Comparison of two commercial DNA extraction kits for the analysis of nasopharyngeal bacterial communities

Authors

Marcos Pérez-Losada, George Washington UniversityFollow
Keith Crandall, George Washington UniversityFollow
Robert J. Freishtat, George Washington UniversityFollow

Document Type

Journal Article

Publication Date

4-2016

Journal

AIMS Microbiology

Volume

2

Issue

2

Inclusive Pages

108-119

DOI

10.3934/microbiol.2016.2.108

Abstract

Characterization of microbial communities via next-generation sequencing (NGS) requires an extraction ofmicrobial DNA. Methodological differences in DNA extraction protocols may bias results and complicate inter-study comparisons. Here we compare the effect of two commonly used commercial kits (Norgen and Qiagen)for the extraction of total DNA on estimatingnasopharyngeal microbiome diversity. The nasopharynxis a reservoir for pathogens associated with respiratory illnesses and a key player in understandingairway microbial dynamics.
Total DNA from nasal washes corresponding to 30 asthmatic children was extracted using theQiagenQIAamp DNA and NorgenRNA/DNA Purification kits and analyzed via IlluminaMiSeq16S rRNA V4 ampliconsequencing. The Norgen samples included more sequence reads and OTUs per sample than the Qiagen samples, but OTU counts per sample varied proportionallybetween groups (r = 0.732).Microbial profiles varied slightly between sample pairs, but alpha- and beta-diversity indices (PCoAand clustering) showed highsimilarity between Norgen and Qiagenmicrobiomes. Moreover, no significant differences in community structure (PERMANOVA and adonis tests) and taxa proportions (Kruskal-Wallis test) were observed betweenkits. Finally, aProcrustes analysis also showed low dissimilarity (M² = 0.173; P< 0.001) between the PCoAs of the two DNA extraction kits.

Contrary to what has been observed in previous studies comparing DNA extraction methods, our 16S NGS analysis of nasopharyngeal washes did not reveal significant differences in community composition or structure between kits. Our findingssuggest congruence between column-based chromatography kits and supportthe comparison of microbiomeprofilesacross nasopharyngeal metataxonomic studies.

Comments

Reproduced with permission of AIMS Press. AIMS Microbiology.

Creative Commons License

This work is licensed under a Creative Commons Attribution 4.0 License.

APA Citation

Pérez-Losada, M., Crandall, K., & Freishtat, R. J. (2016). Comparison of two commercial DNA extraction kits for the analysis of nasopharyngeal bacterial communities. AIMS Microbiology, 2 (2). http://dx.doi.org/10.3934/microbiol.2016.2.108

Peer Reviewed

1

Open Access

1