Establishment of a Sensitive and Reliable Droplet Digital PCR Assay for the Detection of Bursaphelenchus xylophilus

Plants, Год журнала: 2024, Номер 13(19), С. 2701 - 2701

Опубликована: Сен. 26, 2024

Pine wilt disease (PWD), which poses a significant risk to pine plantations across the globe, is caused by pathogenic agent Bursaphelenchus xylophilus, also referred as wood nematode (PWN). A droplet digital PCR (ddPCR) assay was developed for quick identification of PWN in order improve detection sensitivity. The research findings indicate that ddPCR demonstrated significantly higher analysis sensitivity and comparison traditional quantitative (qPCR). However, it had more limited dynamic range. High specificity shown both qPCR techniques diagnosis PWN. Assessments reproducibility revealed lower coefficients variation at every template concentration. Inhibition tests showed less susceptible inhibitors. There strong linear association between standard measurements obtained using (Pearson correlation = 0.9317; p < 0.001). Likewise, there agreement 0.9348; 0.001) evaluation samples. Additionally, samples from symptomatic (100% versus 86.67%) asymptomatic (31.43% 2.9%) trees were diagnosed with greater rates ddPCR. This study’s conclusions highlight advantages over method has lot potential ecological on PWD use quarantines.

Язык: Английский

Detection of Porcine Circovirus (PCV) Using CRISPR-Cas12a/13a Coupled with Isothermal Amplification

Viruses, Год журнала: 2024, Номер 16(10), С. 1548 - 1548

Опубликована: Сен. 30, 2024

The impact of porcine circovirus (PCV) on the worldwide pig industry is profound, leading to notable economic losses. Early and prompt identification PCV essential in managing controlling this disease effectively. A range detection techniques for have been developed primarily divided into two categories focusing nucleic acid or serum antibody identification. methodologies encompass conventional polymerase chain reaction (PCR), real-time fluorescence quantitative PCR (qPCR), situ hybridization (FISH), loop-mediated isothermal amplification (LAMP), immunofluorescence assay (IFA), immunohistochemistry (IHC), enzyme-linked immunosorbent (ELISA). Despite their efficacy, these are often impeded by necessity substantial investment equipment, specialized knowledge, intricate procedural steps, which complicate application field detections. To surmount challenges, a sensitive, rapid, specific method using Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas12a/13a coupled with amplification, such as enzymatic recombinase (ERA), (RPA), has developed. This novel undergone meticulous optimization detecting types 2, 3, 4, boasting remarkable sensitivity identify single copy per microliter. specificity technique exemplary, no observable interaction other viruses PEDV, PRRSV, PRV, CSFV. Its reliability validated clinical samples, where it produced perfect alignment qPCR findings, showcasing 100% coincidence rate. elegance merging CRISPR-Cas technology assays lies its on-site testing without need expensive tools trained personnel, rendering exceptionally suitable applications, especially resource-constrained swine farming environments. review assesses compares process characteristics inherent utilization ERA/LAMP/RPA-CRISPR-Cas12a/Cas13a PCV, providing critical insights practicality effectiveness.

Язык: Английский

Digital PCR in Virology: Current Applications and Future Perspectives

Molecular Diagnosis & Therapy, Год журнала: 2024, Номер unknown

Опубликована: Ноя. 2, 2024

Язык: Английский