Lumbosacral Spinal Tap

Collection of CSF from the lumbosacral cistern is preferred for most large animal patients and is always done when the lesion is located in the spinal cord. Because the predominant flow of CSF is caudal, fluid collected from the cisterna magna reflects only changes within the brain and the most cranial parts of the spinal cord. However, fluid collected from the lumbosacral site is altered by either brain or spinal cord disease. Once a neuroanatomic diagnosis has been made, therefore, the site most suitable for collection can be chosen.

For collection of CSF by lumbosacral puncture, the animal is lightly sedated, and the skin of the dorsal midline over the junction of the sixth lumbar (L6) and first sacral (S1) vertebrae is surgically prepared (Fig. 35-1). A variety of standard sedative protocols are suitable, although xylazine has been shown to reduce CSF pressure.¹¹ Correct placement of the spinal needle is more easily achieved with the animal standing. The proper anatomic site for insertion of the spinal needle is between the dorsal spinous process of L6 cranially and S1 caudally and the two tuber sacrales laterally. The overlying skin forms a depression that can be recognized by palpation, although this may be difficult in well-muscled horses. The correct site for needle placement can also be located by determining (1) the dorsal midline at the “highest point” of the quarters or (2) the point where a line drawn between the caudal aspects of the two tuber coxae intersects the midline.

Fig. 35-1 Close-up view of landmarks for a lumbar cistern puncture in a horse. The large dark ovals (A) represent the position of the two tubers sacrale. The smaller curved lines (B) represent the respective caudal and cranial aspects of the L6 and S1 dorsal spinal processes. The dotted line represents the optimum area for placement of a skin stab.

The skin is anesthetized with 2% lidocaine, and a 1-cm incision is made with a No. 15 scalpel blade. A 6- to 9-inch, 18- to 20-gauge spinal needle is inserted perpendicularly through the incision and advanced until the tip punctures the lumbosacral cistern (a 3½-inch needle can be used in small ruminants, foals, and most cattle). A “snapping” sensation sometimes is felt as the needle passes through the interarcuate ligament. The patient may reflexly contract the tail, anus, and gluteal muscles. Some patients, particularly horses, may respond with violent motor activity. For this reason, a lumbosacral spinal tap performed on a conscious horse should be done only when the animal is restrained in stocks; people have been severely injured when this rule was not followed. The average depth of insertion is 17.64 cm (7 inches) in horses, 8.26 cm (3⅓ inches) in ponies,¹⁰ and about 7.5 cm (3 inches) in adult cattle.

The spinal needle is advanced gently to the floor of the spinal canal. Passage of the needle through the terminal spinal cord or the cauda equina does not cause subsequent neurologic abnormalities. When the needle is seated in the spinal canal, very gentle negative pressure can be applied by withdrawing spinal fluid into a series of 3-mL syringes. If frank blood is obtained, the tip of the needle probably is in one of the ventral vertebral sinuses. The needle should be withdrawn a few millimeters and a clean syringe attached to the hub. Compression of the jugular vein causes engorgement of the ventral vertebral plexus, which increases CSF pressure in the lumbosacral cistern. Failure of the CSF to flow from the needle after compression of the jugular vein could indicate incorrect needle placement or an obliterative lesion of the thoracolumbar spinal cord.

Cisterna Magna Tap

General anesthesia is required for a cisterna magna tap. After the patient has been anesthetized, the dorsal area of the neck overlying the atlantooccipital joint is surgically prepared. The patient’s head is held flexed at a right angle to the neck, with the sagittal plane of the head parallel to the floor or table on which the patient is lying. The head must not be allowed to move while the needle is inserted. The needle is inserted at 1 to 2 cm caudal to a point corresponding to the intersection of the dorsal midline and a line drawn between the cranial aspects of the wings of the atlas. This point usually is 6 to 9 cm (2½ to 3½ inches) from the poll (Fig. 35-2). A 3½-inch, 18-gauge spinal needle is inserted perpendicular to the skin and aimed toward the nose. The needle is advanced slowly with the stylet seated. After the needle has been advanced a few millimeters, the stylet is removed and the hub of the needle examined for CSF flow. The stylet is replaced if it is dry and CSF is not spontaneously dripping from the hub, and the needle is advanced another few millimeters and checked again for CSF flow. Entry of the tip of the needle into the cisterna magna may be accompanied by the sensation of “popping” through a tissue plane or by a sudden decrease in resistance to the advancement of the needle. In other cases, however, no such sensation is perceived, thus the precaution of checking for CSF flow every time the needle is advanced a few millimeters.

Fig. 35-2 View of the ideal area for a cisterna magna cerebrospinal fluid tap in a cow. The two lines (a) represent the wings of the atlas. The large spot in the center (b) is the optimum area for needle insertion. It is essential to prevent head movement by use of anesthesia and restraint.

In most large animals the needle is seated at approximately 5 to 8.75 cm (2 to3½ inches). While advancing the needle, the heel of the hand should be held firmly against the animal’s neck to minimize the possibility of spinal cord injury. The mean depth of insertion is 6.16 cm (2½ inches) in horses¹⁰ and 5.08 cm (2 3½ inches) in cattle. In a cisterna magna tap, the needle entry site is close to the cervical spinal cord and the brainstem. To minimize the danger of central nervous system (CNS) damage during a cisterna magna tap, the animal should be adequately anesthetized and ventilated, because an increased partial pressure of carbon dioxide (PCO₂) results in elevated intracranial pressure.¹² Removal of fluid from the cisterna magna is contraindicated in a patient with increased intracranial pressure because a possibly fatal herniation of the brain through the foramen magnum and under the tentorium cerebelli may occur. Signs of increased CNS pressure include a moderate to marked decrease in mentation, mydriatic pupils, opisthotonos, extensor rigidity, ventrolateral strabismus, and papilledema.

Ultrasonography can be used as an aid to needle placement for collection of CSF from both the atlantooccipital cistern and the lumbosacral subarachnoid space.^13,14 In the authors’ experience, however, CSF collection can usually be effected without such assistance.

Myelography

Contrast material for myelography is injected into the cisterna magna in large animal patients. Withdrawal of CSF before injecting the contrast medium is unnecessary, and reinjection of CSF after it has been withdrawn is inadvisable. The turnover of CSF is rapid and under strict homeostatic control; therefore, withdrawal of CSF through a spinal tap does not have deleterious effects that require its replacement.^15-17 In horses undergoing myelography, two spinal needles can be placed, one at the lumbosacral space and one into the cisterna magna. As the contrast medium is injected through the needle in the cisterna magna, CSF is allowed to drain freely from the lumbar needle. This technique facilitates injection of large volumes of contrast material but is not absolutely necessary to obtain a good-quality myelogram.

Definition and Etiology

Maedi-visna virus, or ovine progressive pneumonia virus (OPPV), infection is a chronic disease of sheep caused by a retrovirus (subfamily Lentivirinae).^38-49 Visna virus is closely related to the many lentiviruses that cause immunodeficiency and neurologic disease in other species; these viruses include equine arteritis virus, caprine arthritis-encephalitis virus, and simian, feline, and human immunodeficiency viruses.^50,51 Caprine and ovine lentiviruses constitute a single virus group, termed the small ruminant lentiviruses (SRLVs), which can readily cross between species.^52-54 These agents are enveloped ribonucleic acid (RNA) viruses that contain reverse transcriptase.^55,56 The respiratory aspects of the viral infection are discussed in Chapter 31.

Clinical Signs

Neurologic disease caused by maedi-visna virus is relatively rare.⁵¹ Nervous system signs may be characteristic of a diffuse encephalitis and include ataxia, twitching of the facial muscles, conscious proprioceptive deficits, normal gait along a straight path, staggering or stumbling when turned or forced to perform a complex maneuver, circling, and blindness. Coma, convulsions, and hyperexcitability may be seen in terminal stages. Other sheep have gradually progressive limb weakness and ataxia, often worse in the rear limbs; myelitis may be the sole clinical manifestation of maedi-visna.^56,57 Some sheep merely show emaciation without neurologic signs. The time between the onset of clinical signs and death may be as long as 1 to 2 years. Because of the slowly progressive nature of OPPV infection and the high probability that affected sheep eventually will develop chronic disorders of the nervous, musculoskeletal, mammary gland, or respiratory system, the presence of antibody in the serum usually is considered evidence of active infection.^58,59

Clinical Pathology

The CSF of affected animals is characterized by pleocytosis, with the cell counts per microliter ranging from 1012 to 1478 for animals infected for 1 month and 4 years, respectively.⁶⁰ The protein concentrations of CSF range from 50 to 100 mg/dL for 30 days after infection. Antiviral antibody and virus can be detected in CSF specimens. The CSF/plasma ratio of immunoglobulin G (IgG) in the CNS is normal (<0.2) during the first month after infection but rises to over 0.4 after 1 month.⁶¹

Pathophysiology

The lesions of maedi-visna virus infection are partly induced by the host’s inflammatory response. Experimental immunosuppression of infected sheep ameliorates the severity of the clinical signs and reduces the pathologic lesions of the cerebral cortex without altering the amount of viral shedding.^62,63 The virus is immunosuppressive. Viral infections usually lead to a variety of secondary bacterial infections.⁶⁴ Spread of OPPV within the tissues and secretion of the virus may be facilitated by concurrent diseases, such as Brucella ovis.⁶⁵ The chronic viremia may be caused by repeated antigenic changes of the virus; by intermittent expression of proteins by persistent proviral deoxyribonucleic acid (DNA) in cells; or by protection of the virus in circulating immune cells. Natural resistance to maedi-visna virus infection may be mediated partly by a nonimmunoglobulin inhibitory substance that is present in ovine plasma.⁶⁴

Pathology

Gross lesions of visna infection are seen only rarely, when inflammation and malacia are extensive. In such cases, areas of yellowish tan discoloration are present in white matter.⁶⁶ The microscopic lesions of visna are predominantly those of a diffuse, nonsuppurative, perivascular inflammation throughout the neuraxis, affecting white matter in particular, although gray matter also is involved. The lesions include demyelination, gliosis, lymphocytic choriomeningitis, round cell infiltration of the choroid plexus, and focal necrotic areas that are infiltrated by macrophages.^67,68 Inflammatory lesions are predisposed to develop in a periventricular location, including around the central canal of the spinal cord.

After exposure to the virus, sheep develop an asymptomatic infection for as long as 6 weeks. During this time the virus can be isolated from the brain and other tissues; later the virus can be isolated from peripheral blood neutrophils but not from tissue homogenates, indicating that replication of the virus occurs in circulating cells during this stage of the infection.⁶⁹ Once the disease has been recognized, the affected sheep should be culled or slaughtered. Rigid control measures that may be partly successful⁴⁶ are discussed in Chapter 31.

Diagnosis and Epidemiology

Spread of maedi-visna virus occurs through ingestion of infected milk by neonates, through in utero transmission, and horizontally within a herd. Recent data suggest that the role of colostrums and milk in the spread of disease may be less important than previously thought, and that seropositivity in young lambs may be caused by colostral antibodies rather than true seroconversion by the lambs.^48,70 Low prevalence of the virus within herds managed extensively rather than intensively further supports the belief that horizontal transmission is the most important mode of infection.⁷¹

The diagnosis of visna infection is made initially by recognition of the clinical neurologic disease in groups of animals in which the pulmonary form of the disease also is present. Agar gel immunodiffusion (AGID) testing has been superceded by more accurate enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR) tests.^51,72-75 The diagnosis and epidemiology of maedi-visna virus infection are discussed in detail in Chapter 31.

Treatment and Prevention

No effective treatment for maedi-visna virus infection is available for field use, although the disease has been used as an animal model system for the study of drugs effective against lentiviruses.^76,77 Prevention is based on herd hygiene and culling of affected animals. Recently, a novel vaccine strategy using a plasmid encoding for viral envelope glycoproteins has shown some promise, although routine clinical use of such vaccines is probably many years away.⁷⁸

Definition and Clinical Signs

The caprine arthritis-encephalitis (CAE) virus belongs to the retrovirus group and is closely related to the virus that causes maedi-visna and ovine progressive pneumonia in sheep (“small ruminant lentiviruses”).⁷⁹ The systemic manifestations of the disease are thoroughly discussed in Chapter 38. The leukoencephalomyelitis form of CAE is predominantly seen in young goats but may occur in goats as old as 22 years.⁸⁰ The clinical signs of leukoencephalomyelitis include ataxia, paraparesis, paraplegia, tetraparesis, tetraplegia, hemiparesis, hemiplegia, head tilt, nystagmus, tremors, torticollis, trismus, salivation, depression, coma, and opisthotonos.^81-87 The neurologic deficits may be either symmetric or asymmetric. Goats with high cervical spinal cord lesions (L1 to L4) are recumbent and unable to raise their heads from the floor. They may show resistance to passive neck flexion. Vision and pupillary light reflexes may be diminished.^83,85 The specific gait disturbances depend on the areas of the spinal cord involved. Signs of neurologic dysfunction may range from paraparesis to tetraplegia. The spinal reflexes range from hypertonia and hyperreflexia to hypotonia and hyporeflexia.^81-86 This diversity of signs is related to the variable location of lesions in the central nervous system (CNS).

Other clinical signs also are variable. In one study, affected goat kids remained afebrile, whereas in another study, 61% of affected animals had rectal temperatures ranging from 38.9° C to 41.3°C (103.6° F to 106.4°F).^81-86 Other signs that could be associated with systemic viral infection include enlarged joints, vague and shifting leg lameness, weight loss, and tachypnea without significant auscultatory abnormalities.

The major differential diagnostic considerations for the neurologic form of CAE include listeriosis or chlamydial and mycoplasmal infections. The CAE virus causes neurologic lesions in numerous regions of the CNS, whereas the lesions of listeriosis are generally restricted to the brainstem. Mycoplasmal infections typically affect kids ranging from 1 to 6 months of age, and affected animals develop polyserositis. They are systemically ill, and the joints are hot, grossly swollen, and painful. Goats with a mycoplasmal infection show extreme pain when the neck is passively flexed because of meningeal and vertebral articular inflammation. Fluid from the body cavities of goats with mycoplasmal infection has a high protein concentration and an increased number of polymorphonuclear cells.

Diagnosis

Previously, an AGID test was used for the diagnosis of CAE, but this has been superseded by ELISA and PCR tests.^88,89 The virus also may be cultured from infected tissue.⁹⁰ One study has reported an increase in total plasma protein and hypergammaglobulinemia in affected goats.⁸² The results of other hematologic and blood chemistry analyses are normal. The changes in the CSF are characteristic only of a chronic granulomatous inflammation. Specific changes include an increased protein concentration and pleocytosis.⁹¹ Cell counts in the CSF of affected goats range from 5 to 1800/dL, and the protein concentration ranges from 0 to 700 mg/dL.⁸⁰

Pathogenesis

Being a member of the retrovirus group, the CAE virus contains RNA-dependent DNA polymerase. When inoculated into goats, the virus causes a chronic infection characterized by demyelinating encephalomyelitis, arthritis, and interstitial pneumonia. The pathologic changes resemble those of an autoimmune process and are probably caused by interactions between the host’s immunologic responses, denatured myelin, and the virus.⁹² Macroscopic pathologic changes in the CNS of naturally infected goats include cloudiness of the meninges and tan discoloration of the white matter.⁸¹ Microscopic changes include disseminated perivascular accumulations of mononuclear cells, demyelination originating in the subependymal region, astrocytosis, and mononuclear leptomeningitis.^{81,85,88,92,93} The inflammatory foci are predominantly composed of macrophages containing material that tests positive on periodic acid—Schiff (PAS) staining. Neuronophagia and neuronal necrosis are not seen.⁹³ Lesions are most severe in the periductular, periventricular, and submeningeal regions of the white matter. Spinal cord lesions are most frequently observed in the thoracolumbar segments.⁸⁰

Treatment

No treatment is available for goats with leukoencephalomyelitis. Control measures are discussed in Chapter 38.

Definition and Etiology

Border disease is a congenital infection of sheep and goats caused by a noncytopathic togavirus (genus Pestivirus).⁹⁴ The border disease agent is antigenically similar to the bovine viral diarrhea (BVD) and hog cholera viruses. Once introduced into a flock, the border disease virus causes a devastating syndrome characterized by abortion, infertility, and deformed lambs. The virus infects naive ewes during pregnancy and causes a variety of fetal malformations, including early embryonic death, abortion and stillbirth, and small, malformed lambs. A seroprevalence rate as high as 29% has been seen in a newly infected flock; however, lambs infected in utero become immunotolerant and remain viremic.⁹⁵ In an infected flock the incidence of fetal malformations and abortions declines over time because of the growing population of ewes with persistent active immunity. Adult sheep that become infected with the border disease virus develop an inapparent, short-lived viremia and become immune to reinfection.

