? Yes Natural Natural Natural 0/2 2/10 6/6 Amniotic fluid Pasture Trough water Sheep Sheep Sheep ? ? ? Natural Natural Natural 1/1 0/2 0/2 Positive isolation from salivary gland mentioned (41) Positive isolation from goat foetal cotyledon mentioned (48) Negative attempt at isolation from contaminated building manure mentioned (48) 836 the small number of attempts to isolate the agent from potentially contaminated environmental sources have all been negative. ROUTE OF ENTRY TO RECIPIENT ANIMALS The oral route is a likely means of entry to the bodies of recipient animals. Experimental studies have shown that the intra-gastric route of scrapie infection is effective in mice (61). Oral administration of infected brain tissue has been shown to transmit infection in both sheep and goats (75). Infected placental tissues have also transmitted scrapie to sheep by the oral route (78, 79). The isolation of the scrapie agent from the alimentary tract of pre-clinically, naturally infected lambs also suggests that this may be the natural route of entry (44). Scarification of the skin and conjunctival inoculation are other suggested routes of entry for infection (15, 81). GENETIC CONTROL OF SUSCEPTIBILITY There is now considerable evidence that scrapie can be transmitted between unrelated animals and the suggestion that the disease is an autosomally recessive inherited condition seems untenable. But there is strong evidence that genetic factors in the host influence susceptibility to the disease. This idea was first suggested by the familial pattern of occurrence of natural scrapie (71), and confirmed during experimental inoculation studies, in which exposure to the scrapie agent could be controlled. These experimental studies were designed to create 'susceptible' and 'resistant' lines of sheep, by selective breeding of a foundation flock according to the response of related animals to inoculation with SSBP/1. The incidence of experimentally induced scrapie in selected lines of Cheviot and Herdwick sheep suggested that susceptibility to inoculation with SSBP/1 was controlled by a single gene known as Sip (a sheep gene exerting control over Scrapie incubation period) (15, 20, 68). The Sip gene has two alleles, sA and pA, and the sA allele is dominant for susceptibility to disease after inoculation with SSBP/1. A further study using Swaledale sheep examined the incidence of experimentally induced scrapie in the offspring of sheep which had not developed clinical disease after inoculation with pooled field strains of the scrapie agent. The study compared this incidence to the incidence in control animals. This study also concluded that susceptibility to scrapie was a dominant inherited trait (46). It is believed that the same gene is responsible for genetic control in all three breeds (49). Further work showed first, that the sA allele may be only partially dominant, and secondly, that experimentally challenged, apparently resistant, homozygous pA sheep may develop scrapie after a long incubation period. Partial dominance was indicated by the division of susceptible Swaledale sheep into two distinct groups according to the length of the incubation period of the disease (13), suggesting that homozygous susceptibles had a shorter incubation period. 837 The absolute resistance of homozygous pA animals was brought into question by the occurrence of scrapie (after a very long incubation period) in a few of the Herdwick sheep that had been selectively bred for resistance to subcutaneous inoculation with SSBP/1 (67). Scrapie also occurred in some of the homozygous pA Cheviot sheep that were resistant to subcutaneous inoculation with SSBP/1, after these sheep had been inoculated intra-cerebrally (20). These results suggest that the Sip gene controls the incubation' period of scrapie in experimentally inoculated sheep. This could give the appearance that the Sip gene influences the susceptibility of sheep to inoculation with the scrapie agent, because those animals with an incubation period longer than the observation period, or longer than their natural lifespan, will appear to be resistant to inoculation. The mechanism by which the Sip gene controls the incubation period of scrapie in sheep may be similar to the way in which the homologous gene in mice, known as Sinc (Scrapie incubation period), controls the incubation period of experimental murine scrapie. The Sinc gene has two alleles, p7 and s7, so called because homozygous s7 mice inoculated with the ME7 strain of the scrapie agent develop scrapie with a short incubation period, whereas the incubation period is prolonged in homozygous p7 mice (19). When mice are inoculated with scrapie, using the intra-peritoneal route, the initial replication of the disease takes place in the lymphoreticular system (LRS), before replication in the central nervous system (CNS) produces the lesions that result in clinical disease (60). Variations in the Sinc genotype have only a small effect on the onset of detectable replication of the ME7 strain of scrapie in the LRS after intra peritoneal inoculation. But there is a profound effect on the time of onset of replication in the brain. Such replication occurs earlier in homozygous Sinc s7 mice than in homozygous p7 mice (59). Other studies, using the 87V strain of the scrapie agent, showed that it is possible to produce a life-time persistence of infection in the LRS of homozygous Sinc p7 mice without the occurrence of clinical disease, by using intra-peritoneal inoculation. The scrapie agent was able to replicate itself in the brain, but neuroinvasion was severely impaired (3, 9). Similar results were obtained when C57BL mice were inoculated by the intra-peritoneal route with a low dose of the 22A strain of the scrapie agent (22). These results show that the specific interactions between different strains of the agent and the alleles of the Sinc gene greatly influence neuroinvasion and the subsequent replication of the scrapie agent in the CNS. The growing evidence