Aims and Objectives: Sampling nasal lining fluid (NLF) via nasosorption is minimally invasive and well tolerated, but the feasibility of assessing the nasal microbiome using these samples is unknown. We aimed to optimise DNA extraction from NLF and determine microbial community profiles.

Methods: Microbial DNA was extracted from a pooled NLF sample in parallel using silica column (ZymoBIOMICS) and precipitation (Qiagen) kits, and a published precipitation method (Saladie et al. Front. Microbiol; 11:2020). A mock community, with enzymatic and/or mechanical lysis, was used to assess extraction bias. Extracted DNA was quantified, and fragment size assessed using Qubit and Tapestation. Amplification of the V3-4 region (341F/806R primers) and 16s rRNA sequencing was performed, and data analysed using the nf-core/ampliseq pipeline, decontam and mixOmics R packages.

Results: All methods extracted high quality DNA from the mock community. Only the precipitation methods extracted sufficient high-quality DNA from NLF and successfully amplified for 16s rRNA sequencing.  Sparse partial least squares discriminant analysis identified bias against gram positive bacteria with enzymatic lysis only in the mock community and identified extraction method as the main source of variability in microbial composition of NLF, although overall composition was similar. The NLF microbiome contained known airway commensals and opportunistic pathogens, with Corynebacterium spp. dominant in the pooled sample.

Conclusions: DNA was successfully extracted from a mock community and NLF, with mechanical lysis vital to reduce extraction bias, and 16s profiling of NLF identified common airway commensals and opportunistic pathogens