Clinical Signs

The severity of clinical signs in affected lambs varies. Changes are most marked in newborn lambs infected in early gestation (before 50 days). The central nervous system (CNS), skin, and skeleton are the most seriously affected. The hairs of congenitally affected lambs are coarse, straight, and elongated and stand out from the body like a halo.⁹⁶ The coat is abnormally pigmented and may have a dark-gray appearance or hyperpigmented spots that are especially prominent over the top of the neck.⁹⁷ The combination of pigmentary abnormalities and long, coarse hair shafts gave rise to the descriptive term “steel wool coat.” Animals that survive shed the abnormal hairs at 9 to 12 weeks of age and replace them with normal hair fibers. Affected lambs also have a short, thickened body, shortened legs, smaller orbital size, and doming of the frontal bone.^98,99 Arthrogryposis occasionally may be seen. Some infected lambs show neurologic symptoms such as ataxia and uncontrollable tremors. The tremors are coarse, involve the trunk and head, and disappear when the animal is asleep.^100,101 The lambs often are alert and appetent but initially need assistance to stand and nurse. Some animals walk normally but hop on the rear limbs when forced to run. Over time the lambs become stronger but continue to show impaired locomotion for months. The CNS signs usually disappear by 20 weeks of age, but the animals appear stunted. Affected animals have greatly decreased viability compared with uninfected herdmates and may die suddenly without showing premonitory symptoms. Aside from neonatal death losses, economic burdens imposed by the viral infection include low birth weights, diminished weaning weights, lowered carcass quality, and infertility.^99,102

Abortions that occur 9 to 106 days after inoculation are seen in 30% of experimentally infected sheep.^97,98,103 In field outbreaks the average gestational age of the aborted fetus is 63 days. Teratogenic effects are most often observed when lambs are infected at 50 to 90 days of gestation.¹⁰⁴ Fetal mummification occasionally may be seen.

The border disease virus is also pathogenic in goats.¹⁰⁵ As in sheep, inoculation of pregnant does results in fetal mummification and abortion. The spinal cords of infected kids are hypomyelinated, but the characteristic hair changes usually observed in sheep fetuses are not seen in goat kids.

The border disease virus has low pathogenicity for cattle. Abortions can be induced in cows inoculated with the virus at approximately 50 days of gestation, and affected calves have cerebral cavitations.¹⁰⁶ The condition is not recognized as naturally transmitted to cows.

Diagnosis

Identification of the viral antigens in tissues using fluorescent antibody tests is the most accurate method of diagnosing border disease. Tissues that most consistently contain viral antigens are those of the abomasum, pancreas, kidneys, thyroid, and testicles.^107,108 Serodiagnostic methods, including serum neutralization (SN), AGID, and complement fixation tests, have been developed.¹⁰⁹ In most cases the BVD virus has been used as the indicator antigen. Serodiagnosis of infected lambs is difficult because the lambs tend to be immunotolerant and therefore do not develop strong serologic responses. Sheep infected as adults develop SN titers ranging from 1:20 to 1:320, whereas the SN titers of animals with congenital infections are consistently below 1:10.¹⁰³ The presence of viral antibodies in the CSF suggests border disease virus infection.

Pathophysiology

Hypomyelination probably is caused by a combination of virus-induced degeneration of the oligodendroglial cells (dysmyelinogenesis), persistent viral infection, and diminished secretion of the thyroid hormones L-3,3′,5′-triiodothyronine and thyroxine.¹¹⁰ A deficiency of these hormones probably results in a lowered concentration of 2′,3′-cyclic nucleotide-3′-phosphodiesterase, which contributes to the hypomyelination. The diminished production of thyroid hormones is thought to be related to direct inhibition of the thyroid gland, because the pituitary activity of these lambs appears to be normal. Hypomyelination appears at approximately the same time as specific antiviral delayed-type hypersensitivity, indicating that an immunopathologic event may be partly responsible. Morphometric measurements of the spinal cords of lambs infected in utero show a permanent reduction of both white and gray matter in a cross-sectional area as a result of the decreased myelin content.¹¹¹ Depressed blastogenic activity of lymphocytes, a decrease in T4 helper cell function, and an increase in T4 suppressor cell function have been demonstrated in affected lambs between 4 and 7 months of age, indicating a viral immunosuppression.^112-114 Such immunosuppressed lambs succumb to parasitism, diarrhea, and bronchopneumonia.

Epidemiology

Border disease is transmitted both vertically and horizontally.^112,115 The agent can efficiently infect sheep through the intact mucous membranes. The major reservoir in infected herds is the asymptomatic, congenitally infected seronegative animal.^95,116 Sheep exposed to the virus as adults develop antibody responses and are able to clear the infections within weeks. Asymptomatically infected animals may shed the virus through the placenta, infected offspring, saliva, respiratory secretions, urine, or feces.^103,112 In one seroepidemiologic study of infected ewes in the western United States, lambs were more often seropositive than ewes.⁹⁵ A large percentage of the seropositive lambs were born to seronegative ewes, indicating the presence of a large amount of virus cycling from asymptomatic carriers. Strain-related differences appear to exist in viral pathogenicity.¹¹⁷

Necropsy Findings

The macroscopic changes associated with border disease virus infection are hydranencephaly, porencephaly, microcephaly, cerebellar hypoplasia, abnormal curvature of the ribs, brachygnathia, doming of the frontal bones of the skull, narrowing of the distance between the orbits, a decrease in orbital size, shortening of the crown-to-rump and diaphyseal lengths, retention of secondary hair fibers, and abnormal skin pigmentation.

Microscopic changes in lambs with congenital infection include hypomyelinogenesis and hypercellularity of the white matter with abnormal-appearing glial cells.¹¹⁸ The CNS shows dysmyelinogenesis, secondary demyelination, and a nodular periarteritis. Viral antigen can be demonstrated in the adventitia of the CNS arterioles.¹¹⁰ The microscopic lesions of the placenta include endothelial swelling, thrombotic occlusion of the vessels, and fibrinonecrotic cellular debris in the fetomaternal space.¹¹⁸

Treatment and Control

In herd situations, blood cultures and examination of skin biopsies by fluorescent antibody tests should be performed concurrently to identify carriers.¹⁰⁶ The SN test is not a reliable indicator of infection. Noninfected pregnant sheep should be kept separated from others in the flock for the first 60 days of gestation to ensure that in utero infections do not occur.

Clinical Signs

Two herpesviruses, bovine herpesvirus type 1 (BoHV-1) and type 5 (BoHV-5), have been associated with encephalitis in cattle.¹¹⁹ BoHV-1 infection typically results in an acute upper respiratory tract disease characterized by fibrinonecrotic white plaques of the nasal, pharyngeal, and tracheal mucosa and abortions (see Chapter 31). Other clinical conditions associated with infectious bovine rhinotracheitis (IBR) infection include epizootic conjunctivitis (Chapter 39) and infectious balanoposthitis or vulvovaginitis (Chapter 43). Infection with BoHV-5, in contrast, usually results in meningoencephalitis with few or no respiratory signs. BoHV-1 infections also can result in meningoencephalitis, but much less frequently. The clinical signs of this encephalitis include depression, mild nasal and ocular discharge, conscious proprioceptive deficits, head pressing, aimless circling, bellowing, salivation, bruxism, paralysis of the tongue, head tilt, nystagmus, convulsions, blindness, coma, and death.^120,121 The seizure activity is characterized by a tonic-clonic convulsion with violent spasms or tremors of the head, with all four legs flexed and the head in opisthotonos.^120,121 Rectal temperatures of 41° C to 42°C (106° F and 107°F) have been reported.¹²² The case-fatality rate of encephalitic IBR is almost 100%; however, recovery occurs in rare case. In experimental infection, signs develop 1 to 2 weeks after infection.

Differential diagnoses for encephalitic herpesvirus infections in cattle include almost all encephalitic, encephalopathic, and neurotoxic diseases of cattle, including rabies, polioencephalomalacia, salt poisoning, and lead toxicity. Accompanying respiratory disease raises the index of suspicion for herpesvirus. A rising serum neutralizing antibody titer in surviving animals can be used to confirm infection, although this does not distinguish between BoHV-1 and BoHV-5. Specific diagnosis is usually made at necropsy using a number of modalities, including virus isolation, immunohistochemistry, and PCR.^123-125 Many of these tests do not distinguish between BoHV-1 and BoHV-5; more specific PCR and restriction fragment analysis techniques can differentiate between these two virus types.¹²⁶

Epidemiology

Bovine herpesviruses cause disease in cattle worldwide, although the encephalitic form of infection is relatively rare in the United States. Epidemiologic factors that appear to favor dissemination of herpesviruses among cattle include a high stocking rate, repeated introduction of animals from diverse backgrounds, and mass weaning of calves at a time when the passively acquired anti-IBR antibodies are waning.¹²⁰ BoHV-1 may survive in the environment for up to a month; cooler ambient temperatures and higher humidity promote virus survival.¹²⁷ Calves less than 6 weeks old are most susceptible, but infection and resultant neurologic disease also have been described in adult cattle.^{121,122,128-134} Animals that survive the disease become persistently infected; virus survives in the nasal and tracheal muscosa and the trigeminal ganglion. Reactivation and virus shedding may occur during periods of stress.^127-129132 Although clinical disease caused by reactivation of latent virus is usually mild and may go unnoticed, shedding of virus during such episodes provides a source for infection of in-contact animals. The viruses grow in the nasal and pharyngeal mucosa. Brain involvement results from centripetal spread along the sensory neurons of the trigeminal and olfactory nerves.¹³² Hematogenous dissemination to the brain is believed to be relatively unimportant.

Pathophysiology

Encephalitic IBR infections cause a nonsuppurative meningoencephalitis that is widely distributed in the gray matter of the brain. Pathologic abnormalities affect predominantly the forebrain, although all areas of the brain can be involved. Findings include marked perivascular cuffing with mononuclear cells, diffuse gliosis, neuronal degeneration and chromatolysis, hemorrhage, edema, necrosis, and neuronophagia. White matter lesions include myelitis with mononuclear cell infiltration and demyelination. Extensive lymphocytic meningoencephalitis is seen. Intranuclear inclusion bodies are rarely seen in bovine herpesvirus encephalitis. The virus usually can be isolated from brain homogenates of affected calves.

Treatment and Control

No adequate therapy exists for the encephalitogenic form of BHV-1 infection. Treatment is symptomatic and supportive and should include oral or intravenous fluid support, nonsteroidal antiinflammatory drugs (NSAIDs), antibiotics in animals with respiratory signs, and nursing care. Diazepam or phenobarbital may be used for seizure control when necessary (Table 35-3). Animals with severe clinical signs may be euthanized for humane reasons.

Table 35-3 Recommended Drug Dosages for Treatment of Cerebrocortical Disease

Vaccines that protect against BoHV-1 also may prevent BoHV-5 infection because of the close antigenic relationship between the two viruses.¹²⁸ Modified live intranasal vaccines may be most effective in preventing clinical disease and also can reduce shedding of virus by infected animals. In some European countries, eradication programs that employ testing and culling of infected animals have been successful in eliminating BoHV-1. Bovine herpesviruses can be spread through fomites and by aerosol transmission for up to 4 miles (6.4 km).¹²⁷ Strict biosecurity procedures are therefore essential to developing and maintaining virus-free herds.

Definition and Etiology

Bovine spongiform encephalopathy (BSE) is a transmissible spongiform encephalopathy (TSE) of cattle that was first described in 1987 in Great Britain (GB).¹³⁵ The TSEs are a group of slowly progressing, invariably fatal neurodegenerative diseases that can affect humans and animals. They are also called “prion diseases” because of the accumulation of prion in the central nervous system (CNS). Since the “prion hypothesis” was first formulated in 1982 by Stanley Prusiner, prions have been widely accepted as the etiologic infectious agents of the TSEs.¹³⁶ The prion is an isoform (PrPSc) of the normal prion protein (PrPc), a host-encoded membrane protein that does not carry any nucleic acid. PrPSc replicates by inducing a posttranslational conformational change in the normal protein PrPc to form the abnormal prion PrPSc.^136-138

Clinical Signs

Video clips of various clinical signs of BSE are available on the Internet.¹³⁹ As with all TSEs, BSE has a long incubation phase. It occurs mostly in 4- to 6-year-old cattle (age range, 20 months to 18 years). The clinical signs of BSE are usually insidious in onset.^140,141 They may be precipitated by stressful situations such as transportation, concurrent illnesses, or states of increased metabolic consumption, such as late pregnancy and parturition.

The detection of clinical signs of BSE and the array of clinical signs reported are influenced by how closely the cattle are being observed and the handlers’ awareness of the insidious and nonspecific nature of the clinical signs. Subtle early behavioral changes are unlikely to be observed in large herds. Cattle may become apprehensive, stay apart from their herdmates, and become fearful of their handlers. As the disease progresses, cattle often become extremely excitable, especially when restrained or placed in confined quarters or an unfamiliar environment. BSE cattle may refuse to move through previously familiar doorways and become impossible to restrain. Unprovoked aggression is less frequently seen. Hyperesthesia is most marked around the head and neck. A light touch on the hindlimbs may induce violent, “ballistic,” and often repetitive kicking, which may also be observed during milking. Animals may overreact, startle, or “panic” in response to sudden visual, tactile, or auditory stimuli. The startle reaction may be gauged by a hand test (punching fist abruptly toward the animal’s head without touching it), flashlight test, a hand clap, or a loud metallic “bang.” Normal animals rarely startle in response to such stimuli, or they may startle once or twice when the stimulus is repeated. BSE cases will frequently startle in a violent and repeatable fashion, with no habituation to a repeated stimulus. In some cases the response may escalate to an extreme “aberrant” startle, with head shaking and seizure-like head bobbing, which may be followed by violent turning, running around, and falling.¹³⁹

Other possible clinical signs of BSE include ptyalism, increased head rubbing, muscle fasciculations, and excessive vocalization. Other behaviors seen in BSE may include frequent and repetitive head tossing, licking of the nostrils, yawning, flehmen, head tossing and head butting, and restlessness.^139,142 A relative bradycardia (≤60 beats/min) may be observed in animals when a tachycardia is expected because of their hyperexcited state. Affected animals often lose weight and have reduced milk output. Ataxia and tremors are seen in more advanced cases. Tremors most often involve the head and neck and may become generalized, especially with exercise. Ataxia is most marked on turns, when going over steps, or on uneven terrain; it may also be accentuated with exercise. BSE animals may become recumbent and unable to feed if the disease is left to run its course. Recumbency may also be triggered by an intercurrent illness or an injury during a fall. The clinical signs of BSE may be more difficult to recognize in animals that have become recumbent than in less advanced cases,¹⁴³ and systematic targeted surveys of “fallen stock” have allowed for the detection of previously unsuspected cases of BSE in various countries.¹⁴⁴

Pathophysiology

The exact function of the prion protein is unknown, as are the mechanisms of neurodegeneration and CNS dysfunction in TSEs.

The presence of BSE infectivity in extraneural tissues has not been identified in natural BSE cases. In pathogenesis studies of cattle orally inoculated with BSE, infectivity was detected during the incubation period in the distal ileum from 6 months after challenge and in the CNS from 32 months after challenge (clinical signs first observed at 36 months).¹⁴⁵ Similar studies in sheep revealed infectivity within Peyer’s patches by 4 months after experimental inoculation and in a wide range of tissues, including the CNS, by 16 months after inoculation.¹⁴⁶ PrPSc was detected in lymphoid tissue (Peyer’s patches) of the distal ileum and in the bone marrow during the clinical phase of the illness.¹⁴⁷ Mice bioassays were negative for infectivity of milk, placenta, tonsils, and lymph nodes from BSE-affected and incubating animals. However, the more sensitive cattle bioassay showed that tonsils do carry small amounts of infectivity.¹⁴⁸ The full results of cattle bioassay studies have not yet been published.

Epidemiology and Zoonotic Potential

BSE may have derived from feedstuffs containing rendered carcasses of scrapie-infected sheep, or it might have been a preexisting sporadic and rare disease unrecognized in the bovine population. The BSE epidemic was traced to the recycling of BSE-infected carcasses into cattle feed, which occurred mostly in the 1980s in the United Kingdom (UK).¹⁴⁹ Cattle in other countries were affected as a result of the importation of infected feedstuffs or cattle, followed by later recycling the agent in their own rendered ruminant feedstuffs.