that Sip and Sinc genes are the same as the ovine and murine PrP genes, respectively (50), justifies extrapolating these findings to sheep scrapie. GENETIC CONTROL OF THE NATURAL DISEASE IN SHEEP The role of Sip in the control of natural scrapie was suggested by the occurrence of major outbreaks of natural scrapie in those Cheviot and Herdwick flocks that had been selected for susceptibility to experimental inoculation with the scrapie agent (14, 57), but not in those flocks selected for resistance. This was confirmed in an experiment, using some of the unaffected female offspring of affected rams from a Suffolk flock which had been bred to maximise the occurrence of natural scrapie. These ewes were assumed to be heterozygous sApA. They were 838 mated with Cheviot rams, which had been bred for resistance to experimental inoculation and were assumed to be homozygous pA. The incidence of disease in the offspring of these matings, following subcutaneous inoculation with either the Cheviot-derived SSBP/1 or a Suffolk strain of the scrapie agent, known as" SUF81, was as one would expect, assuming that a single gene from these Suffolk ewes and Cheviot rams conferred dominant susceptibility to scrapie in the cross-bred offspring (28). The pattern of inheritance in naturally affected flocks suggests that susceptibility to scrapie is a recessive trait (28, 71), not a dominant trait as has been indicated in experimental studies (20, 46, 68). This could be explained by a difference between the effective exposure of naturally exposed sheep, and the effective exposure of experimentally exposed animals. The effectiveness of the exposure is likely to depend on the dose of the agent received and the route of infection. The route of experimental inoculation in mice is known to determine the effective dose of the agent received (58), in that the oral route of exposure is less efficient than intra-cerebral inoculation and other parenteral routes (61). Heterozygous susceptible sheep, exposed to a low dose of the scrapie agent by the oral route, may not develop clinical disease within their lifetimes and the natural disease may appear to occur in an autosomal recessive pattern. BIOLOGICAL MARKERS FOR GENETIC SUSCEPTIBILITY It may now be possible to determine whether sheep are genetically susceptible to scrapie without the need for prolonged breeding experiments and inoculation of the scrapie agent. It has recently been shown that specific variations in the nucleotide sequence in and around the PrP gene are associated with alleles of the Sip gene. This was first demonstrated in experiments using Cheviot sheep bred for susceptibility or resistance to inoculation with SSBP/1 (51). This work has provided evidence that Sip and PrP may be the same gene, and that polymorphisms in the PrP gene may provide biological markers for Sip alleles that could predict susceptibility to scrapie. In addition, these markers were used to confirm the partial dominance of the Sip allele by dividing susceptible animals into those that were homozygous and those that were heterozygous for the PrP polymorphism associated with the Sip susceptibility allele. Homozygous susceptible Cheviot and Cheviot crosses, when inoculated subcutaneously with SSBP/1, develop the disease after a shorter incubation period than heterozygous susceptible animals (31, 65). Several studies have now shown that these PrP polymorphisms are associated with the occurrence of scrapie in naturally affected flocks (52, 53, 54, 62). The pattern of genetic susceptibility may be more complex than has been suggested so far, with different strains of the scrapie agent producing different patterns of susceptibility. In experimental studies mice that are apparently resistant to inoculation with one strain may develop clinical disease when inoculated with another strain (23). Differences between the genetic susceptibilities of sheep that have been inoculated with different strains of the scrapie agent have also been demonstrated (29, 33). These results suggest that disease occurrence is determined by an interaction between the host genotype and the strain of the scrapie agent. This interaction may explain why different PrP polymorphisms are not consistently associated with the occurrence of natural scrapie in different breeds of sheep (53, 63). 839 INCIDENCE OF SCRAPIE IN THE OFFSPRING OF AFFECTED ANIMALS An increase in the incidence of scrapie in the offspring of affected animals may simply reflect the genetic influence on the occurrence of the disease, although it could also indicate that the infectious agent is transmitted from parent to offspring (17). Several studies have investigated the incidence of scrapie in the offspring of affected and unaffected sheep. These data have been summarised in Table IV. These results are presented as an indication of likely effects rather than as an accurate quantitative assessment. The most important problems in interpreting the data from these studies are, first, the possible misclassification of infected animals which showed no clinical signs of the disease as unaffected and, secondly, the confounding effects introduced by the management of the flocks. The misclassification of infected animals is likely to be a problem in the work of Parry, as not all brains were examined histopathologically to determine whether culled animals were pre-clinically infected. The confounding effects of flock management are likely to be important in the early work of Dickinson et al. in which the offspring of affected and unaffected animals were separated (17). The data from the 1974 paper of Dickinson et al. (21) and the 1979 paper of Hourrigan et al. (48) are included because they provide probably the least biased estimates of the effects of these risk factors. The data from the 1965 paper of Dickinson et al. (17) and Table 5 of the 1962 paper of Parry (71) are included as they are often quoted as evidence that the dam has a greater influence on the risk of disease than the sire (92, 