Confidence in total or viable microbial cell counting methods is lacking, due in part to a dearth of relevant reference materials. NIST is developing a living yeast cell reference material (RM 8230) characterized for total cell count and colony forming units. This material serves as a known input for assessing and comparing cell counting and nucleic acid detection approaches, and also supports minimal risk field training in biothreat detection.
Quantification of total and viable microbial cells is critical for applications ranging from assessment of DNA extraction efficiency in support of microbial metagenomics to characterization of antimicrobial efficacy. In addition, safe biological materials are needed to substitute for true biothreat agents during responder training on the field response to suspicious biological materials.
The goal of this project is to develop the first living, whole cell NIST RM characterized for total cell count to challenge microbial count and detection technologies. The material consists of a strain of Baker’s yeast, Saccharomyces cerevisiae NE095, engineered to contain a stable genomic insertion. The insert represents a ≈ 500 bp sequence derived from the genome of extremophile Methanocaldococcus jannaschii (the sequence in taken from NIST SRM 2374) and provides for specific detection of the yeast via nucleic acid based detection technologies. RM 8230 is expected to have broad application, not only as a training material for biological detection in the field, but also as a homogenous, microbial cell material to challenge and compare cell counting approaches.
The candidate RM is in a lyophilized format and has passed extensive homogeneity and stability testing at NIST. The production batch is undergoing characterization and is expected to be released in 2020.
This work was supported by Interagency Agreements between the Department of Homeland Security Science and Technology Directorate (DHS S&T) and NIST.