Rapid detection of monkeypox virus by multiple cross displacement amplification combined with nanoparticle-based...

Area of Science:

• Virology
• Biotechnology
• Infectious Diseases

Background:

• The current monkeypox virus (MPXV) outbreak necessitates rapid and sensitive diagnostic methods.
• Existing diagnostic tools may not offer the speed or specificity required for effective outbreak management.
• Differentiating between West and Central African MPXV isolates is crucial for epidemiological tracking.

Purpose of the Study:

• To develop a rapid, sensitive, and specific diagnostic assay for MPXV infection.
• To enable differentiation between West and Central African MPXV isolates.
• To provide a tool for timely diagnosis during public health emergencies.

Main Methods:

• Combined isothermal multiple cross displacement amplification (MCDA) with a nanoparticle-based lateral flow biosensor (LFB).
• Designed two MCDA primer sets targeting specific genes (D41L and ATI) of MPXV isolates.
• Optimized MCDA reaction conditions at 64°C for 30 minutes, followed by LFB detection.

Main Results:

• The MPXV-MCDA-LFB assay provides results in under 50 minutes, including template preparation.
• Demonstrated high sensitivity, detecting as low as five copies of plasmid DNA or 12.5 copies of pseudotyped virus.
• Confirmed specificity through lack of cross-reactivity with non-MPXV templates.

Conclusions:

• The developed MPXV-MCDA-LFB assay is a valuable tool for rapid and reliable MPXV diagnosis.
• The assay facilitates the differentiation of MPXV isolates, aiding epidemiological surveillance.
• This diagnostic approach addresses the urgent need for effective tools during MPXV outbreaks.