Abstract
Introduction Molecular sequencing techniques offer a new approach to culture in profiling the airway microbiome. A detailed comparison has not been reported and we aimed to perform this.
Methods 138 spontaneously expectorated sputum samples from 71 COPD patients underwent quantification of all cultured isolates and 16S rRNA gene sequencing using Illumina Miseq.
Results Culture identified a total of 299 isolates (median 2/sample) comprising 33 species from 15 different genera. Conversely, sequencing produced 9113146 sequence reads (median 61088/sample) comprising 2259 distinct operational taxonomic units (OTUs) from 114 different genera. Using both methods Streptococcus and Haemophilus species dominated the overall profile (73% and 61% respectively); Veillonella, Actinomyces, Granulicatella and Prevotella comprised 17% of sequencing reads but were not detected by culture. Where identified on culture (n=117), the dominant genus was the same as sequencing in 82% of samples (Cohen's kappa=0.58, p<0.001). The number of cultured isolates per sample correlated with bacterial diversity (observed OTUs [rho=0.30, p <0.001] and Shannon index [rho=0.22, p=0.010], Figure).
Conclusions Both techniques produced comparable population profiles for aerobic organisms and reflect within-sample dominance and species diversity in COPD. 16S rRNA gene sequencing offers superior per-sample resolution and detects many genera missed by standard culture.
- Copyright ©the authors 2016
