6 ways to eliminate viruses during micropropagation

6 ways to eliminate virus during micropropagation

Viruses are one of the major causes of yield losses in crops around the world. They can enter the tissue culture process through different pathways. However, did you know about different ways to eliminate viruses?

Viral infections have been a critical constraint in the development of sustainable agriculture production. In some cases, a single viral infection can lead to a yield loss of up to 60%. When a plant is infected with multiple viruses, it can lead to even more significant losses than a single virus infection in field conditions. These infections lead to similar results in in vitro conditions as well.

One of the most important requirements for successful plant tissue culture procedures is to have sterile plants. Any kind of contamination can lead to the failure of the whole process. Viral infections can prove to be lethal for plantlets in tissue culture. You can control viral infections to some extent by treating mother plants with chemicals. However, there is a probability that these chemicals are not as effective as needed. Also, there is always a case of chemical buildup on the plant surface. So let's avoid this method!

What can we do to eradicate viruses?

When the question of obtaining virus-free plants arise, the best methods are different in vitro thermotherapy-based methods. Let us briefly talk about them:

Different in vitro methods for virus elimination are:

  • Thermotherapy with shoot tip culture
  • Low-temperature therapy
  • Meristem culture
  • Micrografting
  • Chemotherapy
  • Cryotherapy

You can use these methods in combination such as thermotherapy with meristem culture, chemotherapy with shoot tip cryotherapy, etc. You can obtain much more efficient results to eradicate viruses when you combine different methods instead of using just one of these methods.

And if you want to know more about different contaminations that can occur in your tissue culture experiment, then do consider reading our article on "Different types of contaminants in plant tissue culture"

Thermotherapy

This method involves exposing your explants or callus cultures to higher temperatures for a pre-defined duration to eliminate viruses. Each plant species has a suitable temperature range that can be exploited for thermotherapy. Oftentimes, you can use thermotherapy of 35-42 °C for 4-6 weeks for your target plants. You can experiment with different temperatures to find out which temperature would suit your chosen plant species the best. This range will also depend on the type of virus infecting your chosen plant species.

However, the duration for which you expose the explants plays a critical role. You need to decide the thermotherapy duration in such a way that your treated plants survive the higher temperatures and also, have the virus inactivated at the same time. This should give you your desired virus-free plants.

Other important factors to keep in mind before starting thermotherapy are the size of the explants, the position of the explants, and the viral infection status.

Different studies have shown successful viral eradication using thermotherapy in potatoes, grapevines, citrus, and strawberries.

Low-temperature therapy

According to scientists, treating your infected cultures at higher temperatures can only help in eradicating viruses. However, there is another class of viruses that causes severe growth disorders in plants; these are called 'viroids'. It is not possible to eliminate viroids from plant material at higher temperatures. Therefore treating your plant material at temperatures below 10 °C is effective for eradicating viroids such as potato spindle tuber viroid, chrysanthemum chlorotic mottle viroid, etc.

Meristem culture

It is one of the most popular methods of virus elimination in the plant science world. Not only this, but it also has the potential to eradicate fungal and bacterial infections from mother plants. Several ornamental plants such as orchids and fruits like bananas are cultured using meristem tissue culture for large-scale production.

Meristems are the centers of plant growth present in apical and lateral buds and even roots. Meristematic tissue possesses an undifferentiated group of cells that are capable of dividing. These cells can produce new cells that can differentiate into different kinds of plant organs as they mature. The important fact is that these cells are free of viruses and thus serve as a healthy starting material for tissue culture. You can read more about meristem culture here!

You can also combine thermotherapy with meristem culture. This method is mostly done for infected in vitro shoots. It includes thermotherapy of infected in vitro shoots followed by shoot tip culture. This method has been successful for the elimination of potato virus Y, cassava common mosaic virus, onion yellow dwarf virus, etc.

Micrografting

This method involves placing a small meristem or a section of micro shoot onto the top of a rootstock, both cultured in vitro. This method mimics the process of grafting, popularly used for vegetative propagation.

This micrografting procedure builds cells over a period of time and joins both the shoot and root parts to form a single plant having features of both micro shoot and rootstock.

This method has been useful to eradicate viruses in different crops such as tomato, citrus, raspberry, garlic, etc.

Chemotherapy

As the name suggests, you can use chemotherapy by using certain chemicals in the tissue culture media that will prevent virus replication or its movement from infected to healthy parts of your plantlets. Some of the chemicals that are commonly used are acycloguanosine and azidothymidine.

This technique is more effective in combination with other methods. For instance, if you have infected in vitro shoots, then you can culture them on an antivirus chemical containing medium, followed by treatment at higher temperature (thermotherapy) and finally performing shoot tip culture. This combination of methods has been used for the eradication of potato mosaic virus, chrysanthemum B virus, pear vein chlorosis virus, etc.

Shoot tip cryotherapy

When different industries want to preserve plant germplasms, they resort to this method. This involves the preservation of shoot tips from different in vitro cultures in liquid nitrogen at -196 °C. Hence called cryopreservation. As shoot tips possess meristematic cells, they are free from viruses and you can preserve virus-free material. These tips also have lower water content as compared to other plant parts. Due to this reason, ice crystals formed by preserving these tips at extremely low temperatures do not damage the cells. This technique has been successfully used to eliminate viruses like the banana streak virus, potato leafroll virus, etc.

These are the different methods to eliminate viruses from selected plants. These methods will lead to the hassle-free process of micropropagation. You can have a better production of virus-free plants when you know the limits of your chosen plant species such as temperature tolerance and also combine different methods.

We hope that many of you enthusiasts found this article interesting. For more informational posts on different aspects of plant tissue culture, keep checking this space!

Also if you like this article, do share it with others as well. Let us build a wonderful community of plant tissue culture enthusiasts.

By Nancy Bhatia | 24-August-2021

References

  1. https://safety.fsu.edu/safety_manual/supporting_docs/Understanding%20and%20Managing%20Cell%20Culture%20Contamination.pdf
  2. https://study.com/academy/lesson/cell-culture-contamination-types-identification.html
  3. https://www.technologynetworks.com/cell-science/how-to-guides/how-to-prevent-cell-culture-contamination-htg-299231
  4. https://plantmethods.biomedcentral.com/track/pdf/10.1186/s13007-018-0355-y.pdf
  5. Varveri, C., Maliogka, V. I., & Kapari-Isaia, T. (2015). Principles for Supplying Virus-Tested Material. Control of Plant Virus Diseases - Vegetatively-Propagated Crops, 1–32. doi:10.1016/bs.aivir.2014.10.004
  6. Kyte, Kleyn, et al (2013) Plants from test tubes: An introduction to micropropagation. Timber press, Inc.
  7. Bhojwani, S.S., & Dantu, P.K. (2013). Plant Tissue Culture: An Introductory Text. Springer India