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Border disease

Border disease

P F NETTLETON

GENERAL INTRODUCTION: FLAVIVIRIDAE

Introduction

Border disease (BD) is a congenital virus disease of sheep and goats initially reported in 1959 from the border region of England and Wales and since recorded in other parts of Europe, North America, Australia, New Zealand, the Middle East and North Africa. The disease has also recently been confirmed by immunohistochemical demonstration of Border disease virus (BDV) in the brains of hairy-shaker lambs in South Africa.15 Some of these lambs also showed cerebellar hypoplasia. Border disease in sheep is characterized by barren ewes, abortion, stillbirths and the birth of small weak lambs, some of which show tremor, abnormal body conformation and hairy fleeces (’hairy-shaker’ or ‘fuzzy’-lambs) sometimes with excessive pigmentation. In some outbreaks no hairy-shaker lambs are born and the disease is difficult to distinguish from other causes of abortion. Occasionally the number of lambs born is apparently normal, and veterinary advice is sought only when a group of lambs fails to thrive and the number of scouring and dying lambs is abnormally high. Infection in goats is rare with abortion being the main presenting sign.

Aetiology

The cause of BD is a virus serologically related to bovine viral diarrhoea (BVD) virus and classical swine fever (CSF) virus, the three viruses being grouped in the genus Pestivirus within the family Flaviviridae. They are named after the diseases from which they were first isolated. Traditionally, pestiviruses isolated from pigs have been termed CSFV, those from cattle BVDV and those from sheep BDV. Cross-infection between these animal species occurs readily and the viruses are now grouped by their reactivity to monoclonal antibodies and their nucleotide sequences at selected genomic regions. Pestiviruses are enveloped, spherical particles approximately 50 nm in diameter in which the genome is a positive single-stranded RNA molecule. Virtually all pestivirus isolates from sheep and goats are noncytopathic in cell culture, but occasional cytopathic viruses have arisen following integration of cellular sequences into the viral genome.4

From studies with monoclonal antibodies and phylogenetic analysis of genomic sequences a consensus is emerging that there are four principal genotypes of pestiviruses: BVDV-1, BVDV-2, BDV and CSFV.7, 20, 24, 28 Other pestiviruses from wild ruminants do not fit into any of these groups and constitute separate virus types. A giraffe (Giraffa camelopardalis) isolate represents a single type, while pestiviruses isolated in 1996 from a reindeer (Rangifer tarandus) and a bison (Bison bonasus) kept in the same zoo form another separate type.5 While CSF viruses are predominantly restricted to pigs, examples of the other three principal genotypes have all been recovered from sheep. Examination of seven goat isolates has shown them to be BVDV-1 types.6, 23

Comparison of the available complete genomic sequences pestiviruses has revealed that the BD viruses are closer to CSF viruses than they are to BVD viruses.7, 24

There are relatively few reports of cross-neutralization experiments involving pestiviruses from sheep and goats. Nevertheless, four principal serological groups have been identified, which correlate with the monoclonal antibody and genotype groupings.21 A recent comparison of ten sheep pestiviruses from the UK showed two groups of serologically distinguishable BDV isolates, one group related to Moredun BDV and the other to the Weybridge and BVD viruses.19 These antigenic differences correlate well with genotyping results since all the Moredun BDV related isolates type as true BDVs whereas representatives of the other group all fit into the BVDV-1 genogroup. Genotyping studies on a total of 38 UK sheep isolates showed that 23 (60 per cent) were true BD viruses and ten (26 per cent) belonged to the BVDV-1 group. A further five isolates (13 per cent) belonged to the BVDV-2 group.28

It is known that representatives of BD viruses and BVD-1 viruses are serologically distinct and do not cross-protect,which implies that any BD vaccine should contain at least one representative from each of the BDV and the BVDV-1 groups.27 Cross-protection between BVDV-2 and BVDV-1 and BD viruses needs to be studied further. Recent foetal cross-protection studies in pregnant ewes have shown that foetuses of ewes experimentally infected with BVDV-1 were protected from challenge with BVDV-2, whereas foetuses of ewes experimentally infected with the BVDV-2 virus were infected after challenge with BVDV-1.22

Epidemiology

Sheep-to-sheep contact is the principal way in which BDV is transmitted and the most potent source of virus is the persistent excretor. Flocks with no previous experience of the disease are particularly vulnerable. Purchased, persistently infected, young ewes have been shown to introduce BDV into such flocks.8

