Dataset Information

Large-Scale RT-qPCR Diagnostics for Seed Potato Certification

ABSTRACT: Every year, Agroscope examines nearly 300,000 tubers for the presence of viruses, as regulated for the certification of seed potatoes intended for Swiss growers. Since 2016, this examination has been performed via RT-qPCR on dormant tubers directly after harvest. This method offers fast results and eliminates the need for the use of Rindite, which is a toxic and polluting gaseous compound previously used in Switzerland to break the dormancy of seed tubers. The implementation of this molecular analytical method for the routine diagnosis of regulated viruses makes it possible to conduct additional analyses via Illumina sequencing to assess the conformity of the primers and probes used with the sequences of the different viral isolates. This form of quality control in routine diagnosis is a source of information that can answer more fundamental scientific questions related to the epidemiology of viral strains related to certification. The datasets produced in this framework can also be used to explore the diversity of rare or unknown virus species in potato crops.

Supplementary Information

The online version contains supplementary material available at 10.1007/s11540-021-09491-3.

SUBMITTER: Schumpp O

PROVIDER: S-EPMC8572825 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Coronavirus disease 2019 (COVID-19) is an infectious, acute respiratory disease caused mainly by person-to-person transmission of the coronavirus SARS-CoV-2. Its emergence has caused a world-wide acute health crisis, intensified by the challenge of reliably identifying individuals likely to transmit the disease. Diagnosis is hampered by the many unknowns surrounding this disease, including those relating to infectious viral burden. This uncertainty is exacerbated by disagreement surrounding the clinical relevance of molecular testing using reverse transcription quantitative PCR (RT-qPCR) for the presence of viral RNA, most often based on the reporting of quantification cycles (Cq), which is also termed the cycle threshold (Ct) or crossing point (Cp). Despite it being common knowledge that Cqs are relative values varying according to a wide range of different parameters, there have been efforts to use them as though they were absolute units, with Cqs below an arbitrarily determined value, deemed to signify a positive result and those above, a negative one. Our results investigated the effects of a range of common variables on Cq values. These data include a detailed analysis of the effect of different carrier molecules on RNA extraction. The impact of sample matrix of buccal swabs and saliva on RNA extraction efficiency was demonstrated in RT-qPCR and the impact of potentially inhibiting compounds in urine along with bile salts were investigated in RT-digital PCR (RT-dPCR). The latter studies were performed such that the impact on the RT step could be separated from the PCR step. In this way, the RT was shown to be more susceptible to inhibitors than the PCR. Together, these studies demonstrate that the consequent variability of test results makes subjective Cq cut-off values unsuitable for the identification of infectious individuals. We also discuss the importance of using reliable control materials for accurate quantification and highlight the substantial role played by dPCR as a method for their development.

Dataset Information

Large-Scale RT-qPCR Diagnostics for Seed Potato Certification

Supplementary Information

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