Danny Bravo Padilla

Danny Bravo Padilla

Nutrición Vegetal y Suelos, Escuela Superior Politecnica del Litoral, Guayaquil, Guayas, Ecuador

Research Focus & Metrics

Academic Career Length

6Years (2021-Present)

(Calculated once Scilit publication data is synced: current year − year of first publication + 1)

Representative Publications

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24 June 2021 in Viruses
Verified
Journal Article
Open access
PDF file
ACAndres Espindola Camacho
Andres Espindola Camacho
DSDaniela Sempertegui-Bayas
Daniela Sempertegui-Bayas
DPDanny Bravo Padilla
Danny Bravo Padilla
VFViviana Freire-Zapata
Viviana Freire-Zapata
FOFrancisco Ochoa-Corona
Francisco Ochoa-Corona
KCKitty Cardwell
Kitty Cardwell
High-throughput sequencing (HTS) is becoming the new norm of diagnostics in plant quarantine settings. HTS can be used to detect, in theory, all pathogens present in any given sample. The technique’s success depends on various factors, including methods for sample management/preparation and suitable bioinformatic analysis. The Limit of Detection (LoD) of HTS for plant diagnostic tests can be higher than that of PCR, increasing the risk of false negatives in the case of low titer of the target pathogen. Several solutions have been suggested, particularly for RNA viruses, including rRNA depletion of the host, dsRNA, and siRNA extractions, which increase the relative pathogen titer in a metagenomic sample. However, these solutions are costly and time-consuming. Here we present a faster and cost-effective alternative method with lower HTS-LoD similar to or lower than PCR. The technique is called TArget-SPecific Reverse Transcript (TASPERT) pool. It relies on pathogen-specific reverse primers, targeting all RNA viruses of interest, pooled and used in double-stranded cDNA synthesis. These reverse primers enrich the sample for only pathogens of interest. Evidence on how TASPERT is significantly superior to oligodT, random 6-mer, and 20-mer in generating metagenomic libraries containing the pathogen of interest is presented in this proof of concept.