Limits and Opportunities of SARS-CoV-2 Antigen Rapid Tests: An Experienced-Based Perspective

Background: Due to the steadily rising case numbers of SARS-CoV-2 infections worldwide, there is an increasing need for reliable rapid diagnostic devices in addition to existing gold standard PCR methods. Actually, public attention is focused on antigen assays including lateral flow tests (LFTs) as a diagnostic alternative. Therefore, different LFTs were analyzed regarding their performance in a clinical setting. Material and Methods: A pilot sample panel of 13 bronchoalveolar fluids (BALFs) and 60 throat washing (TW) samples with confirmed PCR results, as well as eight throat washes invalid by PCR, were tested with the BIOCREDIT test (RapiGEN), the Panbio(TM) assay (Abbott), and the SARS-CoV-2 rapid antigen test (Roche). Conclusion: The analyzed antigen test showed an interassay correlation of 27.4%, with overall specificities ranging from 19.4% to 87.1%, while sensitivities of the respective tests ranged between 33.3% and 88.1%. Because these assays did not entirely meet all high expectations, their benefit has to be carefully evaluated for the respective test strategy and setting

RT-qPCR based determination of SARS-CoV-2 in health care workers, Cologne, Germany-A pre-delta/pre-omicron proof of principle field study

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), causing coronavirus disease 2019 (COVID-19), and SARS-CoV-2 variants pose an increased risk to global health. Therefore, monitoring of SARS-CoV-2 variants of concern (VOCs) is of high importance for the implementation of disease control methods, for timely public health decisions, and the development of vaccines against SARS-CoV-2 variants. In this study, which was performed before the delta and omicron variants of concern became dominant, a total of 111 SARS-CoV-2 positive samples from our hospital staff in Cologne, Germany, collected from March 2020 to May 2021 were analysed for VOCs. For determination of VOCs, mutation genotyping analysis (MGA) using mutation-specific simple (MSS) probes based on quantitative reverse transcription-polymerase chain reaction (RT-qPCR) of ten spike protein variants (SPVs) was performed. The MGA focuses on the detection of the spike protein mutation (SPM) of SPVs belonging to VOCs. By successful determination of SPV, the work concludes that 24.66 % of the samples belong to VOC B.1.1.7 and 1.37 % of the samples belong to VOC B.1.351. Based on these results, MGA proves to be a suitable alternative to sequencing technologies as it is a rapid, cost-effective, widely available, and feasible method that allows high sample throughput for the determination of circulating and monitored SARS-CoV-2 VOCs. With focus on the novel variants such as SARS-CoV-2 omicron BA.4 and BA.5 similar approaches could be used for a rapid initial screening, while, however, due to the increasing number of single nucleotide polymorphisms that determine the variants of concern in depth screening becomes more cost efficient by next generation sequencing

Analysis of external quality assessment samples revealed crucial performance differences between commercial RT-PCR assays for SARS-CoV-2 detection when taking extraction methods and real-time-PCR instruments into account

In limelight of the ongoing pandemic SARS-CoV-2 testing is critical for the diagnosis of infected patients, contacttracing and mitigating the transmission. Diagnostic laboratories are expected to provide appropriate testing with maximum accuracy. Real-time reverse transcriptase PCR (RT-PCR) is the diagnostic standard. However, only a handful of studies have reviewed their performance in clinical settings. The aim of this study was to compare the performance of the overall analytical matrix including the extraction kit (BD MAX, Promega, Qiagen), the PCR instrument (Agilent Mx3005 P, BD MAX, Qiagen Rotor-Gene, Roche Cobas z 480) and the RT-PCR assay (Altona Diagnostics, CerTest Biotec, R-Biopharm AG) using predefined samples from proficiency testing organizers. The greatest difference of the cycle threshold values between the matrices was nine cycles. One borderline sample could not be detected by three out of twelve analytical matrices and yielded a false negative result. We therefore conclude that diagnostic laboratories should take the complete analytical matrix in addition to the performance values published by the manufacturer for a respective RT-PCR kit into account. With limited resources laboratories have to validate a wide range of kits to determine appropriate analytical matrices for detecting SARS-CoV2 reliably. The interpretation of clinical results has to be adapted accordingly