Bovine spongiform encephalopathy has predominantly affected dairy cattle because of their longer lifespan and more intensive feeding and wider use of concentrate than beef cattle. From 1986 to August 2006, a total of 179,140 cases of BSE were confirmed in GB. These cases have been distributed among 35,411 farms, representing about 60% of the farms holding adult cattle.^150,151 The cattle cohorts born in 1986 to 1989 had the highest incidence of BSE cases, and the peak of the outbreak occurred in 1992, when 36,680 cases were detected through scanning of suspect clinical cases (passive surveillance) in GB. A sustained and sharp decrease in the number of cases has occurred each year since 1992. In 2005, only 39 BSE cases were detected through passive surveillance (of 186 animals investigated), and another 164 (of 547,366 cattle tested) were detected through active (targeted) surveillance programs in GB.¹⁵¹ The decline in the GB epidemic is directly related to the successive bans on the feeding of ruminant-derived meat and bonemeal to cattle. The initial 1988 ban took effect in 1989 in the UK and has been tightened and refined since then. Similar bans have been adopted by many countries around the world. A map of countries that have reported home-born or imported cases of BSE and the number of cases reported each year is available and regularly updated on the website of the Office International des Epizooties (OIE).¹⁵²

No clear genetic predisposition to BSE has been demonstrated in cattle, unlike for scrapie in sheep. Calves born from dams in the later stages of the incubation of BSE are at most 10% more likely to develop BSE than their herdmates. This may be a result of an unidentified genetic predisposition or a low degree of vertical transmission. Mathematical modeling of the rate of reduction in the number of BSE cases in response to the initial feed bans introduced in the UK showed that vertical and lateral transmission of BSE, if they do occur, do so at a very low rate that could not sustain an outbreak of BSE. At the turn of the century, mathematical modeling experts had initially predicted that BSE would be eradicated from the UK by the year 2005.¹⁵⁰ This prediction is being revised, especially because a previously unrecognized BSE phenotype has been identified in Italy, France, and Germany.^153,154

Since the onset of BSE epidemic, new TSEs have occurred in domestic cats, various exotic species, and humans.¹⁵¹ Strain typing in mice showed that these new TSEs were caused by the BSE agent.^155,156 A new variant Creutzfeldt-Jakob disease (vCJD) was first identified in 1996 in the UK, where it has affected 162 people to date.¹⁵⁷ Other countries have detected vCJD cases (two in United States) in people who had lived in the UK between 1980 and 1996, when the strictest measures were enforced to prevent human infections. France, Italy, Republic of Ireland, The Netherlands, Portugal, Spain, and Saudi Arabia have detected vCJD cases in residents who never traveled to the UK. The emergence of vCJD has put worldwide emphasis on the prevention and detection of TSEs in animals. The route of infection of people by BSE has not been clearly traced, but it is generally accepted that the consumption of food that included cattle nervous system was the main cause of vCJD in people. Successive “Specified Risk Materials” bans have been enforced to prevent contamination of human food with nervous system from ruminant animals.

Diagnosis

No in vivo test is available for the diagnosis of BSE. Clinical suspicion can be confirmed only by postmortem examination. The diagnosis of BSE is based on microscopic brain examination or tests that identify prion in brain or spinal cord tissue.^{135,153,154,158} Rapid prion tests include Western blot, paraffin-embedded tissue (PET) blot, and ELISAs.^159-162 Rapid tests have enabled the detection of BSE though surveillance in targeted at-risk populations, such as fallen cattle.

Histopathologically, BSE is characterized by neuronal degeneration and intraneuronal vacuolation in specific brain areas. The vacuolation is accompanied or preceded by the accumulation of PrPSc.^149,163,164 The uniformity of the pathology among affected cattle along the UK epidemic and in various other countries has generally supported the theory that all BSE cases were caused by a single strain of TSE agent, which has been defined by its specific BSE histopathologic phenotype.^149,164-167 Since 2004, atypical BSE cases have been described in Italy, France, and Germany.^153,154,168 Further studies are ongoing to determine whether these cases reflect the existence of a previously unidentified strain of BSE.

Differential Diagnosis

Nervous system diseases that could be confused with BSE include viral encephalopathies (pseudorabies, rabies, Borna encephalitis, bovine immunodeficiency virus encephalitis), listeriosis, polioencephalomalacia, lead poisoning, CNS parasitic migration, brain tumors and abscesses, and vitamin A deficiency. Hepatic encephalopathy and other metabolic imbalances may also be confused with BSE. Because of the insidious nature of BSE, affected cattle may appear to recover coincidentally with blanket therapy for suspected metabolic imbalances and later relapse. This must be kept in mind whenever therapy for metabolic imbalances initially appears to be successful but is followed by one or more relapses.

Treatment and Prognosis

BSE is always fatal, and no treatment is available.

Control

Various measures have been taken to protect public health by removing potentially infected animals and tissues from the food chain. The disposal of BSE-infected tissues and the sourcing of bovine tissues and body fluids for the preparation of medicinal products must take into account the extreme resistance of TSE agents. All TSE agents remain infectious after exposed to a wide range of inactivating treatments and environmental changes, such as autoclaving, rendering, storage for months to years, exposure to ultraviolet light, freezing, thawing, prolonged boiling, and incubation with formalin.^169-172 Because of the public health risk, the carcasses of animals suspected of having BSE should be incinerated.

In countries that have had an outbreak of BSE, the complete exclusion of ruminant tissues from ruminant feed has aided progress toward eradication of BSE, as previously noted. In the UK the initial feed ban of 1988 was found to be insufficient and was reinforced by successive measures. The 1996 ban on feeding mammalian meat and bonemeal to all farmed animals was introduced to prevent contamination of ruminant feed at feed mills and down the supply chain to the farm. Because of the long incubation period of BSE, there is a lag time in the effect of feed control measures, and the full effect of such measures can be assessed only when all animals born after a feed ban reach 4 to 8 years of age.

Definition and Etiology

Scrapie naturally affects sheep, goats, and mouflons. It is the oldest known TSE. It has affected sheep in various countries for more than 250 years. Strain typing in mice has demonstrated the existence of a variety of scrapie agents, none of which are identical to the agent of BSE.

Clinical Signs

Video clips of various clinical signs of scrapie are available on the web.¹³⁹ Most animals with scrapie are 1 to 5 years old. The clinical course varies but is typically slow and may last several months. Shepherds who work closely with their sheep can best recognize the early signs of scrapie. Early behavioral changes are often accompanied by weight loss. Scrapie cases may stay apart from the flock and become nervous and restless.¹⁷³ Pruritus of increasing intensity is a common feature; sheep may rub on fixed objects, scratch with their horns or hooves, and bite or lick at themselves excessively. The head, withers, flanks, back, rump, base of the tail, and lower limbs are typically affected, showing secondary wool loss, dermatitis, and skin infections or excoriations.^173,174 Head and face rubbing and shaking may also cause corneal chemosis or aural hematomas. When scratching the pruritic areas (mostly the back and rump), the animal may display a “scratch reflex” (or “nibble reflex”), which may include reflex nibbling, lip licking, and rhythmic head movements. This reflex is often but not always present in scrapie cases. It may also be observed in other CNS diseases and in pruritic skin diseases, especially ectoparasitism.

Other clinical signs of scrapie include bruxism (tooth grinding) and ptyalism. Less frequently, rumen fluid (cud) may leak through the nostrils or the mouth. Tremors may start with the head and generalize to the whole body. As the disease progresses, some sheep with scrapie may develop signs of apathy, exercise intolerance, and ataxia. The most severe and advanced cases may show convulsions, collapsing episodes, and stupor. If scrapie is left to run its course, death may occur during a convulsion or as a result of starvation. The clinical signs of scrapie in goats are broadly similar to those in sheep.^174-178 Scrapie has been identified in fallen stock, especially in flocks that are not closely supervised.

Pathophysiology

The exact mechanism of CNS degeneration in TSEs has not been identified. The oral route is the most likely route of entry of the scrapie agent,^179,180 which probably enters the gastrointestinal (GI) tract and is transported to peripheral lymphoid tissue.^181,182 Follicular dendritic cells are infected early in the disease course, and there may be an interface between the lymphoreticular system and the nervous system.¹⁸³ Transport to the CNS likely occurs along the vagus nerve from the GI tract.¹⁸⁴ PrPSc has been identified by immunocytochemistry in the GI tract (in Peyer’s patches and in autonomic plexuses) and in various lymphoid tissues of sheep with scrapie.^185,186 Replication of the agent takes place in lymphoid tissues, including the spleen, but circulating lymphoid cells do not appear to be involved in scrapie replication.^181,182,187

Epidemiology

Scrapie occurs endemically in sheep flocks worldwide.^188-190 Sheep are the natural host of scrapie, but the infection can be maintained in goats that have no direct contact with sheep, indicating both lateral and vertical transmission of the agent in that species.^{174,175,188,191} Scrapie cases occur sporadically in infected flocks. Only one or a few animals are affected at any given time. Outbreaks with up to 40% of animals affected in a flock have been linked to the use of infected vaccines¹⁹² or to the introduction of scrapie by an infected animal in a previously uninfected and genetically sensitive flock. The route of natural infection is presumed to be oral. The route of excretion and the means of transfer of the agent between sheep are unknown.¹⁷⁹ Lambing time is known to be a particularly high-risk period for infection for the young. The contamination of pastures with infected placentas is likely to be an important reservoir of the infectious agent in nature.^173,193,194

Although there is no known breed predisposition to scrapie, the genetic makeup of the host controls differences in susceptibility and resistance to scrapie in various breeds.^172,195,196 The molecular basis of this resistance is largely controlled by the PrP gene.¹⁹⁷ The gene responsible originally was thought to determine the incubation period of scrapie and was called sip. This gene is now known to be the prion protein gene. Polymorphisms at three codons of this gene appear to be the main determinants of the susceptibility of sheep to scrapie.^198,199 According to current evidence, the genotype VV₁₃₆RR₁₅₄QQ₁₇₁ (or AA₁₃₆RR₁₅₄ QQ₁₇₁ in some breeds) is most susceptible, and the genotype AA₁₃₆RR₁₅₄RR₁₇₁ is totally resistant.²⁰⁰ The potential relevance of other polymorphisms and other genetic factors is unknown. Selection of scrapie-resistant sheep flocks may be based on pedigree, phenotypic expression of the disease,¹⁷² or genetic testing to select sheep of “resistant” PrP genotypes.²⁰⁰ Studies are ongoing to determine whether “genetic resistance” may confer resistance to disease but not resistance to infection, in which case some genotypes of sheep could act as asymptomatic carriers of scrapie.¹⁵¹

Treatment and Prognosis

Scrapie is a fatal and irreversible disease for which there is no known treatment.

Diagnosis and Pathology

Control

Because there is no effective treatment for scrapie, control measures designed to prevent the spread of the disease are especially important. Scrapie is a reportable disease. Eradication measures vary from country to country.^149,215 Animals suspected of having scrapie are slaughtered and destroyed. Contaminated pastures or paddocks may be left empty of livestock, but the scrapie agent is unlikely to be inactivated in the environment. Contaminated stalls, corrals, and sheds should be disinfected with sodium hypochlorite diluted at 4% available chlorine. In the United States and Canada, scrapie-affected and related animals are destroyed and their flocks quarantined.²¹⁵ Lines of sheep genetically resistant to the development of clinical scrapie have been selected.²¹⁶ These sheep do not develop the clinical signs of scrapie, but they may have a chronic asymptomatic infection that could spread to sheep of susceptible genotypes.^172,216,217

PUBLIC HEALTH CONSIDERATIONS

Although scrapie has been endemic in various areas of the world for more than two and a half centuries, epidemiologic studies have not shown any correlation between the incidence of Creutzfeldt-Jakob disease in humans and that of scrapie in sheep.²¹⁸ It is nevertheless wise to take precautions to minimize the potential for human exposure to scrapie because some sheep could have been infected by BSE, which would be identical to scrapie in any infected flock.²¹⁹ In a study of BSE orally transmitted to sheep, the clinical signs of BSE were identical to those of scrapie.²²⁰

Clinical Signs

Acute onset of ataxia and tetraparesis of variable severity most often characterizes the neurologic form of EHV-1.^229,233 Signs usually appear between 6 and 10 days after infection.²²⁹ The severity of clinical signs can range from subtle neurologic deficits to complete recumbency. Other signs also may be seen, including nasal discharge, limb edema, colic, ocular lesions, and anorexia.^{226,228-231,233} The animal may be febrile at presentation, but most are normothermic.^226,229 Pyrexia is the most consistent premonitory sign before the onset of neurologic disease, although not all pyrexic horses will develop myeloencephalopathy.^232,234 Coughing and nasal discharge sometimes accompany the neurologic deficits or may have been present in the preceding 2 weeks.²³¹

The neurologic signs reflect damage to the white matter of the spinal cord and include ataxia, paresis, conscious proprioceptive deficits, urinary incontinence, flaccid tail and anal tone, and diminished perineal sensation.^226,229,233 Hindlimbs are more severely affected, and deficits usually are symmetric. Bladder atony and dysuria frequently occur, and the associated dribbling of urine leads to secondary perineal scalding. Anal sphincter tone also is diminished, which may result in a distended rectum.

Cranial nerve deficits, including seizures, blindness, and vestibular signs, have been reported with EHV-1 infection.²³¹ Ocular lesions also may be present, such as mydriasis, hypopyon, uveitis, and optic neuritis.^{226,229,235,236}

The clinical signs typically stabilize within 48 hours, although progression varies. Some patients continue to deteriorate and eventually die or are euthanized. Most horses begin to improve within the first 5 to 7 days and ultimately make a full recovery, although the recovery may take months. A patient that does not become recumbent has a much better prognosis.^233,235

Diagnosis

A tentative diagnosis can be made based on characteristic neurologic signs, the history, and supporting findings on the physical examination. A cerebrospinal fluid (CSF) analysis that shows an increase in protein and a normal or slightly increased nucleated cell count (albuminocytologic dissociation) is the classic CSF change seen with EHV-1 infection.^226,229,237 Cerebrospinal xanthochromia may also be seen.²³³ The protein level may be normal, which does not necessarily rule out EHV-1. Occasionally, early in the disease the CSF may be normal or may show only mild abnormalities; CSF protein concentration in the normal range does not rule out EHV-1.²²⁶

A fourfold or greater increase between acute and convalescent virus neutralizing (VN) antibody titers is consistent with a diagnosis of EHV-1 infection.²³⁷ VN titers usually are the easiest and most economic to perform. Because they rise quite rapidly and reach a level much higher with natural infection compared with vaccination, a fourfold rise in the VN titer may not be seen, especially if collection of the acute sample was delayed.^226,235 A single titer of 1:256 or higher is highly suggestive of recent infection.^229,237 The titer of VN antibodies does not correlate with protection against infection or with reduced viremia.²³⁸ Complement fixation (CF) titers decline quite rapidly after infection, requiring sample collection early in the course of the disease. A titer higher than 1:16 is consistent with recent infection.²³⁷ It is important to know which assay the diagnostic laboratory uses, because interpretation of the results differs depending on the test performed.

Viral isolation attempts from the buffy coat of an ethylenediamine tetraacetic acid (EDTA) tube and nasopharyngeal swabs may yield a positive result for EHV-1 for approximately 10 to12 days after infection.²²⁶ It is critical that samples be handled appropriately and transported to the laboratory as soon as possible. A viral transport medium is necessary and should be available from the diagnostic laboratory. Viral isolation from the CSF is unrewarding because there does not appear to be direct viral infection of neurons.²³³

Pathophysiology

An apparent immune complex vasculitis and thrombosis in arterioles of the spinal cord lead to segmental spinal cord ischemia. Although the lesions are characteristic of a type III (Arthus) hypersensitivity, an immune-mediated pathogenesis has not been conclusively demonstrated.²²⁹ However, the theory of an immune-mediated pathogenesis is supported by the finding that horses vaccinated within the previous 12 months were shown to be 9 to 14 times more likely to develop the neurologic form of EHV-1 infection.²²⁹ The vasculitis may be the result of viral infection of the CNS endothelium by circulating infected leukocytes.²³³ The CNS vascular endothelium appears to be the primary site for infection, and the development of neurologic disease may depend in part on the endotheliotropism of the virus. Viral replication within neural tissue has not been definitively demonstrated.^226,229

Epidemiology

Herpesvirus infection is enzootic in the horse population. Infection usually occurs through the respiratory or intestinal epithelium after the animal comes in contact with the virus in fluids from an abortion or in ocular, nasal, or respiratory tract secretions.^229,235 Most horses become infected before 1 year of age. As with most herpesvirus infections, the virus is capable of evading the horse’s immune system and can develop latency.^229,235,239 Sites considered likely for latency are the lymphoid tissue and the trigeminal ganglion.^239,240

Equine herpes myeloencephalopathy is rare, but cases have been reported worldwide.²²⁹ The existence of a neurovirulent form of the virus has been suspected, but the differences in the strains isolated from neurologic syndromes are not significant enough to warrant additional classification.²²⁶ However, equine herpes myeloencephalopathy outbreaks have been reported, lending support to the idea of a neurovirulent form.^230,233,235 During outbreaks, the virus appears to be spread readily, and both morbidity and mortality can be high.²³⁴

Animals of any age or gender are susceptible; pregnant or lactating mares are most often affected.^226,229 Risk factors for the development of neurologic signs include female gender, older age, and fever. A higher prevalence of neurologic disease was found in standardbred, Hispanic breeds, and draft horses in one study of multiple outbreaks of herpesvirus myeloencephalopathy.²²⁹ Foals are rarely affected with the neurologic form of EHV-1 infection. There appears to be some seasonal variation because more cases are seen during the spring and winter months.²³² Stress-associated recrudescence of latent infections and shedding without clinical signs are important in the development of equine herpes myeloencephalopathy in a closed population. The virus may be shed for 3 weeks or longer after infection. A morbidity rate of up to 90% and a mortality rate of up to 40% have been reported.^229,233 There are no reports of equine herpes myeloencephalopathy associated with any modified live virus vaccine currently approved for use in horses.¹⁷⁵ Frequent vaccination (up to four times a year) may render animals more susceptible to neurologic manifestation of EHV-1 infection.²³⁴

Necropsy Findings

Gross and histologic lesions are not always limited to the CNS.^226,233 Ocular lesions have been reported, including hypopyon, iritis, and chorioretinitis. Cystitis and scrotal edema may be present.^226,229 Focal areas of hemorrhage may be found throughout the brain and spinal cord parenchyma and meninges. Vasculitis of the small arteries and veins of the spinal cord white matter and of the gray and white matter of the brain results in ischemic lesions in the CNS.^241,242 Equine herpesvirus is infrequently isolated from the CNS during a postmortem examination.^230,237 Polymerase chain reaction (PCR) and direct immunofluorescence assay in situ can be performed on CNS tissues obtained postmortem to confirm the presence of the virus.²³⁰

Treatment

Supportive care is the most important aspect of treatment for equine herpes myeloencephalopathy.^{226,229,233,237} Measures include bladder decompression twice a day for horses with bladder atony and urinary incontinence, evacuation of the rectum, enteral or parenteral nutritional support, and administration of intravenous or oral fluids. The horse may require support in a sling.

