Absence of myelin basic protein in an improved purified duck embryo rabies vaccine

The encephalitogenic potential of rabies vaccines prepared from nervous tissue is a result of the presence of myelin basic protein. Vaccines prepared from duck embryos are economical and efficient, but, occasionally, cases of allergic encephalomyelitis have been reported. An improved rabies vaccine has been developed that contains the classical Pitman Moore strain of rabies virus grown in embryonated duck eggs. This vaccine has been highly purified and enriched in immunologically effective rabies virus glycoprotein antigen. We have searched for the presence of myelin basic protein using sensitive radioimmunological and immunoblotting techniques. Whereas the classical duck embryo rabies vaccine contained small amounts of myelin basic protein, in the improved purified duck embryo rabies vaccine, none could be detected.     

Rabies encephalitis following fox bite--histological and immunohistochemical evaluation of lesions caused by virus

Rabies caused by fox bite is uncommon, most cases being caused by bite of rabid dogs (95%). We report a 45-year-old lady with rabies encephalomyelitis caused by bite of a rabid wild fox (Vulpes vulpes), a species prevalent in the Deccan plateaus of Central India. Though foxes are known to be susceptible to rabies, literature on the pathological changes caused by fox bite rabies in humans is scarce. Unlike the mild histological alterations described in canine rabies, a florid encephalitic process evolved in fox bite rabies, in our case, with intense microglial reaction, neuronophagia and perivascular inflammatory infiltrates despite clinical manifestation as a paralytic rabies. Immunostaining using polyclonal antibodies to the rabies viral nucleocapsid antigen and to the whole virion demonstrated high viral load within neurons with extensive spread along dendritic arborization and axonal tracts. Genomic sequence analysis demonstrated close homology with canine virus strain with only minor variations.     

Immunohistochemical study of human rabies

Rabies is a communicable disease that is almost always fatal. In its classic form, rabies is well recognized, but cases presenting with a paralytic illness mimic Landre's Guillain–Barre syndrome and in such cases the diagnosis remains in doubt. This problem is further compounded when the history of dogbite is not forthcoming. At autopsy rabies can be diagnosed by subjecting fresh tissue to virologic investigations or examining formalin‐fixed paraffin‐embedded tissue sections for the presence of characteristic inclusions; that is, the Negri bodies. However, these inclusions are not present in all cases. Hence, the need arises for a better method for diagnosis. The present study utilized immunohistochemistry as a diagnostic tool using both monoclonal and polyclonal antirabies antibodies in 20 cases of rabies encephalomyelitis. The diagnosis of rabies could be confirmed in 17 cases (85%) based on neuropathologic findings alone. In contrast, immunohistochemistry yielded positive results in all cases. Moreover, the amount of rabies viral antigen was much more abundant than could be expected from the histopathologic findings. Thus immunohistochemistry is a rapid, safe, sensitive and specific technique for the diagnosis of rabies.     

Encephalomyelitis,brain tumors,neuromuscular diseases and miscellaneous disorders

Japanese neuropathologists have accomplished and contributed to a considerable number of achievements, and some of these are cited in other articles in this issue. Several of these achievements as well as other miscellaneous discoveries are briefly summarized in the present paper. Specifically these relate to rabies postvaccinal encephalomyelitis, experimental allergic encephalomyelitis, brain tumor research, neuromuscular disorders, schizophrenia, viral infections, glial inclusion body in multiple system atrophy, and the neurobiology of glia.     

Delayed onset of post–rabies vaccination encephalitis

A 31-year-old veterinarian developed seizures, left hemiparesis, loss of memory, and behavioral disorders 5 months after intensive antirabies vaccination, the longest incubation period yet recorded. Computed tomographic scan revealed a right frontal contrastenhanced mass that extended to the left frontal lobe through the corpus callosum. Brain biopsy showed foci of primary demyelination largely confined to the white matter. The lesions were characteristic of the demyelinating encephalomyelitis that follows treatment with certain vaccines against rabies. The hemiplegia improved, but seizures, memory impairment, and abnormal behavior persisted.     

The causes of the syndromes of Landry (1859) and of Guillain, Barré and Strohl (1916)

Post-infectious and post-vaccinal peripheral neuritis or encephalomyelitis have frequently been considered the human equivalents of experimental allergic neuritis (EAN) or encephalomyelitis (EAE). The major basis for these comparisons between diseases in humans and experimental animals rests on the classical observations of "paralytic accidents" of Pasteur-type vaccination against rabies. These old observations in humans injected with brain tissue indicate a remarkable heterogeneity of periphéral as well as central nervous system syndromes, quite in contrast with the remarkable specificity for either peripheral or central involvement in most experimental animals. The syndromes of Landry (1859) and of Guillain, Barré and Strohl (1916) differ clinically and pathologically, the latter a purely peripheral neuritis and the former a mixture of transverse myelitis and facial neuritis. Each can be caused by many different factors, including 1) direct infection by wild or attenuated rabies virus, 2) direct auto-sensitization by myelin antigens in the vaccine and 3) indirect or cross-reactive sensitization by viral or bacterial antigenic determinants (epitopes) with sufficient chemical homology with aminoacid sequences in central or peripheral myelin antigens to be recognized as immunological homologies.     

