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cultivation of virus
- 2. • Cultivation of virus Virus are considered as the primary infection causing agents in human Viruses are obligatory intracellular parasite Host for viruses may be animals ,plants, human, bacteria,fungus, protozoan and algae. Hence virologist use any one of the host system for viral culture Better treatment of infections needs proper diagnosis .Isolation or cultivation of virus is one of part of diagnosis
- 3. TECHNIQUES IN CULTIVATING AND IDENTIFYING ANIMAL VIRUSES Viruses require living cells as their “ medium” In vivo-laboratory bred animal and embryonic bird tissue In vitro-cell or tissue culture method
- 4. THE PRIMARY PURPOSE OF VIRUS CULTIVATION To isolate and identify virus in clinical samples To do research on viral structure, replication, genetics and effects on host cell To prepare viruses for vaccine production
- 5. Viruses are cultured by using three methods Inoculation of virus into animal Embryonated Egg Tissue culture
- 6. INOCULATION OF VIRUS INTO ANIMAL In 1932 used mice for viral cultivation Here after animals like rabbit , guinea pig ,rat , suckling mice, hamsters ,and monkeys are used for viral cultivation Inoculation site may vary depends up on the type of virus and target site of infection Eg ; intra cerebral inoculation is performed on mice to cultivate Arboviruses After inoculation observe morphological and physiological changes of animals
- 7. ADVANTAGES All types of animal viruses are cultivated. Used to understand pathogenesis. Used to understand the immune response of the animal. Used to study the efficiency of the vaccines Used to develop vaccines Used to study interaction of drugs
- 8. DISADVANTAGES Expensive Difficult to handle Show biological diversity May leads to latent infections Require efficient maintainence Objection from blue cross members
- 10. EMBRYONATED EGG Woodruff and goodpasteur (1931)used fertilized chicken egg for viral cultivation this is a simpler technique than animal inoculation ,are inexpensive and easily available . Eggs usually not interfere with virus multiplication due to absence of immune response. Suitable cells for the growth of viruses are available in embryo and its membrane ,which may facilitate the growth of virus. 8-11day old chick embryo is used for cultivation. Incubate for 2-9days after inoculation . Duration of incubation is also depends on the type of virus and the route Of inoculation .
- 12. VIRUSES ARE INOCULATED VARIOUS SITES OF EMBRYONATED EGG To prepare the egg for viral cultivation The cell surface is disinfected with iodine Make hole with the help of drill Inoculate viruses in any one of the method mentioned After inoculation is sealed with gelatin and the egg is incubated Viruses reproduce only certain parts of the egg
- 13. TABLE SHOWS THE SITE OF RESPECIFIC VIRUS FOR CULTIVATION SITE OF INOCULATION Chorio allontoic membrane inoculation (CAM) Aminotic inoculation Allontoic inoculation Yolk sac inoculation VIRUS Herpes simplex virus pox, virusrous, sarcoma Influenza virusMumps virus Influenza virusmumpus virus. New castledisease virus Herpes simplex virus
- 14. VIRUS ARE INOCULATED INTO VARIOUS SITE OF EMBRYONATED EGG Viral suspension or virus containing tissue is injected into the fluid of the egg . Virus is injected into the proper location in the egg( virus are able to reproduce only in certain parts of embryo Death of the embryo or formation of typical pocks (pock assay) Haemagglutination or lesion on the membranes of the egg that result from viral growth After growth viruses are identified with the help of serological techniques . Eg ; pox virus produce pock or lesion . Influenza virus produce haemagglutination.
- 16. TISSUE CULTURE Cultivation of tissue or organ for the growth of viruses. Three types of tissue culture Organ culture Explant culture Cell culture
- 17. TYPES OF TISSUE CULTURE ORGAN CULTURE are mainly clone for highly specialized parasite of organ Eg;tracheal ring culture is clone for isolation of corona virus EXPLANT CULTURE fragments of tissue can grown as explant in plasma clot this method is rarely used cell cuture CELL CUTURE it is mostly used for cultivation of viruses.
- 18. CELL CULTURE Now a days cell cultures are mainly employed in cultivation /isolation of virus Growth of cells disassociated from the parent tissue by mechanical process is called cell culture. Cell cultures are sole system for virus isolation Cell culture was first successfully under taken by ROSS WARRION in 1907 ROUX In 1885 for the first time maintained embryonic chick cells in a cell culture
- 20. TYPES OF CELL CULTURE Primary cell culture Diploid cell lines Continuous cell
- 21. PRIMARY CULTURE Primary cultures are derived from the normal tissues of an animal. Can be sub cultured only once or twice These are widely used as the best culture system because they support the widest range of virus Eg ; monkey kidey cells Once the primary is subcultured it becomes knows as cell line A characterized cell line derived by selection or cloning are called cell strains
- 22. DIPLOID CELL LINE After the first subculture the pimary subculture and becomes known as diploid cell line Derived from human fatal tissue and can be subcultured 20-50times .eg; human embroyonic kidney These diploid cells are the most employed host of choice for the the production of human vaccine virus
- 24. Continuous cells Continuous cell lines are capable of an infinite number of doublings. Such lines may arise with the mutation of a cell strain or more commonly from the established cell cutures from malignant tissue .many viruses which are difficult or impossible to grow have been cultured in continuous lines.
- 26. CUTURED VIRUS ARE IDENTIFIED BY Cytopathic effect Haemadsorption Interference Immuno fluorescence test