Administration of antiinflammatory drugs soon after the onset of neurologic signs may be beneficial. Corticosteroids have been used because of the possible immune-mediated pathogenesis, but no objective data are available evaluating the efficacy of this treatment. Dexamethasone (0.05 to 0.1 mg/kg IV) can be given every 12 to 24 hours for 3 to 5 days, with the dosage then tapered for 1 to 3 days.²³⁷ The possibility of viral reactivation is unlikely at the recommended dosage.²²⁹ Dimethyl sulfoxide (DMSO; 0.25 to 1 mg/kg by slow IV infusion) every 12 to 24 hours is routinely used when treating neurologic disease, although its efficacy has not been documented.²²⁶

Use of antibiotics should be considered if the horse is recumbent or has urinary tract involvement, or if respiratory tract signs are present. Antiviral agents, particularly acyclovir, have been recommended based on their use in humans for herpes simplex virus encephalitis. There is insufficient evidence to recommend the use of antiviral agents for EHV-1 infection in horses.²³⁷

Prevention

Currently, no prophylactic measures for or methods of preventing equine herpes myeloencephalopathy are available. The vaccines currently used to prevent EHV-1 respiratory and abortion syndromes do not claim to prevent equine herpes myeloencephalopathy. Findings in vaccinated horses naturally exposed to EHV-1 during disease outbreaks are mixed regarding protection against myeloencephalopathy.^232,238 Vaccination may reduce the incidence of the other EHV-1—related diseases and thereby reduce exposure to the virus and the risk of developing equine herpes myeloencephalopathy. A recent study concluded that modified live virus vaccine provided significantly better protection against EHV-1 myeloencephalopathy than a killed vaccine,²³⁸ although the duration and robustness of such protection are not yet clear. Vaccination in the face of an outbreak is controversial²²⁶; viremia may be reduced or prevented as a result of vaccination, but an increase in antibody levels may be partly responsible for, or may play a role in the development of, the neurologic form.

Management practices may reduce the risk of introducing or disseminating EHV-1 infection. Such practices include isolating all new arrivals for at least 3 weeks and maintaining distinct herd groups based on age, gender, and occupation. Pregnant broodmares should be kept apart from the general population as much as possible. Minimizing stress may reduce the likelihood of recrudescence of a latent infection.

Definition and Etiology

Pseudorabies is caused by an encapsulated DNA virus, suid herpesvirus type 1 (Su-HV1), a member of the genus Varicellovirus, subfamily Alphaherpesvirinae, family Herpesviridae.^243,244 This virus is able to infect the CNS and other organs (e.g., respiratory tract) of virtually all mammals except humans and most primates. Domestic and feral swine are the natural hosts and may become latent carriers. The virus can also infect cattle, sheep, cats, dogs, and goats as well as wildlife (e.g., raccoons, opossums, skunks, rodents) and rarely, horses.^245-248 In domestic and wild ruminants, pseudorabies is an acute and usually fatal encephalitis.

Clinical Signs

The incubation period ranges from 90 to 156 hours, and the illness may last 8 to 72 hours, although some affected animals have been found dead without any observed clinical signs.^249,250 The initial clinical sign is usually a manifestation of paresthesia, with acute and severe pruritus inducing self-trauma, dermal abrasions, swelling, and alopecia.²⁴⁹ Other signs may include fever, bellowing, bloat, feet stamping, excessive salivation, chewing of the tongue, ataxia, circling, nystagmus, and strabismus.^251-253 Aggression may be seen, but most affected animals become depressed. As the disease progresses, there may be twitching, hyperesthesia, tenesmus, spasmodic kicking, excessive licking of the nostrils, continuous mastication, sweating, vocalization, semicoma, coma, convulsions, opisthotonos, hyperpnea, tachypnea, and slow irregular respiration.²⁵⁴ The clinical signs of pseudorabies in cattle closely resemble those of rabies, polioencephalomalacia, salt poisoning, meningitis, lead poisoning, hypomagnesemia, and enterotoxemia.

Clinical Pathology

The Aujeszky’s disease virus (ADV) can be isolated from the pharyngeal or nasal secretions of affected animals and can be easily cultured from infected nervous tissues. Strains of ADV are antigenically distinct from other herpesviruses but share common antigens with the infectious bovine rhinotracheitis (IBR) virus. Heterospecific antibodies to the IBR virus may cross-neutralize the pseudorabies virus and confound serologic tests. For virus culture, tissues should be collected from sensory ganglia or the sensory parts of the spinal cord. Segments serving the pruritic sites should be collected preferentially because these areas contain the highest concentration of virus.

Pathophysiology

Ruminants are susceptible to pseudorabies infection after intradermal, subcutaneous, intranasal, or oral exposure to the virus. After subcutaneous, oral, or nasal infection, the virus spreads centripetally to the CNS by axonoplasmic transport. During the acute infection the virus may be present in the nasal mucosa, secretions, and saliva.

Epidemiology

Aujeszky’s disease has a worldwide distribution and, although cases occur only sporadically, is an economically important disease because of the regulatory quarantine and other restrictions imposed on animals from affected herds (it is a reportable disease). Outbreaks have occurred in cattle in the United States, Europe, Australia, New Zealand, Latin America, and South America. Until recently, Aujeszky’s disease was endemic in the United States, but it now has been successfully eradicated from domesticated swine. As of December 2006, all states are classified as free of pseudorabies in domesticated swine (OIE status V), although ADV remains present in feral pigs in parts of the United States.

The viral infection in cattle is perpetuated partly by the occurrence of latent infections in pigs, which can be recrudesced by stressful conditions.²⁵⁵ Occasional spillover of the virus from swine into ruminants occurs because of the proximity of the two species in many livestock operations.^250,256 The natural routes of transmission include nose to nose, fecal oral, and venereal.^245,257 Indirect transmission usually occurs by inhalation of aerosolized virus. The virus may travel via aerosols up to 2 km (1.2 miles) in certain weather conditions and may survive up to 7 hours in nonchlorinated well water; for 2 days in anaerobic lagoon effluent and in green grass, soil, feces, and shelled corn; for 3 days in nasal washings on plastic and pelleted hog feed; and for 4 days in straw bedding. It is inactivated by drying, sunlight, and high temperatures (≥37°C). Dead-end hosts, such as dogs, cats, and wildlife, can transmit the virus between farms, but aside from feral swine, these animals survive only 2 to 3 days after becoming infected. Sheep have also been infected by modified live virus vaccines targeted for use in swine.^258,259

Latency of the viral infection in ruminants is not an important mechanism for perpetuation of an outbreak; most animals die within 48 hours of disease onset and may shed ADV in their saliva, oral secretions, and mucous membranes for only up to 6 days after infection, and direct spread of the virus between infected and uninfected cattle is unlikely.²⁵⁶ Wild mammals such as raccoons may play a role in pseudorabies survival and transmission; rats, however, develop transient infections and do not appear to transmit or perpetuate the virus.^260-262 The pseudorabies virus may survive in contaminated meat products for up to 7 weeks. The role of this prolonged survival in perpetuating outbreaks in ruminants is unknown. Differences in virus pathogenicity for cattle have been correlated with the type of syncytium formation in tissue cultures.²⁶² The role of these strain differences in the perpetuation and dissemination of the disease is unclear.

Necropsy Findings

The macroscopic changes that occur in animals infected with the pseudorabies virus are alopecia, edema, and hemorrhage at the pruritic site. The neuropathologic changes include perivascular cuffing, interfascicular edema, nonsuppurative encephalitis, gliosis, neuronal degeneration, and eosinophilic intranuclear (Cowdry type A) inclusion bodies. The lesions are most pronounced in the dorsal nerve rootlets and the dorsal horn.

Treatment

There is no treatment for pseudorabies, which is usually fatal in livestock, although rare recoveries have been reported.²⁵²

Prevention

The most effective method of preventing pseudorabies virus infection in ruminants is eliminating their exposure to swine. Contaminated pens can be disinfected with 10% sodium hypochlorite solution, quaternary ammonium compounds, tamed iodine, or phenolic compounds.²⁶³ At least 5 minutes of contact time should be allowed before the disinfectant is rinsed from the contaminated surfaces. Fumigation with formaldehyde for 6 hours effectively kills the virus, as does 360 minutes of contact with ultraviolet light.

Etiology

Togaviruses are single-stranded, linear, positive-sense RNA viruses with an envelope that makes them susceptible to drying and ultraviolet light. EEE, WEE, and VEE viruses are the most frequently isolated togaviruses from epidemics of encephalitis in horses and humans in the Western Hemisphere.²⁶⁵ The first recorded epidemic of EEE in horses occurred in Massachusetts in 1831; the virus was first isolated from a horse in 1933.²⁶⁵ It was soon determined that epidemics of encephalitis in horses in North America were caused by different alphaviruses with regional geographic biases.^266,267

The virus is composed of a nucleocapsid within the envelope that has icosahedral symmetry consisting of peplomers arranged as trimers.²⁶⁵ Viral species are differentiated by hemagglutination inhibition (HI) activity and neutralizing specificity.^267-269 The common group-specific antigenic determinants are still mainly defined by serologic techniques, such as fluorescent antibody, complement fixation, and ELISA.^270-274 Genetic sequencing has application in molecular epidemiology.^275-277

For WEE virus, WEEV and Highlands J virus (HJV) constitute the two main subgroups.^278-281 WEEV predominates west of the Mississippi River, whereas HJV is primarily detected east of the Mississippi.^282,283 Both these viruses are capable of causing encephalitis in horses, with HJV likely less pathogenic for mammals.^282,283 Other subtypes of WEE virus have been identified in the United States, and most WEE infections in western states are likely the result of infection with one of the several antigenic variants of the actual WEE subtype, although minimal disease has been reported in humans and horses in the last decade.

The organization of VEE is much more complex. Six distinct subtypes (I through VI) and numerous varieties of viruses (designated by letter) within these subtypes are classified within the VEE virus complex.^284-288 The “epidemic types” of VEE virus (types IAB, IC, and IE) are responsible for the large outbreaks of encephalitis in horses in the Western Hemisphere in the past 20 years.^289,290 “Endemic types” of VEE virus are considered to be of low pathogenicity for equids under most circumstances.^291,292 These include type ID and IF variants from Central America and Brazil, respectively; type II (Everglades) virus found in Florida; three known variants of type III (Mucambo) virus; type IV (Pixuna) virus; type V (Cabassou) virus; and type VI virus.

Epidemiology

The classic life cycle of equine alphaviruses involves transmission between birds or rodents and mosquitoes.²⁷⁴ In some cases, other domestic and wild animal species, especially species exotic to North America such as the emu, have been affected during these outbreaks.²⁹³ Understanding the antigenic and genetic relationships among the viruses in the WEE complex has proved more challenging than for the viruses in the EEE and VEE virus complexes. WEE virus is a member of the WEE antigenic complex that includes several Old World viruses in addition to the New World viruses previously described. Phylogenetic analyses of isolates from North and South America indicate that regional WEE lineages appear to evolve independently for several years to a few decades (e.g., genotypes in South America are apparently absent from North America).^283,294-296 However, relatively homogenous genotypes of WEE are dispersed across both North America and South America. This contrasts with EEE and VEE viruses, in which certain virus genotypes appear to be restricted to either North America or South America. WEE virus has been reported in several countries in South America (Argentina, Guyana, Ecuador, Brazil, and Uruguay), but only in Argentina has it been associated with human disease and significant epidemics in horses.^281,297-299

Pathogenesis

The virus genome is 9.7 to 11.8 kilobases in length and encodes both nonstructural and structural proteins.³³³ Alphaviruses replicate to high titer in the cytoplasm of infected cells and exit the cell by the budding of preassembled nucleocapsids through the plasmalemma.³³⁴ They cause cytopathic effects in a wide range of vertebrate cells in vitro. Virulence of VEE virus depends on systemic virus load rather than specific neurovirulence.^335,336 Both EEE and VEE have efficient intracellular replication and are pathogenic when inoculated intracerebrally.³³⁷ Studies in mice demonstrate that neural invasion is likely to occur secondary to vascular infection or invasion through the olfactory epithelium.³³⁸ In young mice, there is intense replication in osteoclasts of developing bone, possibly explaining why young animals and humans are much more susceptible to severe disease.

The regional lymph node is presumed to be the site of primary viral replication after the bite of a mosquito infected with EEE, WEE, or VEE virus.²⁶⁴ The reticuloendothelial system is a major target in epidemic VEE infections. The viruses cause encephalitis after hematogenous or neuronal spread. Immunity, after both inapparent infection and clinical disease, as caused by EEE, WEE, and VEE, is long lasting in all species. Horses infected with EEE and WEE do not excrete infectious virus, and recovered animals are not persistently infected with virus.

Clinical Findings

Different strains of EEE, WEE, and VEE viruses may differ in their virulence not only for horses, but also for humans, certain domestic and wild animals, birds, and laboratory animals.³²⁸ Clinical observations during epidemics of VEE indicate that the disease is generally less severe in donkeys (burros) and mules.³³⁹ None of these viruses appears to cause clinical disease in their reservoir hosts indigenous to North and South America. Many human and equine infections, apart from those caused by highly virulent strains, are subclinical. When disease does occur, broad differences exist in the clinical manifestations produced by the three virus complexes in horses and humans.

Eastern equine encephalitis and epidemic VEE viruses are generally more neuroinvasive than WEE and endemic VEE viruses. Children and young animals of all susceptible species are more likely than adults to develop clinical signs referable to infection of the central nervous system.^{264,301,323,340,341} The incubation periods of EEE, WEE, and VEE vary from 2 to 3 days to, rarely, as long as 3 weeks. Inapparent infections in horses may or may not be accompanied by fever. In clinical cases, pyrexia is the first clinical manifestation of infection. Temperature has usually abated or is only moderately elevated by the time signs of encephalitis become evident. Neurologic signs are variable, but obtunded mentation, ataxia, paralysis, anorexia, and ultimately stupor occur in clinical cases. Irregular gait, grinding of teeth, incoordination, circling, staggering, head pressing, and hyperexcitability are also observed; clinical signs are progressive in nature. Severely ataxic animals may stand by leaning against walls or other objects and sometimes stand with their hindlegs crossed. Partial or even total blindness may be evident. In severely depressed horses the head hangs low with drooping ears, and the eyelids may be slightly swollen and partly closed, while the lips are flaccid and the tongue may protrude from the mouth. The profound depression associated with these virus infections give rise to the common name of “sleeping sickness.” Esophageal paralysis, as manifested by repeated unsuccessful attempts to drink, has also been described.

The course of disease in severely affected horses varies between 2 and 14 days. Almost all horses with EEE encephalitis die, regardless of the quality and intensity of clinical care. Horses with disease caused by VEE or WEE are more likely to survive than those with EEE-induced disease. At terminal stages, horses become recumbent, become comatose, and frequently exhibit seizure activity.

Diagnosis

Clinical signs and antemortem clinicopathologic findings are not specific for alphavirus infection. Viral and other encephalitides can cause abnormal cerebrospinal fluid (CSF), which usually consists, in the horse, of a moderate mononuclear pleocytosis with an increased CSF protein. Eastern equine encephalitis is unique in that acute infection frequently results in a neutrophilic pleocytosis. Because there is a high mortality rate associated with this disease, identification of a neutrophilic pleocytosis indicates probable EEE infection and offers the veterinarian a chance to prognosticate regarding the horse’s survival.

It is paramount to obtain a definitive diagnosis for clinical signs of encephalitis in the horse, to justify and institute effective control measures, because of the risk of these viruses to the health and well-being of both humans and equine livestock. The viruses of EEE, WEE, and VEE frequently, but not always, can be isolated or detected after death in brain material of diseased horses by the use of cell cultures (e.g., Vero cells), intracerebral inoculation of suckling mice, and by detection of specific nucleotide sequences using reverse-transcriptase polymerase chain reaction (RT-PCR) technology.^342,343 Blood is an inappropriate specimen for virus recovery because no circulating organism is usually present when neurologic signs become apparent. In an epidemic situation, however, it may be possible to isolate the virus from nonencephalitic horses in the affected group, particularly if they have increased body temperature. The cytopathic or lethal effects of the virus in cell cultures or experimental animals can be inhibited by the use of specific antisera, and in this way the virus involved may be specifically identified.

