Targeting the 16S rRNA Gene by Reverse Complement PCR Next-Generation Sequencing: Specific and Sensitive Detection and Identification of Microbes Directly in Clinical Samples
Simone J.C.F.M. Moorlag, Jordy P. M. Coolen, Bart Van den Bosch, Elisabeth Hui-Mei Jin, Jochem B. Buil, Heiman Wertheim, Willem J. G. Melchers
Abstract
The identification of the causative infectious pathogen in patients suspected of having a bacterial infection is essential for diagnosis and the start of appropriate treatment. Over the past 2 decades, molecular diagnostics have improved the ability to detect and identify bacteria. However, novel techniques that can accurately detect and identify bacteria in clinical samples and that can be implemented in clinical diagnostics are needed. Here, we demonstrate the clinical utility of bacterial identification in clinical samples by a novel method called 16S RC-PCR. Using 16S RC-PCR, we reveal a significant increase in the number of clinical samples in which a potentially clinically relevant pathogen is identified compared to the commonly used 16S Sanger method. Moreover, RC-PCR allows automation and is well suited for implementation in a diagnostic laboratory. In conclusion, the implementation of this method as a diagnostic tool is expected to result in an increased number of diagnosed bacterial infections, and in combination with adequate treatment, this could improve clinical outcomes for patients.