|Structure of Rabies Virus|
– rod or bullet- shaped (75 x 180 nm) ; one end conical, other planar ( concave).
– Membranous envelope with protruding spikes or peplomers (10 nm) which are composed of trimers of viral gp (G) spikes don’t cover the planar end.
– Beneath the envelope is the membrane or matrix (M) protein layer
– membrane may project outwards from planar end of some virion forming a bleb.
– The core of the virion consists of helical RNP ( group specific antigen).
– Genome is unsegmented, linear, ss(-)sense RNA ( 12kb, MW 4.6 X 106).
– virion contain an RNA dependent RNA polymerase.
– survives at 40C for weeks and at -700C for years.
– sensitive to ethanol, iodine- preparations, 40– NH4+ compounds, soap, detergents and lipid solvents ( ether, chloroform, acetone).
– inactivated by phenol, formalin, ß- propiolactone( BPL), UV or sunlight; thermal inactivation at 500C in 1 hr, and at 600C in 5 minute.
– inactivated by CO2, so, for storage in dry ice, it should be sealed in glass vials.
– virus attaches to cells via its gp spikes; nicotinic acetylcoholine receptor may serve as cellular receptor ( in neurons).
– Entry is by endocytosis.
– genome transcribed by viral RNA plymerast to 5 mRNA species; the monocistronic mRNA species code for 5 virion proteins; nucleocapsid (N); polymerase proteins (L,P); matrix (M), and gp (G).
– Genome RNP is a template for complementary(+) sense RNA, which is responsible for generation of (-) sense progeny RNA
– same viral protein serves at polymerase for viral RNA replication as well as transcription.
– newly replicated genomic RNA associates with viral transcriptase and nucleo- protein to form RNP cores in the cytoplasm; M- protein important in packaging the RNA and protein N and linking it to the envelope.
– The particles acquire an envelope by budding through the plasma membrane.
– rabies virus has a wide host range; all warm- blooded animals including human can be infected.
– Susceptibility varies; high (fox, wolves, bats, cats), to moderate ( dogs, sheep) to low ( opossums).
– Virus is widely distributed in the nervous system, saliva, urine, lymph, milk and blood of infected animals.
– when freshly isolated in lab, strains are referred to as “ street virus”; these cause fatal encephalitis in lab animals following inoculation by any route, after a long and variable incubation period ( 1-12 weeks) and regularly produce intracytoplasmic inclusion bodies (Negri bodies).
– serial brain to brain passage in rabbits of the street virus yields a fixed virus that no longer multiplies in extraneural tissues; after intracerebral inoculation, it produce fatal encephalitis after a short and fixed incubation period of 6.7 days; negri bodies usually not demonstrable.
– there is a single serotype of rabies virus; however there are strain differences among viruses isolated from different species.
– substitution at AA position 333 of gp results in loss of virulence, where Arg has replaced Gln or Ile in the more pathogenic variant.
– purified spikes containing viral gp elict neutralizing( protective) antibody; thus provide a safe and effective subunit vaccine.
– antiserum prepared against purified nucleocapsid Ag used in diagnostic IFT, the antibody are not protective and are group specific.
– virus passesses haemagglutinating activity ( property of gp spike), optimally seen with goose erythrocytes at 0-40C and PH 6.2.
– gp spikes are inactivated by heat ( 560C for 30-60 min), ether, trypsin, deoxycholate or Tween 80, but not by BPL.
1. short prodrome. 2. acute neurologic( encephalitic) phase 3. coma / death
• prodrome lasting 2-10 days may show malaise, anorexia, headache, photophobia, nausea and vomiting, sore throat, fever; there is profound sense of apprehension, anxiety, agitation, irritability, insomnia or depression.
During the acute neurologic phase( 2-7 days), there are sign of nervous system dysfunction which begins with hyperactivity → bouts of bizarre behaviour, agitation or seizures.
• pathognomic feature is difficulty in drinking, together with intense thirst; attempts to drinking may bring painful spasms of pharynx / larynx producing choking and gagging, patients develop a dread of even the sight or sound of water ( hydrophobia).
• generalized convulsions follow, death occurs with in 1-6 days due to respiratory arrest during convulsion.
– furious rabies→ predominantly encephalitic disease with neurologic dysfunction.
– dumb rabies → paralytic illness; with symmetrical ascending paralysis followed by coma and death, occurs in about 20% patients.