Currently, there are no reliable antemortem diagnostic tests to detect virus in clinically affected horses, and serology provides the mainstay of presumptive antemortem diagnosis. The demonstration of specific IgM antibody (dilution of 1:400) suggests recent infection.³⁴⁴ The detection of IgM antibody in CSF (if available) is even more conclusive. Rising antibody titers to EEE, WEE, or VEE viruses in the sera of horses that survive can be detected by testing of acute-phase and convalescent-phase sera. Even in endemic areas, it is not possible to diagnose/differentiate EEE, WEE, or VEE in the horse with any certainty based on clinical signs and epidemiologic circumstances.

Rabies, hepatic encephalopathy, and equine protozoal myeloencephalitis are the major diseases that must be considered in the differential diagnosis in the Western Hemisphere. Other diseases that should be considered are equine herpesvirus 1 infection of the central nervous system, leukoencephalomalacia (a neuromycotoxicosis caused by the ingestion of maize infected with Fusarium moniliforme), and ataxia resulting from cervical vertebral malformation.

Pathologic Findings

In horses, brain lesions are thought to be the direct result of viral replication and are characterized by necrotizing encephalitis with neuronal dysfunction.³⁴⁵ There are no consistent gross lesions in horses that die of EEE, WEE, or VEE. Histologically, neuronal necrosis with neurophagia, marked perivascular cuffing with both mono- and polymorphonuclear leukocytes, and focal and diffuse microglial proliferation are evident. The lesions are more pronounced in the gray matter than in the white matter of the brain. Lesions are most marked in the cerebral cortex, thalamus, and hypothalamus, whereas the spinal cord is mildly affected. More frequent and severe lesions are usually present in the cervical spinal cord than in lumbar cord segments.

Treatment

No known antiviral medications have demonstrated reliable activity against alphaviruses, and treatment of disease in affected horses is supportive. The survival rate for EEE infection is low compared with other infectious encephalitides. In most cases, horses die 3 to 5 days after onset of signs.

Corticosteroids should be considered as a component of therapy for horses with neurologic signs consistent with viral encephalitis and neutrophilic CSF. If administered early, corticosteroids (to reduce brain edema) and intravenous fluids may aid recovery. In human patients, treatment with methylprednisolone at 1000 mg/100 kg is often recommended. Administration of flunixin meglumine (1.1 mg/kg every 12 hours IV) or other antiinflammatory medications to horses with EEE does not often result in the dramatic response frequently observed in horses with West Nile virus. Mannitol (0.25 to 2.0 g/kg every 24 hours IV) may assist in the control of brain edema. Detomidine hydrochloride (0.02 to 0.04 mg/kg IV or IM) is effective for prolonged tranquilization.

Intravenous immunoglobin therapy has been used in people for its proposed anticytokine effect. Interferon-alpha (IFN-α) is a relatively common therapy. The recommendation for IFN-α therapy is based on anecdotal reports in the human and veterinary literature. Limited information regarding efficacy in the horse is available.

Prevention

The alphaviruses are not stable in the environment and are easily inactivated by common disinfectants. Mosquito control and immunization of horses are both important in control of EEE and VEE epizootics. Vector control can be achieved through reducing the breeding activities of mosquitoes by implementing appropriate water management systems, although this can be difficult in extensive areas.^346,347 The widespread dispersal (usually achieved by aerial spraying) of insecticides has been used successfully, although several critical factors need to be considered before embarking on such a step. Concern over indiscriminate spraying of insecticides can be mitigated in some circumstances if the biology of the mosquito vector is well known. For example, in northern Florida, where EEE is endemic, treating the pools of water in which Culiseta melanura breeds with a larvicide is often practical and economically feasible on those farms with valuable horses. Swamps with soil types that support the breeding of C. melanura can often be recognized by the nonentomologist by the presence of the loblolly bay tree (Gordonia lasianthus); this broadleaf evergreen tree grows to a height of about 30 feet (9 m) and can easily be recognized by its white magnolia-like flowers and serrated leaf.^264,300,301 Risk assessment can be assisted by geographic mapping of large areas where mosquitoes breed by using thematic mappers, such as those on orbiting satellites.

Immunization of horses has proved highly effective as an adjunct to other control measures, particularly in outbreaks of VEE in which horses may serve as a source of infection for mosquitoes.^348,349 Currently, bivalent vaccines (usually consisting of formaldehyde-inactivated virus) are commercially available against EEE and WEE. These vaccines require a primary and a secondary immunization schedule about a month apart, followed by biannual boosters. The vaccines are not particularly effective in protecting foals and yearling horses, for unknown reasons.³⁵⁰

The 1969–1971 VEE epidemic in Central America and the southern United States was controlled partially by immunizing large numbers of horses with an attenuated VEE virus strain, TC-83.^348,349 This vaccine strain was produced by serial passage of an epidemic variant in guinea pig cell cultures. Because of concerns over the presence of low-level viremia in some horses and the possible transmission of vaccine virus between horses by mosquitoes and reversion to virulence, inactivated vaccines against VEE are now available for use in horses. These vaccines are not widely used in North America because they compromise the international movement of horses for competition and breeding.

Outbreaks of VEE might be completely prevented if sustained and widespread vaccination with live attenuated VEE vaccines was performed in Central and South America.²⁹⁰ Public health and animal industry officials should consider maintaining vast quantities of the live attenuated TC-83 vaccine. The use of the formalin-inactivated vaccine (usually marketed as a multivalent antigen) is discouraged in VEE endemic areas because of the need for multiple vaccinations (and thus delayed onset of protection), short-lived immunity to VEE, lack of long-term compliance from agricultural officials and the horse-owning public in endemic locales, and concern of limited response to the live vaccine in horses immunized recently with a killed product.

Surveillance for encroachment of alphaviruses in new geographic locales is also paramount to control. Most southern U.S. states have encephalitis testing programs that offer subsidized testing for horses with suspected viral encephalitis. Enhanced passive surveillance for alphaviruses should be undertaken when environmental conditions are favorable. For example, hurricanes were implicated in the VEE outbreak in Mexico in 1995. Given that the United States has experienced intense hurricane activity in 2004 and 2005, enhanced surveillance for epizootic VEE should logically be undertaken.

Alphaviruses are pathogenic for people but require a vector for infection. The horse does not develop a sufficient level of viremia with EEE and WEE to act as a reservoir or amplifying host for these viruses. In contrast, horses are considered the most important species for amplification of virus in epizootic VEE. Therefore, control measures should be implemented to limit new exposure of horses in locales that are undergoing epizootic VEE. This includes restriction of horse movement; clinically normal horses may be viremic, and the disease can be translocated. Horses should be vaccinated and sprayed frequently with permethrin-based products. Mosquito abatement efforts should be pursued. Blood and tissues from VEE horses should be handled as infectious and biohazardous materials, with tissues from EEE and WEE horses handled as such.

Getah/Ross River Viruses

The Semliki Forest complex of the Togaviridae are widespread throughout their geographic range, either in the Orient (Getah) or South Pacific (Ross River).^351,352 Although closely related, there is little overlap of these viruses. As with the North American alphaviruses, these viruses are transmitted by mosquitoes (Aedes, Anopheles, and Culex species).^353-360 With Getah virus (GV) the reservoir is not known, although swine appear important in its transmission to mosquitoes.^360-363 Other feral species are not firmly established. With Ross River virus (RRV), marsupials are important reservoirs.³⁶⁴ With GV, horses might develop sufficiently high viremia to transmit virus, but this is unlikely with RRV.^365-367 Many animals have been found to be seropositive to GV, including ruminants, and seroprevalence in horses varies from less than 10% to over 90%.^360,368-370 RRV is endemic in horses and widespread, with seroprevalence ranging from 50% to 80% in Australian horses.^351,371-373

Getah virus is likely more pathogenic in horses than RRV.^365,368,369 Clinical signs include pyrexia, edema of the limbs, and stiff gait. Horses may also develop urticaria and submandibular lymphadenopathy. The course of GV disease is 7 to 10 days. With RRV, horses also develop fever, lameness, swollen joints, inappetence, generalized stiffness, and even colic.^351,371,373 High rates of mortality are not a feature of either virus.

Serologic detection of antibody is the mainstay of diagnosis of these viruses, although a vaccine against GV complicates diagnosis.^374-376 RRV can be isolated from the blood of horses during the disease course. Pathologic changes associated with these viruses are seldom described because of their low mortality. Horses primarily develop perivascular inflammation in the brain.

Treatment is supportive; no antiviral is specific for alphaviruses. A vaccine for GV is available in Japan.

Borna Disease

Borna disease is a cause of encephalitis first reported in horses in Borna, Germany, in the nineteenth century, with identification in other Northern European countries, such as Switzerland, Liechtenstein, and Austria.³⁷⁷ Unconfirmed outbreaks have been reported in the Middle East and Japan.³⁷⁸ The Borna disease virus (BDV) is linked to other regions through serologic testing and identification of antibodies in horses from Turkey, Israel, Iran, and the United States.^379-382 Other species noted with clinical signs of BDV include small ruminants, cattle, rabbits, cats, and dogs.³⁸³

The etiologic agent of Borna disease is a nonsegmented, negative-sense, single-stranded RNA virus of the order Mononegavirales, family Bornaviridae.^384-386 The viruses of Borna disease and Near Eastern encephalitis are indistinguishable. Importantly, with complete sequencing of the genome, several distinct virus clusters occur geographically.^381,387,388 In Germany, Borna has endemic foci centrally and south.^377,381,385 Subclinical exposure in horses is between 10% and 20%, with seroprevalence as high as 50% in stables with disease.

The Borna disease virus is shed through nasal and lacrimal secretions and in the urine of infected animals, so direct contact is suspected.^383,385,389 BDV is resistant to drying and other adverse environmental conditions. Outbreaks in horses in the Middle East may represent transmission from a dense population of infected wild birds, and recurrent outbreaks in Germany have been thought to originate from birds that have migrated from the Near Eastern countries.^377,381,385 Most outbreaks in horses occur in the early spring or autumn.

It is assumed that BDV enters the horse though intranasal infection and migrates to the brain transaxonally.^383,390,391 Direct replication occurs in neurons and glial cells, with spread to peripheral nerves and retina. Infection with BDV is persistent, and Borna disease has a slow onset thought to be caused by chronic inflammatory reaction in the brain. The lesion resembles that of lymphocytic choriomeningitis virus (LCMV) infection, in which the virus induces a cell-mediated response that disrupts and replaces functional aspects of the neuropil.

The clinical signs of Borna disease in large animals are similar to those of the other equine encephalitides.³⁷⁷ In moderate to severe cases, horse die 1 to 4 weeks after onset of disease. In mild cases, recovery can be spontaneous; however, a chronic disease can occur with recurrent exacerbations. Typical clinical signs include ataxia, head tilts, muscle fasciculations, hindlimb paresis, and localized cutaneous hyperesthesia or hypoesthesia. Aggressive behavior may be noted. In ruminants the clinical signs include head tremors, hyperesthesia, ataxia, anorexia, propulsive walking, coma, and convulsions.

For antemortem diagnosis, antibodies to the agent may be found in the serum and CSF of most infected animals by ELISA, indirect fluorescent antibody (IFA), and Western blot.^377,382 Some work indicates that CSF antibodies are more likely to be found in clinically affected horses than normal horses. There is often a mononuclear pleocytosis with high protein concentration. Enhanced-sensitivity PCR techniques have demonstrated virus in the CSF and peripheral mononuclear cells.³⁹² The criteria for diagnosis include a horse with neurologic abnormalities testing antibody positive for BDV in serum or CSF, or a horse with neurologic signs and appropriate histopathologic changes.^377,383

Pathologic abnormalities resemble those of a viral encephalomyelitis, and confirmatory diagnosis is by immunohistochemistry (IHC), virus isolation, and detection of viral nucleic acids.^389,392-395 The characteristic microscopic lesion of Borna disease is the Joest-Degen inclusion body in the neuronal nucleus, but this is not always observed. Virus is detectable easily with monoclonal or polyclonal antibodies. Histopathologic changes are those of a typical polioencephalomyelitis, with a particular loss of neurons in the hippocampus. There is involvement of the gray matter of the olfactory bulb, basal cortex, caudate nucleus, thalamus, hippocampus, and medulla oblongata.

No antiviral agents are available for BDV, and use of the amantadine sulfate (developed for treatment of influenza infection) is controversial.^396-398 Likewise, vaccination as a protective strategy is controversial. It is widely assumed that a modified live vaccine would offer more protection; however, the live vaccine available until the early 1990s in Germany was removed from the market because of questionable efficacy.³⁹⁶

Controversy also surrounds the zoonotic potential of BDV. Mental health patients, in particular schizophrenics, have been associated with infection through serologic testing.^399-401 However, normal unaffected humans have also been found to be antibody positive. Thus the causality of this virus and the neurologic dysfunction, mainly characterized by altered behavior and perception in people, remain to be firmly elucidated.

Hendra/Nipah Viruses

In 1994 an outbreak of a previously unreported illness occurred in horses and humans in Hendra, a suburb of Brisbane, Australia.^402,403 A single mare brought into a stable died 2 days after arrival. In the following 2 weeks, 17 horses at the same facility developed respiratory and neurologic signs; several died or were euthanized. Two people that had close contact with the original mare also developed a flulike illness; one survived, but the other died from multiorgan involvement and cardiac arrest. Originally classified as a morbillivirus, Hendra virus was found to be similar to Nipah virus, was isolated from tissues of affected horses and people, and was believed to be the etiologic agent, as proved by development of similar disease in healthy horses after challenge with tissue homogenates from affected animals.^402,404,405 Nipah virus primarily causes respiratory disease in pigs and respiratory and neurologic disease in humans.^404,406 Infection of horses with Nipah virus has been indicated by serologic testing only in a few horses and in the brain of one horse. These two viruses occupy a new genus, Henipavirus, family Paramyxoviridae, subfamility Paramyxovirinae, order Mononegavirales.

These viruses are primarily bat viruses (genus Pteropus, fruit bat), with approximately half the bats testing positive in an endemic focus, whereas other mammalian hosts are comparatively seronegative.^407,408 Transmission is thought to be by direct contact of horses, likely with secretions such as urine, aborted fetuses, and other reproductive fluids.^407-409

The predominant clinical signs of equine hendravirus infection are those of respiratory disease: tachypnea, dyspnea, frothy nasal discharge, tachycardia, fever, anorexia, and death.^405,410 Signs of neurologic disease that may accompany the respiratory signs include dullness and ataxia.

Sera from infected horses are positive for antibodies to the virus.

After infection by direct contact, Hendra/Nipah virus appears to localize in the respiratory tract epithelium.^{404,405,411,412} Experimentally, however, infections can occur by all routes, resulting in respiratory shedding of the virus. As with other paramyxoviruses, there is widespread dissemination of the virus in lung, liver, kidney, lymph nodes, and blood after infection. Infection primarily involves the vascular endothelial cells with pulmonary edema, although CNS pathology is similar to that caused by distemper or measles.

The infection results in an interstitial pneumonia with focal necrotizing alveolitis, subpleural edema, and syncytium formation in the vascular endothelium.^403,410

Treatment is supportive and symptomatic care only. No specific agents have been shown to combat the Hendra/Nipah viruses.

California Serogroup Diseases

Bunyaviruses are mainly mosquito-transmitted viruses and are found worldwide. Some of these viruses are known to be transmitted by Culicoides flies as well.^361,413-427 Approximately 12 serotypes of these viruses, collectively known as the California serogroup, have been isolated in North America, South America, Africa, Europe, and Asia. The viruses have various names, depending on location or host and possibility for equine infection, including snowshoe hare, Jamestown Canyon, Cache Valley, and Main Drain. Transmission is mainly by Aedes mosquitoes.

Clinical signs are consistent with encephalitis and include ataxia, weakness, stiff neck, head pressing, and dysphagia. Systemic signs include tachycardia and fever. Horses that recover generally do so in 1 week.

Encephalosis

Th virus for encephalosis is closely related to bluetongue virus and African horse sickness (AHS) virus and is an Orbivirus.^428,429 Culicoides species transmit this virus to horses of South Africa, Kenya, and Botswana.^429-432 Because this virus is morphologically similar in cell culture to AHS virus, correct identification of this virus sometimes is not made. There is widespread subclinical exposure of this virus to horses throughout endemic regions, and several different serotypes exist.

Although subclinical disease is the most likely manifestation of encephalosis, the virus was originally named for isolation from a horse with typical signs of encephalitis.⁴³³ In addition to signs of acute encephalitis, horses will demonstrate systemic signs of fever, depression, and facial edema. Cardiac failure has been described, and pregnant horses may abort.