94). Further examples of the data of Parry, in addition to Table 2 of the 1962 paper, have been included to show the variable dam and sire effects obtained in analysing the data from different groups of animals (71, 72). The data presented in Table IV have been analysed to provide answers to two important questions. First, relative risks (RR) have been calculated to determine whether the risk of disease is greater in the offspring of affected animals than in the offspring of unaffected animals born into the same flock. Furthermore, if the risk of disease is greater in the offspring of affected animals, then are the offspring of affected dams and the offspring of affected sires at equal risk? Secondly, population attributable risk percentages (PAR%) have been calculated to determine the proportion of all cases of scrapie occurring in these populations that occur because these sheep are the offspring of affected animals. The relative risks (RR) were calculated by dividing the incidence of disease in the offspring of affected animals by the incidence of disease in the offspring of asymptomatic animals. An RR significantly greater than one indicates that the offspring of affected animals are at increased risk of developing scrapie. In all of the studies presented, the offspring of affected dams have a significantly increased risk of developing scrapie, with the RR varying from 1.85 to 4.80. The effect of the sire on the risk of disease is not so clear. In only some of the studies did the offspring of affected sires have a significantly increased risk of developing scrapie. However, in all of the studies, the offspring of an infected dam and an infected sire are at greater risk than the offspring of an infected dam and an uninfected sire. The data from the early study of Dickinson suggest that the offspring of affected sires are not at increased risk of developing scrapie (15); however, the confounding effect RR: relative risk Suffolk Suffolk S3 60.5 S Ä vi cu 50.0 (4/8) 8.0 (26/325) 27.1 (23/85) 6.6 (18/273) 85.7 (12/14) 14.7 (25/31) (15/102) 80.6 18.5 (5/27) (46/76) 13.5 8.9 19.8 11.9 4.4 38.8 (24/75) 32.0 (10/22) 46.0 (7/32) 21.9 (2/11) 18.2 (24/65) 36.9 (50/129) 1.64 (2.13-4.72) 3.17 (1.01-2.17) 4.61 (0.84-1.70) 1.19 (0.96-1.31) 1.15 (0.93-1.52) 1.19 (1.19-2.25) PAR%: population atlributable risk percentage (2.34-4.38) 3.20 (1.85-2.85) 2.30 (3.12-7.40) 4.8 (2.30-9.56) 4.69 (1.49-2.43) 1.86 35 Suffolk cu E Suffolk CU' 35.3 & (54/153) « «¡£ >n X Blackface 5.2 21.5 35.5 17.9 3.14 (16/18) 88.9 (74/79) 93.9 (18/21) 85.7 (14/18) 93.8 7.5 68.5 27.1 31.6 (8.07-17.23) 11.79 (8.18-20.5) 30.4 (3.68-9.92) 6.04 (2.30-11.20) 5.07 (1.84-3.20) 2.43 (2.07-4.76) 8.8 CI: confidence intervals (31/32) (48/56) 86.0 13.00 77.8 eu (1.37-2.51) PÄ 35 1.85 i# 41.9 Risk in offspring of two affected parents IT) Suffolk -Ç3 •S CU (13/31) RR (95% CI) HS 24.8 Incidence (%) (cases/at risk) Risk in offspring of affected sire is (26/105) Incidence (%) (cases/at risk) Risk in offspring of affected dam u Suffolk Breed Incidence in offspring of two unaffected parents The incidence of scrapie in the offspring of affected and unaffected sheep TABLE IV (Assumes parental outcome from genotype) (71: Table 2) (Assumes parental outcome from genotype (72: Table Al-A) management information from (41) (71: Table 5) (15: Table 10.2) (21: Table 1) (48: Table 2) Reference 840 35 S Ä 841 of flock management in this study has already been mentioned. The affected and unaffected flocks were initially separated, so that the offspring of affected ewes were kept in an affected flock and the offspring of unaffected ewes were kept in an unaffected flock (17). This means that horizontal transmission could be an important confounding factor, which would exaggerate the effect of the disease status of the dam, and underestimate the effect of the status of the sire on the risk of disease. In the later study of Dickinson et al. (21), the affected and unaffected flocks were not separated, but data from three generations of matings were combined to produce the results given in Table IV. Although there was no significant increase in the risk of developing scrapie for the offspring of affected sires using the combined data from all generations, in the final F generation the offspring of affected sires were at significantly increased risk of developing disease (RR = 1.48, 9 5 % confidence interval ± 1.01-2.17). In summary, these results suggest that both the dam and the sire have an effect on the risk of the offspring developing scrapie and, although the dam may have more effect than the sire, the extent of that difference is not as great as has been suggested in the past. 2 Any difference between the effect of the dam on the risk of disease and the effect of the sire could be used to indicate if the increased risk in the offspring of affected animals is the result of genetic susceptibility, transmission of infection from the dam or the sire to the embryo, or transmission of infection from the dam during gestation or post-partum. Lambs do not usually have any contact with the sire after conception. An increase in the risk of disease occurrence in the offspring of affected sires would suggest that at least part of the increased risk for the offspring of affected animals is due to a genetic susceptibility to the disease or transmission of infection to the embryo. Given the previous evidence for a genetic influence on susceptibility to scrapie, and the experimental evidence that the scrapie agent has never been isolated from semen or male reproductive tissues (Table V), any effect of the sire on the incidence of disease is most likely to be a genetic effect. Assuming that the increased risk in the offspring of affected sires is because these animals are genetically susceptible to scrapie, it is likely that the risk of disease in the offspring of affected dams is increased by a similar amount as a result of increased genetic susceptibility. A greater risk of disease in the offspring of affected