Rabies

Rabies is an important disease in wildlife in the United States and Canada, and dog rabies is still a major public health problem in many developing countries of the world. Rabies virus is transmitted in saliva by animal bites. Bats transmitted most recent cases of human rabies in the United States, often without known exposures. There have been recent developments in our understanding of rabies pathogenesis. Characteristic clinical features should raise the possibility of a diagnosis of rabies and initiation of appropriate diagnostic tests. Therapy of human rabies has been futile except in four patients who were immunized with rabies vaccine prior to the onset of their disease. Rabies can be prevented after an exposure in unimmunized patients with local wound cleansing and administration of rabies vaccine and human rabies immune globulin.     

Pleomorphism of fine structure of rabies virus in human and experimental brain

Identification of the Negri bodies in the brain of an 8-year-old boy who died 8 days after a paralytic illness and 20 days after a dog bite, and who had received 9 injections of Semple's anti-rabies vaccine, provided evidence that he died of acute rabies encephalitis and not of post-vaccinal allergic encephalomyelitis. The Negri bodies in the human subject and those seen in the inoculated mouse differed in their morphological structure: the former consisted of a matrix of very fine granular material bearing larger granules or strands of higher electron-density resembling nucleic acids and representing products of host cell-virus interaction; and the latter showed better defined areas of granular matrix containing tubular, bullet-shaped and elongated forms of viral structures, and nucleocapsids or capsule-deficient cores, representing the virions, emerging from them. Fine structural examination of the patient's brain and of the inoculated mouse has provided evidence of the pleomorphism of the Negri bodies and the various stages of formation of viral material and virions in them, the animal alone showing the mature virions of rabies, and proving the infectivity of the Negri bodies of the human brain.     

SPONTANEOUS RECOVERY FROM THE ENCEPHALOMYELITIS IN MICE CAUSED BY STREET RABIES VIRUS

Recovery from rabies was studied in an experimental model. Young adult mice were inoculated in a hindlimb footpad with street rabies virus (fox salivary gland isolate). In a group of 62 mice, 97% developed clinical rabies with paresis of the extremities and spasticity, and 37% recovered with neurological sequelae. There was an acute inflammatory reaction in the brainstem and grey matter of the spinal cord, and degeneration of myelinated axons in the white matter of the cord and in the dorsal roots. Rabies virus antigen was found in the central nervous system of all mice examined between day 5 and 13, and also in trigeminal and dorsal root ganglia. Surviving mice had neutralizing antibodies in serum and brain tissue, and 90% survived an intracerebral challenge with the CVS strain of fixed rabies virus. Spontaneous recovery from rabies encephalomyelitis was demonstrated with evidence of viral replication and pathological changes in the central nervous system.     

Neural complications of cell culture rabies vaccine prepared from hamster kidney. Clinicopathological report of a case]

A case died of cell culture rabies vaccine prepared from hamster kidney was reported. Autopsy showed invasion of both the peripheral and the central nervous systems. In the CNS there were not only acute disseminated encephalomyelitis, but also changes similar to acute viral encephalitis.     

Semple rabies vaccine: presence of myelin basic protein and proteolipid protein and its activity in experimental allergic encephalomyelitis

Myelin basic protein (MBP) as a cause of postvaccinal encephalomyelitis (PVE) due to Semple rabies vaccine (SRV) has been suggested in previous reports. No actual measurement of MBP in SRV was done. In this study we detected MBP and PLP in the vaccine using immunological methods. The vaccine was found to contain 28 micrograms MBP per ml vaccine. Inoculation with SRV plus adjuvant resulted in the development of experimental allergic encephalomyelitis (EAE) in 2 of 3 guinea pigs. For control, chick embryo vaccine (CEV) was used and MBP was not detected. EAE was not induced in animals inoculated with it. These results suggest that MBP in vaccines may play a decisive role in the production of PVE.     

