Dr.

E Srikaanth Reddy
Associate professor –G.S.L {community medicine}
Ex- national consultant ,world health organization , 2019-
2021.[posted at Odisha ]
Joint-secretary Indian public health association-Andhra
Pradesh 2025-2027.
 Presentation Outline
1. Some facts about rabies
2. History of the rabies virus
3. Transmission
4. Symptoms
5. Prevalence
6. Morphology and genome
7. Important viral proteins
8. Replication cycle
9. Diagnostic Techniques
10. Prevention, including pre- and post-exposure
treatments
11. Conclusion
 Some Facts about Rabies

• Belongs to the family Rhabdoviridae

• Genus Lyssavirus (Greek root “lyssa”)
• Infects warm-blooded mammals, including
humans
• Causes encephalitis of the central nervous
system
• Results in death of the host
 History of Rabies Virus

• Man described the disease in 2300 B.C. in

the Eshnunna Code of Mesopotomia.
• The origin “rabhas”, meaning “to do
violence” comes from ancient Indian
Sanskrit dating 3000 B.C.
• Many cultures have incorporated myths
and legends that can be connected to the
rabies virus.
 History of the Rabies Virus
•Outbreak of rabies in Europe
in 20th century spawned the
Vampire myth- linked to bats.
•Rabies epidemic in 13th
century France may have
spurred Werewolf myth as
well- linked to wolves.
•Rabies is commonly spread
by bats and wolves and other
wildlife like foxes and coyotes.
[Link]
 History of the Rabies Virus

• Since Roman times, man established the

link between the infectivity of a rabid dogs
saliva and the spread of the disease.
• Because there is no cure for rabies, those
that had been bitten by a rabid dog would
commonly commit suicide to avoid the
painful death that would inevitably follow.
 History of the Rabies Virus

• Louis Pasteur was the first person to

diagnose that rabies targeted the CNS.
• Also determined that nervous tissue of an
infected human or animal also contained
the virus.
• In 1890 created the rabies vaccine and
saved 9 year old Joseph Meister after he
had been bit by a rabid dog.
 Transmission of Rabies

• Is zoonotic, meaning is transmitted from

animal-animal or animal-human.
• Rabies is most commonly contracted from
the bite of an animal that is infected with
the virus.
• The virus is found in the saliva of the
infected animal.
 Transmission of Rabies

• Can also enter via any mucous

membrane, such as the eye, nose or
mouth.
• Handling and skinning of infected animal
carcasses.
• Few cases of human to human
transmission.
• Can be transmitted via aerosol, but that’s
also uncommon.
 Transmission of Rabies
• Inside the U.S., important reservoirs for the
transmission of the disease are wild animals
such as raccoons, skunks, foxes, bats, and
coyotes.
• The time it takes for rabies to affect the animal
averages 1-3 months. Sometimes may only be
a few days, and rarely longer than a year after
exposure.

[Link]/
 Transmission of Rabies

• The rabies infection and the symptoms

that accompany it is classified by five
stages:
1. Incubation (1-3 months)
2. Prodromal, where first symptoms occur
3. Acute neurological phase
4. Coma
5. Death or recovery
 Transmission of Rabies

1. Incubation
-The animal or human is bitten by rabid animal.
-Rabies enters the body via the saliva of the infected
animal.
-The virus spreads through the nerves via retrograde
axoplasmic flow. The virus makes it’s way to the dorsal
root ganglion where it replicates.
- No symptoms are experienced at this time, and the
disease can’t be transmitted to another animal yet.
 Transmission of Rabies

2. Prodromal Stage
-Virus replicates in the dorsal root ganglion of the spinal
cord where the blood-nerve barrier is thin.
-This is where the immune system detects the virus and
starts creating antibodies.
-First symptoms are felt: fever, muscle ache, agitation,
anxiety, nausea, vomiting, ache in the bitten extremity.
-From this point the virus replicates and travels up to the
brain, and the immune system can’t keep up with fighting
the virus.
 Transmission of Rabies

3. Acute Neurological Stage

- There is rapid proliferation of the virus in the brainstem,
thalamus, basal ganglia, and spinal cord
- Brain begins to swell, encephalitis
- Fluctuation between consciousness and confusion
- Aggressive behavior, violent episodes
- Convulsions and seizures
- Hallucinations
- Hydrophobia (foaming at the mouth) induced by
pharyngeal spasms (the most common image of rabies)
 Transmission of Rabies
3. Acute Neurological Stage, continued
- Aerophobia
- Sensitivity to lights, sounds and smells
- Respiratory spasms.

