Viral Transport Media (VTM)- Principle, Preparation, Uses, Limitations

Last Updated on October 11, 2020 by Sagar Aryal

Viral Transport Media (VTM) Definition

A viral transport media (VTM) is a nutrient substance used to carry and maintain the viability of specimens (viruses) to a laboratory for the identification and further processing of the sample.

  • Transport media for viruses are especially important as they are more labile than other infectious agents.
  • The probability of successful isolation increases when the time interval between collection and inoculation of the culture is less and when the specimen contains as many virus particles as possible.
  • In order to maximize the amount of virus in the specimen, the sample collection should be done early in the patient’s illness.
  • Successful and reliable isolations of viruses can be accomplished when a suitable transport medium is used.
  • The isolation count also increases when viral specimens are sent to either a local laboratory or a reference laboratory.

Viral Transport Media (VTM)

Figure: Viral Transport Media (VTM). Image Source: BD.

Principle of Viral Transport Media (VTM)

  • Viral agents vary widely in composition, stability, structure, morphology, and size. The loosely packaged enveloped viruses obtained from human samples are generally more labile than the non-enveloped dense viruses.
  • Because they are labile to the environmental conditions, the viability of these viruses must be protected by using different solutions.
  • Some enveloped viruses, such as herpes simplex viruses (HSV), are surprisingly stable in the non-frozen state when placed in a suitable holding environment. However, most of the viruses do require a colder temperature.
  • The infectivity of viruses decreases over time, and generally, the decay rate is a function of temperature, so that stability is enhanced by cooling
  • Thus, viral transport media are prepared with the idea of providing appropriate conditions of temperature, pH, and nutrients.
  • The constituents of an appropriate viral transport media are selected to create an isotonic solution containing proteins to protect the viral structure, antibiotics to control microbial contamination, and one or more buffers to control the pH.
  • Viral transport media are prepared with the idea of sustaining the viability of the viral culture or specimens for tests like the nucleic acid amplification test (NAAT) while preventing the drying of the sample.

Composition of Viral Transport Media (VTM)

An ideal viral transport medium would possess many of the following characteristics;

  1. It would preserve the activity of the virus, even at room temperatures.
  2. It would be nontoxic to cell cultures and not obscure the appearance of viral cytopathic effects.
  3. It would have a long shelf life (either in a frozen or non-frozen state).
  4. It would be applicable for both culture isolations and direct tests such as enzyme immunoassays or immunofluorescence.
  • In order to achieve all the above-mentioned characteristics, viral transport media usually have a protective protein component, antimicrobial agents to prevent the chances of contamination, and buffers to maintain the pH of the solution.
  • Some VTMs also have additional ions and minerals that aid in maintaining the viability of such specimens.

Preparation of Viral Transport Media (VTM)

  • A number of viral transport media are commercially available, which can directly be bought and used under the conditions mentioned.
  • Some such media include the COPAN Universal Transport Media and the Eagle Minimum Essential Medium (E-MEM).
  • However, some other media can be prepared locally while maintaining suitable conditions and composition.
  • The composition and preparation of different viral transport differ depending on whether the sample is taken from humans or other animals.

For specimen from humans:

The following is the method of preparation for locally made VTMs for the nasal and throat swabs from humans:

  • Add 10g veal infusion broth and 2g bovine albumin fraction V to sterile distilled water (to 400 ml).
  • Add 0.8 ml gentamicin sulfate solution (50 mg/ml) and 3.2 ml amphotericin B (250 µg/ml) –
  • Sterilize by filtration.

The following is a more elaborate procedure for the preparation of a larger volume of VTMs for humans:

  1. Inactivate 500mL of fetal bovine serum (FBS) by heating for 30 minutes in a water bath at 56.0°C +/- 1.0°C. Use commercially inactivated FBS if available.
  2. Add 50mL of amphotericin B, add 50mL of gentamicin to the inactivate FBS and then filter sterilize the solution through a 0.20 to 0.45 µm filter unit (150mL filter unit).
  3. Add 10mL of the FBS to 500mL of Hanks Balanced Salt Solution (HBSS).
  4. Add 2mL of the Gentamicin/Amphotericin B mixture to the previously formed HBSS with FBS.
  5. Disperse the solutions to bottles and ca the bottles. Mix by inverting the bottle.
  6. Label the bottle along with the date of production, additives, and expiration date.
  7. Aliquot 3mL of the medium into individual sterile conical screw-capped tubes (such as 16x100mm tubes). Keep lids tightly closed after the medium is dispensed. Label the containers with the necessary information.
  8. Store at 2-8°C.

For specimen from animals:

  • For the collection and transport of specimens in animals, two types of VTMs can be utilized.
  • Transport medium 199 is widely used for the collection and transport of clinical specimens from all species.
  • A second medium, PBS-Glycerol transport medium, which is the glycerol-based medium, provides longer-term stability of specimens where cooling is not immediately possible.
  • It is suitable for egg inoculation but not suited for tissue culture inoculation.

Preparation of Transport medium 199

  1. Take a tissue culture medium 199, containing 0.5% bovine serum albumin (BSA).
  2. To 1 litre of above add: benzylpenicillin (2 x 106 IU/litre), streptomycin (200 mg/litre), polymyxin B (2 x 106 IU/litre), gentamicin (250 mg/litre), nystatin (0.5 x 106 IU/litre), ofloxacin hydrochloride (60 mg/litre), and sulfamethoxazole (0.2 g/litre).
  3. Sterilize by filtration and distribute in 1.0–2.0 ml volumes in screw-capped tubes.

Preparation of PBS-Glycerol transport medium

  1. Prepare the phosphate-buffer saline by adding 8 g of NaCl, 0.2 g of KCl, 1.44 g of Na2HPO4, and 0.24 g of KH2PO4. Add distilled water to this mixture to make 1 liter.
  2. Autoclave the prepared mixture and mix 1:1 with sterile glycerol to make 1 liter.
  3. To 1 litre of above add: benzylpenicillin (2 x 106 IU/litre), streptomycin (200 mg/litre), polymyxin B (2 x 106 IU/litre), gentamicin (250 mg/litre), nystatin (0.5 x 106 IU/litre), ofloxacin hydrochloride (60 mg/litre), and sulfamethoxazole (0.2 g/litre).
  4. Dispense 1.0–2.0 ml of transport medium into sterile plastic screw-cap vials.
  5. Store these vials at –20 °C until used or at room temperature for short periods of 1–2 days.

Video: A New Viral Transport Media Manufacturing Plant By Thermo Fisher Scientific

Uses of Viral Transport Media (VTM)

  • Viral transport media are used for the collection and transport of specimens containing viruses.
  • These are also used to maintain the viability of other organisms like mycoplasma, chlamydiae, and ureaplasma.
  • VTMs also help to maximize the number of viral particles in a sample.
  • Liquid transport media are used primarily for transporting swabs or materials released into the medium from a collection swab.
  • It facilitates the collection and transport of samples in areas where the facilities of refrigeration are not available.

Limitations of Viral Transport Media (VTM)

  • Some growth of contaminants might be observed during the long period of transport.
  • After transportation, the specimen should be inoculated in the proper medium as soon as possible. For optimum results, the time-lapse between sample collection and inoculum onto the culture medium should be reduced to the minimum.
  • Some viruses might be more labile than others, and the VTMs might be as effective.
  • Some antibiotics and chemicals used in VTMs might change the pH of the solution, which affects the viability of the organism.

References and Sources

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