This study evaluated automated and manual commercial DNA extraction methods for their ability to recover DNA from Brucella species in phosphate-buffered saline (PBS) suspension and from spiked swab specimens. Six extraction methods, representing several of the methodologies which are commercially available for DNA extraction, as well as representing various throughput capacities, were evaluated: the MagNA Pure Compact and the MagNA Pure LC instruments, the IT 1-2-3 DNA sample purification kit, the MasterPure Complete DNA and RNA purification kit, the QIAamp DNA blood mini kit, and the UltraClean microbial DNA isolation kit. These six extraction methods were performed upon three pathogenic Brucella species: B. abortus, B. melitensis, and B. suis. Viability testing of the DNA extracts indicated that all six extraction methods were efficient at inactivating virulent Brucella spp. Real-time PCR analysis using Brucella genus-and species-specific TaqMan assays revealed that use of the MasterPure kit resulted in superior levels of detection from bacterial suspensions, while the MasterPure kit and MagNA Pure Compact performed equally well for extraction of spiked swab samples. This study demonstrated that DNA extraction methodologies differ in their ability to recover Brucella DNA from PBS bacterial suspensions and from swab specimens and, thus, that the extraction method used for a given type of sample matrix can influence the sensitivity of real-time PCR assays for Brucella.

Quality standards as part of an effective quality management system (QMS) are the cornerstone for generating high-quality test results. Next-generation sequencing (NGS) has the potential to improve both clinical diagnostics and public health surveillance efforts in multiple areas, including infectious diseases. However, the laboratories adopting NGS methods face significant challenges due to the complex and modular process design. This document summarizes the first phase of quality system guidance developed by the Centers for Disease Control and Prevention (CDC) NGS Quality Workgroup. The quality system essentials of personnel, equipment, and process management (quality control and validation) were prioritized based on a risk assessment using information gathered from participating CDC laboratories. Here, we present a prioritized QMS framework, including procedures and documentation tools, to assist laboratory implementation and maintenance of quality practices for NGS workflows.

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