4. Coma
- Spasms decrease because paralysis occurs
- Heart arrhythmias and irregular heartbeat persist
- Cerebral encephalitis causes major organs to collapse
- Fixed pupils that are unresponsive to light
 Transmission of Rabies

5. Death or Recovery
- Death results from encephalitis of the brain.
- Heart failure along with major organ failure are the
main causes of death.
- Only four known cases of human recovery, and they
were permanently brain damaged.
 Prevalence of Rabies

• In the U.S., dogs were the main reservoir

previous to the 1950’s., where now
raccoons are the main reservoir.
• Vaccines in domestic house pets
drastically reduced the number of human
rabies cases.
• Now less than 10% of reported rabies
cases come from domestic animals.
 Prevalence of Rabies

• In the U.S. in 1955, raccoons accounted

for 1% of rabies cases, where in 2001
raccoons accounted for 40% of over 7,000
reported cases.

[Link]/rabies
Data Summaries by the United States Public Health
Service Regarding Prevalence of Rabies in the U.S.

5,540 Mammals tested

In 1955- reported by the
US Public Health Service

3,900 Animals 6,947 Animals tested

Tested in 1960 In 1994

Cockrum, 1997
 Prevalence of Rabies
• In this century, the number of human
deaths in the United States attributed to
rabies has declined from 100 or more each
year to an average of 1 or 2 each year
after the 1960’s. Most recent rabies cases
are linked to exposure to rabid wildlife.
• Animal control and vaccination programs
begun in the 1940's have practically
eliminated domestic dogs as reservoirs of
rabies in the United States.
 Prevalence of Rabies

• Although human deaths in the U.S. are

very low, the World Health Organization
reports that 35,000 to 50,000 people die
every year from rabies in developing
countries, mainly India and Asia.
• Currently, dogs are the number one
reservoir of rabies in the developing world.
• China is suffering from more rabies deaths
than SAR’s (Chinese Ministry of Health).
Morphology,

Replication and

Diagnostic

Techniques
 Morphology of rabies virus
•Negative single-stranded RNA genome
•Rod shaped with a flexible nucleocapsid tapering
at one end
–Nucleocapsid contains important viral proteins

180nm\75nm

[Link]
• A membranous envelope surrounds the
nucleocapsid
– Envelope contains both viral and glycoproteins
– Derived from the lipids of the host cell membrane
– Viral proteins of envelope form spikes that aid in viral
attachment

[Link]
 Important viral proteins
• N-protein
– makes up most of the nucleocapsid
– catalyzes replication process

• G-protein
– glycoprotein, come together to form the spikes of the
viral envelope
– mediates fusion
– its presence triggers immune response of host
– determines pathogenic ability of the virus
 • L-protein
– Small part of nucleocapsid
– Codes for RNA polymerase
– Functions in polymerizing activity

• P-protein
– Also makes up a small portion of the nucleocapsid
– Contributes to the ability of the L-protein to both bind
and move down template strand during the
attachment of nucleotides

Voyles 1993
Voyles 1993
 Infection Cycle
1. Attachment to host cell
- Main site of attachment is
at the neuromuscular
Nerve terminal
junction site of nerve cells
- Main receptor is the
nicotine acetylcholine
receptor
- These receptors have a
high affinity for the viral G-
protein
[Link]/depts/physiology

Lewis 2000
 2. Penetration via endocytosis
-the viral membrane attaches to the host cellular membrane
and enters into acidic endosomes by endoycytosis
3. Uncoating of the viral envelope
-uncoating separates the nucleic acid from the envelope
and nucleocapsid
-this step is necessary before transcription and
replication can take place
-takes place in the cytoplasm

Gaudin 2000
4) Transcription
-each gene is transcribed, separately, into its
complementary mRNA

5) Translation
-the five viral proteins are synthesized
6. Replication
-the RNA polymerase binds to mRNA and begins
synthesis of the complementary positive strand
-the newly synthesized N-protein binds to the termination
synthesis of each mRNA so that a complete strand
can be synthesized
-the full-length positive RNA strand then serves as a
template for the synthesis of the negative viral
genome
7. Assembly
-the N,L, and P-protein form the nucleocapsid around
the RNA strand and attaches to the cell
membrane
6. Budding
-Virus buds from the cell membrane, taking some of the
Wagner 1996
glycoprotein from the host to form the envelope
 Diagnostic Techniques
1. Histological examination for Negri bodies
-negri bodies are cytoplasmic masses of viral
nucleocapsids found in the brain tissue
-problem is that negri bodies are only present in 50-80%
of rabies cases
2. Direct flourescent antibody test
-uses tissue from suspected host and labeled
antibodies, specific to the viral antigen
-if the rabies antigen is present, the antibodies will
attach; monitored by flourescent microscope
-the technique poses the risk of infection to technician,
but is a quick diagnosis
Jogai 2002
 Negri body

[Link]
 3. Immunohistochemical technique
-confers the presence of viral antigens in
organs outside the NS (GI-tract, heart, etc.)
-biopsies are stained with immunoperoxidase,
to expose antigens, and treated w/ labeled
rabies-specific antibodies to detect antigens
-benefits
▪ reduces risk to technician because tissue sample
are embedded in formalin-fixed paraffin

