Impact of sampling and DNA extraction methods on skin microbiota assessment

Autor: Christine Mielcarek, Thomas Z. Lerch, Frédérique Changey, Anne Roynette, Yanisse Boulesnane, Julie Leloup, Anne-Marie Pensé-Lhéritier
Přispěvatelé: Institut d'écologie et des sciences de l'environnement de Paris (iEES Paris ), Institut de Recherche pour le Développement (IRD)-Sorbonne Université (SU)-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-Centre National de la Recherche Scientifique (CNRS)-Université de Paris (UP)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), École de biologie industrielle (EBI), Institut de Recherche pour le Développement (IRD)-Sorbonne Université (SU)-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-Centre National de la Recherche Scientifique (CNRS)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)
Rok vydání: 2020
Předmět:
Zdroj: Journal of Microbiological Methods
Journal of Microbiological Methods, Elsevier, 2020, 171, ⟨10.1016/j.mimet.2020.105880⟩
ISSN: 0167-7012
1872-8359
DOI: 10.1016/j.mimet.2020.105880
Popis: The skin microbiota is characterized by high intra- and inter-variability among individuals, due to a multitude of intrinsic and extrinsic parameters such as genetics, lifestyles or pollution. This variability may be heightened due to sampling method as the skin is a multilayer organ and its outermost layer consists of dead cells. In order to investigate this biological variability in a reproducible way, we studied how sampling procedure and DNA extraction methods influence the qualitative and quantitative gathering of bacterial communities. Here, we tested a new sampling procedure that consists in exerting a slight abrasion (scrubbing) on the skin prior to swabbing and extracting DNA in order to remove squames and access deeper ecological niches. Scrubbed and non-scrubbed samples were collected from a panel of six volunteers, and four DNA extraction methods were performed on the samples. The abundance, diversity and structure of the bacterial communities were measured using qPCR technics and 16S rDNA gene-metabarcoding. Bacterial community abundance was significantly impacted by the DNA extraction method (in favor of a method designed for tissues) but not by sampling procedure, as scrubbing does not increase bacterial biomass gathered. Bacterial α- and β-diversities were both affected by DNA extraction methods and sampling procedure. Scrubbing reveals different microbial composition by gathering bacteria living in deeper skin layer, resulting in a lower intra-personal variability. The taxonomic analysis showed that more bacteria belonging to anaerobes groups were present in scrubbed samples. We conclude that DNA extraction methods designed for tissue are not necessarily associated with high qualitative efficiency and slight scrubbing prior DNA extraction reduces intrapersonal variability and give access to a new microbial diversity.
Databáze: OpenAIRE
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