Cultivation Of Virus

Viruses are obligate parasites that are much smaller than bacteria. They are considered as ultra microscopic as they cannot be seen through the light microscope. The medical importance of viruses is due to their ability to cause large varieties of human infections. Diseases range from simple common cold to serious AIDS infection. 

They cannot be grown on artificial culture media, so special techniques have to be employed to grow them outside the host. Commonly three main methods are employed for the cultivation of viruses.

1. Animal inoculation; in the initial days, human beings who volunteered were used as means of inoculation in order to study their characteristics. But this has been adopted only for very serious issues as it created serious risk.
   Monkeys were used for the isolation of pox virus by Landsteiner and Popper in 1909. The application of monkeys in virology was also limited due to the cost and difficulty in handling.

This paved way to the use of white mice (pioneered by Theiler in 1903). Mice are being used widely still now for various study purposes. Infant or suckling mice are used for the cultivation of Arbovirus and Coxsackie virus, as they do not grow in any other systems. Mice may be inoculated by routes like; intracerebral, sub-cutaneous, intraperitoneal or intranasal. Animals like guinea pig, rabbit, etc are also used in some cases.

Death of the animal, disease or lesions may be a sign of the growth of virus in these animals. The main drawback of this method is that, viral growth may be interfered by the immune responses and the animal becomes a harboring agent of the latent virus. Pathogenesis, immune response, epidemiology and oncogenesis also utilize the animal inoculation method for study.

2. Embryonated eggs; the Embryonated eggs provide a variety of sites for the virus cultivation. Chorioallantoic membrane inoculation produces visible lesions known as pocks. The pock morphology varies with viruses. One pock is formed by each infectious particle under optimum conditions. These are employed for variola and vaccinia. Allantoic cavity inoculation is used for the cultivation of influenza virus and paramyxovirus. Yolk sac inoculation is used for chlamydiae and rickettsiae. Duck eggs are employed for the preparation of inactivated non-neural rabies vaccine.

3. Tissue culture; cultivation of small pieces of tissues and organs in-vitro for virus cultivation has been practiced long back. Vaccinia virus was cultivated in the corneal fragments of rabbit by Steinhardt and colleagues in 1913. Bacterial contamination on these tissues is prevented by the use of antibiotics. Three types of tissue culture are available:

1. Organ culture: small bits of organs are maintained in-vitro for the isolation of some viruses. Example, Corona virus is isolated from tracheal ring organ culture.
2. Explant culture: fragments of shredded tissues are grown as explants inserted in plasma clots. Example, Adenovirus isolation from adenoid tissue explants cultures.
3. Cell culture: this is the most commonly employed method. Tissues are dissociated into smaller cell components by proteolytic enzymes like trypsin. By a series of treatment steps, along with the addition of antibiotics, the cell suspension is used for growing the virus particles. Methods like cytopathic effect, metabolic inhibition, hemadsorption, immunfluorescence, etc are employed to detect the growth in cell cultures.
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