Development of reference material from Rift Valley fever virus RNA.

Area of Science:

• Virology
• Molecular Diagnostics
• Metrology

Background:

• Rift Valley fever virus (RVFV) is a significant pathogen primarily found in Africa.
• Molecular diagnostic tests, such as nucleic acid amplification tests, are crucial for RVFV detection.
• Currently, metrologically certified reference materials for RVFV nucleic acids are unavailable, hindering accurate molecular testing.

Purpose of the Study:

• To develop and characterize metrologically certified RNA reference materials for Rift Valley fever virus (RVFV).
• To provide essential tools for the performance evaluation and quality assessment of RVFV molecular diagnostic assays.

Main Methods:

• In vitro transcribed RNA, encompassing the L, M, and S segments of the RVFV genome, was used to create the reference materials.
• Reverse-transcription droplet digital PCR (RT-ddPCR) was employed for the absolute quantification of viral RNA copy numbers in the developed reference materials.

Main Results:

• The developed RVFV RNA reference materials (KRISS 111-10-537, 538, and 539) exhibited excellent homogeneity, with copy number variations between bottles ranging from 4.0% to 6.7%.
• These reference materials demonstrated robust stability, remaining stable for up to 14 days at 4°C and for 12 months at -80°C.
• Quantified RNA copy numbers ranged from 7.82 × 10⁴ to 2.51 × 10⁶ copies/µl.

Conclusions:

• The newly developed RVFV RNA reference materials provide accurate reference values for viral gene copy numbers.
• These materials are expected to reduce false positives and negatives in RVFV molecular tests.
• The availability of these reference materials will contribute to improved diagnosis and control of Rift Valley fever.