Recovery of live virus after storage at ambient temperature using ViveST™

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Abstract

Background

A major impediment to performing virological field studies in developing nations is the lack of ultra-low freezers as well as the expense and difficulty of shipping frozen samples. A commercially available product, ViveST™, was developed to preserve nucleic acids at ambient temperature for use in specimen storage and transportation. However, its applications as a viral storage, transport and recovery device have not been evaluated.

Objective

To examine the ability of ViveST to preserve live virus following storage at ambient temperature.

Study design

A panel of six viruses was stored at ambient temperature (∼22 °C) in ViveST with fetal bovine serum (FBS), or ViveST with minimal essential media (MEM) and compared with virus stored in universal transport media (M4RT), MEM, and FBS alone. Stored viruses included: human adenovirus (14p), dengue virus 2 (16608), echovirus 3 (Morrisey), human rhinovirus 15 (1734), Coxsackie virus B5 (Faulkner), and herpes simplex virus 1 (HF). After 7 days storage at ambient temperature, virus recovery was measured via titration using viral plaque assays or focus-forming unit assays.

Results

Viral titer studies indicate that ViveST with either FBS or M4RT preserved/recovered 5 different viruses for 1 week at ambient temperature. MEM preserved 4 viruses while FBS and ViveST with MEM preserved 3 viruses each. Statistical analyses indicate that M4RT and ViveST with FBS preserved significantly more virus than the other treatments.

Conclusions

These data suggest that ViveST with either FBS or M4RT may be useful in field specimen collection scenarios where ultra-cold storage is not available.

Section snippets

Background

The most accurate approaches to viral characterization require the collection of viable virus which is amplified in culture. Often, field collection and preservation of circulating virus requires ultra-low freezers or dry ice. Because such cold-chain infrastructure is not available in many developing countries, viral characterization data in these countries can be sparse. Even if virus can be successfully preserved and shipped from the field site to a reference laboratory, there are often long

Objective

Using in vitro viral titer studies, we sought to evaluate the ability of the ViveST device to preserve live virus at ambient temperature in comparison with other viral transport media.

Viruses

A panel of six viruses was used in this study in order to determine the range of viruses that could be preserved with ViveST. Viruses of various sizes were evaluated and included those with DNA or RNA genomes, with or without envelopes. This panel included enteric cytopathic human orphan virus 3 (ECHOV), Morrisey; human rhinovirus 15 (HRV), 1734; human Coxsackievirus B5 (CBV), Faulkner and herpes simplex virus 1 (HSV), HF. These viruses are prototype strains of the Center for Disease Control

ViveST preserves virus at ambient temperature for 7 days

Viable HAdV, CBV, ECHOV, HSV, and HRV were recovered from the ViveST with FBS treatment after 7 days storage at ambient temperature (Fig. 2). Viruses showed an average reduction in infectious units (i.u.) by 2 orders of magnitude when compared to the starting titer of the respective virus (Fig. 2). ECHOV exhibited the highest rate of preservation of i.u. when compared with HAdV, HSV, and HRV (p = 0.00006). While there was a significant loss of i.u. for all viruses (p = 0.00035), at least 15,000

Discussion

Currently there are no standard methods for the storage and recovery of live virus at ambient temperature and investigations in this subject are sparse. Studies investigating the use of cell culture media and viral transport media for preserving virus at room temperature have documented a low recovery rate for HSV, HAdV, and other viruses tested.4, 5 Recovery of live HIV from ViveST has been attempted, but was unsuccessful using the standard drying protocol.12 This may have been a result of the

Funding

This work was supported by grants from the US Department of Defense Armed Forces Health Surveillance Center's Global Emerging Infections Surveillance and Response Program (I0179_10_UN Dr. Gray) and from the National Institute of Allergy and Infectious Diseases (R01 AI068803 – Dr. Gray).

Competing interest

None declared.

Ethical approval

Not required.

Acknowledgments

We thank Mimi Healy of Vivebio, for her support, advice, and encouragement concerning this work. We are also grateful to Thomas Waltzek for his insights and recommendations on viruses.

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