Persistently infected lambs that reach maturity often have reduced fertility. Ewes that conceive either abort or produce persistently infected lambs sometimes over a period of years. Persistently infected rams usually have small soft testicles but can transmit virus in their semen as well as nasal/ocular secretions and saliva. The speed of virus spread in a susceptible flock exposed to one or more persistent excretors will depend on the contact between sheep. Under field conditions, intimate contact at mating time or gathering for any purpose will allow the virus to spread widely, whereas during normal grazing with no close contact virus spread will be slow. Any intensification of husbandry, particularly housing during early pregnancy, increases the risk of an explosive outbreak of BD. In flocks where the disease is endemic, the older ewes having shed the infection are immune and their lambs will be uninfected, and it is the progeny of primiparous ewes that are most commonly affected.

Pestiviruses from other domestic ruminants particularly cattle and pigs can also cause BD in sheep. The most serious threat comes from cattle10 since they are the principal hosts of pestiviruses; serological survey data from several countries indicate that about 70 per cent of mature cattle have antibody to BVDV whereas in the same regions the prevalence of pestivirus antibody in sheep is much lower, varying from about 5 to 50 per cent. Furthermore, cattle persistently infected with BVDV are ubiquitous; surveys in several countries have shown the prevalence of such cattle to be 0,4 to 0,9 per cent in randomly selected animals and between 1,7 and 10,5 per cent in herds experiencing disease.

Among free-living ruminants pestiviruses have been isolated from red (Cervus elaphus), roe (Capreolus capreolus) and fallow deer (Dama dama), African buffalo (Syncerus caffer), giraffe, wildebeest (Connochaetes spp.) and eland (Tregelaphus oryx), and serological surveys in Europe, North America and Africa have shown that many species have detectable antipestivirus antibodies.2, 12, 16, 25 Where sheep are grazed extensively in contact with free-living ruminants the possibility of infection cannot be excluded.

Pestiviruses are important contaminants of modified live virus vaccines. All such vaccines produced in ovine, bovine or porcine cell cultures, or in media supplemented with serum from these species, risk being contaminated with pestivirus. Sheep pox and orf virus vaccines administered to sheep, and an orf virus vaccine to goats, have been incriminated as vectors of BDV infection.17, 26

Pathogenesis, clinical signs and pathology

Healthy new-born and adult sheep exposed to pestivirus isolates from different countries experience only mild or inapparent disease. Slight fever and a mild leukopenia are associated with a short viraemia detectable between 4 and 11 days post-infection after which serum neutralizing antibody appears. Two isolates, however, have been shown to produce high fever, profound leukopenia, anorexia, respiratory signs, enteritis and death in 50 per cent of experimentally infected lambs. Necropsies revealed the presence of haemorrhagic enteritis and fibrinous pneumonia.11, 30

The most serious consequences occur when BDV infects susceptible pregnant ewes. Viraemia leads to an acute necrotizing placentitis detectable about ten days post-infection. The placentitis may be severe enough to contribute to early foetal death and abortion in some cases, but if the pregnancy is sustained, the lesions heal in about 25 days. Virus can cross to the foetus within one week of infection, and while the immune response of the ewe rapidly eliminates all virus from maternal tissues, it has no effect on that replicating in the foetus.

The ultimate outcome of foetal infection depends on several factors including the strain and dose of virus, the breed of foetus and its ability to repair damage. The most important factor, however, is the stage of foetal development at which infection occurs. The age at which the foetus gains immunological competence is critical in determining the distribution and persistence of virus, which, in turn, influences the extent of foetal damage. The ovine foetus can first respond to an antigenic stimulus between approximately 60 and 85 days of its 150-day gestation period. The possible fate of a foetus infected before or after this crucial period is summarized in Figure 87.1

If a foetus is infected before the onset of immune competence, viral replication is uncontrolled and its death is likely. In experimental infections, deaths of 50 to 75 per cent of foetuses occur depending on the virus strain used. In lambs that have survived infection in early gestation, virus is present in all organs. Such lambs appear to be tolerant to the virus and have a persistent infection usually for life. Typically, on pathological examination, they do not show any evidence of inflammatory reactions, but there are characteristic changes in the central nervous system (CNS) and skin.

