Trustworthy Identification of Resistance Biomarkers of Bacillus weihenstephanensis: Workflow of the Quality Assurance Procedure

Omics databases have exploded, opening the avenue to take strain diversity or physiological variability into consideration in microbiological risk assessment (MRA). However, one obstacle to the integration of omics data in MRA is the production of quantitative data that may be used to build mathematical models. Gene expression is recognized as relevant biomarker to describe bacterial behavior and reverse transcription quantitative PCR (RT-qPCR) is considered as the gold standard for accurate, sensitive, and fast measurement of gene expression. However, numerous critical points may arise throughout the entire workflow of RT-qPCR data acquisition influencing accuracy of the results and reliability of the conclusions. Although recommendations about the minimum information that should be found in publications about quantitative real-time PCR experiments, heterogeneity in the reporting of RT-qPCR quality controls in publications remains. Herein, the step-by step RT-qPCR quality controls established for the selection of Bacillus weihenstephanensis resistance biomarkers were described. Throughout this example, appropriate quality procedures and quality controls that shall be set up and carefully assessed to ensure reliable interpretations in RT-qPCR were depicted.