Role of clinical laboratories in response to the COVID-19 pandemic

The outbreak of COVID-19 (caused by SARS-CoV-2) is now a pandemic that has caused a global socio-economic disorder. Since its identification in the region of Wuhan, China, 2,629,579 confirmed cases with over 182,900 COVID-19-related deaths have been reported globally [1]. In response to the outbreak, several state authorities and commercial companies have developed diagnostic assays to test individuals for the SARS-CoV-2 infection. Currently, over 40 diagnostic assays have received Emergency Use Authorization (EUA) from the Federal US FDA for COVID-19 testing. In the US, clinical laboratories are required to perform ‘bridging studies’ on FDA-approved SARS-CoV-2 diagnostic assays to implement testing under the EUA regulation. Absence of adequate testing due to various factors, most significant of which being supply chain issues, is most likely contributing to community spread. In midst of numerous challenges, clinical laboratories have a critical role to play in response to the current COVID-19 pandemic. In addition to ensuring the testing requirements of the population in the present hour, laboratories have an unprecedented responsibility to prepare for the aftermath of the pandemic. Although, the reverse transcription-polymerase chain reaction (RT-PCR)-based assays for the detection of SARS-CoV-2 nucleic acid regions might be the most practical approach at present, qualitative assays are far from providing insights into the evolution of the virus and the varied immune response in different populations. Herein, we discuss the three main categories of diagnostic assays available for the identification of SARS-CoV-2 infection, their utility and a way-around the challenges associated with each assay. Further, laboratory management issues are highlighted that might be considered by laboratories for optimal functioning

Next-Generation Sequencing (NGS) in COVID-19: A Tool for SARS-CoV-2 Diagnosis, Monitoring New Strains and Phylodynamic Modeling in Molecular Epidemiology

This review discusses the current testing methodologies for COVID-19 diagnosis and explores next-generation sequencing (NGS) technology for the detection of SARS-CoV-2 and monitoring phylogenetic evolution in the current COVID-19 pandemic. The review addresses the development, fundamentals, assay quality control and bioinformatics processing of the NGS data. This article provides a comprehensive review of the obstacles and opportunities facing the application of NGS technologies for the diagnosis, surveillance, and study of SARS-CoV-2 and other infectious diseases. Further, we have contemplated the opportunities and challenges inherent in the adoption of NGS technology as a diagnostic test with real-world examples of its utility in the fight against COVID-19

COVID-19 screening in a healthcare or community setting: complexity of saliva as a specimen for PCR-based testing

Testing for SARS-CoV-2 has highly significant clinical and epidemiological implications in the current COVID-19 pandemic. Reverse transcription PCR (RT-PCR)-based assays are the predicate method for detecting the virus, primarily from nasopharyngeal swab (NPS) samples. However, collection of NPS samples poses certain challenges that include exposure risk to healthcare workers, supply chain constraints pertaining to swabs and personal protective equipment and self-collection being difficult and less sensitive. Furthermore, several reports have highlighted the relatively poor sensitivity of NPS samples in early infection and longitudinal testing [1–3]. Amid these challenges, several other sample types are under investigation for COVID-19 testing, of which saliva samples are of significant interest owing to their ease of collection and alleviation of some of the challenges with NPS sampling. In the US, the FDA has approved saliva-based collection methods for laboratories submitting for emergency use authorization

Clinical performance and utility of a comprehensive next-generation sequencing DNA panel for the simultaneous analysis of variants, TMB and MSI for myeloid neoplasms

The extensively employed limited-gene coverage NGS panels lead to clinically inadequate molecular profiling of myeloid neoplasms. The aim of the present investigation was to assess performance and clinical utility of a comprehensive DNA panel for myeloid neoplasms. Sixty-one previously well characterized samples were sequenced using TSO500 library preparation kit on NextSeq550 platform. Variants with a VAF ≥ 5% and a total read depth of \u3e50X were filtered for analysis. The following results were recorded-for clinical samples: clinical sensitivity (97%), specificity (100%), precision (100%) and accuracy (99%) whereas reference control results were 100% for analytical sensitivity, specificity, precision and accuracy, with high intra- and inter-run reproducibility. The panel identified 880 variants across 292 genes, of which, 749 variants were in genes not covered in the 54 gene panel. The investigation revealed 14 variants in ten genes, and at least one was present in 96.2% patient samples that were pathogenic/ likely pathogenic in myeloid neoplasms. Also, 15 variants in five genes were found to be pathogenic/ likely pathogenic in other tumor types. Further, the TMB and MSI scores ranged from 0–7 and 0–9, respectively. The high analytical performance and clinical utility of this comprehensive NGS panel makes it practical and clinically relevant for adoption in clinical laboratories for routine molecular profiling of myeloid neoplasms