Figure 87.1 Possible outcomes of Border disease virus infection before and after the foetus gains immunological competence between approximately 60 and 85 days’ gestation

In all parts of the CNS there is a deficiency of myelin which is associated with an increased density of interfascicular glia. The lesions are most obvious in new-born lambs with tremor, and the dysmyelination is thought to result from direct viral action on oligodendrocyte precursors leaving a deficit of mature myelin-forming cells at critical stages of CNS development,3 but it has also been attributed to depressed levels of circulating thyroid gland hormones.1 In older lambs, the myelin defect is less obvious and has usually resolved by the time the lamb is 20 weeks of age, but the density of interfascicular glia remains high and cells with swollen nuclei can persist for up to three years.

The fleece abnormalities of BD lambs result from an increased size of primary wool follicles and decreased numbers of secondary wool follicles in the skin. These result in a greater than normal proportion of large fibres, many of which are medullated, and a reduced number of fine fibres. Although gross fleece changes become less apparent as the affected lamb matures, serial skin biopsies have shown that the follicular alterations are permanent.

The effect of these changes is to produce an abnormally coarse and straight birthcoat, so-called hairy lambs. The effect is most obvious in fine-fleeced or medium-fleeced breeds, and is least obvious in coarse-fleeced breeds where changes may only be noticeable on the nape of the neck. In some field outbreaks the birthcoat changes can be difficult to assess because of breed and age variations in fleece type.3

If foetal infection occurs between approximately 60 and 85 days, the period when the immune system is developing, the outcome is unpredictable. Some lambs will be born antibody-positive and virus-negative while others will be viraemic and antibody-negative. Infections at this stage can produce a severe necrotizing and inflammatory reaction within the foetal CNS leading to an extensive destruction of the germinal layers of the brain. The ensuing lesions of cerebellar hypoplasia and dysplasia, hydranencephaly and porencephaly are responsible for severe nervous signs and locomotor disturbances which are manifested by affected lambs after birth. Some may show skeletal abnormalities such as arthrogryposis and excessively long legs. The severe destructive lesions in the CNS appear to be immune-mediated, and affected lambs frequently have high concentrations of serum antibody to BDV.

Foetal infection after approximately 85 days of gestation is met by an effective immune system. Foetal death is rare and virtually all lambs will be born apparently normal with demonstrable antibody against the virus. Nevertheless, microscopic lesions can be detected in them, principally in the CNS, which consist of a disseminated nodular periarteritis, suggestive of a cell-mediated allergic reaction, affecting small and medium-sized arterioles. These lesions can remain detectable for at least the first year of post-natal life.

The fate of lambs infected in early gestation and which are persistently infected is variable. Clinically affected lambs have a low chance of survival; many die early in life while survivors have a poor growth rate and an increased susceptibility to other diseases. Less severely affected lambs and apparently normal persistently infected lambs can survive for years. After BDV colostrum-derived antibody has waned, persistently infected lambs become viraemic with no, or low titres of BDV neutralizing antibody. They readily yield noncytopathic virus from blood and bodily secretions and appear to be immunotolerant to the infecting virus. Normal concentrations of serum protein and immunoglobulins, and the ability of the animals to produce antibody to other agents, confirm that persistently infected sheep can have a normal immune responsiveness in spite of their specific tolerance. Development of antipestivirus antibodies by some persistently infected sheep in later life shows that the immune tolerance may not be absolute.18

The mechanisms involved in the establishment and maintenance of persistent BDV infections are complex and much remains to be learned. An unstable equilibrium appears to be established between the host and the virus. Evidence for breakdown of this equilibrium has been seen in some sheep among groups of persistently infected sheep housed apart from all other animals. Spontaneous development of intractable scour, wasting, excessive ocular and nasal discharges sometimes accompanied by respiratory distress have been encountered in sheep aged between 2 and 21 months. At necropsy, such sheep have gross thickening of the mucosa of the distal ileum, caecum and colon resulting in focal hyperplastic enteropathy. Histologically there is ulceration and hyperplasia of the mucosa with foci of inflammation that penetrate the tunica muscularis and are accompanied by mononuclear cell infiltration and necrosis. Cytopathic BDV can be recovered from the gut of these lambs and, with no obvious outside source of cytopathic virus, it is most likely that such virus originates from the lamb’s own virus pool. Other persistently infected lambs in the groups did not develop the syndrome which has also been recognized in occasional field outbreaks of BD and has several similarities with bovine mucosal disease.18

