Optical Biosensor Platforms Display Varying Sensitivity for the Direct Detection of Influenza RNA

Area of Science:

• Virology
• Biosensing Technology
• Bioinformatics

Background:

• Non-nucleic acid amplification detection methods offer rapid viral diagnostics.
• Influenza virus's propensity for mutations necessitates advanced diagnostic and surveillance tools.
• Existing diagnostic assays may have limitations in detecting diverse influenza strains.

Purpose of the Study:

• To evaluate the Fast Evaluation of Viral Emerging Risks (FEVER) pipeline for influenza diagnostics.
• To assess ultrasensitive detection platforms, including waveguide biosensors and flow cytometry assays, for influenza RNA detection.
• To compare the in silico performance of FEVER probes against CDC diagnostic assays.

Main Methods:

• In silico analysis of the FEVER pipeline for sequence coverage and probe mismatch tolerance.
• Experimental evaluation of waveguide-based optical biosensors and flow cytometry bead-based assays.
• Development and testing of molecular beacon and exonuclease selection methods for RNA detection.

Main Results:

• FEVER probes demonstrated higher mismatch tolerance and detection rates for influenza sequences compared to CDC probes.
• Ultrasensitive detection of influenza RNA was achieved, with limits as low as 50 nM (waveguide) and 1 nM (flow cytometry) using molecular beacons.
• An exonuclease selection method improved detection sensitivity to 500 pM.

Conclusions:

• The FEVER pipeline, combined with ultrasensitive optical biosensors, presents a promising approach for influenza diagnostics.
• This method enhances viral detection capabilities, supporting both accurate diagnosis and pandemic surveillance.
• The developed probes and detection platforms show potential for improved identification of emerging influenza strains.