dams than in the offspring of affected sires could be due to transmission of infection from the mother to the embryo or foetus during gestation, or to the lamb post-partum. There is some evidence that infection may be transmitted to the embryo. An embryo transfer experiment has recently been performed. Unwashed embryos from inoculated donor ewes (which subsequently developed clinical disease) were transferred to genetically resistant recipent ewes. Six of the twenty lambs produced in this experiment developed scrapie, which suggests that transmission to the embryo may occur (32). This conflicts with the results of a previous study in which washed embryos were transferred from inoculated ewes (which subsequently developed clinical disease) to recipient ewes from scrapiefree flocks. None of these twenty-seven lambs developed scrapie (27). These results are difficult to reconcile but the genetic susceptibility of the lambs born, the origins of the scrapie-free embryo recipients, the interval of time between the inoculation of embryo donors and the collection of the embryos, and the washing of collected embryos are all important variables which differed in these studies. The positive results of the most recent study suggest that infection of the embryo is a possibility that requires further investigation. In addition, the scrapie agent has been isolated from the 842 TABLE V Isolation of scrapie agent from reproductive and mammary tissues Number of donors Assay used , Natural or in which experimental infection Species Route infection detected Reference Donor animals Tissue or excretion Species C h m c a c a s e s l Sheep Yes Natural 2/7 Mice Cerebral (44) Mammary gland Sheep Yes Natural 0/7 Mice Cerebral (44) Mammary gland Goats Yes Natural 0/3 Mice Cerebral (43) Colostrum Sheep 0/1 Mice Cerebral Sheep No ? Natural Colostrum Natural 0/3 Mice Cerebral (44) (47) Milk Sheep ? Natural 0/6 Mice Cerebral (47) Milk Goats Yes Natural 0/3 Mice Cerebral (43) Milk Goats Yes Experimental 0/1 Goats Cerebral (75) Ovary Sheep 0/9 Mice Cerebral (44) Sheep Yes ? Natural Ovary Natural 4/14 Mice Cerebral (47) Ovary Goats Yes Natural 0/3 Mice Cerebral (43) Uterus Sheep 0/3 Mice Cerebral (44) Sheep Yes ? Natural Uterus Natural 4/13 Mice Cerebral (47) Uterus Goats Yes Natural 0/3 Mice Cerebral (43) Testis/seminal vesicles Sheep Yes ? Natural 0/1 Mice Cerebral (44) Natural 0/6 Mice Cerebral (47) 0/21 Mice Cerebral (47) (44) (47) Supra-mammary lymph node « Testis/seminal vesicles Sheep Semen Sheep ? Natural Foetal tissue Sheep 0/4 Mice Cerebral Sheep Yes ? Natural Foetal tissue Natural 1/13 Mice Cerebral . ? Information unknown ovary, uterus and foetal tissues from infected animals in some studies (Table V). This suggests that infection may be transmitted in the female reproductive tract. The possibility that infection is transmitted from mother to offspring in utero is also supported by the occurrence of scrapie, at a young age, in the non-inoculated offspring of inoculated ewes (18, 36, 37). 843 The increased risk of disease in the offspring of affected dams could also be due to the post-partum transmission of infection, possibly as a result of contact with placental tissue. Post-partum transmission of infection in milk is also a possibility but the scrapie agent has never been isolated from the milk of affected ewes in experimental studies (Table V). The theory of post-partum transmission of infection is supported by the increased risk of disease in those naturally exposed animals that remain in a potentially contaminated environment for longer periods (Table II). The second question that was addressed using the data in Table IV was the proportion of the cases in these populations that occurred because these animals were the offspring of infected sheep. The population attributable risks percentages (PAR%) were calculated using the equation given in the Appendix. Such percentages provide an estimate of the proportion of cases that occurred within the study population because they were the offspring of affected animals. The remaining cases in these populations cannot be attributed to parental transmission of disease and would have occurred whether or not the parents of these sheep were affected. If infected animals are misclassified as uninfected animals, then the RR and the proportion of the population exposed will be underestimated. This, in turn, means that the PAR% will also be underestimated. Such a misclassification will occur if animals incubating scrapie are culled before they develop clinical signs, or if animals can carry and transmit infection without developing clinical signs. As both these sources of misclassification are likely to occur in scrapie-affected flocks, the PAR% have been calculated to give an indication of the proportion of cases that may be due to increased genetic susceptibility or transmission of infection from infected animals to their offspring, but cannot be used to provide an accurate quantitative assessment. The results from the two most reliable studies at the top of Table IV suggest that a total of between 2 1 % and 3 2 % of cases can be attributed to parental status. This figure was obtained by combining the proportion of cases attributable to affected sires, to affected dams and to two affected parents. For example, using the data from the first study listed, 17.9% of cases occurred because the sire was affected, 5.2% because the dam was affected and 8.8% because both parents were affected, giving a total of 31.9% of cases that could be attributed to parental status. However, as part of the increased risk in the offspring of affected animals is thought to be due to an increased genetic susceptibility of these animals, only a proportion of these cases could be attributed to maternal transmission of infection. These results suggest that a considerable proportion of the cases occurring in these flocks are the result of horizontal transmission of infection between unrelated animals. These estimates apply only to these