Border disease also occurs in goats. There appears, however, to have been only one single report of the natural disease in them.11 Experimental infections of pregnant goats with BDV produces severe placentitis and abortion. Kids surviving to term manifest clinical disease similar to that of BD lambs and rarely survive.3

Diagnosis and differential diagnosis

The diagnosis of BD presents little difficulty in typical ‘hairy-shaker’ lambs. Nevertheless, laboratory confirmation is advisable since swayback caused by copper deficiency, daft lamb disease (a poorly defined, inherited, cerebellar degenerative disease), bacterial meningoencephalitis, polioencephalomalacia (thiamine deficiency), and other viral teratogens (see Diseases caused by Akabane and related Simbu-group viruses), will have to be considered in the differential diagnosis. In South Africa pregnant ewes that are allowed to graze maize residues contaminated with the fungus Diplodia maydis may produce stillborn or weak lambs with varying degrees of status spongiosus of the white matter.13 As ovine placentas and foetuses aborted due to BDV infection have no pathognomonic lesions, laboratory confirmation will be necessary to differentiate it from the other known causes of ovine abortion such as brucellosis, Q fever, toxoplasmosis, leptospirosis, chlamydiosis and salmonellosis.

In cases where teratology is involved investigations should include certain other virus infections.

Histological examination of the CNS can confirm BD but should be supported by the demonstration of viral antigen in tissues by specific immunohistochemical-staining or virus isolation/detection from blood and tissues. Aborted foetuses are often unrewarding for the demonstration of virus since they have usually died several days before expulsion. Whenever possible, recently dead or severely affected lambs should be submitted to the laboratory for diagnostic purposes.

Alternatively, the best specimens to submit should be taken in duplicate from the thyroid, kidneys, brain, spleen, gut and several lymph nodes. One set should be submitted fresh for antigen detection and the other placed in virus transport medium for virus isolation. As well as virus isolation, reverse transcription polymerase chain reaction (RT-PCR) can be used to detect viral RNA and to genotype the causative virus.29 For serology, a foetal heart blood sample and a blood sample from the dam should be collected and placed in sterile bottles without preservative or anticoagulants.

From live new-born ‘hairy-shaker’ or weak lambs confirmation of BD is based on the demonstration of virus in heparinized blood samples. The presence of maternally-derived colostral antibody can mask the presence of virus in the blood plasma of such lambs; therefore these samples should be collected before the lambs have suckled. To detect the antibody-negative, virus-positive, persistently infected sheep specimens of heparinized blood from all animals in a suspected group should be taken. Serological examination of individual sheep for BD antibodies is rarely helpful, but antibody testing of a ten per cent sample of different age groups of animals can be useful for demonstrating the presence and extent of BDV infection in a flock.

Control

The control of BD will depend on the level of infection in a flock. Sporadic outbreaks can be controlled by removing for slaughter, before the commencement of the following breeding season, the entire lamb crop and sheep suspected of introducing the disease. In endemically infected flocks susceptible animals retained for breeding should be deliberately exposed, while they are not pregnant, to known persistently- infected lambs. This is achieved by close herding them for at least three weeks, preferably indoors, to allow the BDV to spread effectively. The exposure period should terminate two months before the start of the breeding season.

On farms with no history of BDV, the introduction of new breeding stock needs careful consideration. Ideally, replacement females should be home-bred and the blood of all purchased rams should be tested for the presence of virus before the rams are brought onto the farm to ensure that they are not persistently infected. Where females are also purchased the feasibility of similarly testing them should be considered. Recently purchased females should always be mated and kept isolated from the rest of the flock until they have lambed. Because of the risk of infection of sheep from cattle it is essential that pregnant ewes are not mixed with cattle.

There is only one commercial vaccine available for the control of BDV but it is not licensed in all countries. It is a killed adjuvanted vaccine that contains representative strains of BDV and BVD-1 viruses, and is administered to young animals before they reach breeding age. It was reported to protect the progeny of all seven vaccinated ewes when the progeny of five of eight non-vaccinated control ewes were infected with virus following challenge of all 15 ewes between the 54th and 65th day of pregnancy.9 Further vaccine development is required; candidate vaccines should be tested for efficacy in pregnant sheep before being released for general use.

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