Diabolical effects of rabies encephalitis

Rabies is an acute encephalomyelitis in humans and animals caused by rabies virus (RABV) infection. Because the neuropathological changes are very mild in rabies, it has been assumed that neuronal dysfunction likely explains the severe clinical disease. Recently, degenerative changes have been observed in neuronal processes (dendrites and axons) in experimental rabies. In vitro studies have shown evidence of oxidative stress that is caused by mitochondrial dysfunction. Recent work has shown that the RABV phosphoprotein (P) interacts with mitochondrial Complex I leading to overproduction of reactive oxygen species, which results in injury to axons. Amino acids at positions 139 to 172 of the P are critical in this process. Rabies vectors frequently show behavioral changes. Aggressive behavior with biting is important for transmission of the virus to new hosts at a time when virus is secreted in the saliva. Aggression is associated with low serotonergic activity in the brain. Charlton and coworkers performed studies in experimentally infected striped skunks with skunk rabies virus and observed aggressive behavioral responses. Heavy accumulation of RABV antigen was found in the midbrain raphe nuclei, indicating that impaired serotonin neurotransmission from the brainstem may account for the aggressive behavior. We now have an improved understanding of how RABV causes neuronal injury and how the infection results in behavioral changes that promote viral transmission to new hosts.     

Association of HLA and T-cell receptor gene polymorphisms with Semple rabies vaccine-induced autoimmune encephalomyelitis.

Semple rabies vaccine is derived from brain tissue infected with rabies virus that is subsequently inactivated with phenol. Semple rabies vaccine-induced autoimmune encephalomyelitis (SAE) occurs in 1 in 220 immunized individuals. The immune response to myelin basic protein and pathological changes of demyelination in SAE suggest that this disease is the human homologue of experimental autoimmune encephalomyelitis (EAE). SAE and EAE are frequently studied as models for the human demyelinating disease multiple sclerosis. Major histocompatibility complex (MHC) class II and T-cell receptor (TCR) gene polymorphisms play important roles in rodent susceptibility to EAE and were analyzed to determine if the same was true in humans with SAE. HLA-DRB1, HLA-DQB1, and TCRBV gene polymorphisms were studied in Thai individuals with SAE (n = 18), with vaccination without neurological complications (n = 43), and without vaccination (n = 140). The allele frequencies of HLA-DR9 (DRB1*0901) and HLA-DR17 (DRB1*0301) were increased in SAE patients (DR9 = 22%, DR17 = 14%) compared with vaccinated controls (DR9 = 13%, DR17 = 6%) and with unvaccinated controls (DR9 = 9%, DR17 = 4%). The allele frequency of HLA-DQ7 (DQB1*0301) was decreased in SAE patients (8%) compared with vaccinated controls (15%) and with unvaccinated controls (25%). These susceptibilities are distinct from those associated with multiple sclerosis. The frequencies of TCRBV alleles and haplotypes were similar in SAE patients and vaccinated controls. These data suggest that genetic susceptibility associated with MHC class II alleles may have a role in the pathogenesis of SAE and its mechanism may be different from those involved in multiple sclerosis.     

Apoptotic cell death in experimental rabies in suckling mice

A fatal encephalomyelitis developed after intracerebral inoculation of 6-day-old ICR mice with the challenge virus standard (CVS) strain of fixed rabies virus. The brains of CVS-infected mice showed widespread morphologic changes of apoptosis, which were particularly prominent in pyramidal neurons of the hippocampus and in the cerebral cortex. Evidence of oligonucleosomal DNA fragmentation was sought in situ using the TUNEL method. TUNEL staining was observed in many neurons, and rabies virus antigen was usually demonstrated with immunoperoxidase staining in similar regions. Neurons in the dentate gyrus of the hippocampus demonstrated expression of viral antigen, apoptotic changes, and positive TUNEL staining. This region normally demonstrates little infection in CVS-infected adult mice. Double labeling of neurons with TUNEL and viral antigen indicated that infected neurons actually underwent apoptosis. Increased immunoreactivity against the Bax protein was demonstrated compared to uninfected mice. Purkinje cells expressed viral antigen, but did not show significant morphologic changes of apoptosis or TUNEL staining. In contrast, neurons in the external granular layer of the cerebellum did not express viral antigen, but demonstrated greater morphologic changes of apoptosis and positive TUNEL staining than uninfected controls. Apoptotic cell death likely plays an important role in the pathogenesis of rabies virus infection in suckling mice. There was evidence of more apoptosis in the brains of suckling mice than in those of adult mice and this finding explains the greater neurovirulence of rabies virus in younger mice. Rabies virus likely induces apoptosis in vivo by both direct and indirect mechanisms. Received: 15 July 1997 / Revised: 25 August 1997 / Accepted: 8 September 1997     

The role of immune responses in the pathogenesis of rabies

In the absence of treatment, infection with a variety of rabies virus strains most often results in a lethal outcome. This can be averted by prompt immunization following exposure demonstrating that the development of anti-rabies viral immunity prior to extensive infection of neurons is protective. Otherwise it might be expected that immune clearance of the virus would result in neurological sequelae. Thus, the capacity of a rabies virus to induce a protective immune response is a major, negative determinant of its pathogenicity and highly pathogenic rabies viruses have characteristics that avoid triggering protective immune responses. On the other hand, there is evidence that certain aspects of immunity may contribute to the pathogenesis of rabies under certain circumstances. The relationship between rabies virus and the immune system of the host is the focus of this review.     

