**Human Rabies Epidemiology and Diagnosis**

Brett W. Petersen and Charles E. Rupprecht *Centers for Disease Control and Prevention, Atlanta*, *GA USA*

#### **1. Introduction**

246 Non-Flavivirus Encephalitis

Zlotnik, I., Peacock, S., Grant, D.P. & Batter-Hatton, D. (1972). The pathogenesis of western

*Pathol*, 53, 1, 59-77,

equine encephalitis virus (W.E.E.) in adult hamsters with special reference to the long and short term effects on the C.N.S. of the attenuated clone 15 variant. *Br J Exp* 

> Rabies is a fatal viral infection that is most commonly spread to humans through the bite of an infected animal. The disease is an acute progressive encephalitis caused by highly neurotropic zoonotic viruses belonging to the *Lyssavirus* genus in the *Rhabdoviridae* family (Kuzmin, 2009). Of the twelve species of lyssaviruses, rabies virus (RABV) is the most important with respect to its impact on public health. RABV is distributed globally and found on all continents except Australia and Antarctica. In the United States, multiple RABV variants circulate in wild mammalian reservoir populations including raccoons, skunks, foxes, and bats. Rabies has the highest case fatality rate of any infectious disease and kills an estimated 55,000 people annually, primarily in developing countries within Africa and Asia (Knobel, 2005). However, rabies is a preventable disease. Postexposure prophylaxis (PEP) consisting of rabies immune globulin and rabies vaccine is successful in preventing the disease when administered promptly after an exposure to the virus has occurred. Additionally, vaccination of domestic animals against rabies and stray animal control programs greatly reduce the risk of RABV transmission to humans. Implementation of these measures in developed countries such as the United States has led to drastic declines in the incidence of human rabies. Despite this success, rabies remains a significant public health issue. Each year approximately 7,000 rabid animals are reported in the United States (Blanton, 2010). Up to 35,000 people annually are estimated to receive PEP due to exposures to suspect rabid animals (Christian, 2009). Given the high cost of rabies PEP, this represents a substantial economic burden as well. A clear understanding of the epidemiology of human rabies in the United States can help to manage these human exposures using the best available evidence. In this way, the risk of infection can be assessed more precisely and ensure rabies PEP is administered more judiciously. The identification of epidemiologic patterns can also be used to focus educational messages for human rabies prevention and thereby increase public awareness of rabies and the importance of seeking medical care after a potential exposure occurs. Furthermore, providing accurate descriptions of the clinical presentation of human rabies is essential in recognizing and diagnosing the disease in a timely fashion. Delayed or missed diagnoses place others at risk of exposure if appropriate infection control precautions are not instituted, exposures are not treated appropriately, or organs or tissues from an infected individual are used for transplantation (Houff, 1979; Javadi, 1996; Hellenbrand, 2005; Kusne, 2005; Srinivasan, 2005). An early diagnosis also

Human Rabies Epidemiology and Diagnosis 249

Data analyses were performed using SAS version 9.2 (SAS Institute Inc., Cary, North Carolina). Data were summarized using descriptive statistics and comparisons between human rabies cases and cases of encephalopathy with negative rabies diagnostic testing were made using Chi-square or Fisher's exact test. Some variables were dichotomized before statistical comparisons for determination of odds ratios (OR) and 95% confidence intervals

RABV antigens were detected using the direct fluorescent antibody (DFA) test of skin biopsy specimens, touch impressions of corneal epithelial cells, or fresh brain tissue as described (CDC, 2006). Skin biopsy specimens were taken from the nuchal area of the neck where viral antigens can be present in hair follicles containing cutaneous nerves, as

RABV antibody testing for cases reported before 1973 utilized the mouse neutralization test (Jackson, 2003). After 1973, serology was determined using the rapid fluorescent focus inhibition test (RFFIT) or the indirect fluorescence assay (IFA), as described previously (Noah, 1998). The RFFIT measures RABV neutralizing antibodies while the IFA detects serum reactive with RABV antigen in infected cell cultures. Antibodies in serum were considered diagnostic if there was no history of rabies immunization prior to sample collection. Antibodies in cerebrospinal fluid (CSF) were considered diagnostic regardless of

RABV was isolated through intracerebral inoculation of suckling mice or by addition of suspensions of brain or saliva specimens to cultured mouse neuroblastoma cells, as

Viral nucleic acids were obtained using standard extraction procedures and reagents. Samples used for nucleic acid extraction included saliva, fresh brain, paraffin-embedded brain, and nuchal skin. Reverse transcription polymerase chain reaction (RT-PCR) was performed using primers targeting the sequence of the nucleoprotein gene. Standard dideoxynucleotide sequencing methods were utilized to determine the nucleotide sequences

RABV variants were identified through antigenic and/or molecular typing. Antigenic typing uses a reference panel of monoclonal antibodies directed against the nucleoprotein to determine the variant of RABV isolates. Molecular typing methods identify the RABV variant by comparing the nucleotide sequence obtained by RT-PCR with a database of sequences from known reservoirs within the United States as well as foreign countries

of all PCR products obtained, as described previously (Noah, 1998).

(CI). Associations were considered statistically significant at p-values less than 0.05.

**2.3 Statistical analysis** 

**2.4 Diagnostic laboratory testing 2.4.1 Detection of antigen** 

described previously (Noah, 1998).

rabies immunization history.

described previously (Noah, 1998).

**2.4.5 Identification of rabies virus variants** 

**2.4.3 Virus isolation** 

**2.4.4 RNA detection** 

throughout the world.

**2.4.2 Serology** 

provides the patient with the opportunity for treatment and possible survival. Insights gained from each attempt at treatment further our understanding of the disease and add to the body of knowledge that can be applied to future cases. When rabies is ruled out, efforts can be focused on identifying more treatable causes of encephalitis. With these goals in mind, this review will describe the epidemiology of human rabies, examine the signs and symptoms of disease, and review the laboratory diagnostic testing and results for all reported human rabies cases in the United States between 1960 and 2010.