populations. The proportion of cases attributable to maternal and horizontal transmission in other flocks will depend on the incidence of disease in the flock and the amount of contact between unrelated animals. It is possible that in low-incidence flocks, and in flocks where there is not much contact between unrelated animals, maternal transmission of infection may account for a much greater proportion of cases. It has been suggested that maternal status is important in determining whether lambs develop scrapie when the disease is first introduced to a flock, but that, as levels of environmental contamination increase, horizontal transmission becomes more important (24). This could be the reason why the offspring of affected sires appear to be at increased risk only in the later generations in the 1974 study of Dickinson et al. (21), when the level of environmental contamination had increased. 844 CONTROL OF DISEASE The evidence for an increased risk of disease in the offspring of affected animals has been used as the basis of selective culling policies to control the occurrence of scrapie in affected flocks. This approach has been effective in some flocks, especially if combined with lambing management practices to reduce contact between animals, and thus reduce the horizontal spread of the disease (66). But the culling of maternal bloodlines has not been completely effective in controlling the disease in the USA (12), which, as is consistent with the evidence presented here, suggests that horizontal transmission of infection is important. Culling animals that are born at about the time that an infected ewe has lambed has been suggested as an additional control procedure. A research project is in progress at the Central Veterinary Laboratory (CVL) to determine whether there is any evidence to support the hypothesis that these animals are at increased risk of developing scrapie. However, any such culling policy could be undermined by the possible transmission of infection from pre-clinically infected animals and from sources other than the placenta. The use of careful lambing management and disinfection procedures may aid in controlling the spread of infection from pre-clinically infected animals, but environmental contamination from other sources would be very difficult to control. A more effective method of controlling clinical disease may become available with the identification of genetically resistant rams, which could be used to increase the proportion of genetically resistant breeding females in a flock. However, it will be important to determine whether these genetically resistant animals are capable of carrying the infection in peripheral tissue, as has been suggested by the experimental studies in mice (3, 9, 22), before this becomes a widely used method of controlling scrapie. Research projects are also in progress at CVL to determine the effect of using genetically resistant rams on the incidence of clinical disease in affected flocks, and to discover whether genetically resistant animals from affected flocks are infected with the scrapie agent. CONCLUSIONS This review of the available data on the occurrence of scrapie suggests that scrapie is not an inherited condition but an infectious disease with a genetic influence on the incubation period. The route by which the infectious agent is transmitted is still unknown. While there is experimental evidence to suggest that placental tissues are a possible source of infection, it is impossible to be certain whether other tissues, discharges or excretions contain lower concentrations of the agent that may lead to accumulation on pasture. The data presented in Table IV show that the offspring of affected animals are definitely at increased risk of developing scrapie. This increase is thought to be largely the result of increased genetic susceptibility in these animals. The data also show that a considerable proportion of the cases occurring in these high-incidence flocks cannot be attributed to parental status and must, therefore, be the result of horizontal transmission of infection. Further epidemiological investigations of the independent effects of maternal infection and genetic susceptibility on the incidence of disease are being conducted, using recently discovered biological markers for genetic susceptibility. This would 845 allow the proportion of cases that could be attributed to maternal transmission of infection, and those that could be attributed to horizontal transmission, to be determined. Management factors which increase the likelihood of the infection being transmitted between animals, both from mother to offspring and horizontally, are also being investigated. The effect of using genetically resistant rams on the incidence of clinical disease in these flocks will also be determined, as will the possibility that genetically resistant animals are carrying the infection. These studies should help to clarify the means of natural transmission of scrapie and prompt new and more effective recommendations to aid in controlling the disease. ACKNOWLEDGEMENTS The author is very grateful to R.H. Kimberlin, M. Dawson and J.W. Wilesmith for their constructive comments during the preparation of this review. * * ÉTUDE SUR L'ÉPIDÉMIOLOGIE DE LA TREMBLANTE DU MOUTON. L.J. Hoinville. - Résumé : L'objet de cette étude est de résumer et d'évaluer les informations disponibles sur l'étiologie de la tremblante du mouton dans les troupeaux d'ovins atteints d'une infection naturelle, notamment celles concernant les possibilités de transmission entre animaux apparentés. L'auteur examine les résultats de plusieurs études majeures consacrées à la question au cours des trente dernières années. Les principales conclusions sont que la tremblante du mouton est une maladie infectieuse et qu'il existe une influence génétique sur la période d'incubation. Le risque accru de maladie chez les descendants d'animaux atteints s'expliquerait par une plus grande sensibilité génétique, les cas