Correct testing that is group specific for encephalosis virus, but not for AHS or bluetongue virus, should be performed on serum from affected horses.^428,434-437 Final, definitive diagnosis is difficult even postmortem; cerebral edema, enteritis, cardiac myodegeneration, and fibrosis are seen.

There is no recognized treatment or prevention for encephalosis.

Epidemiology

The life cycle of WNV consists of transmission by hematophagous mosquitoes to an avian host; with amplification occurring in both these hosts.⁴⁵⁵ For maintenance, the virus must be cycled between biologic vectors, and presumably, vertical transmission occurs within the vector for season-to-season maintenance.⁴⁵⁶ Passerine birds are likely the most important nonvector reservoir hosts. Horses and humans are dead-end hosts because they do not amplify the virus in quantities sufficient to infect mosquitoes.

Other modes of transmission include oral infection of both avian and mammalian hosts, most notably predator birds and felids. Birds can shed WNV both orally and cloacally.^457-461 Virus is high enough in the blood of humans to be borne through blood and organ contamination by subclinical WNV-positive donors and can be transmitted vertically through placenta and milk.^462-468

Initial manifestations of WNV disease occurred in humans and were characterized as flulike or febrile illness.⁴³⁸ Subsequent outbreaks of illness were focal and sporadic, with children more susceptible to actual disease.⁴⁵⁵ In endemic areas, most humans and horses demonstrate increased risk of spontaneous seroconversion with age.^{457,461,469,470} In the last decade, WNV outbreaks began increasing in both frequency and severity, with neurologic disease and higher mortality in birds, horses, and humans. Before 1999, WNV caused focal intense outbreaks in horses in the Middle East and Europe.^461,470-472 The first year the virus occurred in New York, several dozen cases of equine WNV occurred by 2001. The outbreak spread westward and reached epizootic proportions, infecting large numbers of immunologically naive horses.^473-475

Equine disease in the United States caused by WNV peaked in 2002 with the report of 15,257 cases that year (U.S. Department of Agriculture [USDA], 2003). Birds have developed natural immunity to WNV, so relatively fewer human and equine cases have been reported, especially in the Northeast and Southeast. However, overt WNV clinical disease still occurs. There were 3000 human cases and 119 human fatalities from WNV reported to the Centers for Disease Control and Prevention (CDC) in 2005. The average number of equine cases has been about 1000/year. As of January 2006, the official CDC count of equine WNV was more than 23,000 U.S. cases (Jennifer Lehman, personal communication). Horses with clinical signs of WNV disease reportedly have a 28.4% to 38% mortality rate.^473,476,477

By 2005, WNV had been identified in all the 48 continental U.S. states.⁴⁷⁸ Canadian provinces reporting disease include Quebec, Ontario, Manitoba, Saskatchewan, and Alberta, with New Brunswick and Nova Scotia reporting evidence of WNV-positive birds.⁴⁷⁹ In Latin America, serologic evidence of WNV has been detected in the Dominican Republic, Mexico, Guadeloupe, El Salvatore, Puerto Rico, Cayman Islands, Jamaica, Belize, and Cuba.^{461,470,480-484} The incidence of equine and human disease appears low for Central America, South America, and the Caribbean compared with the United States.⁴⁸⁵ A decrease in the virulence of the North American strain has been reported, however, and other flaviviruses occur in Central and North America. Cross-protective immunity may also account for the decreased incidence in these locales.

The Culex mosquito is considered the most important vector of WNV in North America, although the virus has been detected in more than 60 species of North American mosquito.^{461,462,486-489} Culex pipiens—positive pools are most frequently detected in the northeastern United States, with Cx. tarsalis constituting the majority of positive pools in the West, and Cx. quinquefasciatus and Cx. nigripalpus in the Southeast.^486,490-503 In the Southwest, epidemics in 2004 were associated, in decreasing frequency, with Cx. quinquefasciatus, Cx. tarsalis, and Cx. pipiens.

Little is known regarding the actual vector of transmission to the horse itself. Blood meal analysis supports Cx. pipiens as primarily avian feeders, whereas mammalian feeders include mainly Anopheles quadrimaculatus, Coquillettidia perturbans, and Aedes albopictus. Recent analysis of Cx. quinquefasciatus has detected both human and bird feeding, and Cx. salinarius reportedly has widest range of host feeding.⁵⁰⁴ Culex mosquitoes require a blood meal of approximately 10⁵ to 10⁷ PFU/mL of virus for 30% to 100% of mosquitoes to obtain infection with WNV, respectively.⁵⁰⁵ Experimental infection of horses by mosquito transmission studies is accomplished with Aedes aegypti and A. albopictus.^506,507

In ticks, transtadial transmission has been inconsistently documented in Ixodes species.^508,509 Under experimental conditions, Carios capensis transmitted WNV to ducklings, and Ornithodoros moubata transmitted WNV to mice.^509,510

More than 300 species of birds have been reported “WNV positive” in the United States, with 16 new species identified during the 2005 season.⁵¹¹ Passeriformes and Charadriiformes obtain the highest titers with the longest persistence and viremia, and tissue levels can consistently exceed 10⁸ PFU/mL.⁴⁶⁰ The house sparrow is considered the most important amplifying host.⁴⁶⁰ Although susceptible to fatal infections, corvids develop very high viremia and still are likely efficient reservoirs.^{457,459,460,512} Corvid susceptibility to WNV is notable in the North American outbreak. Early studies with the Egyptian WNV strain, however, produced high mortality in crows.^513,514

With its vector requirements, WNV in horses and humans is seasonal in temperate regions (with expected year-round activity in subtropical regions).^{497,504,515,516} Late September and October are times of peak incidence of disease, with July representing the start of intense WNV activity. Temperature-dependent spatial modeling supports these disease dynamics, with risk increasing from 25% in late August to more than 75% by the second week of September.^517-519 A drop in ambient temperature usually results in a rapid decrease in reporting activity.^520,521

Age is a factor in susceptibility to neuroinvasive disease, as reported for both people and horses.^{461,472,522-525} Although men are more frequently affected with neuroinvasive disease, there appears to be no breed or gender predilection in the equine. In fact, in one study examining outcome parameters, female horses were 2.9 times more likely to die than male horses with neurologic signs.^522-524 This may reflect pasture-associated exposure with broodmares or longer lifespan due to reproductive value.

Multiple free-ranging and domesticated mammals, including the big brown bat, little brown bat, eastern chipmunk, eastern gray squirrel, eastern striped skunk, white-tailed deer, and brown bear, have demonstrated spontaneous seroconversion to WNV.^{461,492,505-511,526} Neurologic disease has been diagnosed in WNV-positive gray squirrels and fox squirrels, and febrile disease has been described in cats.^527,528 Oral infection has been induced in experimental infection of cats. Both farmed and free-ranging alligators can obtain extremely high viremia and may be an important reservoir in the Southeast.⁵²⁶ In farm-raised alligators, cloacal shedding has been demonstrated, with oral infection likely. Serologic evidence of infection has been demonstrated in domestic dogs. Llamas have been reported to develop neurologic disease.^529,530

Pathogenesis

The syndrome of WNV in the horse consists of neurologic signs, with abnormalities in mentation, the spinal cord, and cranial nerves, primarily of the midbrain and hindbrain.^474,531-540 Virus localization in the motor neurons of the thalamus, medulla, and pons likely accounts for changes in behavior.^532,541,542 Change in consciousness occurs frequently, likely caused by lesions in the midbrain and rostral pons affecting the reticular formation.^540,543 The reticular formation projects to the thalamus, which in turn sends diffuse projections and is the source of cholinergic stimulation to the entire cerebral cortex.⁵⁴⁰ Thus, disturbances of the reticular formation and the midbrain may induce the behavioral changes observed, which range from aggression to somnolence.* Spinal deficits are characterized as multifocal, asymmetric, with weakness and ataxia likely reflecting direct infection of the spinal cord, interruption of motor tracts in the hindbrain, and loss of fine motor control through infection of the large nuclei of the thalamus and basal ganglia.^546,547 Loss of fine motor control is evidenced by involuntary skin and muscle fasciculations, tremors, and hyperesthesia, and a predilection for the basal ganglia likely results in Parkinson-like syndrome. Infection in the pons and medulla oblongata can explain clinical deficits of cranial nerves (CNs) VII, XII, and IX.⁵⁴⁸

It is unknown how WNV invades the central nervous system (CNS); WNV is proposed to cause minimal viremia, with replication in the lymph nodes, followed by invasion into the CNS across the blood-brain barrier.⁴⁴⁶ Alternatively, there may be transaxonal transmission.^438,519 In rodent models the virus is capable of invading the CNS after peritoneal challenge, demonstrating a primary predilection for neural tissues.^549,550 Viremia occurs 3 to 5 days after experimental challenge and lasts 24 to 72 hours, with dissemination into the CNS at 4 to 6 days after inoculation. Intrathecal infection of horses results in clinical signs 7 to 10 days after inoculation. WNV directly infects nerve cell bodies, and in rodents, initial replication occurs in the basal ganglia, with dissemination later to cortex, cerebellum, and hippocampus. The large neurons of the ventral or anterior horns are infected late in infection, and in mammalian hosts, neuronal viral load is low. As noted earlier, cell lysis occurs with viral replication, but WNV also induces apoptosis in neurons, which likely accounts for most of the pathophysiology.^549,551

Clinical Findings

When infected with lineage type I WNV, horses develop neurologic disease, whereas infection with lineage type II viruses is subclinical in nature.^† Both systemic and neurologic abnormalities are observed in horses. Systemic clinical signs occur initially and include a mild to moderate increase in rectal temperature (38.6° C to 40°C; 101.5° F to 104.0°F), anorexia, and depression. Presenting complaints can be insidious and can manifest as abdominal pain or lameness, including a bizarre gait, as in rabies infection in the horse. The occurrence of neurologic symptoms is frequently sudden and progressive, and the exact course of disease in any one animal is unpredictable. The most unique manifestations of equine WNV encephalomyelitis are changes in personality and development of fasciculations. Periods of hyperexcitability and apprehension are common and can be severe, to the point of aggression. Change in personality is manifest by a quiet horse becoming hyperexcitable and an abnormally aggressive horse becoming submissive. Some horses develop attention deficits, even with bouts of sleeplike behavior during activity, resembling cataplexy or narcolepsy. Fasciculations, although not pathognomonic, are so notable that many clinicians base their preliminary diagnosis on their occurrence. These tremors usually involve the face and neck muscles but can involve all four limbs and the trunk so that normal activities such as walking, eating, and interaction with handlers and other horses are interrupted. Rapid blinking of eyelids is common (and stimulated with a penlight), and horses likely are photophobic.

One of the initial signs of motor abnormality is bradykinesia, characterized as a short, slow-stilted gait described by observers as “lameness,” with laminitis being a common differential at this stage. Spinal abnormalities are characterized by ataxia and paresis that can be highly asymmetric or can involve only one or both forelimbs or hindlimbs. Cranial nerves are frequently abnormal, with weakness of the tongue, muzzle deviation, and head tilt most common. The tongue abnormalities can also be associated with dysphagia and choking. Abnormalities of the cauda equina also occur and consist of stranguria and rectal impaction. All these clinical signs are variable, and after initial signs abate, about one third of clinically affected horses experience an increase in severity of clinical signs within the first 7 to 10 days of onset. Overall, about 30% of the horses progress to complete paralysis of one or more limbs. Most of these horses are euthanized for humane reasons or die spontaneously.

In many horses, clinical signs generally improve within 3 to 7 days of displaying onset. After 3 to 5 days, horses that are recovering or stabled may exhibit a sudden recurrence of signs. Clinical signs may be of short duration, or horses may become suddenly recumbent and either die or recover only with prolonged treatment. Horses that become recumbent often need aggressive supportive care. Once the horse has demonstrated significant improvement, full recovery within 1 to 6 months can be expected in 90% of patients.⁵⁵⁴ Residual weakness and ataxia appear to be the main problems; however, personality changes were reported as well.⁵⁵⁴ In humans, some patients have experienced long-term inflammation of the meninges, and others have experienced the long-term loss of the use of one or more limbs. In addition, mild to moderate, persistent fatigue on exercise and chronic headache have also been noted.

Diagnosis

In general, complete blood count (CBC) and serum biochemistry profiles of WNV horses are normal.⁵³¹ Horses may present with a mild absolute lymphopenia, elevated muscle enzymes, and hyponatremia, which has also been described in humans with viral encephalitis. Cerebrospinal fluid cell counts are normal to elevated and mononuclear in nature, with protein levels also normal to elevated.⁵⁵⁵

Confirmation of WNV infection with encephalitis in horses begins with assessment of whether a horse meets the case definition based on clinical signs occurring in an area in which WNV has been confirmed in the current calendar year in mosquito, bird, human, or horse.⁴⁷³ Serologic testing relies on the development of immunoglobulin M (IgM) and neutralizing-antibody responses in acute-phase serum, as tested by an IgM-capture enzyme-linked immunosorbent assay (MAC-ELISA) and neutralization testing (NT), respectively. In serologic confirmation of flaviviruses and most arboviruses, the neutralization format is considered the “gold standard” because of the specificity of neutralizing antibody generated in response to these viruses. Until fall 2001, a positive IgM titer of 1:400 or higher and a plaque reduction neutralizing-antibody test (PRNT) titer of 1:10 or higher on a single serum was considered confirmatory for serodiagnosis of WNV encephalomyelitis in a horse exhibiting appropriate clinical signs. Since 2001, reliance on the PRNT for serologic confirmatory diagnosis in horses has diminished because of the availability of WNV vaccination for horses. Vaccination induces formation of neutralizing antibody and likely confounds interpretation of the PRNT. Most diagnostic laboratories use MAC-ELISA for confirmation of WNV disease (because consistently detected IgM rarely occurs after vaccination) in the horse, and the sensitivity and specificity of this test are approximately 81% and 100%, respectively.⁵⁵⁶

Other means of confirmation include postmortem detection of WNV by PCR, culture, and IHC in CNS tissues. Several methods of WNV nucleic acid testing in equine tissue have been described, including nested PCR targeting the E protein, which demonstrated sensitivity for relatively low viral load, and real-time PCR. The E-protein target appears less sensitive; however, the NS5 target has detected WNV nucleic acids in CNS tissues, heart, and intestine of clinically affected horses.

Differential Diagnoses

Even if WNV is suspected based on time of year and vaccination history, other infectious CNS diseases should be considered in the differential diagnoses, including alphaviruses, rabies virus, equine protozoal myeloencephalitis (EPM), equine herpesvirus type 1 (EHV-1), botulism (less likely), and verminous meningoencephalomyelitis (Halicephalobus gingivalis, Setaria, Strongylus vulgaris). Alphaviruses, rabies, H. gingivalis infection, hepatoencephalopathy, and leukoencephalomalacia are rapidly progressive cortical signs characterized by behavioral alterations, depression, seizure, and coma. The appearance of seizure and coma is rare in WNV horses but can occur. Cranial nerve signs in EEE and WEE are also common, including head tilt, pharyngeal/laryngeal dysfunction, and paresis of the tongue. The incidence of WEE in horses is fairly low in the United States, but mortality and severity of clinical signs would more closely resemble WNV. Differentiation from rabies is problematic because clinical signs in horses with rabies frequently include ataxia, weakness, or gait abnormalities. Although there are periods of somnolence, blindness, and some cranial nerve deficits, WNV horses appear to become rapidly recumbent or stabilize over several days, as opposed to rapid deterioration with EEE, rabies, and H. gingivalis. Spinal disease caused by EPM is a more difficult differential if horses with WNV are not febrile and do not exhibit excessive muscle fasciculations. Noninfectious causes to consider include hypocalcemia, tremorigenic toxicities, hepatoencephalopathy, and leukoencephalomalacia.

Serum titers should be evaluated for recent exposure to other encephalitides, including EEE, WEE, and EHV-1. An IgM-capture format is appropriate for EEE and WEE, but for the NT format, paired titers are necessary in vaccinated horses. Because WNV can present with asymmetric weakness and ataxia, Western blot testing for EPM should also be performed on CSF.

Pathologic Findings

Flaviviruses cause polioencephalomyelitis (inflammation of gray matter), with lesions that increase in number from the diencephalon through the hindbrain and frequently increase in severity caudad in the spinal cord.* Gross pathologic findings are limited in WNV infection in the horse. The meninges may be congested. Small to moderate-sized foci of hemorrhagic discoloration can be observed in the brain and spinal cord. The most common areas observed in the brain include the basal ganglia, rostral colliculus, and both pons and medulla. The most common sign of spinal changes appears in the lumbar cord. Edema and softening of tissues are also common.

Histopathologic changes are consistent with viral infection and neural cell death. The basal ganglia, thalamus, pons, and medulla have the highest numbers of lesions, characterized by two to several cell layers of mononuclear perivascular cuffing. Predominantly confined to the gray matter, there are also collections of mononuclear cells within the parenchyma (gliosis). By contrast, this is limited in the cortex and cerebellum. Neuronal damage includes chromatolytic neurons and neuronophagia. In long-standing disease, areas of neuronal dropout may be seen. In the spinal cord, there is perivascular cuffing, gliosis, and damaged neurons. The inflammation associated with the neuropil is confined mainly to the gray matter.

