Comparison of self-collected oral swabs for supragingival microbiome characterization using 16S rRNA gene amplicon...

The oral microbiota is a microbially dense and complex environment with links to local and systemic health outcomes such as periodontitis and cardiovascular disease respectively. Multiple factors may influence microbiome composition, where pre-analytical influence, such as the sample collection method should be minimized. Our aim was to compare three different commercial oral swabs to assess ease-of-use, bacterial DNA yield, and microbiome composition. The Isohelix buccal swab with Dri-Capsules (I), the Omnigene oral swab (O), and the Zymo Research DNA/RNA Shield SafeCollect swab (Z) were evaluated in parallel. Fifteen anonymized volunteers collected supragingival samples. Questionnaires regarding ease-of-use and comfort were completed. Swab samples (n = 45) and negative controls (n = 8) were subject to DNA extraction with the Maxwell RSC Buccal Swab DNA kit and 16S rRNA gene qPCR was performed to measure bacterial DNA yield (limit of detection: 10^-6 ng/μL). DNA was subject to 16S rRNA gene amplicon and ITS sequencing using the QIAseq 16S/ITS Screening Panel on the Illumina MiSeq. The 16S V3-V4 regions were analyzed for genus-level alpha- and beta-diversity and were performed using QIIME2 v.2024.10. Participants preferred O and Z swabs for ease-of-use and comfort, while the I swab had generally negative assessments. The O swab resulted in the highest median bacterial DNA concentration (1.0 ng/μL) followed by the the Z (0.1 ng/μL) and I (0.02 ng/μL) swabs. There was some taxonomic abundance variation between the I and Z swab but these observations were not supported by genus-level differential abundance analysis. There were also no differences in diversity measures at the genus level between swabs. Considering pre-analytical quality is key for oral microbiome studies.