Method of Virus Elimination
i) Thermo-therapy and Meri-stem Culture:
Conveniently viruses are eliminated by thermo-therapy of whole plants in which the plants are exposed to temperature between 35 to 40 0C for a few minutes to several weeks depending on the host-virus combination. Thermo-therapy is based on the fact that most viruses are killed at temperature much below those which kill their host plants. Thermotherapy is usually effective against iso-metric and thread –like viruses. Thermotherapy is often combined profitably with meri-stem culture to obtain virus- free plants in general; shoot-tips are excised from heat treated plants since larger ex. Plant can be safely taken from heat treated plants.
Prolonged exposure of shoot tip culture to a low temperature ( 5 0C) has proved quite successful in virus elimination. This is often called Cryo-therapy. E.g. In case of chrysanthemum, shoot tip culture exposed with 5 0C four (4 ) months yieldest 67 % plant free from chrysanthemum stunt virus (CSV) and 22% plants were fre from chrysanthemum chloritic mottle virus ( CCMV). The frequency of virus free plants increased to 73 % in case of CSV and to 49 percent in CCMV after 7.5 months exposure to 5 0C. The cold treatment is preferred as the heat treatment is less injurious to the plants and often more effective in virus elimination.
Some chemicals viz. Virazole, Actinomycin-D, Cyclohexamide, etc. which affect the virus multiplication may be added into the culture medium for curing the shoot tips virus.
4. Virus Indexing:
Testing of plants for the presence or absence of concerned viruses is called virus indexing. Every meri-stem tips or callous derived plants must be tested before using it is a mother plant to produce virus-free stocks. This necessitates indexing of plants several times at periodical intervals and only those individuals which give consistency negative results should be labelled as virus tested for specific viruses.
a) Sap Transmission Test:
In which the sap from test plants may be used to inoculate highly sensitive and healthy indicator plants of a specific virus or group of viruses. An indictor plant of a specific virus is that plant species on variety which is highly susceptible to the virus and readily develop the symptoms. The inoculated indicators plants are maintained in a green house or aphid- proof cages.
b) Elisa Test (Enzyme Linked Immuno Sorbant Assay) :
This test is performed by adding a drop of centrifuged sap of a test plant to a drop of anti-serum taken from the blood of rabbit, if the virus is present, the precipitation will take place due to the presence of specific anti-bodies in the blood. The Elisa is one of the serological methods used to identify virus based on anti-body reaction. It is most convenient, rapid and effective test system especially when a large number of samples are to be handled.