survenant dans les troupeaux à forte incidence étant en grande partie dus à une transmission horizontale de l'infection. MOTS-CLÉS : Epidémiologie - Génétique - Transmission - Tremblante du mouton. * # * REVISION DE LA EPIDEMIOLOGÍA DEL PRURIGO LUMBAR EN LA OVEJA. L.J. Hoinville. Resumen: El objetivo de esta revisión es el de sintetizar y evaluar los datos disponibles sobre la etiología del prurigo lumbar en rebaños de ovejas infectados de forma natural. Se presta una especial atención a los datos relativos a una posible transmisión entre animales emparentados. El autor examina datos procedentes de diversos estudios realizados sobre el particular durante los últimos treinta años. Las principales conclusiones establecen que el 846 prurigo lumbar es una enfermedad infecciosa, y que existe una influencia genética sobre el período de incubación. Se piensa que el mayor riesgo de enfermedad entre la descendencia de animales infectados obedece en gran medida a una mayor susceptibilidad genética; en rebaños con una elevada tasa de incidencia, una gran proporción de los casos son resultado de la transmisión horizontal de la infección. PALABRAS CLAVE: Epidemiología Transmisión. Genética - Prurigo lumbar - * * Appendix Calculation of population attributable risk percentages The population attributable risk percentages were calculated by combining the following three formulae from Henekens and Burring (45). 1. PAR = AR X 2. AR = I - I e P e 0 3. PAR% = PAR H- I T To give PAR% = ( [ I - I ] P ) ^ I e PAR: AR: P: I: I: I: PAR%: e e 0 x 0 e T Population attributable risk Attributable risk Proportion of cases in population exposed Incidence in exposed group Incidence in unexposed group Overall incidence in the population Population attributable risk percentage * * * 847 REFERENCES 1. BROTHERSTON J.G., RENWICK C.C., STAMP J.T., ZLOTNIK I. & PATTISON I.H. (1968). - Spread of scrapie by contact to goats and sheep. J. comp. Pathol, 78, 9-17. 2. BROWN P. & GAJDUSEK D.C. (1991). - Survival of scrapie virus after 3 years' interment. Lancet, 337, 269-270. 3. BRUCE M.E. (1985). - Agent replication dynamics in a long incubation period model of mouse scrapie. J. gen. Virol, 66, 2517-2522. 4. BRUERE A.N. (1977). - Scrapie: a point of view. N.Z. vet. J., 25, 259-260. 5. BULL L.B. & MURNANE D. (1958). - An outbreak of scrapie in British sheep imported into Victoria. Aust. vet. J., 34, 213-215. 6. CHANDLER R.L. (1961). - Encephalopathy in mice produced by inoculation with scrapie brain material. Lancet, 1, 1378-1379. 7. CHATELAIN J., BARON H., BAILLE V., BOURDONNAIS A., DELASNERIE-LUAPRETRE N. & CATHALA F. (1986). - Study of endemic scrapie in a flock of 'Ile de France' sheep. Eur. J. Epidemiol, 2, 31-35. 8. CHELLE PL. (1942). - Un cas de tremblante chez la chèvre. Bull Acad. vét. Fr., 15, 294-295. 9. COLLIS S.C. & KIMBERLIN R . H . (1985). - Long-term persistence of scrapie infection in mouse spleens in the absence of clinical disease. FEMS Microbiol. Letters, 29, 111-114. 10. CUILLE J. & CHELLE P L . (1936). - La maladie dite tremblante du mouton est-elle inoculable? C.R. Acad. Sci, Paris, 203, 1552-1554. 11. CUILLE J. & CHELLE PL. (1939). - Transmission expérimentale de la tremblante à la chèvre. C.R. Acad. Sci., Paris, 208, 1058-1060. 12. DAVIES D.C. & KIMBERLIN R.H. (1985). - Selection of Swaledale sheep of reduced susceptibility to experimental scrapie. Vet. Rec, 116, 211-214. 13. DETWILER L.A. (1992). - Scrapie. In Transmissible spongiform encephalopathies of animals (R. Bradley & D. Matthews, eds). Rev. sci. tech. Off. int. Epiz., 11 (2), 491-537. 14. DICKINSON A.G. (1974). - Scrapie. Nature, 252, 179-180. 15. DICKINSON A.G. (1976). - Scrapie in sheep and goats. In Slow virus diseases of animals and man (R.H. Kimberlin, ed.). North Holland Publishing Company, Amsterdam, 209241. 16. DICKINSON A.G., YOUNG G.B., STAMP J.T. & RENWICK C.C. (1965). - A note on the distribution of scrapie in sheep of different ages. Anim. Prod., 6, 375-377. 17. DICKINSON A.G., YOUNG G.B., STAMP J.T. & RENWICK C.C. (1965). - An analysis of natural scrapie in Suffolk sheep. Heredity, 20, 485-503. 18. DICKINSON A.G., YOUNG G . B . & RENWICK C.C. (1966). - Scrapie: experiments involving maternal transmission in sheep. In Report of scrapie seminar, Washington, D.C, 27-30 January 1964. Agricultural Research Service (ARS), United States Department of Agriculture (USDA), 244-247. 19. DICKINSON A.G., MEIKLE M.H. & FRASER H. (1968). - Identification of a gene which controls the incubation period of some strains of the scrapie agent in mice. /. comp. Pathol, 78, 293-299. 848 2 0 . DICKINSON A.G., STAMP J.T., RENWICK C.C. & RENNIE J.C. ( 1 9 6 8 ) . - Some factors controlling the incidence of scrapie in Cheviot sheep injected with a Cheviot-passaged scrapie agent. J. comp. Pathol, 78, 3 1 3 - 3 2 1 . 2 1 . DICKINSON A.G., STAMP J.T. & RENWICK C.C. ( 1 9 7 4 ) . - Maternal and lateral transmission of scrapie in sheep. J. comp. Pathol, 84, 19-26. 2 2 . DICKINSON A.G., FRASER H. & OUTRAM G.W. ( 1 9 7 5 ) . - Scrapie incubation time can exceed natural lifespan. Nature, 256, 7 3 2 - 7 3 3 . 2 3 . DICKINSON A.G. & FRASER H. ( 1 9 7 9 ) . - An assessment of the genetics of scrapie in sheep and mice. In Slow transmissible diseases of the nervous system, Vol. 1 (S.B. Prusiner & W.J. Hadlow, eds). Academic Press, New York, 3 6 7 - 3 8 5 . 2 4 . DICKINSON A.G. & OUTRAM G.W. ( 1 9 8 2 ) . - Genetic aspects of unconventional virus infections: the basis of the virino hypothesis. In Novel infectious agents and the central nervous system (G. Bock & J. Marsh, eds). Ciba Foundation Symposium 1 3 5 . John Wiley, Chichester, 6 3 - 8 3 . 2 5 . DRAPER G.J. ( 1 9 6 3 ) . - 'Epidemics' caused by a late-manifesting gene: application to scrapie. Heredity, 18, 1 6 5 - 1 7 1 . 2 6 . FITZSIMMONS W.M. & PATTISON IH. ( 1 9 6 8 ) . - Unsuccessful attempts to transmit scrapie by nematode parasites. Res. vet. Sci., 9, 2 8 1 - 2 8 3 . 