Epidemiology of driver mutations in lung cancer in a German tertiary hospital in patients with testing indication

Background: KRAS, BRAF, EGFR and ALK-mutation testing is a prerequisite for non-small-cell lung cancer treatments, but there remains limited epidemiological information about such mutations in German cohorts. Materials & methods: Between February 2010 and June 2015, a total of 1080 tumor samples from 1019 non-small-cell lung cancer patients were analyzed for KRAS, BRAF, EGFR and ALK-mutations by Therascreen-pyrosequencing and FISH. Results: Mutation patterns differed dependent on the histological subtype and sex. Mainly, adenocarcinomas were mutated and formed the major histological group. Double mutations were observed also not explicitly screened for. In our German cohort, female adenocarcinoma patients had statistically significantly higher rates EGFR mutations than male patients. Discussion: The different mutation patterns dependent on histological phenotypes warrant further epidemiological studies while suggesting different mechanisms of cancerogenesis

Epidemiology of EML4-ALK translocations in a small, German non-small-cell lung cancer patient cohort

Background: Fusions and translocations of the ALK gene are an important genetic marker in different types of cancer and are therapy relevant, especially in lung cancer, as they predict the patient's response to crizotinib therapy. Thereby, EML4-ALK is assumed to be the most frequent fusion of ALK in lung cancer, although ALK is known to be able to fuse with approximately 20 different genes. Patients & methods: Formalin-fixed paraffin-embedded lung tissue from non-small-cell lung cancer patients previously tested positive for EML4-ALK were reanalyzed with three different FISH probe systems that are commercially available. Results: We describe a short series of patients with ALK translocations in which a surprisingly high percentage (66%) of non-EML4-ALK fusions were identified in non-small-cell lung cancer despite an estimated low percentage (similar to 20%) of such translocations. Conclusion: Depending on the diagnostic method used, those patients could receive a false-negative diagnosis and would miss the chance to benefit from novel crizotinib therapy

Direct comparison of Altona-SARS-CoV-2 dual target RT-qPCR Assay with commercial LAMP Assay using throat washes in health care staff testing

Background: Rapid molecular diagnostics by PCR has a crucial role in handling the global SARS-CoV-2 pandemic. As diagnoses are time-sensitive and global supply chains are susceptible to various factors alternative detection methods would be an important backup. Objectives: During the study the performance of a commercially available isothermal LAMP method for SARSCoV-2 detection was compared to a IVD RT-PCR Assays using throat wash specimens that were routinely taken in our hospital setting. Study design: Throat wash specimens of hospital staff (n = 174) previously tested positive for SARS-CoV-2 by the Altona Diagnostics RealStar SARS-CoV-2 RT-PCR (Altona Diagnostics, Hamburg, Germany) was tested for SARS-CoV-2 also by the SARS-CoV-2 Rapid Colorimetric LAMP Assay (NEB Germany GmbH, Frankfurt a.M., Germany). Results: The sensitivity of the colorimetric LAMP Assay compared to RT-qPCR was 78.74%, and the specificity was determined to 88.24% with a positive predictive value of 0.986 and a negative predicitve value of 0.882. The positive and negative likelihood ratio for LAMP was 6.693 and 0.241, respectively, while the diagnostic odds ratio was 27.77. Conclusions: In times of limited PCR test ressources and in settings with limited PCR capacities, the colorimetric LAMP Assay could serve as an alternative, if a calculable loss of sensitivity is acceptable from the Public Health perspective in certain settings

Detailed overview on the mutations detected by and the sensitivity of the GeneReader NGS sequencing platform

This article presents additional next generation data from our preclinical validation study. In total 121 samples (clinical specimen and interlaboratory test samples) were tested successfully with next generation sequencing. 38 different mutations in six different genes were detected. Next to the detection of different mutations, the reproducibility of the NGS test was analyzed. Three samples were analyzed five times and the results were compared. Several mutations classified as non-pathogenic so far, have been detected repeatedly. (C) 2018 The Authors. Published by Elsevier Inc