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Pilot Assessment of RNA Stabilization Methods for Influenza A Virus in Swine Oral Fluids
Berenice Munguía-Ramírez1, Betsy Armenta-Leyva1, Luis Giménez-Lirola1
1Department of Veterinary Diagnostic and Production Animal Medicine, College of Veterinary Medicine, Iowa State University, 1800 Christensen Drive, Ames, IA 50011-1134, USA.
Pathogens (Basel, Switzerland)
|May 4, 2026
Summary
Stabilizing influenza A virus (IAV) RNA in swine oral fluids is crucial for molecular surveillance. Carbohydrates like trehalose, sorbitol, and sucrose, along with commercial stabilizers, effectively preserved IAV RNA under thermal and freeze-thaw stress, offering a low-cost solution.
Area of Science:
- Veterinary Virology
- Molecular Diagnostics
- Biochemistry
Background:
- Swine influenza A virus (IAV) surveillance depends on molecular detection methods.
- RNA integrity in oral fluid samples is compromised without consistent cold-chain storage.
- Maintaining RNA stability is critical for accurate diagnostic results in field settings.
Purpose of the Study:
- To evaluate the efficacy of carbohydrates and commercial stabilizers in preserving IAV RNA in swine oral fluids.
- To assess RNA stability under simulated field-relevant thermal and freeze-thaw stress conditions.
- To identify cost-effective, non-inactivating solutions for IAV RNA stabilization.
Main Methods:
- Pilot study using swine oral fluids from experimentally infected pigs (H1N2, H3N2).
- Samples were treated with molecular-grade carbohydrates (trehalose, sorbitol, sucrose, mannitol) or commercial stabilizers (PrimeStore® MTM, RNA later®).
- Exposure to storage at 25 °C for up to 144 hours and multiple freeze-thaw cycles.
- IAV RNA detection quantified using RT-qPCR, with results analyzed as efficiency standardized Cq values (ECq).
Main Results:
- Carbohydrates (trehalose, sorbitol, sucrose) and commercial stabilizers significantly maintained higher ECq values compared to untreated samples.
- Effective RNA preservation was observed under both thermal stress (25 °C) and freeze-thaw cycling.
- The tested stabilizers demonstrated superior performance in protecting IAV RNA integrity.
Conclusions:
- Carbohydrates and commercial stabilizers show promise for stabilizing IAV RNA in swine oral fluids.
- These agents can mitigate RNA degradation under challenging field storage conditions.
- Carbohydrates offer a potentially low-cost, non-inactivating alternative for improving swine IAV surveillance.
