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bioRxiv · 10.1101/2025.01.24.634829

Evaluating Seqstant LiveGene Analysis in Real-Time Assessment of Metagenomic Next-Generation Sequencing (mNGS) Data from Respiratory Samples

Boutin, S.·Klein, S.·Untergasser, G.·Loka, T. P.·Jakob, S.·Khatamzas, E.·Wrettos, G.·Knobloch, H.·Nurjadi, D.
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Abstract

BackgroundThe detection of pathogens causing infections by conventional diagnostic methods can be challenging and next-generation sequencing (NGS) technology offers a promising alternative method. In this study, we evaluated the performance of real-time metagenomic next-generation sequencing (rt-mNGS) for the detection of pathogens in respiratory samples. MethodWe used rt-mNGS, using the Seqstant LiveGene Analysis platform, on 335 respiratory samples in comparison to conventional culture results. ResultsWe observed an overall good concordance in 71.64% (240/335) of the methods. The rt-mNGS outperformed the gold standard culture in 16.12% (54/335) of the samples, while the culture was superior in detecting the clinically relevant pathogen in 12.24% (41/335) of the samples. The non-inferiority of rt-mNGS was statistically significant ({delta} = 10, = 0.05, 1 - {beta}= 0.8). We also observed that the real-time analysis of NGS data is beneficial in obtaining reliable timely results as the initial report at cycle 46 exhibits a Positive Predictive Value (PPV) of 93.75% at the species-level with a sensitivity of 32.09%. ConclusionOverall, our study showed the non-inferiority of rt-mNGS compared to the standard-of-care microbiology for respiratory samples with statistical significance. Moreover, the rt-mNGS method exhibited superior sensitivity and superior overall performance. It also uniquely detected certain organisms that are typically hard to culture. However, rt-mNGS reported a higher number of false positives and faced limitations in detecting Aspergillus spp. In conclusion, the study highlights the potential of rt-mNGS as a powerful tool in clinical diagnostics of respiratory infections and beyond.

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Boutin, S., Klein, S., Untergasser, G., Loka, T. P., Jakob, S., Khatamzas, E., Wrettos, G., Knobloch, H., Nurjadi, D.. 2025-01-28. Evaluating Seqstant LiveGene Analysis in Real-Time Assessment of Metagenomic Next-Generation Sequencing (mNGS) Data from Respiratory Samples. https://doi.org/10.1101/2025.01.24.634829

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