1. Development and validation of a one-step reverse transcription real-time PCR assay for simultaneous detection and identification of tomato mottle mosaic virus and tomato brown rugose fruit virusAntonio Tiberini, Ariana Manglli, Anna Taglienti, Ana Vučurović, Jakob Brodarič, Luca Ferretti, Marta Luigi, Andrea Gentili, Nataša Mehle, 2022, original scientific article Abstract: Tobamovirus species represent a threat to solanaceous crops worldwide, due to their extreme stability and because they are seed borne. In particular, recent outbreaks of tomato brown rugose fruit virus in tomato and pepper crops led to the establishment of prompt control measures, and the need for reliable diagnosis was urged. Another member of the genus, tomato mottle mosaic virus, has recently gained attention due to reports in different continents and its common features with tomato brown rugose fruit virus. In this study, a new real-time RT-PCR detection system was developed for tomato brown rugose fruit virus and tomato mottle mosaic virus on tomato leaves and seeds using TaqMan chemistry. This test was designed to detect tomato mottle mosaic virus by amplifying the movement protein gene in a duplex assay with the tomato brown rugose fruit virus target on the CP-3’NTR region, which was previously validated as a single assay. The performance of this test was evaluated, displaying analytical sensitivity 10−5–10−6-fold dilution for seeds and leaves, respectively, and good analytical specificity, repeatability, and reproducibility. Using the newly developed and validated test, tomato brown rugose fruit virus detection was 100% concordant with previously performed analyses on 106 official samples collected in 2021 from different continents.
Keywords: real-time PCR, tomato mottle mosaic virus, tomato brown rugose fruit virus, leaves detection, seeds detections, performance criteria
Published in DiRROS: 16.07.2024; Views: 5; Downloads: 5
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2. Diagnostic accuracy of (1-3)-[beta]-D-glucan to predict Pneumocystis jirovecii pneumonia in non-HIV-infected patientsPetra Rogina, Miha Skvarč, 2020, original scientific article Abstract: Background Pneumocystis jirovecii pneumonia (PCP) is a common and potentially fatal opportunistic infection in immunocompromised non-HIV individuals. There are problems with clinical and diagnostic protocols for PCP that lack sensitivity and specificity. We designed a retrospective study to compared several methods that were used in diagnostics of PCP. Patients and methods One hundred and eight immunocompromised individuals with typical clinical picture for PCP and suspicious radiological findings were included in the study. Serum samples were taken to measure the values of (1-3)-[beta]-D-glucan (Fungitell, Associates of Cape Cod, USA). Lower respiratory tract samples were obtained to perform direct immunofluorescence (DIF, MERIFLUOR Pneumocystis, Meridian, USA) stain and real-time PCR (qPCR). Results Fifty-four (50%) of the 108 patients in our study had (1-3)-[beta]-D-glucan > 500 pg/ml. Patients that had (1-3)-[beta]-D-glucan concentrations < 400 pg/ml in serum, had mean threshold cycles (Ct) 35.43 +- 3.32 versus those that had (1-3)-[beta]-D-glucan concentrations >400 pg/mL and mean Ct of 28.97 +- 5.27 (P < 0.001). If we detected P. jirovecii with DIF and qPCR than PCP was proven. If the concentration of (1-3)-[beta]-D-glucan was higher than 400 pg/ml and Ct of qPCR was below 28.97 +- 5.27 than we have been able be certain that P. jirovecii caused pneumonia (odds ratio [OR] 2.31, 95% confidence interval [CI] 1.62-3.27, P < 0.001). Conclusions Measurement of (1-3)-[beta]-D-glucan or qPCR alone could not be used to diagnose PCP. Diagnostic cut-off value for (1-3)-[beta]-D-glucan > 400pg/ml and qPCR below 30 Ct, allow us to conclude that patient has PCP. If the values of (1-3)-[beta]-D-glucan are < 400 pg/ml and qPCR is above 35 Ct than colonization with P. jirovecii is more possible than PCP.
Keywords: Pneumocystis jirovecii pneumonia, real-time PCR, non-HIV-infected patients
Published in DiRROS: 12.07.2024; Views: 55; Downloads: 33
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3. Microarray-based uncovering of reference genes for quantitative real-time PCR in potato tuber infected with PVYBarbara Gerič Stare, Aleš Sedlar, Vladimir Meglič, 2021, original scientific article Keywords: potato, potato tuber, tuber infection, potato virus Y, real-time PCR, PCR, polymerase chain reaction, reference genes, stored ppotato tubers, pathogens
Published in DiRROS: 31.05.2021; Views: 1554; Downloads: 344
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