Flexible upscaling of laboratory PCR testing capacity at the Robert Koch Institute during the SARS-CoV-2 pandemic

Krause, Eva; Michel, Janine; Puyskens, Andreas; Hofmann, Natalie; Rinner, Thomas; Biere, Barbara; Dorner, Brigitte G.; Skiba, Martin; Schaade, Lars; Nitsche, Andreas

doi:10.1186/s12985-023-02088-x

Cited by 2 publications

(1 citation statement)

References 24 publications

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“…Automation can also contribute to accurate testing of pooling procedures by providing accurate mixing, sample tracking, and avoidance of cross-contamination. In previous reports, the pooling and testing stages were usually performed manually (or methods not specified), although some studies utilized automated extraction devices or fully automated platforms for the testing of pooled samples ( 4 , 16 – 19 ). Barak et al utilized a liquid handler for automated media pooling but its performance was not validated by comparison with individual testing ( 7 ).…”

Section: Introductionmentioning

confidence: 99%

Development and evaluation of three automated media pooling and molecular diagnostic systems for the detection of SARS-CoV-2

Matsumura,

Noguchi,

Shinohara

et al. 2024

Microbiol Spectr

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Pooled testing combined with molecular diagnostics for the detection of SARS-CoV-2 is a promising method that can increase testing capacities and save costs. However, pooled testing is also associated with the risks of decreased test sensitivity and specificity. To perform reliable pooled testing, we developed and validated three automated media pooling and molecular diagnostic systems. These pooling systems (geneLEAD-PS, Panther-PS, and Biomek-PS) comprised existing automated molecular detection platforms, corresponding automated media pooling devices, and laboratory information management systems. Analytical sensitivity analysis and mock sample evaluation were performed, and the obtained data were used to determine the sizes of the pool for the validation study. In the validation study, a total of 2,448, 3,228, and 6,420 upper respiratory samples were used for geneLEAD-PS, Panther-PS, and Biomek-PS, respectively, and the diagnostic performances were compared with the reference RT‒PCR assay. A pool size of 6 for geneLEAD-PS and a pool size of 4 for Panther-PS and Biomek-PS were selected for the validation studies. All three systems showed high positive percent agreement values of ≥90.5% and negative percent agreement values of ≥99.8% for any specimen type. Pooled testing resulted in a 65%–71% reduction in cost per sample. The testing capacities of geneLEAD-PS, Panther-PS, and Biomek-PS were 144 samples in 3 hours, 384 samples in 5.5 hours, and 376 samples in 4 hours, respectively. The developed pooling systems showed robust diagnostic performances and will increase the testing capacities of molecular diagnostic tests while saving costs and may contribute to infection control of COVID-19. IMPORTANCE During the COVID-19 pandemic, there have been surges in demand for accurate molecular diagnostic testing and laboratory supply shortages. Pooled testing combined with highly sensitive molecular testing, which entails mixing multiple samples as a single sample, is a promising approach to increase testing capacities while reducing the use of consumables. However, pooled testing is associated with risks that compromise diagnostic performance, such as false negatives due to dilution of positive samples or false positives due to cross-contamination. To perform reliable pooled testing, three different pooling systems (an automated pooling device, an automated molecular detection platform, and a laboratory information management system) were developed to accurately interpret pooled testing results. These three systems were validated using multiple clinical samples and showed high concordance with individual testing. The developed pooling systems will contribute to increasing reliable molecular testing capacities while using fewer consumables and saving costs.

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Section: Introductionmentioning

confidence: 99%

Development and evaluation of three automated media pooling and molecular diagnostic systems for the detection of SARS-CoV-2

Matsumura,

Noguchi,

Shinohara

et al. 2024

Microbiol Spectr

View full text Add to dashboard Cite

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Shaken, not stirred: magnetic bead DNA extraction as a rapid and effective method for the scaling up of bovine tuberculosis diagnosis

Lorente-Leal,

Gomez-Buendia,

Gutiérrez-Tobaruela

et al. 2024

BMC Vet Res

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Background The growing use of real-time PCR (qPCR) as a diagnostic method for bovine TB (bTB) requires rapid and effective DNA extraction methods, which are crucial for its success. Automated DNA extraction methods based on magnetic beads are a promising alternative to conventional silica column-based protocols (COL protocol) due to their high throughput capacity and reduced hands-on time. This study aimed to assess the performance of the MagMax CORE Nucleic Acid Purification kit and the KingFisher Flex instrument (KF protocol) as an alternative for scaling up the use of qPCR in bTB diagnosis. Methodology Performance was evaluated with two different real-time PCR (qPCR) protocols, based on the IS 6110 element and the QuantiFast and VetMAX™ (QF and VM protocols) kits, on 145 frozen tissue homogenates confirmed as either bTB-positive or negative through a composite reference standard based on microbiological culture, column-based extraction, and qPCR, as well as on negative tissue samples spiked with 10 6 to 10 3 CFU/ml of M. bovis BCG. Results The performance of both qPCR protocols was very high on samples extracted using the KF protocol, with positive percent agreement (PPA) values of 89.04% [95% Confidence Interval (CI): 79.54–95.15%] and 93.15% [95% CI: 84.74–97.74%] for the QF and VM protocols, respectively, and negative percent agreement (NPA) values of 100% [95% CI: 95.01–100.00%]. A higher variability was identified in samples analysed with the same qPCR protocol but different extraction methods. Higher Ct values were identified for samples extracted using the KF protocol in both routine and spiked samples, likely due to using the same amount of starting material for both extraction methods, which was lower than recommended by the manufacturer for the KF protocol. Discussion The results of this study indicate that the MagMAX CORE Nucleic Acid Purification kit coupled with a KingFisher Flex instrument is a valuable alternative for the extraction of MTBC DNA from bovine tissues. However, the increased variability and Ct values suggest that a larger amount of starting material is recommended for this methodology, warranting further studies. Supplementary Information The online version contains supplementary material available at 10.1186/s12917-024-04427-8.

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Flexible upscaling of laboratory PCR testing capacity at the Robert Koch Institute during the SARS-CoV-2 pandemic

Cited by 2 publications

References 24 publications

Development and evaluation of three automated media pooling and molecular diagnostic systems for the detection of SARS-CoV-2

Development and evaluation of three automated media pooling and molecular diagnostic systems for the detection of SARS-CoV-2

Shaken, not stirred: magnetic bead DNA extraction as a rapid and effective method for the scaling up of bovine tuberculosis diagnosis

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