Infectious Disease Pathology

Fatal Rabies Infection

Sherif R. Zaki
Centers for Disease Control
Atlanta, GA


Clinical Summary
In February 2003, a previously healthy 25 year old male visited his physician with head and body aches, nausea, abdominal pain, chills, fever of 99°--100° F, dry cough, and listlessness. Upon retrospective questioning, his wife reported that he had showed mild personality changes during the previous days. Six days later, the patient awoke disoriented with unsteady gait and slurred speech. He was evaluated in a local emergency department and admitted to the hospital. Physical examination revealed mild ataxia and confusion. Laboratory values were substantial for decreased sodium. A lumbar puncture revealed a white blood cell count of 24/µL, a red blood cell count of 10/µL, a glucose concentration of 58 mg/dL, and a protein concentration of 81 mg/dL. An electroencephalogram demonstrated generalized slowing. Magnetic resonance imaging of the brain was interpreted with a high T2 signal in the hypothalamus and bilateral mesial temporal lobes.

The patient remained febrile and hyponatremic (range: 119--125 mmol/L) with declining mental status. On the fifth day of hospitalization, the patient was intubated, and twitching on his right side was noted. On day six, he was unresponsive and had near-constant myoclonic activity. On the 11th day, a computerized tomography scan of the head showed sulcal effacement and diffuse cerebral edema. The patient remained comatose and intermittently febrile. Despite aggressive critical care management, the patient died on the 14th hospital day.

At autopsy, histopathologic evaluation showed severe meningoencephalitis involving the cortex and white matter of the cerebral hemispheres, deep gray nuclei, cerebellum, and spinal cord. Brain tissue submitted to a research laboratory was positive by polymerase polymerase chain reaction (PCR) for Naegleria. Material provided: Scanned glass slide of CNS (hematoxylin and eosin)

Images H&Es of CNS pathology and IHC for Rabies (Figures 1-8)


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Diagnosis
Fatal Rabies Infection

Pathologic Findings:
At autopsy, histopathologic evaluation showed severe meningoencephalitis involving the cortex and white matter of the cerebral hemispheres, deep gray nuclei, cerebellum, and spinal cord. Brain tissue submitted to a research laboratory was positive by polymerase chain reaction (PCR) for Naegleria. Initial microscopic examination of brain tissue did not detect any inclusions suggestive of viral infection.

Tissues were forwarded to CDC for pathologic evaluation for Naegleria. No amebic trophozoites were seen in tissues. Immunohistochemical (IHC) and PCR assays for various amoebae, including Naegleria fowleri, were negative. Histopathological evaluation of tissues demonstrated diffuse, predominantly lymphohistiocytic, infiltrates and microglial nodules involving the cerebrum, brain stem, cerebellum, and spinal cord (Figures 1-2). Cytoplasmic inclusions consistent with Negri bodies were identified throughout the central nervous system (CNS), particularly in the Purkinje cells of the cerebellum and in neurons of the frontal cortex, thalamus, hippocampus, midbrain, and pons (Figures 3-5). Intracytoplasmic rabies virus antigens were detected on immunohistochemical staining in neurons from multiple areas of the CNS (Figures 6-8). Electron microscopy of the midbrain demonstrated abundant rhabdovirus particles However, abundant intracytoplasmic inclusions of neurons in several areas of the brain suggested a diagnosis of rabies. Further testing, including reverse transcriptase-PCR of fixed brain tissue, supported the diagnosis of rabies. Nucleotide sequence analysis and antigenic typing with monoclonal antibodies on frozen brain tissue indicated that the specific etiologic agent was a southeastern raccoon rabies virus variant. Genetic sequence analysis indicated 100% homology with a raccoon rabies virus variant from Virginia.

Discussion
Rabies is an acute encephalitis caused by viruses in the genus Lyssavirus, family Rhabdoviridae, that is nearly uniformly fatal in unvaccinated hosts. Although the virus is present in animal reservoirs, infection in humans is rare in the United States, with only two cases reported in 2003 [1, 2] and no more than six cases reported in any year in the past decade [3]. The primary mode of transmission is through the bite of an infected animal, most commonly a bat in the United States [4]. Transmission of rabies virus from corneal transplants and more recently through transplantation of solid organs and vascular material have been reported [5].

In March 2003, a previously healthy man aged 25 years from northern Virginia died from a diagnosed illness of meningoencephalitis of unknown etiology after a 3-week illness. Histopathologic review of central nervous system tissues at CDC revealed viral inclusions suggestive of Negri bodies, and subsequent tests confirmed a diagnosis of rabies. Genetic sequencing identified a rabies virus variant associated with raccoons, but how the patient became infected remains unknown.

