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  1. Home
  3. Amplification-free Detection Of Zoonotic Viruses Using Cas13 And Multiple Crispr Rnas.
  1. Home
  3. Amplification-free Detection Of Zoonotic Viruses Using Cas13 And Multiple Crispr Rnas.

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Amplification-free Detection of Zoonotic Viruses Using Cas13 and Multiple CRISPR RNAs.

Caitlin H Lamb1, Aartjan J W Te Velthuis1, Cameron Myhrvold1,2,3,4

  • 1Department of Molecular Biology, Princeton University, Princeton, NJ 08544.

Biorxiv : the Preprint Server for Biology
|June 4, 2025

View abstract on PubMed

Summary
This summary is machine-generated.

A new CRISPR-Cas13 assay offers a rapid, sensitive, and cost-effective method for detecting zoonotic viruses like hantaviruses and influenza A virus without amplification. This field-deployable tool shows promise for early infection detection in various samples.

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Area of Science:

  • Molecular Biology
  • Virology
  • Biotechnology

Background:

  • Zoonotic viruses, including hantaviruses and influenza A viruses, pose significant risks to human and animal health.
  • Current diagnostic methods for these viruses can be slow, expensive, or require specialized laboratory equipment.
  • There is a critical need for rapid, sensitive, and accessible diagnostic tools for early detection of zoonotic viral infections.

Purpose of the Study:

  • To develop and evaluate an amplification-free CRISPR-Cas13-based assay for detecting hantavirus and influenza A virus RNA.
  • To assess the impact of using multiple CRISPR RNAs (crRNAs) on the sensitivity of the amplification-free assay.
  • To determine the feasibility of using this assay for detecting viral RNA in both laboratory and clinical samples.

Main Methods:

  • An amplification-free CRISPR-Cas13 assay was designed to detect genomic RNA of hantaviruses (Tula Virus - TULV, Andes Virus - ANDV) and influenza A virus.
  • The sensitivity of the assay was tested using varying numbers of crRNAs for different viruses.
  • The assay's performance was validated using viral RNA in tissue culture samples and clinical nasopharyngeal swabs.

Main Results:

  • For TULV and ANDV detection, using two or three crRNAs significantly enhanced the sensitivity of the amplification-free assay.
  • Additional crRNAs did not improve the sensitivity for influenza A virus RNA detection.
  • The CRISPR-Cas13 assay successfully detected TULV and ANDV RNA in cell culture and influenza A virus RNA in clinical samples, showing 85% agreement with RT-qPCR for positive sample identification.

Conclusions:

  • Amplification-free CRISPR-Cas13 technology presents a viable strategy for the rapid and sensitive detection of zoonotic viral RNA.
  • The optimization of crRNA combinations can improve assay sensitivity for specific viral targets.
  • This assay holds potential as a field-deployable diagnostic tool for early identification of zoonotic virus infections, aiding in public health surveillance and response.