▪ can examine the spread of the virus in organs

outside of the nervous system

Jogai 2000
 4. Mouse inoculation test
-brain material from the patient in question is inoculated
into mice to see if it leads to fatality
-this procedure takes at least a week

5. RT-PCR
-Reverse Transcriptase-Polymerase Chain Reaction
-can make a DNA copy of the viral genome and use
PCR, with a primer specific to the rabies genome, to
determine its presence

Meslin 1996
Pre- and Post-exposure
Prophylaxis
 Pre-exposure Prevention

1. Avoid contact with wild animals

2. Do not handle dead animals
3. People that work with wild or domestic
animals should be vaccinated
4. Vaccination of domestic and reservoir
wild animals
 What is an Effective vaccine?
• One that increases the number of antibodies
specific to the viral antigen
– Antigens are expressed on the cell surface and bind
with antibody
– The antibody acts by neutralizing the antigen,
decreasing its pathogenic ability

• Acts as an inhibitor for adsorption and/or

replication of the virus
• Is high in G- or N-viral proteins
[Link]
 Main Types of Vaccines

1. Brain-tissue vaccine
-synthesized from infected brain tissue originally by
Pasteur
-use both adult and suckling mammal brain tissue
(SMB)
-causes neural complications
▪ associated with immune response to neural antigens and
foreign proteins contained in the vaccine
▪ these complications are lower in SMB vaccine
▪ relatively cost-effective
▪ 90% of human vaccination still uses this

Koprowski 1996
 2. Cell-culture vaccine
-prepared from supernatant of virus-infected cells
-two main types
a. chicken-embryo
▪major neurological complications due to embryo
antigens
▪not generally used in U.S. for this reason

b. human-diploid cell vaccine

▪virion preparations grown in human diploid cells
▪requires fewer doses and causes fewer
complications
▪used as standard for preparation of other vaccines
▪very costly

Meslin 1996
 3. Genetically engineered vaccines
-the goal is to conserve the antigenic structure of the G-
protein, while reducing its pathogenic ability
-the recombinant G-protein is modified in such a way
that:
● it is almost identically similar to challenge virus
● it decreases viral uptake, and/or prevents budding, and/or
stimulates high expression of the G-protein
-induces high levels of neutralizing antibodies, allowing
protection against several rabies strands
-safe, potent, cost-effective
-but through recombinant processes in body, wild-type
virus could be regenerated

Morimoto 2001
 Post-exposure Prophylaxis
1. Wash bite wound thoroughly with soap and
water
2. Isolate the animal if possible
3. Seek post-exposure treatment
-same vaccines above are also used in post-exposure
treatment to stimulate the development of
antibodies
-can also use lectins or neurotoxins that are specific to
the nAchR to inhibit viral infection
●these will successfully compete with the receptor, decreasing
viral uptake

Marchetti 1995 and Voyles 1993

Monitoring Post-exposure Vaccines

• Why?
– Efficacy of the vaccine varies with individual
– The generation of high amounts of rabies antibodies
in a short time is imperative for survival

• Methods to monitor vaccine efficacy

1. Mouse Neutralization test (MNT)
– Virus/serum mix at several dilutions are inoculated in mice
and the mortality/survival rate is measured
– Time-consuming, expensive, and need constant supply of
rabies conjugate
2. Rapid Flourescent Focus Inhibition Test
-mix dilute serum with constant dose of CVS
-stain with flourescent antibody to detect presence of non-
neutralized virus
-time-consuming, expensive, and need constant supply of rabies
conjugate

3. ELISA
-dilutions of sera are added to wells coated with G-or N-protein
-detection of rabies antibodies specific to viral protein by
monitoring absorbance
-expensive and equipment may not be readily available
4. Latex agglutination test
-new technique that is less expensive, less time-consuming, and
less laborous
-serum of treated patient is tested on beads that have been coated
with purified glycoprotein of the rabies virus
-amount of agglutination is a direct measure of the efficiency of the
vaccine
 Current Research
• Though the rabies virus is an unyielding,
generally fatal disease, it can be used for the
good
1. The virus is being used to map specific neuronal
pathways in the body
2. Through the molecular characterization of the rabies
virus, the transmission from reservoir species to
non-reservoir species can be monitored
 Future Research
• For any of you future microbiology
researchers, here are some aspects of
rabies that need to be further addressed
1. Need clarification of the mechanism of
transport and how rabies actually targets
cells after uptake and replication
2. The evolutionary force resulting in the
development of two or more dominant
strains in a particular region and why with
time one may become obsolete
 Thank You!!!!!!

Dr. E Srikaanth Reddy

Associate professor –G.S.L