Approximately 125 family members and friends and 173 health-care workers were questioned retrospectively about direct unprotected exposures to the patient's secretions and tissues. After detailed investigation, five family members and three hospital employees received postexposure prophylaxis for potential exposure to patient secretions.

The patient was an office worker who for the previous 6 years had lived, worked, and recreated in areas in which raccoon rabies was endemic. However, extensive interviews with family, friends, and co-workers revealed that he had no specific exposure to terrestrial animals likely to be infected with the raccoon rabies virus variant. The patient did not spend much time outdoors, but the potential existed for encountering a rabid mammal while camping or in a trash can, wood pile, or other outdoor environment.

Approximately 7,000--9,000 cases of animal rabies are diagnosed annually in the United States [6]. This report describes the first documented case of human rabies associated with a raccoon rabies virus variant [1]. Of the 37 human rabies cases reported in the United States since 1990, no history of suspicious animal bite exposure was documented for 28 of the 30 cases presumed to be acquired in the United States. With the isolation of raccoon rabies virus from this patient, human cases have been associated with all of the major reservoirs and vectors of the disease in the United States, including dogs, cats, bats, foxes, skunks, coyotes, and bobcats. Human rabies cases without a definitive history of animal exposure are associated commonly with bat rabies viruses [7]. Challenges to implicating an animal source readily can include failure to seek medical care for perceived minor lesions, non-recognition of the actual exposure event, communication (i.e., language) barriers, and recall bias from memory loss or impaired speech in encephalitic patients. Incubation periods range typically from 1 to 3 months after exposure but in rare cases can exceed 1 year in duration, further complicating collection of an adequate history.

During the late 1970s, rabid raccoons were identified in Virginia and West Virginia after probable translocation of infected animals from the southeastern United States. Raccoon rabies spread throughout the region, with approximately 50,000 rabid raccoons diagnosed to date. During 2003, Tennessee became the twentieth affected state, and the enzootic area now stretches from eastern Canada to Florida [8].

Rabies should be considered in the differential diagnosis of any acute, rapidly progressive encephalitis, regardless of documented history of animal bite. Prompt ante- or postmortem diagnosis is necessary for accurate reporting of human rabies to public health officials and implementation of appropriate infection-control measures, including prompt administration of prophylaxis to exposed persons. Microscopically, inflammatory cells are seen in the brain stem and spinal cord and to a lesser extent, in the basal ganglia and the thalamus. Perivascular cuffing is primarily lymphocytic with some histiocytes and plasma cells. Meningeal infiltrates are sparse of absent and glial nodules are present in about half the cases. About 70-80 % of cases show typical eosinophilic cytoplasmic inclusions. Without cytoplasmic inclusions (Negri bodies), rabies encephalitis appears as a non-specific encephalomyelitis and other causes of viral and other infectious encephalitides should be included in the differential diagnosis. IHC and other molecular assays that can be performed on formalin-fixed tissues can be helpful to diagnose otherwise unexplained cases of encephalitis or to confirm presumptive cases of fatal rabies, particularly when samples for mouse inoculations, culture or PCR are not available. The diagnosis of rabies in humans can be confirmed by demonstrating neutralizing antibodies in the serum of persons who have never been vaccinated against rabies [5, 9, 10, 11, 12, 13, 14, 15, 16].

The Advisory Committee on Immunization Practices publishes guidelines for human rabies prevention [17], and recommendations have been published for the management of suspected cases [18]. Human rabies postexposure prophylaxis is effective when administered promptly and properly. Human-to-human transmission is a concern, but no cases among health-care workers exposed to a rabies patient have been reported [19, 20]. In the case described in this report, careful risk assessment based on identifiable contact with the patient's secretions limited the number of persons receiving prophylaxis. Emergency medicine physicians, infectious-disease consultants, and state and national public health officials can provide advice on rabies prophylaxis for complicated or unusual exposure scenarios to prevent this fatal disease and aid in its diagnosis.

Take Home Bullet Points:
Approximately 7,000--9,000 cases of animal infections are diagnosed annually in the United States.

In comparison, only around 40 human infections have been reported in the United States since 1990.

Characteristic CNS cytoplasmic inclusions are highly suggestive of the diagnosis and can be seen in about 70-80% of cases .

IHC and PCR on formalin-fixed tissues are organism-specific useful diagnostic assays and are used to confirm the diagnosis.

References:
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  2. Human death associated with bat rabies - California, 2003. MMWR Morb Mortal Wkly Rep 2004;53:33-5.

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