Rhabdo Viruses (Rabies virus)

Rhabdoviridae
Rabies Virus

1. Rhabdoviruses
(Rabies Virus)

2. Family Rhabdoviridae
•Comprises of two genera:
oGenus Lyssavirus-Contains Rabies virus and Rabies related
viruses
oGenus Vesiculovirus-Contains Vesicular stomatitis viruses
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3. RABIES VIRUS-
Morphology
•Bullet-shaped(75 nm in width &180 nm in length)
•Enveloped-lipid envelope (10 nm long)
hemagglutinatingpeplomerspikes (glycoprotein-G) are
embedded.
•Envelope is lined internally by a layer of matrix protein
•Nucleocapsid-helical symmetry & comprises of-
single-stranded, negative-sense RNA plus
nucleoprotein and polymerase proteins
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4. Major antigens of Rabies virus
Glycoprotein G Nucleoprotein
Peplomersor spikes embedded in envelope Capsid proteins associated with viral RNA
Species-specific Group-specific and cross-reactions are seen with rabies related
viruses.
Role in pathogenesis-It binds to acetyl choline receptors in
neural tissues, which is the first step of pathogenesis
(attachment)
Does not have any role in pathogenesis
Diagnostic role-It induces hemagglutinationinhibiting
antibodies which can be detected in patient's serum by
hemagglutinationinhibition test
1.CFT-It induces complement fixing antibodies which can be
detected in patient's serum by complement fixation test
2. Antiserum prepared against the purified nucleocapsid is used in
immunofluorescence test
Role in immunity-It induces neutralizing antibodies which are
protective in nature & also stimulates cytotoxic T cells
Antibodies are not protective.
Role in vaccination-Because it is protective in nature, the
purified glycoproteins may therefore provide a safe and
effective subunit vaccine.
Not used for vaccination
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5. Animal Susceptibility
•Infects all warm-blooded animals, including humans.
•Susceptibility varies among various animals.
oVery highly susceptible animals-Foxes, jackals, wolves and cotton rats
oHighly susceptible animals-Rabbits, cattle, cats, hamsters, raccoons and bats
oModerately susceptible animals-Dogs, goats, sheep, horses & non-human primates
oLow susceptible animals-Opossums
•Rabies virus also undergoes certain changes when it is serially propagated in
animals.
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6. Street viruses& Fixed viruses
Street viruses Fixed viruses
Freshly isolated Isolated after serial intracerebral passage in
rabbits/cell lines
Produce Negri body Do not produce Negri body
Affect salivary glands Do not affect salivary glands
Pathogenic Not pathogenic (except-if not inactivated
properly)
Incubation Period:1-3 months (20-
90days)
Incubation Period:
4-6 days
Produce disease Used for vaccine
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7. Transmission
•Dog biteis the most common mode
•Others -monkey, sheep, goat, cat, buffalo and horse (except rat bite and
human bite)
•Bat bite(mostly goes unnoticed)-Migrating fruit-eating bats
•Human-to-human transmission -extremely rare.
•Non-bite exposuresare rare such as-
oLick on abrasion or mucosa
oInhalation of virus containing aerosols generated from infected bats.
oCorneal transplantation
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8. Pathogenesis of rabies
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9. Histopathological changes
•Mononuclear cell infiltration
•Perivascular cuffing of lymphocytes
•Babes nodules consisting of glial cells
•Negribodies
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10. Negribodies
•Intracytoplasmic eosinophilic inclusions in brain neurons
•Composed of rabies virus proteins and viral RNA
•Site -Purkinje cells of the cerebellum and in pyramidal neurons of the
hippocampus, and are less frequently seen in cortical and brainstem neurons.
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11. Clinical Manifestations
•Incubation period-prolonged and variable, ranges from 1 week to 19 years
•Incubation period inversely related to the distance for the virus to travel from
the site of inoculation to CNS. Hence it is usually shorter in-
oChildren than in adults
oBites on head, neck & upper limbs than legs
oShort people
oSevere lacerations
oPresence of genetic predisposition
oLow host immunity
oVirus-High dose of inoculum and from the site of inoculation to CNS and ↑virulence of
the strain
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12. Clinical spectrum
•Prodromal phase
•Acute neurologic phase
•Coma and death.
