Rabies Virus

Introduction

• Fatal viral zoonotic diseases and can occur in many warm-blooded animals.
• Preventable viral diseases of mammals, most often transmitted through the bite of a rabid animal.
• Domestic animals account for less than 10% of the reported case (rabies case) with cats, cattle, and dogs.
• Rabies virus infects the central nervous system (CNS) causing encephalopathy and ultimately death.
• Death usually occurs within days of the onset of symptoms.

Rabies epidemiology

• Globally 20 minutes post exposure treatment cases and 30000 hydrophobia cases are estimated.
• Over 90% of those cases occur in Asia.

Epidemiology of Rabies in Nepal

• Annually 25000-30000 post-exposure treatment cases of human & 100 cases of hydrophobia are estimated.
• Among rabies cases male 63% and female 37%.
• Most of the cases 64% adult aged more than 15 years.

The rabies virus

Family :Rhabdoviridae

Genus : Lyssavirus

Structure

• Bullet shaped(180*75nm) negative sense signal stranded RNA virus.
• RNA codes for 5 protein(M), glycoprotein(G), polymerase(L).
• ‘G’ forms the trimeric spikes(10nm) on the surface of the virus.
• ‘M’ associates with 2 structural components.
• Unsegmented signal stranded RNA is tightly encased(bound) with the nucleoprotein(N).
• The genome is of negative polarity the virus contains an RNA-dependent. RNA polymerases(L) protein.
• A phosphoprotein(P) is also found in the RNP core.
• The replication cycle of rabies virus takes place entirely in the cytoplasm of infected cells & results in the formation of numerous virus particles.
• Masses of nucleocapsids accumulate in the cytoplasm of neurons to form inclusions called Negri bodies.
• Rabies virus is mainly could be found in milk & other body discharges.
• Considering infectivity only the virus present in the saliva is potential to causes infection.

Pathogenesis

• During replication period, the rabies virus may move into the nervous system or remain at close the bite.
• When the virus eventually reaches the peripheral nerves, it advances to the spinal ganglia, spiral cord, and brain.
• The virus moves passively within the axoplasm of peripheral nerves to the central nervous system (CNS) in a retrograde fashion.
• From the brain virus often returns to the periphery using the same axoplasmic route as used for centripetal movement.
• A favored peripheral site is a salivary gland but towards the end of disease course, the virus may be found in many other tissues.
• In some cases, the virus never reaches the salivary gland or any exit route, and the host can die tissue from acute encephalitis.
• The levels of virus in the salivary gland of a rabid animal may be 1000 or more times the level found in the brain , suggesting that the virus replication at these sites as well.
• A limited amount of Histopathological changes occurs in the CNS of animal or animals dying of rabies.
• Neurons of infected animals and humans contain typical intracytoplasmic inclusions, the Negri bodies.
• Peri-vascular cutting and limited neuronal necrosis also occur in few cases.

Clinical pictures

Furious rabies

• The virus reaches to CNS.
• Cause a headache, fever, irritability, restless, anxiety
• Muscle pain, salivation, vomiting
• Stage of excitement starts with painful muscle spasm triggered by swelling of saliva or water.
• Hydrophobia
• Final stage: coma and death

Dumb rabies

• Same as furious rabies without the stage of excitement.

Transmission

• The commonest mode in man is by the bite of a dog.
• Rabies is transmitted by contamination of mucous membrane, aerosol transmission, corneal transplantation.

Incubation period

• Two weeks to 6 months.
• The incubation period of rabies in Nepal as per reporting is found to be 12 days to 2 years.
• Depends on the site of the bite, a severity of bite, class of wound, a number of a mound, category of exposure, type and amount of virus inoculated, species of biting animals, protection provided by clothing, age, immune status of victim as well as the animal and wound treatment immediately after bite etc.

Diagnosis

1. Direct fluorescent antibody test (DFA): Determination of rabies virus in the brain tissue or anti-rabies antibody.
2. Histopathology: Determination of Negri bodies from biopsy tissue.
3. Immuno-histochemistry: determine the virus similar to DFA.
4. Ultrastructure: using electron microscopy
5. Virus culture: in mouse neuroblastoma cells or baby hamster kidney cells.
6. Real- Time Polymerase Chain Reaction (RT-PCR)

Prevention and control

Vaccination

• Post-exposure vaccination: possibly due to long incubation period
• Pre- exposure vaccination: used in risk group
• Vaccines: human diploid cell vaccine (100% efficiency), Sample vaccine, duck embryo vaccine.

Post-exposure prophylaxis

• Wound treatment: washing and cleaning the wound
• Passive immunization: Human rabies immunoglobulin (HRIG), Equine rabies immunoglobulin (ERIG)
• Active immunization: Injection of rabies vaccine

Apart from vaccination and some prophylaxis, there are some other factors to consider. Such as:

Responsibilities after exposure

• The geographic location of the incident
• The type of animal that was involved
• How the exposure occurred (provoked or unprovoked)
• The vaccination status of animal
• Whether the animal can be safely captured and tested for rabies.

Be a responsible pet owner

• Keep vaccinations up to date for all dogs and cats
• Keep your pets under direct suspension so they do not come in contact with wild animals
• Call your local animal control agency to remove any stray animals from your neighborhood.
• Avoid direct contact with unfamiliar animals.

Laboratory diagnosis of Rabies

Laboratory diagnosis of rabies can be done in following ways:

A. Direct detection methods

Electron microscopy

• The specimen (eg CSF) is negatively stained with potassium phosphotungstates.
• The stained preparation is examined for the bullet-shaped rabies virus using an electron microscope.

Antigen detection and histopathology

• A biopsy specimen is usually taken from the skin of neck at the hairline. Impression preparation of brain or corneal tissue may be used.
• Rabies virus antigens in the infected tissues are identified most rapidly and accurately by immunofluorescence or immunoperoxidase staining using antibodies monoclonal antibodies.
• Negri bodies are demarcated, oval or round, having a distinctive internal structure with basophilic granules in an eosinophilic matrix.
• Viral antigens in Negri bodies can be demonstrated by immunofluorescence.

Nucleic acid detection

• RT-PCR test can be used to amplify pans of the viral genome from fixed or unfixed brain tissue.
• Sequential of amplified products allows identification of infecting virus strain.

B. Virus isolation

• The available tissue is inoculated intracerebrally into suckling mice infection in mice results in encephalitis and death.
• CNS of the inoculated animal is examined for Negri bodies and antigen.
• In specialized laboratories, hamster kidney cell lines and mice neuroblastoma cells can be inoculated for rapid (2-4 days) growth of rabies virus.
• An isolated virus is identified by fluorescent antibody tests with specific antiserum.

C. Serology

• Serum antibodies to rabies can be detected by immunofluorescence or neutralization tests.
• Such antibodies develop slowly in infected persons or animals during the progression of the disease but promptly after vaccination with cell derived vaccines.
• Antibodies in CSF are produced in rabies-infected individuals but not in response to vaccination.

REFERENCE

Cheesbrough, M.Medical Laboratory Manual for Tropical Countries.Vol. Vol 2. ELBS London, 2007.

Tille, P.Diagnostic Microbiology.13th. Elsevier, 2014.

D, Greenwood, Slack RCB, and Peutherer J.Medical Microbiology.Dunclude Livingstone: ELBS, 2001.