– now that survival from established rabies has been demonstrated in rare instances, it is necessary to be able to make lab distinct between rabies and other form of encephalitis.
•tissues infected identified rapidly by IFT or immunoperoxidase staining using anti- rabies monoclonal antibody.
•specimens: salivary, corneal or conjunctival smears or skin biopsy from nape of the neck ( ante mortem).
or impression smears of cut surface of salivary gland, hippocampus, brain stem or cerebellum ( post mortem).
– RT- PCR to amplify parts of rabies virus genome; useful in molecular epidemiology to trace the source of virus in cases with out a history of exposure.
available tissue inoculated intracerebrally into suckling mice this results in encephalitis and death; CNS of inoculated animal examined for Negri bodies and rabies antigen ; cytopathic effect is minimal.
isolated virus identified by fluorescent Ab- tests with specific antiserum
– serum antibody detected by IFT or Nt tests; such antibody develop slowly (7-10 days) since the onset of illness).
– Antibody in CSF present in rabies – infected individuals, but not in response to vaccination.
– a latex agglutination test on saliva from rabid dogs gives 99% specificity and 95% sensitivity when compared with IFT on brain smears.
– All animals considered rabid or suspected rabid should be sacrificed immediately for lab examination of neural tissues; other animal should be held for observation for 10 days.
– The brain of infected animal may be removed carefully and 2 portions
•one in 50% glycerol saline ( for biological test) and
• other in jenkers fixative ( for microscopy).
– Examination of salivary glands by IFT for demonstration of viral antigen.
– impression smears of brain stainded by sellers technique ( basic fuchsin and methylene blue in methanol) to find Negri bodies.
IMMUNITY AND PREVENTION
• local treatment : prompt cauterization of wound destroys the virus
– wound should be thoroughly cleaned with soap solution / detergent and running water for 5 minute, followed by application of 40-70% alcohol or tincture of iodine or 40– NH4 compound ( cetavlon).
– scrubbing should be avoided and suturing delayed if possible.
•passive treatment with antirabies immunoglobulin (Ig) preferably human, 20 IU/ Kg body weight, given half in an around the wound and half in the gluteal muscle.
active immunization: with inactivated whole virus vaccine ( cell – culture grown), containing atleast 2.5 IU/ dose, given into deltoid muscle.
all vaccines for human use contain only inactivated rabies virus.
– associated with serious risk of neurological complications ( postvaccination encephalitisP due to the presence of encephalitogenic factor ( a basic protein associated with myelin)
– low potency per dose; poor immunogen.
– complete treatment involves upto 23 painful injections.
1.egg vaccine: duck embryo vaccine → discontinued due to poor immunogenicty
live attenuated vaccine → prepared in chick embryos ( eg flury strain)
– used for animals but not for humans
– low egg passage of 40-50 (LEP) for dogs.
– High egg passage of 180 (HEP) for cattles and cats.
a. Human diploid cell vaccine (HDCV): Virus adapted to growth in WT- 38 human normal fibroblast cell line.
– virus preparation concentrated by ultrafiltration and inactivated with BPL
– highly antigenic and free from serious side effects; high cost.
b. rabies vaccines, adsorbed ( RNA): Made on diploid cell line derived from fetal rhesus monkey lung cells; inactivated with EPL and concentration by adsorption to aluminum phosphate.
c. purified chick embryo cell vaccine ( PCEC) prepared from fixed rabies virus strain flury LEP grown in chicken fibroblasts; inactivated with BPL and further purified by zonal centrifugation.
– prepared by cold ethanol fractionation from plasma of hyperimmunized humans
– concentrated serum from horses hyper immunized with rabies virus.
- indicated for persons at high risk ( researchers, lab workers, veterinarians )
- To attain an antibody level presumed to be protective by means of vaccine administration prior to any exposure.
- Two injections of 1 ml vaccine given into deltoid muscle 4 weeks apart; re- inforcing dose given at 12 months.
POST EXPOSURE PROPHYLAXIS
decision to administer rabies antibody, rabies vaccine or both depend on
→ aerosols released during centrifugation of infected materials in lab.
→ Ingestion of flesh of rabid animals ( high dose would be necessary)
– Only documented cases involve transmission by corneal transplants.
– no successful treatment for clinical rabies; symptomatic treatment may prolong life, but outcome is almost always fatal.
– preexposure vaccination is desirable for all persons who are not at high risk of contact with rabid animals, but this doesnot eliminate the need for prompt post exposure prophylaxis if an exposure to rabies occur.