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13. Prodromal phase
•Lasts for 2-10 days
•Characterized by non-specific symptoms such as fever, malaise, anorexia,
nausea, vomiting, photophobia, sore throat, abnormal sensation (paresthesia,
pain, or pruritus) around the wound site.
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14. Acute neurologic phase
•Either encephalitic (80%) or paralytic type (20%)
•Encephalitic rabies (2-7days) :
oHyperexcitability
oLucid interval
oAutonomic (sympathetic) dysfunctionfeatures may be seen such as
↑lacrimation, ↑salivation (leads to foamingat the mouth), ↑ perspiration,
gooseflesh, cardiac arrhythmia and priapism.
oHydrophobia(fear of water) or aerophobia (fear of air)
•Paralytic or dumb rabies (20% of cases)
oEspecially in people who are partially vaccinated or infected with bat rabies virus.
oCharacterized by flaccid paralysis quadriparesiswith facial paralysis.
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15. Coma and death
•Patient develops coma that eventually leads to death within 14
days
•Patients with paralytic rabies may survive longer up to 30 days.
•Death is almost certain. Recovery and survival are extremely
rare.
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16. Laboratory diagnosis-
Rabies Antigen Detection
•Direct immunofluorescence test (direct-IF); also called as direct fluorescent
antibody (DFA) test:
oPerformed to detect rabies nucleoprotein antigens in specimens by using specific
monoclonal antibodies tagged with fluorescent dye
oHigh sensitivity and specificity
o“Gold standard”
oBest specimen -hair follicle of nape of neck (most sensitive)
oCorneal impression smear can also be used. It is usually positive in late stage with a
sensitivity of 30%.
•Immunohistochemistry
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17. Viral Isolation
•Mouse inoculation: Intracerebral inoculation into suckling mice can cause
encephalitis and death.
•Brain biopsies -Examined for Negribodies and rabies antigen
•Cell lines: Mouse neuroblastoma cell lines and baby hamster kidney (BHK) cell line
oYield virus (2–4 days) much faster than that of mice inoculation
oViral growth -detected by direct-IF test using specific antiserum.
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18. Antibody detection
•Detection of CSF antibodies is more significant than serum antibodies.
•CSF antibodies appear early and they are produced only in rabies-infected
individuals but not in response to vaccination.
•Various antibody detection tests include:
oMouse neutralization test (MNT)
oRapid fluorescent focus inhibition test (RFFIT)
oFluorescent antibody virus neutralization (FAVN)
oIndirect fluorescence assay (IFA)
oHemagglutinationinhibition test (HAI)
oComplement fixation test (CFT)
oImmunoperoxidaseinhibition assay.
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19. Viral RNA detection
•Reverse transcription-polymerase chain reaction (RT-PCR) can be used to
amplify genes of rabies virus RNA from fixed or unfixed brain tissue.
•Most sensitive and specific assay available
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20. Negribody detection
•Useful for post mortem diagnosis of rabies.
•They are intracytoplasmic eosinophilic inclusions with
characteristic basophilic inner granules.
•Sharply demarcated, spherical to oval, and about 2-10 µm in
size.
•Most common sites of Negribodies are neurons of
cerebellum and hippocampus.
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21. Negribody detection
•Commonly used stains are-histological stains such as
H&E and Sellers stains (basic fuchsinand methylene
blue in methanol).
•Immunohistochemistry-Peroxidase labelled specific
antibodies are used to detect the viral inclusions in
formalin-fixed tissues.
•Negribody detection is pathognomonic of rabies.
(may not be detected in 20% of cases)
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Negribodies in brain biopsy by
H and E stain

22. Treatment
•No specific treatment for rabies.
•Symptomatic treatment may prolong life, but the outcome is almost always
fatal.
