Assuring the Quality of Next-Generation Sequencing in Clinical Microbiology and Public Health Laboratories

Abstract

Clinical microbiology and public health laboratories are beginning to utilize next-generation sequencing (NGS) for a range of applications. This technology has the potential to transform the field by providing approaches that will complement, or even replace, many conventional laboratory tests. While the benefits of NGS are significant, the complexities of these assays require an evolving set of standards to ensure testing quality. Regulatory and accreditation requirements, professional guidelines, and best practices that help ensure the quality of NGS-based tests are emerging. This review highlights currently available standards and guidelines for the implementation of NGS in the clinical and public health laboratory setting, and it includes considerations for NGS test validation, quality control procedures, proficiency testing, and reference materials. N ext-generation sequencing (NGS) is transforming the landscape of clinical microbiology and public health laboratories. The applications of NGS are wide-ranging and include wholegenome sequencing, microbiome analysis/metagenomics, transcriptome profiling, infectious disease diagnosis, pathogen discovery, and public health surveillance. For example, NGS has recently been used to better understand hospital outbreaks and inform infection control practices (1), and it can be used in the clinical microbiology laboratory to identify unknown organisms, predict antimicrobial resistance, assess virulence gene content, and inform molecular epidemiology efforts (2). Metagenomic "unbiased" NGS applications, coupled with recently developed bioinformatics solutions (3-5) that enable the identification of all pathogens directly from a clinical sample based on sequence homology, have the potential to complement or even replace current standard clinical laboratory tests. For example, the use of metagenomics combined with a rapid bioinformatics pipeline recently facilitated a clinically actionable diagnosis of neuroleptospirosis when conventional testing was initially unable to identify the causative organism (6). A number of agencies are working to bring NGS into the public health laboratory setting. For example, through the U.S. Centers for Disease Control and Prevention (CDC) Advanced Molecular Detection (AMD) Initiative, national, state, and local partners are beginning to incorporate NGSbased methods into disease surveillance systems. AMD initiatives include broad applications of NGS to address public health problems, including vaccine improvement, identification of emerging threats, and tracking diseases and outbreaks (http://www.cdc.gov /amd/). The CDC, Food and Drug Administration (FDA), National Institutes of Health (NIH), National Center for Biotechnology Information (NCBI), National Library of Medicine, and the U.S. Department of Agriculture/Food Safety and Inspection Service (USDA/FSIS) have established an Interagency Collaboration on Genomics and Food Safety (Gen-FS), with the goal of fostering timely access to genomic data for foodborne pathogen surveillance and ou...

Validation and Implementation of Clinical Laboratory Improvements Act-Compliant Whole-Genome Sequencing in the Public Health Microbiology Laboratory

“…The importance of reference materials for the validation and QC of wet- and dry-lab WGS processes was noted previously (28, 31, 33). Unlike human genomics (43), there is no well-established resource of reference materials for the validation of WGS in public health microbiology laboratories.…”

Clinical metagenomics

“…In laboratory test development, well-characterized, standardized and validated reference materials or databases that enable measurement of performance characteristics of an assay, including sensitivity, specificity and accuracy. The details for the specific steps vary by laboratory and are described extensively elsewhere [33][34][35][36][37] .…”

INNUENDO: A cross‐sectoral platform for the integration of genomics in the surveillance of food‐borne pathogens

“…Using these procedures, the operator ensures that no sequence data move forward in the process without meeting the minimum quality standards. Few QC matrixes have been defined for NGS-based testing and several software applications have been developed for this scope (Gargis et al, 2016). However, these tools are either too challenging for non-bioinformaticians to use or simplified down to a one-button-click black box.…”