16S rRNA Amplicon Sequencing of Nasal Microbiome

Nasal swabs were placed in 500 μL de PBS, vortex for 30 s and stored at −80 °C until used for DNA extraction. DNA was extracted from 200 μL of the initial 500 μL PBS where the swabs were resuspended and eluted in 100 μL of PBS using the Nucleospin Blood (Macherey Nagel, Bethlehem, PA, USA) kit. Quantity and quality assessment of the DNA was performed using BioDrop DUO (BioDrop Ltd., Cambridge, UK). Samples were submitted for 16S rRNA gene amplicon sequencing using the Illumina paired-end 2 × 250 bp kit (MS-102-2003 MiSeq^® Reagent Kit v2, 500 cycle) following the manufacturer’s instructions. The library preparation for sequencing was performed within 24 h after the DNA extraction at Servei de Genòmica, Universitat Autònoma de Barcelona. The region amplified was V3–V4 that covers two hypervariable regions of the conserved gene, and it was sequenced according to the Illumina protocol [27 (link)] as previously described [6 (link)]. The entire sequence dataset is available at the NCBI database, SRA accession number PRJNA717778.