Screening metagenomic data for viruses using the e-probe diagnostic nucleic acid assay

Area of Science:

• Plant pathology
• Bioinformatics
• Molecular diagnostics

Background:

• Next-generation sequencing (NGS) is computationally intensive for routine diagnostics.
• Identifying microbial or viral targets in complex metagenomic data requires efficient methods.

Purpose of the Study:

• To develop and validate a rapid diagnostic method using next-generation sequencing (NGS) data.
• To enable the detection and differentiation of plant viruses directly from unassembled sequence reads using pathogen-specific sequences (e-probes).

Main Methods:

• Developed an e-probe diagnostic nucleic acid assay utilizing unassembled NGS data as the search target.
• Tested the assay with mock databases and real-world NGS data from plants infected with Bean golden yellow mosaic virus (BGYMV) and Plum pox virus (PPV).
• Evaluated the ability to detect and differentiate between PPV strains using strain-specific e-probe sets.

Main Results:

• The e-probe method successfully detected both DNA (BGYMV) and RNA (PPV) viruses in plant samples.
• Strain-specific e-probes differentiated between Plum pox virus strains.
• The assay identified a dual infection of BGYMV and Bean golden mosaic virus in one sample.

Conclusions:

• E-probe diagnostics offer a computationally efficient approach for identifying specific pathogens in NGS datasets.
• This method enhances the speed and accuracy of plant virus diagnostics.
• The assay has potential for detecting multiple pathogens and differentiating strains within a single sample.