2 7 . FOOTE W.C., CLARK W., MACIULIS A., CALL J.W., HOURRIGANJ., EVANS R.C., MARSHALL M.R. & D E CAMP M. ( 1 9 9 3 ) . - Prevention of scrapie transmission in sheep, using embryo transfer. Am. J. vet. Res., 54, 1 8 6 3 - 1 8 6 7 . 2 8 . FOSTER J.D. & DICKINSON A.G. ( 1 9 8 8 ) . - Genetic control of scrapie in Cheviot and Suffolk sheep. Vet. Rec, 123 (6), 1 5 9 . 2 9 . FOSTER J.D. & DICKINSON A.G. ( 1 9 8 8 ) . - The unusual properties of CH 1 6 4 1 , a sheep-passaged isolate of scrapie. Vet. Rec, 123, 5 - 8 . 3 0 . FOSTER J.D. & DICKINSON A.G. ( 1 9 8 9 ) . - Age at death from natural scrapie in a flock of Suffolk sheep. Vet. Rec, 125, 4 1 5 - 4 1 7 . 3 1 . FOSTER J.D. & HUNTER N. ( 1 9 9 1 ) . - Partial dominance of the sA allele of the Sip gene for controlling experimental scrapie. Vet. Rec, 128, 5 4 8 - 5 4 9 . 3 2 . FOSTER J.D., MCKELVEY W.A.C., MYLNE M.J.A., WILLIAMS A., HUNTER N., HOPE J. & FRASER H. (1992). - Studies on maternal transmission of scrapie in sheep by embryo transfer. Vet. Ree, 130, 3 4 1 - 3 4 3 . 3 3 . GOLDMANN W., HUNTER N., SMITH G., FOSTER J. & HOPE J. ( 1 9 9 4 ) . - PrP genotype and agent effects in scrapie: change in allelic interaction with different isolates of agent in sheep, a natural host of scrapie. J. gen. Virol, 75, 9 8 9 - 9 9 5 . 3 4 . GORDON W.S. ( 1 9 4 6 ) . - Louping-ill, tick-borne fever and scrapie. Vet. Rec, 58, 5 1 6 - 5 2 2 . 3 5 . GORDON W.S. ( 1 9 5 7 ) . - Scrapie. Vet. Ree, 69, 1 3 2 4 - 1 3 2 8 . 3 6 . GORDON W.S. ( 1 9 5 9 ) . - Scrapie Panel. Proc. U.S. Livestock Sanit. Assoc., 63, 2 8 6 - 2 9 4 . 37. GORDON W.S. ( 1 9 6 6 ) . - Review of work on scrapie at Compton, England, 1 9 5 2 - 1 9 6 4 . In Report of scrapie seminar, Washington, D.C., 2 7 - 3 0 January 1 9 6 4 . Agricultural Research Service (ARS), United States Department of Agriculture (USDA), 19-36. 3 8 . GORDON W.S. ( 1 9 6 6 ) . - Transmission of scrapie and evidence of spread of infection in sheep at pasture. In Report of scrapie seminar, Washington, D.C., 2 7 - 3 0 January 1 9 6 4 . Agricultural Research Service (ARS), United States Department of Agriculture (USDA), 8-13. 849 39. GORDON W . S . (1966). - Variation in susceptibility of sheep to scrapie and genetic implications. In Report of scrapie seminar, Washington, D.C., 27-30 January 1964. Agricultural Research Service (ARS), United States Department of Agriculture (USDA), 53-67. 40. GREIG J.R. (1940). - Scrapie: observations on the transmission of the disease by mediate contact. Vet. J., 6, 203-206. 41. GREIG J.R. (1950). - Scrapie in sheep. J. comp. Pathol, 60, 263-266. 42. HADLOW W.J. (1990). - An overview of scrapie in the United States. JAVMA, 196 (10), 1676-1677. 43. HADLOW W.J., KENNEDY R.C., RACE R.E. & EKLUND C . M . (1980). - Virologic and neurohistologic findings in dairy goats affected with natural scrapie. Vet. Pathol, 17, 187-199. 44. HADLOW W.J., KENNEDY R.C. & RACE R.E. (1982). - Natural infection of Suffolk sheep with scrapie virus. J. infect. Dis., 146, 657-664. 45. HENEKENS C.H. & BURRING J.E. (1987). - Epidemiology in medicine. Little, Brown & Co., Boston, 383 pp. 46. HOARE M., DAVIES D.C. & PATTISON I.H. (1977). - Experimental production of scrapie- resistant Swaledale sheep. Vet. Rec, 101, 482-484. 47. HOURRIGAN J.L. (1990). - Experimentally induced bovine spongiform encephalopathy in cattle in Mission, Tex, and the control of scrapie. JAVMA, 196, 1678-1679. 48. HOURRIGAN J., KLINGSPORN A., CLARK W.W. & D E CAMP M. (1979). - Epidemiology of scrapie in the United States. In Slow transmissible diseases of the nervous system, Vol. 1 (S.B. Prusiner & W.J. Hadlow, eds). Academic Press, New York, 331-356. 49. HUNTER N. (1991). - Natural transmission and genetic control of susceptibility of sheep to scrapie. Curr. Topics Microbiol Immunol, 172, 165-180. 50. HUNTER N. (1992). - Scrapie in sheep and goats. In Progress in sheep and goat research (A.W. Speedy, ed.). CAB International, Wallingford, Oxford, 131-151. 51. HUNTER N., FOSTER J.D., DICKINSON A.G. & HOPE J. (1989). - Linkage of the gene for the scrapie-associated fibril protein (PrP) to the Sip gene in Cheviot sheep. Vet. Rec, 124, 364-366. 52. HUNTER N., FOSTER J.D., BENSON G . & HOPE J. (1991). - Restriction fragment length polymorphisms of the scrapie-associated fibril protein (PrP) gene and their association with susceptibility to natural scrapie in British sheep. J. gen. Virol, 72, 1287-1292. 53. HUNTER N., FOSTER J.D. & HOPE J. (1992). - Natural scrapie in British sheep: breeds, ages and PrP gene polymorphisms. Vet. Rec, 130, 389-392. 54. HUNTER N., GOLDMANN W., BENSON G . , FOSTER J.D. & HOPE J. (1993). - Swaledale sheep affected by natural scrapie differ significantly in PrP genotype frequencies from healthy sheep and those selected for reduced incidence of scrapie. J. gen. Virol, 74, 1025-1031. 55. JOUBERT L., LAPRAS M., GASTELLU J., PRAVE M. & LAURENT D. (1972). - Un foyer de tremblante du mouton en Provence. Bull Soc. Sci. vét. Méd. comp. Lyon, 74, 165-184. 56. KATIYAR R.D. (1962). - A preliminary report on the occurrence of scrapie in Indian sheep. Ceylon vet. J., 10, 93-96. 57. KIMBERLIN R.H. (1979). - An assessment of genetical methods in the control of scrapie. Livestock Prod. Sci., 6, 233-242. 850 58. KIMBERLIN R . H . & WALKER C.A. (1978). - Pathogenesis of mouse scrapie: effect of route of inoculation on infectivity titres and dose-response curves. J. comp. Pathol, 88, 39-47. 59. KIMBERLIN R.H. & WALKER C.A. (1988). - Incubation periods in six models of intra-peritoneally injected scrapie depend mainly on the dynamics of agent replication within the nervous system and not the lymphoreticular system. J. gen. Virol, 69, 2953-2960. 60. KIMBERLIN R.H. & WALKER C.A. (1988). - Pathogenesis of experimental scrapie. In Novel infectious agents and the central nervous system ( G . Bock & J. Marsh, eds). Wiley, Chichester, 37-62. 61. KIMBERLIN R.H. & WALKER C.A. (1989). - Pathogenesis of scrapie in mice after intra-gastric infection. Virus Res., 12, 213-220. 