•Isolation-Patient should be isolated in a quiet room, protected as far as
possible from external stimuli such as bright light, noise, water or cold air
which can precipitate spasms.
•Sedative and anti-anxiety drugs such as morphine can be used.
•Hydration and urination should be properly maintained
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23. Prognosis
•Mortality in Rabies is almost 100%
•Preventable by administration of post-exposure therapy during
the early incubation period.
•There are seven well-documented cases who survived from
rabies –mostly because of taking rabies vaccine in the early
incubation period.
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24. Prevention of human rabies
•Post-exposure prophylaxis (PEP) includes local wound care and both active
and passive immunization.
•Local wound carecan greatly decrease the risk of rabies if initiated
immediately.
oPhysical cleansing
oChemical inactivation-Antiseptics such as povidone iodine or alcohol can be used to
inactivate the residual viruses.
oBiological neutralization of the virus by giving anti-rabies immunoglobulin
oDevitalized tissues should be debrided
oTetanus prophylaxis should be given
oAntibiotic treatment is initiated to prevent secondarybacterial infection
oSuturing is contraindicated
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25. Passive Immunization (Rabies Immunoglobulin)
•Rabies Immunoglobulins (RIG) -causes neutralization and loss of infectivity of
the virus. Hence RIGs are usually administered locally at the site of exposure.
•Two typesof RIGs are available:
oEquine rabies immunoglobulin (ERIG)-It is given at dose of 40 IU/kg . Being
heterologous in origin (horse), it is associated with serum sickness.
oHuman rabies immunoglobulin (HRIG)-It is given in a dose of 20IU/kg. It is devoid of
side effects.
•Maximum volumeof RIGs should be infiltrated into and around the wound(s);
remaining volume if any should be administered by deep IM injection at a site
distant from the vaccine injection site.
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26. Active Immunization –
Neural vaccine
•Neural vaccines-derived from the nervous tissues of animals infected with
the fixed rabies virus.
•No longer in use since 2004 -Encephalitogenic, poorly immunogenic and are
associated with serious risk of neurological complications.
•Examples include-
oSemplevaccine: Derived from infected sheep brain, inactivated with phenol.
oBeta propiolactone(BPL) vaccine: Modified Semplevaccine which is inactivated with
beta propiolactoneinstead of phenol
oInfant mouse brain vaccines: Derived from infected neural tissue of newbornmice.
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27. Active Immunization –
Non –Neural vaccine
•Both cell culture vaccines (CCVs) and purified duck embryo vaccines (PDEV)
are currently used in India.
oEgg derived vaccines-Allantoiccavity of embryonatedeggs provide excellent site for
the preparation of rabies vaccine.
Purified duck embryo vaccine (PDEV)-Duck eggs being larger, give a higher yield than that of hen.
Unlike the neural vaccine, it is less reactogenic, but it is less antigenic too; hence multiple (16–25)
doses have to be given to obtain a satisfactory antibody response. It is no longer manufactured.
Live attenuated chick embryo vaccinesuch as Flurystrain was in use before for vaccinating animals,
but now it is obsolete.
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28. Active Immunization –
Non –Neural vaccine
oRecombinant viral vaccine-Vaccinia virus carrying the rabies surface glycoprotein gene
has been developed.
Given orally, has been successful for immunizing animals, but still in experimental stage for human use.
oCell culture derived vaccinesare the most recommended vaccine for prevention of
rabies. They are highly immunogenic and devoid of neurological complications. Three
vaccines are available in India -
Purified chick embryo cell (PCEC) vaccine is prepared from chicken fibroblast cell line
Purified Vero cell (PVC) vaccine is prepared from Vero cell line
Human diploid cell (HDC) vaccine is derived from WI-38 (human embryonic lung fibroblast cell line).
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29. National Guideline on Rabies Prophylaxis
•Regimen for post exposure prophylaxis-
oIM regimen or Essen regimen (1-1-1-1-1): Five doses of intramuscular injections; one
dose (0.5 or 1ml) each given on days 0, 3, 7, 14 and 28. Day 0 indicates the date of
administration of first dose of vaccine and may not be the date of rabies
exposure/animal bite.
oID Regimen(or Thai Red Cross Schedule) (2-2-2-0-2)-This involves injection of 0.1ml of
reconstituted vaccine on two sites per visit on days 0, 3, 7 and 28.
