MOLECULAR DIAGNOSTICS :: INFECTIOUS DISEASE Automated workflow ~ 20 hrs
Extracted Total RNA
cDNA synthesis
capture PCR Target
Amplicon pooling & Cleanup
mission, it was important for public health officials to understand the distribution in the community. NPHL can track the rise and introduction of new variants such as Omicron through information gained from viral sequencing to implement ap- propriate public health countermeasures for an impacted area or population. In the Fall of 2020, a novel mutation
was detected and clustered to a group of elderly patients who lived in a nursing home. The patients had no symptoms, however, had high viral levels detected suggesting there may be an increase in transmissibility but decrease in virulence. NPHL researchers discovered that a spe- cific mutation in the spike protein was impacting protein structure, conferring decreased virulence. Discoveries like this allow the scientific community to develop a more in-depth understanding of muta- tions that may have an impact on viral infectivity, pathogenicity, and transmis- sion. Thus, NPHL continues to request and sequence samples across the state to actively monitor and track SARS-CoV-2 as part of pandemic preparedness. With the onset of the SARS-CoV-2 pandemic, Clear Labs quickly added clinical sequencing to expand beyond its core business of food safety testing to help with the fight against the COVID-19 pandemic. It used its automated platform as the framework for development of two NGS assays, a SARS-CoV-2 diagnostic/ strain surveillance assay, and a SARS- CoV-2 whole genome sequencing (WGS) assay. Currently, more than 50% of state public health laboratories (PHLs) use the Clear Dx platform for WGS. The assays have significantly increased the PHLs SARS-CoV-2 sequencing efforts and have aided in the identification and surveil- lance of existing and emerging strains, such as the Omicron sub lineage, BA.2. Both assays are fully automated and use integrated bioinformatic pipelines to gen- erate high quality consensus genomes. The Clear Dx platform is custom-built with a PCR instrument and sequencers housed within a robot that automates pipetting, minimizing the requirement for manual pipetting by staff. Once sequences are generated, the data is analyzed through a bioinformatics pipe- line that produces raw sequencing data
Sample-level BC ligation
Sample pooling & Cleanup
(FASTQ) and high quality assemblies (FASTA) that can be exported for further analysis or to public databases such as the Global Initiative for Sharing Avian Influenza Data (GISAID) and National Center for Biotechnology Information (NCBI), allowing for data sharing with the national and international scientific community in near real time. There are numerous benefits associ- ated with adopting an automated system: 1. Minimal personnel time required for sequencing set up and analysis, freeing up staff to perform other required tasks in the laboratory 2. Ease in set up and training so that laboratory staff with varying expertise and training have the ability to run the instrument, resulting in minimal disrup- tion in laboratory operations and testing 3. Fast turn-around time (<20 hours) from starting the instrument to obtaining a result, resulting in a faster response to outbreak investigations, variant tracking, and surveillance 4. Increasing the testing volume to meet the demands of the pandemic 5. Reduction in user errors to ensure accurate and consistent results The clinical community has embraced automation to enhance laboratory capac- ity and workflow efficiencies. Automation of sequencing will allow for all labora- tories to contribute to generating data with regards to pathogens that are seen regularly in the clinical laboratory, such as multi-drug resistant organisms, organ- isms associated with Hospital Acquired Infections (HAI), and many more. Utiliza- tion of NGS on a routine basis will enable collection of data that can help to expand development of more precise clinical di- agnostics, new antibiotic formulations, vaccines, etc. to tackle these organisms that continue to affect the health of our communities. As we are envisioning a future with increased availability of automated solutions, the complement to automat- ing sequencing will be to automate technology-enhanced analytics that have the capability to analyze data in a standardized and accessible fashion. Automation of genomic data analysis can remove the burden of manual analysis to enable faster identification of outbreak
Seq. adapter ligation
Sequencing Seq. data analysis
clusters; clearer understanding of the course of spread of a pathogen through a hospital or community; predicting the evolution of the pathogen as a form of risk modeling for public health policy de- cision-making; and many more benefits. In addition, cloud-connected systems can enable near real-time data extraction and insights, minimizing the lag time that cur- rently exists for data consolidation across states. To have an effective national and international pathogen surveillance system, it is important to recognize and overcome the persisting challenges with multi-jurisdictional data stewardship, data privacy protection, and data sharing.. Academic, clinical, and public health laboratories have been integral in the pan- demic response through their sequencing efforts. Whole genome sequencing has been an important tool for obtaining insight about SARS-CoV-2 that assisted public health officials design targeted strategies to help minimize the spread and curb the reach of the virus across all communities. The utilization of NGS for SARS-CoV-2 surveillance has been a great asset for the public health and scientific communities. Prior to COVID- 19, sequencing was conducted for special projects, and in a limited capacity, due to cost and the extensive time requirement for set up. The pandemic has introduced the necessity for performing sequencing more frequently and for more pathogens. Democratizing genomics through lower- ing costs, automation, integrated bioin- formatics, and a curated public database, paves the way for the establishment of a critical genomic infrastructure that can accelerate rapid data exchange and deliver near real-time pathogen insights to better prepare for future pandemic that may arise.
Ramin Khaksar, Chief Scientific Officer of Clear Labs, a supplier of fully automated, next-generation sequencing (NGS) for genomic surveillance and diagnostics
Baha Abdalhamid, MD, PhD, D (ABMM), Assistant Professor, Department of Pathology and Microbiology at the University of Nebraska Medical Center
MLO-ONLINE.COM MAY 2022 17
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