62. LAPLANCHE J.L., CHATELAIN J., THOMAS S., DUSSAUCY M., BRUGERE-PICOUX J. & LAUNAY J.M. (1992). - PrP gene allelic variants and natural scrapie in French Ile-deFrance and Romanov sheep. In Prion diseases of humans and animals (S.B. Prusiner, J. Collinge, J. Powell & B. Anderton, eds). Ellis Horwood, Chichester, 329-337. 63. LAPLANCHE J., CHATELAIN J., BEAUDRY P., DUSSAUCY M. & BOUNNEAU J.L. (1993). - French autochthonous scrapied sheep without the 136Val PrP polymorphism. Mammalian Genome, 4, 463-464. 64. MCFADYEAN J. (1918). - Scrapie. J. comp. Pathol. Ther, 31, 102-131. 65. MACIULIS A., HUNTER N . , W A N G S., GOLDMANN W., HOPE J. & FOOTE W.C (1992). - Polymorphisms of a scrapie-associated fibril protein (PrP) gene and their association with susceptibility to experimentally induced scrapie in Cheviot sheep in the United States. Am. J. vet. Res., 53, 1957-1960. 66. MINISTRY OF AGRICULTURE, FISHERIES AND FOOD (MAFF) (1982). - Flock recording as an aid to the control of scrapie. Leaflet 823. Her Majesty's Stationery Office (HMSO), Edinburgh, 4 pp. 67. MORGAN K.L., NICHOLAS K . , GLOVER M.J. & HALL A.P. (1990). - A questionnaire survey of the prevalence of scrapie in sheep in Britain. Vet. Rec, 127, 373-376. 68. NUSSBAUM R.E., HENDERSON W.M., PATTISON L.H., ELCOCK N . V . & DAVIES D . C . (1975). - The establishment of sheep flocks of predictable susceptibility to experimental scrapie. Res. vet. Sci., 18, 49-58. 69. PALSSON P.A. (1979). - Rida (scrapie) in Iceland and its epidemiology. In Slow transmissible diseases of the nervous system, Vol. 1 (S.B. Prusiner & W.J. Hadlow, eds). Academic Press, New York, 357-366. 70. PARRY H.B. (1960). - Scrapie: a transmissible hereditary disease of sheep. Nature, 185, 441-443. 71. PARRY H.B. (1962). - Scrapie: a transmissible and hereditary disease of sheep. Heredity, 17, 75-105. 72. PARRY H.B. (1983). - Scrapie disease in sheep. Historical, clinical, epidemiological, pathological and practical aspects of the natural disease (D.R. Oppenheimer, ed.). Academic Press, London, 192 pp. 73. PATTISON I.H. (1964). - The spread of scrapie by contact between affected and healthy sheep, goats or mice. Vet. Rec, 76, 333-336. 74. PATTISON LH. & MILLSON G.C. (1960). - Further observations on the experimental production of scrapie in goats and sheep. J. comp. Pathol, 70, 182-193. 851 7 5 . PATTISON LH. & MILLSON G.C. ( 1 9 6 1 ) . - Experimental transmission of scrapie to goats and sheep by the oral route. J. comp. Pathol, 71, 1 7 1 - 1 7 6 . 76. PATTISON I.H. & MILLSON G.C. ( 1 9 6 1 ) . - Scrapie produced experimentally in goats with special reference to the clinical syndrome. J. comp. Pathol., 71, 1 0 1 - 1 1 0 . 77. PATTISON I.H. & MILLSON G.C. ( 1 9 6 2 ) . - Distribution of the scrapie agent in the tissues of experimentally inoculated goats. J. comp. Pathol, 72, 2 3 3 - 2 4 4 . 7 8 . PATTISON I.H., HOARE M.N., JEBBETT J.N. & WATSON W.A. ( 1 9 7 2 ) . - Spread of scrapie to sheep and goats by oral dosing with foetal membranes from scrapie-affected sheep. Vet. Rec, 90, 4 6 5 - 4 6 8 . 7 9 . PATTISON I.H., HOARE M.N., JEBBETT J.N. & WATSON W.A. ( 1 9 7 4 ) . - Further observations on the production of scrapie in sheep by oral dosing with foetal membranes from scrapie-affected sheep. Br. vet. J., 130, 6 5 - 6 7 . 80. SCHREUDER B.E.C., DEJONG M.C.M., PEKELDER J.J., VELLEMA P., BROKER A.J.M. & BETCKE H. ( 1 9 9 3 ) . - Prevalence and incidence of scrapie in the Netherlands: a questionnaire survey. Vet. Rec, 133, 2 1 1 - 2 1 4 . 8 1 . SCOTT J.R., FOSTER J.D. & FRASER H. ( 1 9 9 3 ) . - Conjunctival instillation of scrapie in mice can produce disease. Vet. Microbiol, 34, 3 0 5 - 3 0 9 . 82. SIGURDARSON S. ( 1 9 9 1 ) . - Epidemiology of scrapie in Iceland and experience with control measures. In Sub-acute spongiform encephalopathies (R. Bradley, M. Savey & B. Marchant, eds). Kluwer Academic Publishers, Dordrecht, 2 3 3 - 2 4 2 . 83. SlGURDSSON B. ( 1 9 5 4 ) . - RIDA, a chronic encephalitis of sheep (with general remarks on infections which develop slowly and some of their special characteristics). Br. vet. J., 110, 3 4 1 - 3 5 4 . 84. STAMP J.T. ( 1 9 6 2 ) . - Scrapie. Trans. Highlands agrie Soc. Scotland, 7, 1 9 - 3 6 . 85. STAMP J.T. ( 1 9 6 2 ) . - Scrapie: a transmissible disease of sheep. Vet. Ree, 74, 3 5 7 - 3 6 2 . 8 6 . STAMP J.T., BROTHERSTON J.G., ZLOTNIK I., MACKAY J.M.K. & SMITH W . ( 1 9 5 9 ) . - Further studies on scrapie. J. comp. Pathol, 69, 2 6 8 - 2 8 0 . 87. STEMSHORN B.W. ( 1 9 7 5 ) . - Histoire de cas - un cas de tremblante naturelle chez une chèvre. Can. vet. J., 16, 8 4 - 8 6 . 88. STOCKMAN S. ( 1 9 1 3 ) . - Scrapie: an obscure disease of sheep. J. comp. Pathol, 26, 317-327. 89. TAYLOR D.M. ( 1 9 8 9 ) . - Scrapie agent decontamination: implications for bovine spongiform encephalopathy. Vet. Ree, 24, 2 9 1 - 2 9 2 . 90. TOUMAZOS P. ( 1 9 9 1 ) . - Scrapie in Cyprus. Br. vet. J., 147, 1 4 7 - 1 5 4 . 9 1 . TOUMAZOS P. & ALLEY M.R. ( 1 9 8 9 ) . - Scrapie in goats in Cyprus. N.Z. vet. J., 37, 160-162. 92. WESTAWAY D. & PRUSINER S.B. ( 1 9 9 0 ) . - Link between scrapie and BSE? Nature (London), 346, 1 1 3 . 9 3 . WILSON D.R., ANDERSON R.D. & SMITH W . ( 1 9 5 0 ) . - Studies in scrapie. J. comp. Pathol, 60, 2 6 7 - 2 8 2 . 94. WRATHALL A.E. & BROWN K.F.D. ( 1 9 9 1 ) . - Embryo transfer, semen, scrapie and BSE. In Sub-acute spongiform encephalopathies (R. Bradley, M. Savey & B. Marchant, eds). Kluwer Academic Publishers, Dordrecht, 2 4 3 - 2 5 3 . 852 9 5 . YOUNG G.B., STAMP J.T., RENWICK C.C. & DICKINSON A.G. ( 1 9 6 6 ) . - Field observations of scrapie incidence. In Report of scrapie seminar, Washington, D.C., 2 7 - 3 0 January 1964. Agricultural Research Service (ARS), United States Department of Agriculture (USDA), 1 9 9 - 2 0 6 . 9 6 . ZLOTNIK I. & KATIYAR R.D. ( 1 9 6 1 ) . - The occurrence of scrapie disease in sheep of the remote Himalayan Foothills. Vet. Rec., 73, 5 4 3 - 5 4 5 . 9 7 . ZLOTNIK I. & RENNIE J.C. ( 1 9 6 3 ) . - Further observations on the experimental transmission of scrapie from sheep and goats to laboratory mice. J. comp. Pathol., 73, 150-162.