•Potency-Single intramuscular dose should have a minimum potency of 2.5 IU.
•Site of injection:
•Deltoid region is ideal site (Gluteal region -not recommended)
•Infants and young children-Antero-lateral part of the thigh is the preferred
site.
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30. National Guideline on Rabies Prophylaxis
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Category of risk Type of exposure Recommended prophylaxis (WHO)
Category I
(No risk)
Touching, or feeding of animal
Licks on intact skin
No treatment needed if history is
reliable
Category II
(Minor risk)
Minor scratches or abrasions without
bleeding or
nibbling of uncovered skin
•Wound management
•Rabies vaccine
•Observe the dog for 10 days
Category III
(Major risk)
Single or multiple transdermal bites
with oozing of blood,
Licks on broken skin (fresh wounds)
or mucous membrane
Bite by wild animals /bat
•Wound management
•Rabies immunoglobulin
•Rabies vaccine
•Observe the dog for 10 days

31. Regimen for pre-exposure prophylaxis
•Recommended for high risk groups:
oLaboratory staff handling the virus and infected material
oClinicians or any person attending to human rabies cases
oVeterinarians
oAnimal handlers and travellers from rabies free areas to rabies endemic areas.
•Three dosesare given at day -0, 7, and 21 or 28 days either by IM (0.5ml) or
ID (0.1ml) schedule.
•Antibody titre -checked every 6 months for 2 years and thereafter every 2
yearly. Booster dose is given if it is less than 0.5 IU/ml
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32. Regimen for post exposure prophylaxis in
previously vaccinated individuals
•Depends on the antibody titer and the severity of the wound
bite:
oSevere bite or titer unknown: 3 doses are given at -0,3 and
7 days
oLess severe bite or titer >0.5 IU/ml: 2 doses given at -0 and
3 days
•Immunoglobulins are usually not needed
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33. Epidemiology
•Rabies is enzootic and epizootic disease of both wild and domestic animals worldwide.
•Source-Infected dog is the source of infection in 99% of cases.
•Age-Though all ages are affected, children aged 5–15 years are at greater risk.
•Rabies free countries-Defined as the areas where no case of indigenously acquired
rabies has occurred in man or animals for 2 years. There are few countries/places
which are rabies free-
oAustralia and Antarctica
oBritain
oIceland & Ireland
oChina -(Taiwan), Cyprus
oJapan, Malta
oNew Zealand
oIndia-Andaman & Nicobar, LakshadweepEssentials of Medical Microbiology

34. Rabies in Dogs
•As 90% of human rabies is caused by dog bites, control of rabies in dogs is the
most important step to prevent human rabies.
•Clinical features:
oRabies in dog has a typical clinical course.
oIncubation period varies 3-8weeks (ranges from 10 days to few years)
oLike in humans, two types of manifestations may be seen
•Furious rabies (or Mad dog syndrome)
•Dumb or paralytic rabies
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35. Rabies in Dogs
•Control of rabies in dogs:
oMost logical and cost effective approach for control of urban rabies is-Elimination of
stray dogs and mass immunization of at least 80% dogs in an area.
•Immunization of dogs: All dogs should receive a primary immunization at 3-4
month of age, followed by boosters as per the type of vaccine used. Vaccines
commonly used in dogs are-
oBPL inactivated neural vaccine
oLive attenuated chick embryo vaccine
oOral recombinant glycoprotein vaccine
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36. OTHER RHABDOVIRUSES
•Other Lyssaviruses (Rabies like viruses)-
oEuropean bat Lyssavirus1 and 2 (in Europe)
oAustralian bat Lyssavirus(in Australia)
oDuvenhagevirus (in Africa)
oLagos bat virus (in Africa)
oMokolavirus (in Africa)-isolated from shrews (not bat)
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