Clinical Utility of Optical Genome Mapping and 523-Gene Next Generation Sequencing Panel for Comprehensive Evaluation of Myeloid Cancers

The standard-of-care (SOC) for genomic testing of myeloid cancers primarily relies on karyotyping/fluorescent in situ hybridization (FISH) (cytogenetic analysis) and targeted gene panels (usually ≤54 genes) that harbor hotspot pathogenic variants (molecular genetic analysis). Despite this combinatorial approach, ~50% of myeloid cancer genomes remain cytogenetically normal, and the limited sequencing variant profiles obtained from targeted panels are unable to resolve the molecular etiology of many myeloid tumors. In this study, we evaluated the performance and clinical utility of combinatorial use of optical genome mapping (OGM) and a 523-gene next-generation sequencing (NGS) panel for comprehensive genomic profiling of 30 myeloid tumors and compared it to SOC cytogenetic methods (karyotyping and FISH) and a 54-gene NGS panel. OGM and the 523-gene NGS panel had an analytical concordance of 100% with karyotyping, FISH, and the 54-gene panel, respectively. Importantly, the IPSS-R cytogenetic risk group changed from very good/good to very poor in 22% of MDS (2/9) cases based on comprehensive profiling (karyotyping, FISH, and 54-gene panel vs. OGM and 523-gene panel), while additionally identifying six compound heterozygous events of potential clinical relevance in six cases (6/30, 20%). This cost-effective approach of using OGM and a 523-gene NGS panel for comprehensive genomic profiling of myeloid cancers demonstrated increased yield of actionable targets that can potentially result in improved clinical outcomes

Optical Genome Mapping: Integrating Structural Variations for Precise Homologous Recombination Deficiency Score Calculation

Homologous recombination deficiency (HRD) is characterized by the inability of a cell to repair the double-stranded breaks using the homologous recombination repair (HRR) pathway. The deficiency of the HRR pathway results in defective DNA repair, leading to genomic instability and tumorigenesis. The presence of HRD has been found to make tumors sensitive to ICL-inducing platinum-based therapies and poly(adenosine diphosphate [ADP]–ribose) polymerase (PARP) inhibitors (PARPi). However, there are no standardized methods to measure and report HRD phenotypes. Herein, we compare optical genome mapping (OGM), chromosomal microarray (CMA), and a 523-gene NGS panel for HRD score calculations. This retrospective study included the analysis of 196 samples, of which 10 were gliomas, 176 were hematological malignancy samples, and 10 were controls. The 10 gliomas were evaluated with both CMA and OGM, and 30 hematological malignancy samples were evaluated with both the NGS panel and OGM. To verify the scores in a larger cohort, 135 cases were evaluated with the NGS panel and 71 cases with OGM. The HRD scores were calculated using a combination of three HRD signatures that included loss of heterozygosity (LOH), telomeric allelic imbalance (TAI), and large-scale transitions (LST). In the ten glioma cases analyzed with OGM and CMA using the same DNA (to remove any tumor percentage bias), the HRD scores (mean ± SEM) were 13.2 (±4.2) with OGM compared to 3.7 (±1.4) with CMA. In the 30 hematological malignancy cases analyzed with OGM and the 523-gene NGS panel, the HRD scores were 7.6 (±2.2) with OGM compared to 2.6 (±0.8) with the 523-gene NGS panel. OGM detected 70.8% and 66.8% of additional variants that are considered HRD signatures in gliomas and hematological malignancies, respectively. The higher sensitivity of OGM to capture HRD signature variants might enable a more accurate and precise correlation with response to PARPi and platinum-based drugs. This study reveals HRD signatures that are cryptic to current standard of care (SOC) methods used for assessing the HRD phenotype and presents OGM as an attractive alternative with higher resolution and sensitivity to accurately assess the HRD phenotype