Neurologic complications of Semple-type rabies vaccine: clinical and immunologic studies

We studied 61 patients with complications of Semple-type postexposure rabies immunization. Thirty-six had neurologic signs, and 25 had only fever, headache, or myalgia. Thirty-two patients had CNS complications, and 4 had an acute peripheral neuropathy. Disease was acute and monophasic in 33, but 3 patients had progressive disease, including 1 patient with a relapsing-remitting course. No clinical features, including CSF content of myelin basic protein, were prognostic indicators. In three of six patients with encephalomyelitis, lymphocytes showed a proliferative response to myelin.     

Perspectives in Diagnosis and Treatment of Rabies Viral Encephalitis: Insights from Pathogenesis

Rabies viral encephalitis, though one of the oldest recognized infectious disease of humans, remains an incurable, fatal encephalomyelitis, despite advances in understanding of its pathobiology. Advances in science have led us on the trail of the virus in the host, but the sanctuaries in which the virus remains hidden for its survival are unknown. Insights into host–pathogen interactions have facilitated evolving immunologic therapeutic strategies, though we are far from a cure. Most of the present-day knowledge has evolved from in vitro studies using fixed (attenuated) laboratory strains that may not be applicable in the clinical setting. Much remains to be unraveled about this elusive virus. This review attempts to re-examine the current advances in understanding of the pathobiology of the rabies virus that modulate the diagnosis, treatment, and prevention of this fatal disease.     

Correlation of clinical and neuroimaging findings in a case of rabies encephalitis

BACKGROUND: Rabies encephalitis is a feared, virtually uniformly fatal form of central nervous system infection. The incidence of rabies encephalitis in the United States is almost certainly underestimated because of the predominance of bat-borne rabies, which can be spread without traumatic exposure. Because of its rarity in developed countries, rabies encephalitis has been seldom studied with modern imaging techniques. SETTING: University-based teaching hospital. PATIENT: A case of pathologically confirmed rabies encephalitis is presented. Diagnosis of rabies was made by seroconversion testing while the patient was alive and was confirmed postmortem by the presence of rabies antigens and Negri bodies in the brain. The patient had 2 magnetic resonance studies done that showed dramatic abnormalities in the medulla and pons that correlated with features of the neurologic examination and hypothalamic-pituitary abnormalities. RESULT: The patient had a fulminant encephalitic course that ended in death. CONCLUSION: Rabies is an uncommon cause of fatal encephalitis. Anatomic imaging studies such as computed tomographic and magnetic resonance scans have generally been negative in confirmed cases of rabies. We report a case of confirmed rabies with extensive brainstem and hypothalamic-pituitary abnormalities on magnetic resonance imaging. Although these findings are nonspecific, they should raise the clinical suspicion of rabies in the setting of aggressive encephalitis of unclear cause, and appropriate diagnostic tests should be performed.     

Acute ascending motor paralysis due to rabies: a clinicopathological report

An unusual presentation of a not so uncommon disease is described. Practical difficulties in arriving at a diagnosis of paralytic rabies are discussed. An enquiry of exposure and suspicion of rabies in atypical Landry's paralysis is stressed. The importance of autopsy to confirm either of the diagnoses is emphasized.     

Presence of specific antigens in neuronal cells infected with fixed and street rabies virus strains

Summary The presence of rabies specific antigens is investigated after infection with different rabies virus strains in neural cell lines and in the central nervous system of laboratory rodents. In fixed rabies infected cells, the rabies glycoprotein is found to be present 48 h after infection, whereas in hamsters this protein was found 5 days after an intracerebral inoculation. In contrast, rabies glycoprotein was not detectable in any of the street rabies-infected cell system by the fluorescent antibody test, although nucleoprotein was present, showing that infection occurred in these cells. Rabies glycoprotein was also undetectable in the central nervous system (CNS) of athymic nude mice which is known to be very sensitive to street rabies infection and to contain large quantities of viral material. Our results suggest that the smaller amount of rabies glycoprotein synthesized during street rabies infection are of consequence for the pathogenesis of rabies disease. The immunopathology of street rabies virus infection is certainly modulated by the failure of the viral glycoprotein to be present in large quantities on the surface of the infected cellular membrane as in the case of fixed rabies.Supported by grants from the Institut Pasteur and from INSERM (CRE 78.4.146.1 and ATP 50.77.82.009)