Treatment

To date, no antiflavivirus compounds are marketed. Therapy is supportive for these infections, although several experimental inhibitors of RNA virus replication are under development.^558-563 Accurate assessment of the direct effect of any pharmacologic intervention in WNV disease is difficult because horses can begin the recovery process within 72 hours of onset of clinical signs. Flunixin meglumine (1.1 mg/kg every 12 hours IV) early in the course of the disease appears to decrease the severity of muscle tremors and fasciculations within a few hours of administration. To date, much of the mortality in WNV horses results from euthanasia of recumbent horses for humane reasons rather than spontaneous fatality. Recumbent horses are mentally alert and frequently thrash, sustaining many self-inflicted wounds and posing risk to personnel. Therapy of recumbent horses is generally more aggressive and may include dexamethasone sodium (0.05 to 0.1 g/kg every 24 hours IV) and mannitol (0.25 to 2.0 g/kg every 24 hours IV). Detomidine hydrochloride (0.02 to 0.04 mg/kg IV or IM) is effective for prolonged tranquilization. Low doses of acepromazine (0.02 mg/kg IV or 0.05 mg/kg IM) provide excellent relief from anxiety in both recumbent and standing horses. Until EPM is ruled out, prophylactic institution of antiprotozoal medications is recommended. Other supportive measures may include oral and intravenous fluids and antibiotics for treatment of infections that frequently occur in recumbent horses (wounds, cellulitis, and pneumonia).

Interferon-alpha is a relatively common therapy,^560,564,565 based on differences in genetic loci to immune response elements that induce nonspecific immunity in mice. The success of IFN-α rests on anecdotal reports in the human and veterinary literature. Intravenous WNV-specific immunoglobulin therapy has also been described, and in a blind controlled placebo trial with low numbers of animals, the risk for development of recumbency was lower. Otherwise, outcomes and severity were the same. In human therapy, high-dose glutamate has been pursued to prevent neuronal cell death.⁵⁶⁶ Another experimental investigation in mice involves β-lactam inhibitors that stimulate GLT1, a chemical that activates glutamate.^567,568

Prevention

Initial epidemiologic studies indicated a point source for WNV infection, demonstrating that the outbreak could be controlled by vaccination.^{474,476,477,544} Presently, three vaccines are licensed for prevention of WNV viremia in the United States. Vaccination before the mosquito season is optimal. Two of these vaccines, a killed product and a recombinant product based on the canarypox vector, require an initial injection followed in 3 to 6 weeks with a booster injection.⁵⁶⁹ Most manufacturers recommend more frequent vaccination in areas with year-round mosquito seasons. However, the recently licensed third vaccine demonstrated 12 months of immunity against severe intrathecal challenge with WNV. Limited information is available regarding long-term immunity with the other vaccines, given that the duration of immunity was tested using the mosquito model of WNV infection, which results in seroconversion and viremia as a measure of efficacy. Horses that have recovered from clinical WNV have long-term immunity against WNV and should not require immunization.

West Nile virus is a zoonotic in that a bird reservoir maintains the virus in an endemic life cycle in the environment, with transmission by mosquito to humans. There is little risk of disease by direct contact with a horse. However, the ecology of horse pastures and stables with standing water, a high degree of biologic debris, and “bridge” vectors that feed on mammalian populations pose a risk to people. The same type of management tactics for prevention are important for humans, except that there is no vaccine and deet-based products are recommended for protection. The North American epidemic of WNV has demonstrated new modes of transmission involving blood-borne and occupational risks. Blood-borne transmission can occur between a viremic host and accidental needle injection. In addition, occupational infection has occurred through necropsy of avian hosts. Veterinarians and horse owners should institute personal protection with appropriate clothing, gloves, and eye protection when coming in contact with animal tissues during the arbovirus season.

Definition and Etiology

Louping ill is an acute, fatal encephalomyelitis of sheep that occasionally infects humans, wild ruminants, horses, and cattle.^570-572 The disease has been reported in England, Scotland, Ireland, Norway, Turkey, and Bulgaria.^573-575 The etiologic agent of louping ill is a neurotropic, single-stranded RNA virus (flavivirus) that is transmitted by primarily the tick Ixodes ricinus, as well as by Rhipicephalus appendiculatus, Ixodes persulcatus, and Hemaphysalis anatolicum. Most outbreaks occur in swampy areas with dense populations of infected ticks and wild animals. Infection through blood contamination of hypodermic needles, other fomites, and blood products has been reported.⁵⁷⁶ Minor genetic variations occur among viruses from different geographic areas, forming four subtypes.⁵⁷⁷ A similar disease of sheep in Spain is caused by a genetically distinct flavivirus.⁵⁷⁸

Clinical Signs

Sheep develop central nervous system (CNS) disorders. Infection in horses often is asymptomatic.⁵⁷⁹ The initial clinical signs of louping ill include fever, anorexia, depression, constipation, and generalized muscular tremors. The ensuing signs are characteristic of CNS disease and include ataxia, conscious proprioceptive deficits, head tremors, hypermetria, and hyperexcitability. The hypermetria results in a characteristic “bunny hopping gait,” which gives the disease its name. Further progression of clinical signs is associated with cerebrocortical dysfunction; these signs include head pressing, hyperesthesia, recumbency, convulsions, coma, and death. Survivors have residual neurologic deficits. The duration of the illness is approximately 12 days.

Clinical Pathology

Both hemagglutination inhibition and complement fixation tests can be used to detect virus in infected animals.⁵⁷⁶ High levels of virus-specific IgG and IgM can be detected in the cerebrospinal fluid (CSF) of affected animals.⁵⁸⁰ Viremia peaks at approximately 3 days after inoculation and disappears by 7 days. Because animals usually are not viremic at the time the nervous system lesions develop, virus recovery is best done from the brain or spinal cord. Isolation of the louping ill virus from the CSF is difficult.

Pathophysiology

Ticks become infected when feeding on a viremic host. The virus survives in the salivary glands of the tick and can overwinter here, being transmitted to a new host when the tick becomes active the following year.⁵⁷⁶ After inoculation into susceptible sheep, the louping ill virus migrates into the regional lymph nodes and spleen and then replicates. Viremia occurs 6 to 20 days after the invasion of the lymphatic tissues.⁵⁷¹ Viral replication in the brain causes nonsupportive inflammation and neuronal degeneration.⁵⁷¹ Rapid antibody production is associated with recovery.⁵⁸¹ Concomitant infection of sheep with the agent of tick-borne fever (Erlichia phagocytophila) results in a greater level of viremia and increased mortality from the louping ill virus.⁵⁸²

Epidemiology

Louping ill principally affects yearling sheep in the spring. Maternal immunity wanes when lambs are about 3 months old. Infection develops weeks to months after sheep have been placed on pastures infected by Ixodes ricinus. In any outbreak the prevalence of clinical louping ill is low, but the seroprevalence of antibodies in adult sheep in endemic areas is high, indicating a continuous, low-level exposure to the virus, and many animals are asymptomatically affected. Factors such as climate, tick population, and immune status of the flock all play a role in the severity of an outbreak.⁵⁷⁶ The case-attack rate may reach 60% of the population, whereas the mortality rate is low, rarely exceeding 15%. The degree of susceptibility of neonatal lambs and adult sheep is similar. Adults tend to be more heavily parasitized by the host ticks and thus play a major role in virus survival and transmission.⁵⁸³ Pigs, cattle, horses, and red deer also become infected with the virus and can develop similar clinical disease. The seroprevalence rate of the virus in horses from one endemic area was 10%.⁵⁷⁹ Milk from infected goats can reach titers high enough to infect suckling kids.⁵⁸⁴

Sylvatic cycles of virus transmission occur in which viral amplification takes place through certain wild mammals and red grouse.^570,585,586 The hare, in particular, appears to play a major role in the propagation and persistence of virus in the red grouse population.^587,588 Certain other small mammals, such as field voles, however, do not have a role in this cycle of virus persistence.⁵⁸⁹ In the case of grouse, it has been demonstrated that the host can be infected by ingesting the infected tick, rather than by being bitten by the tick.⁵⁹⁰

Mild occupational infections can occur in shepherds, veterinarians, and laboratory workers who cultivate the virus. The louping ill virus probably is maintained on pastures through infected sheep because grouse populations become unstable and die out whenever the virus is introduced.⁵⁹¹ The high incidence of louping ill in the spring and summer months probably corresponds to the peak activity of ticks. The virus may persist for long periods in the arthropod vector, but it is unclear if transovarial transmission of the agent occurs. Ixodes ricinus is a three-host tick with a life cycle of 3 years. The ticks do not walk, and dissemination over a range requires animal transport. After a blood meal, the tick molts and then rests on vegetation until the next meal, approximately 12 months later. The activity of the ticks tends to increase greatly in the spring whenever the ambient temperatures are above 7°C (43°F).

Necropsy Findings

Gross lesions are absent at necropsy. The histologic lesions of louping ill include perivascular cuffing with mononuclear cells and neutrophils, gliosis, neuronal necrosis, and mononuclear cell meningeal inflammation.^592,593 Microscopic lesions are most severe in the Purkinje cells, the motor nuclei, and the ventral horn cells. The forebrain is spared.⁵⁹⁴ Virus in tissue can be detected by virus isolation, IHC, and RT-PCR.^576,595,596

Prevention

There is no treatment for louping ill encephalitis, but supportive care should be provided.

A formalin-inactivated vaccine given in the last trimester has been recommended for preventing louping ill.⁵⁹⁷ A single dose of the vaccine provides responses that are protective for at least 2 years. Colostral antibody titers higher than 1:40 are considered protective.⁵⁸³ Other methods of preventing the disease include frequent acaricidal dipping of sheep and clearing of pastures to reduce the population of infected intermediate hosts.

Louping ill has not been detected in the United States, but it is a reportable disease. Any suspicion of this disease should be reported to state and federal authorities. The disease also has a zoonotic potential; infection of humans can occur through tick bites, infected sheep or goat milk, or fomites.

Definition and Etiology

Infection with rabies virus results in severe and usually fatal neurologic disease. Most mammals are susceptible and are infected through bites from other animals during or near the clinical phase of illness. The rabies virus and other highly neurotropic “rabies-related” viruses belong to the genus Lyssavirus, family Rhabdoviridae. (bullet-shaped RNA viruses). Wildlife provides a natural reservoir for rabies, and each rabies strain is maintained in particular reservoir host(s). Although they can readily cause rabies in other species, wildlife-adapted strains of rabies usually die out when passed into species to which they are not adapted. Other, “rabies-related” lyssaviruses can cause a neurologic disease identical to rabies. They include the Lagos and Duvenhage viruses of bats in parts of Africa, the Mokola virus of rodents and shrews in Africa, and the European and Australian bat lyssaviruses.^598-600

Clinical Signs

Livestock cases of rabies have been reported mostly in cattle and occasionally in horses, sheep, and goats.^600,601 Rabies should be included in the differential diagnosis of any unexplained acute, rapidly progressive encephalitis, especially in the presence of autonomic instability, dysphagia, hydrophobia, paresis, or paresthesia.^600,602,603 The incubation period ranges from a few days to 6 months, depending on the pathogenicity of the inoculum and the distance from the site of inoculation to the brain. The shortest incubation periods are seen in animals bitten on the head, whereas the longest incubations have been reported after bites on the extremities of the pelvic limbs.

The disease is usually fatal after a clinical course of 1 to 8 days. Occasionally, an animal may be found dead without any previously observed clinical sign. Early clinical signs of rabies in livestock may be nonspecific sensory and behavioral changes, including anorexia, restlessness, depression, separation from the herd, and mild ataxia. Rumination may stop. Horses often appear colicky. Affected animals may repeatedly attempt to mount inanimate objects or show regional paresthesia with pruritus. Rubbing of the pruritic area results in loss of hair or wool and skin ulcerations. As the disease progresses, animals may become hyperexcitable, fearful, or enraged (furious rabies) or mentally depressed (dumb rabies). Furious and dumb states may alternate in the same animal. Cattle rabies frequently manifests as dumb rabies with flaccid paraparesis, tetraparesis, and tetraplegia (paralytic rabies) as the disease progresses.

In an experimental study, all cattle and sheep infected with the rabies virus exhibited excessive salivation and behavioral changes, and the majority also displayed tremors, bellowing, aggression, hyperesthesia, and pharyngeal paresis/paralysis.⁶⁰⁰ Tetraplegic animals may show frantic motor activity of the legs and bellow when stimulated. Animals with dumb rabies are depressed, inappetent, and usually febrile (>39.4°C [103°F]) and may have a drooped head and neck, ptosis, flaccid facial musculature, profuse salivation, yawning, repeated nibbling motions with the lips, tenesmus, paraphimosis, odontoprisis, head pressing, circling, wide-base stance, difficulty rising, and falling episodes. Other clinical signs associated with dumb rabies include flaccidity of the tongue, tail, anus, and urinary bladder and blindness, strabismus, and nystagmus.

The first clinical sign of paralytic rabies in horses may be an unexplained ataxia or shifting leg lameness, soon followed by paraparesis or paraplegia.⁶⁰⁴ Spinal reflexes and tone in the affected limbs may be decreased or absent. Most affected animals become recumbent in 3 to 5 days. Initially the recumbent animal may be able to eat and drink with help, but it soon becomes anorectic and develops encephalopathic signs, followed by coma and generalized seizures^605,606 (Fig. 35-4).

Fig. 35-4 Ten-month-old mixed-breed calf with terminal rabies virus encephalitis. The calf is comatose and showing opisthotonos.

Regardless of the clinical manifestations, rabies is rapidly progressive and fatal disease leading to cardiorespiratory failure, and death usually occurs within 10 days. In all disease forms of livestock, the clinical signs rapidly worsen over 1 to 3 days, until the patient becomes recumbent and comatose. Animals may develop a pharyngeal-laryngeal paralysis resulting in stertorous breath sounds, inability to drink (thus the common name of “hydrophobia”), and accumulation of frothy saliva at the commissures of the lips.⁶⁰⁷

One summary of 21 cases of rabies in horses enumerated the frequency of particular clinical signs seen as the presenting complaint (Table 35-5). Rabies can only be differentiated from other encephalitic disorders by specific postmortem tests or virus isolation, which is usually not possible before death occurs. In horses the clinical signs of rabies often are indistinguishable from other encephalopathies, such as hepatoencephalopathy, leukoencephalomalacia, togaviral encephalitis, equine herpesvirus type 1 (EHV-1), protozoal and other meningoencephalomyelitides, and space-occupying masses. In ruminants, rabies may resemble herpesv.irus, thromboembolic meningoencephalomyelitides, nervous ketosis, grass tetany, polioencephalomalacia, nervous coccidiosis, or even focal spinal cord or peripheral nerve diseases. Most horses die of fatal encephalopathy within 5 days of the onset of signs; but cases lasting as long as 14 days have occasionally been observed.⁶⁰⁸

Table 35-5 Occurrence of Particular Clinical Signs of Rabies in Horses

From Green S et al: Rabies in horses: 21 cases, J Am Vet Med Assoc 200:1133, 1992.

Clinical Pathology

There is no valid antemortem test to diagnose rabies in livestock. Rabies usually remains immunologically and serologically silent throughout the incubation and clinical phase of the illness.²⁴⁴ Clinicopathologic tests are not specific for rabies but may help rule out other, more common diseases. The CSF may be normal or may show moderate mononuclear pleocytosis (5 to 30 cells/μL) and increased protein (60 to 200 mg/dL), occasionally with neutrophils, eosinophils, or xanthochromia reported.

Pathology and Postmortem Diagnosis

Rabies causes a nonsuppurative meningoencephalomyelitis. Microscopic changes include brain edema, meningeal congestion, focal areas of hemorrhage, perivascular cuffing, gliosis, neuronophagia, and neuronal degeneration. These changes are most severe in the dorsal root ganglia and can be seen in most CNS regions.

A definitive diagnosis of rabies may be based on the finding of Negri bodies (not always present, especially in cases euthanized early in the disease course) or through a positive direct or indirect fluorescent antibody (DFA, IFA) test conducted on fresh smears of CNS tissues (thalamus, hippocampus, brainstem, pons, medulla, cerebellum).^609-611 If these tests are negative or inconclusive and there has been human exposure, additional testing can be done using intracerebral inoculation of mice with CNS tissue from the suspect case. An antigen-capture enzyme immunodiagnostic technique has been developed for salivary gland specimens, to capture antigen onto solid phase using purified polyclonal antibodies. The specificity and sensitivity of this method are similar to the direct immunofluorescence test, but it has not gained widespread clinical use in the United States.⁶⁰⁹ The DFA test on brain tissue has become the standard by which the value of other rabies tests is evaluated.⁶¹⁰ In cases of human exposure, the local public health parties should always be contacted before sample collection and submission. The diagnostic pathologist will usually accept the refrigerated head (or even whole body for small animals and bats), or refrigerated brainstem and cerebellum collected through the foramen magnum in mature cattle and horses. Care should be taken to prevent any human exposure to body fluids and CNS tissue of any animal with rabies on the list of differential diagnoses, especially in areas where rabies is known to be present in any domestic or wildlife species; open wounds or small healing sores may provide portal of entry for the rabies virus.

Pathophysiology

The structure of the rabies virus capsid proteins is similar to that of the neurotoxins of cobra venom and acetylcholine.⁶¹² After subcutaneous or intradermal inoculation, the rabies virus replicates locally. After several days, it binds to the acetylcholine (and other) receptors of the peripheral nerves and then migrates retrogradely to the CNS through peripheral nerves, spinal rootlets, and the spinal cord. After entry into the nerve cell rootlets, the virus travels to the brain along nerve tracts and into CSF. From there it spreads centrifugally along the rootlets of the cranial nerves to the salivary glands and the nasal epithelium. Shedding of the virus in the nasal secretions and saliva may precede the onset of clinical signs but not the presence of virus in the brain, where it needs to replicate before it can reach the salivary and nasal secretions. Therefore, whenever an animal bites a person, the attacking animal can always be sacrificed and tested for the potential of rabies virus transmission, and it not necessary to wait for disease progression before testing an animal for rabies, as recommended earlier when Negri bodies were the main diagnostic test for rabies. In humans, most cases become apparent within 1 to 3 months of infection, but up to 10% of cases have had incubation periods in excess of 6 months, extending to several years in rare cases.⁵⁹⁸ The rabies virus replicates in the cell bodies (gray matter) of the CNS. Dysfunction of these neurons results in behavioral changes and variable abnormalities of the cranial and the peripheral nerves, with multifocal loss of lower motor neuron and autonomic function. Death results from cardiorespiratory paralysis as the virus infects the brainstem medullary centers.

Epidemiology

Rabies is a reportable zoonosis that has been detected worldwide, with some exceptions (especially islands). It does not presently occur in Scandinavia, United Kingdom, Ireland, Australia, New Zealand, or Iceland.⁵⁹⁸ The disease is endemic in other parts of the world, including the United States, Canada, and Europe, but remains epizootic in Central and South America, Africa, and parts of Asia, especially where a tropical climate and a high population of infected feral mammals favor viral propagation and transmission.^599,601 The rabies virus is shed in the saliva and does not survive when dried or exposed to ultraviolet (UV) light. The most common method of viral transmission to domestic animals is through the bite of an infected feral or wild mammal. Humans and laboratory animals have also developed fatal infections after respiratory exposure to viral aerosols. Low levels of rabies virus are also shed in the milk of infected animals and may occasionally infect offspring nursing infected dams.^613,614 Laboratory animals, foxes, and skunks also can be infected experimentally by ingestion of infected tissues from rabies cases.^613,614 The consumption of thorny bushes may also have caused horizontal transmission of rabies in an outbreak in African Kudu.⁶⁰⁰ In the United States, survival of the virus in the wild may depend on cycling of infections in skunks, bats, or raccoons, which also are common sources of livestock infections.^615,616

Certain strains of rabies virus have low virulence in skunks and bats, resulting in asymptomatic carriers serving as reservoirs. The rabies virus is also transmitted directly among bats through aerosols in caves. The bats may die off from rabies during periods of high viral contamination. In the United States the most common wild reservoirs are foxes in Alaska and along the border of Mexico, raccoons in the East, and skunks in the central and western regions.⁶⁰¹ Mandatory vaccination of dogs has virtually eradicated canine rabies from developed countries, where rabies transmission from domestic dogs to livestock is rare.^598,617 Island nations have been able to remain free of rabies by imposing a 6-month quarantine on imported dogs and cats. In South America, rabies outbreaks in cattle often result from vampire bat bites, causing up to 50,000 cattle deaths annually. Human rabies cases may result from bites inflicted by infected pets, feral dogs and cats, bats, or wildlife. Cases resulting from contact with infected cattle or horses are rare.^601,617,618

Animal rabies cases are most common in late summer, possibly because wild animals give birth to offspring during the early spring, and young animals become infected and then spread the disease as they expand into new territory under pressure from selective forces (e.g., predation, starvation, hunting).

The various strains of rabies virus are maintained in different hosts. Serial passage of virus from field cases (“street rabies virus”) into laboratory animals can produce a modified, “fixed virus” with loss of virulence for the field hosts and increased virulence for the laboratory species. Such fixed strains have been used in the preparation of vaccine viruses.

Prevention

Although widespread rabies vaccination of livestock is neither economically feasible nor justifiable on public health grounds, vaccination of valuable livestock, or animals that travel or are in regular contact with humans (e.g., petting zoos, animal shows) should be considered, especially in or near rabies epizootic areas. A list of licensed rabies vaccines for the protection of animals against rabies is regularly updated by state veterinary authorities.⁶¹⁹

The disposition of livestock that have been bitten by rabid animals depends on the animal’s vaccination history, local and national regulations, and the value of the bitten victim. In the United States and Canada, the disease must be reported to the state public health department. Management of exposed livestock must be coordinated with public health officials. Any livestock bitten by a wild animal should be considered to have been exposed to rabies, regardless of the availability of the biting animal for testing. If a bitten horse had been vaccinated before the bite, it should be revaccinated immediately and kept under observation for 90 days. Other exposed, unvaccinated animals with a low economic value should be euthanized, or if the animal is very valuable, the bite wound may be washed with copious amounts of water and iodine or quaternary ammonium disinfectants, after which the animal should be quarantined for at least 6 months and the brain examined if it dies.^619,620

Vaccination of animals shortly after rabies virus infection is not recommended because it is less effective than prophylactic vaccination. In one study, postinoculation vaccination of experimentally infected sheep had no effect on the incubation period, clinical signs, or mortality rate. Moreover, the antibody titers of the vaccinated animals had no predictive value for protection.^621,622 Whenever human exposure to rabies virus is suspected, the suspected animal must be euthanized and its brain examined for evidence of infection. A vaccinated or valuable animal may be quarantined for 6 weeks under veterinary supervision, and the brain must be examined if it develops clinical signs suggesting rabies during that period. The World Health Organization (WHO) publishes and regularly reviews its recommendations for vaccination and prophylaxis in humans after exposure or suspected exposure to the rabies virus.⁵⁹⁹

Control of rabies by immunization of susceptible wildlife using baits containing modified live vaccines has been successful in certain cases.^{601,618,623,624} Such approaches may provide a means for large-scale immunization of wildlife or feral animal populations for eventual eradication of the disease.

Clinical Signs

Affected animals show signs of a multisystemic disease. The initial clinical signs in cattle are fever (39°C to 41.5°C [102.1° F to 106.7°F]), anorexia, depression, and stiffness. The cattle also may show signs of a respiratory disease characterized by nasal discharge, dyspnea, and cough. These animals occasionally have a painful response to percussion of the hoof, as well as swelling of the coronary band or polyarthritis and tenosynovitis.⁶²⁶ Auscultatory abnormalities may include high-pitched wheezes and crackles over the lung fields or pleural and pericardial friction rubs. Because of the fibrinous peritonitis and pleuritis, the clinical signs in some affected animals may resemble those of hardware disease. These animals may grunt or groan when sudden pressure is applied to the xiphoid region. Some animals may respond to soft percussion of the xiphoid region by striking or kicking at the examiner. There is an initial diarrhea. Progression of the disease is related to development of the meningoencephalitis and is characterized by ataxia and conscious proprioceptive deficits, circling, head tilt, opisthotonos, hyperesthesia, stiff neck, convulsions, and coma. Animals may die after 4 to 10 days.

Diagnosis

The detection of elementary bodies in the exudate cells of pleural and peritoneal effusions is highly suggestive of SBE. The chlamydial agent can be cultured from the blood and body fluids of early infections in guinea pigs inoculated intraperitoneally with fresh tissue specimens and held for 6 to 7 days. Inoculation of embryonated eggs is a less sensitive diagnostic technique than animal inoculation. The causative organism has been identified as a distinct species, Chlamydia pecorum, by means of DNA analysis, immunologic assays, and serology.⁶³⁵

Pathogenesis and Pathologic Lesions

The mode of infection and the genesis of pathologic lesions are unknown in natural cases. Growth of the chlamydiae in the arteriolar endothelium causes vasculitis, hemorrhage, edema, and accumulation of fluid in the body cavities. Diffuse fibrinous pleuritis, peritonitis, and meningitis also are seen. Microscopic changes include perivascular mononuclear cell infiltration and neuronal degeneration. The lesions all are composed of networks of neutrophils and mononuclear cells enmeshed in fibrin. Some of the inflammatory cells contain elementary bodies.

Treatment

Early cases of SBE can be treated with oxytetracyclines (20 to 50 mg/kg/day for a minimum of 7 days). Efficacy is indicated by a reduction of fever within the first 24 hours of treatment. The chlamydial agent is also susceptible to penicillin and erythromycin, but the clinical efficacy of these drugs is unknown.

Control

There are no known effective control measures for the prevention of SBE. The chlamydial agent is susceptible to a number of disinfectants, including 2% sodium hydroxide (NaOH), 5% cresol, and 0.3% quaternary ammonium compounds.

Definition and Etiology

Meningitis can occur either from direct extension of infectious agents into the calvarium or from hematogenous infection. Causes of suppurative meningitis include direct extension of pyogenic infections into the calvarium from infected skull fractures,⁶³⁹ osteomyelitis from sinusitis or otitis, osteonecrosis caused by thermal cauterization during dehorning⁶⁴⁰ or by improper placement of trephination holes, cribriform plate fractures, and extension from infected coccygeal vertebrae. In horses, septic meningitis is a common sequela to surgical removal of progressive ethmoidal hematomas and also has been reported as a consequence of local spread of infections of the paranasal sinuses, nasal cavity, periocular tissues, and submandibular lymph nodes.⁶⁴¹

Centripetal migration of Cryptococcus neoformans along peripheral nerve rootlets results in suppurative meningoencephalitis.^642,643 Other bacteria that can cause suppurative meningitis include Streptococcus zooepidemicus in foals and goats⁶⁴⁴; Streptococcus suis and Streptococcus equi in foals⁶⁴⁵; Actinomyces species in horses; Escherichia coli, Pasteurella, Streptococcus, Staphylococcus pyogenes, Mannheimia, and Arcanobacterium (Actinomyces) pyogenes in calves; and Globicatella sanguinis in lambs.^646-649 Embolic showers to the central nervous system (CNS) also occur in cases of left-sided endocarditis. Pseudomonas aeruginosa mastitis of cattle and goats may terminate with septicemia and meningitis. In adult animals, previous surgical procedures may result in sepsis and subsequent meningitis.⁶⁵⁰ Management or environmental factors may predispose to outbreaks of meningoencephalitis within groups of animals, as occurs when inadequate colostrum is supplied to neonates, or when contamination of the water supply occurs, as reported in an outbreak of amebic meningoencephalitis in cattle.⁶⁵¹

Suppurative meningitis of hematogenous origin is common in neonates (see Chapter 18). Gram-negative bacteria, E. coli, and Salmonella species are the dominant organisms involved in neonatal infections. One survey reported a 43% prevalence of septic meningitis in necropsied calves.⁶⁵² Deficient passive transfer of colostral antibodies to the neonate predisposes to hematogenous meningitis (see Chapter 53). Mycoplasma mycoides subsp. mycoides is a common cause of meningitis in goat kids. Underlying immunodeficiency disorders may predispose to the development of suppurative encephalitis in mature animals.⁶⁵³

Clinical Signs

The earliest clinical signs of meningitis are usually those of systemic illness, such as diarrhea, fever, and anorexia, accompanied by stiff neck and hyperesthesia.^654,655 Passive manipulation of the head and neck causes sudden extension and hypertonicity of the limbs. Slight tactile stimulation of the skin may result in strong spasmodic extension of the limbs, fasciculations of the underlying musculature, or even generalized frantic motor activity. The patient’s behavior may vary from extreme depression to hyperexcitability or mania. Animals may display trismus and may vocalize when the head and neck are flexed. Tetraparesis, hyperreflexia, and a tendency to circle or fall toward one side may be noted. A subtle intentional head tremor often is observed in foals. Dysfunction of one or more cranial nerves may result in facial muscular tremors, nystagmus, facial palsy, blindness, anisocoria, or strabismus. These deficits are inconstant. Progression of the clinical signs is associated with a decreased sensorium, propulsive walking, coma, seizures, and status epilepticus. Evidence of infection elsewhere may be apparent, such as swollen joints, omphalophlebitis, or hypopyon.⁶⁴⁷

The clinical signs of purulent meningitis closely resemble those of metabolic encephalopathies, including hypomagnesemia, hypoglycemia (which may occur simultaneously with septic meningitis in neonates), and encephalomalacias (e.g., salt poisoning). The onset of clinical signs may be delayed in horses with Cryptococcus neoformans meningitis.⁶⁴²

Clinical Pathology

Septic meningitis should be differentiated from metabolic encephalopathies by measurement of the plasma concentrations of sodium, glucose, and magnesium and by laboratory evaluation of hepatic function. Diagnosis of meningitis is based on examination of the cerebrospinal fluid (CSF; for normal values, see Table 35-1). The CSF of animals with meningitis may be turbid and white to amber in color, may foam when shaken, and may clot. Xanthochromia may be observed in some specimens. The white blood cell (WBC) counts in CSF of calves with purulent meningitis are typically greater than 100 neutrophils/μL (mean count, 4004 WBCs/mL), and protein concentrations range from 20 to 270 mg/dL.⁶⁵⁴ The differential cell counts in the CSF are either predominantly neutrophilic or mononuclear, with fewer neutrophils. The concentration of glucose in the CSF often is less than 50% of the corresponding concentration in the blood. Because of the lack of opsonic and bactericidal activity in the CSF, bacteria can proliferate to high titers and are often observed in Gram-stained smears of CSF. One study showed intracellular bacteria in 10 of 22 calves with meningitis.⁶⁵⁶ Abnormalities in the blood are inconsistent and reflect secondary conditions such as septicemia, diarrhea, or overaggressive fluid therapy. These changes could include leukocytosis, left shift, toxic changes in the hemogram, hyperkalemia, respiratory acidosis, hypoglycemia, and hyponatremia or hypernatremia. Evidence of bacteremia and sepsis may be found in peripheral blood.

Pathophysiology

Because there is little bactericidal or opsonic activity in the CSF, animals are highly susceptible to meningeal infection by low numbers of bacteria. Bacterial proliferation in the CNS results in production of bacterial endotoxins, cytokines, and other products of inflammation that damage the neural parenchyma. Vascular sequelae of infection may include thrombotic or hemorrhagic infarcts. After several days, inflammation of the arachnoid trabeculae and choroid plexus can result in decreased CSF absorption and hypertensive hydrocephalus.⁶⁴⁷

Necropsy Findings

The meningeal vessels appear to be congested, and the meninges are swollen, opalescent, and petechiated. The CSF is cloudy or amber and may contain fibrin clots. In cases associated with bacteremia, a fibrinopurulent iridocyclitis may be observed. Microscopic changes of CNS tissues include infiltration by neutrophils and lymphocytes, endarteritis of the meningeal vessels, choroiditis, scattered leptomeningeal hemorrhages, and bacterial colonies around the blood vessels of the meninges and the brain parenchyma.

Concomitant pathologic lesions, including omphalophlebitis, septic arthritis, anterior uveitis, and panophthalmitis, result from dissemination of the septic process. These lesions may be helpful in differentiating meningitis from metabolic encephalopathies. When meningitis occurs secondary to trauma, the site of organism entry may be detectable. Fungal meningitis caused by C. neoformans often is accompanied by granulomatous lesions of the lips, nasal mucosa, and peripheral nerves.

Treatment

Treatment of bacterial meningitis is difficult, and the mortality rate is high. Early recognition and treatment are essential for adequate recovery. Antimicrobial sensitivity tests performed on isolates from CSF may provide valuable information on ideal drugs. However, these tests frequently are unavailable because of the difficulty in isolating primary pathogens from CSF in most cases of purulent meningitis. Because prompt treatment of meningitis is critical, antibiotic therapy usually must be based on the Gram-staining characteristics of sedimented bacteria and the initial 24-hour cultures.

Domestic animals have a well-defined blood-CSF barrier, and antibiotics that reach high plasma concentrations may not necessarily reach bactericidal concentrations in the brain. To ensure an adequate antibacterial efficacy, antibiotic concentrations in the CNS should range from 10 to 30 times the minimum inhibitory concentration (MIC) of the infecting bacteria.^656-664 Antibiotics that are inherently bactericidal tend to produce superior responses compared with agents that are primarily bacteriostatic. The major factors influencing CSF penetration of an antimicrobial agent are the lipid solubility, degree of ionization, and molecular weight of the drug. In general, broad-spectrum drugs with a nonpolar basic character tend to have the greatest CNS penetration and efficacy for treatment of meningitis. Antibiotics tend to diffuse into CSF to a greater extent when it is inflamed. Box 35-1 shows the relative penetrability of antibiotics and antimicrobials into the CSF.